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Full text of "Malacologia"



HARVARD UNIVERSITY 

Library of the 

Museum of 

Comparative Zoology 



/OL. 34, NO. 1-2 1992 



MALACOLOGIA 



nternational Journal of Malacology 
Revista Internacional de Malacologia 
Journal International de Malacologie 
Международный Журнал Малакологии 
Internationale Malakologische Zeitschrift 



Publication dates 

Vol. 28, No. 1-2 19 January 1988 

Vol. 29, No. 1 28 June 1988 

Vol. 29, No. 2 16 Dec. 1988 
Vol. 30, No. 1-2 1 Aug. 1989 

Vol. 31, No. 1 29 Dec. 1989 

Vol. 31, No. 2 28 May 1990 
Vol. 32, No. 2 7 June 1991 
Vol. 33, No. 1-2 6 Sep. 1991 



'OL 34, NO. 1-2 1992 

MCZ 
LIBRARY 

SEP 1 5 1992 

HARVARD 
UNIVERSITY 

MALACOLOGIA 



iternational Journal of Malacology 
Revista Internacional de Malacologia 
Journal International de Malacologie 
Международный Журнал Малакологии 
Internationale Malakologische Zeitschrift 



MALACOLOGIA 

Editor-in-Chief: 
GEORGE M. DAVIS 



Editorial and Subscription Offices: 

Department of Malacology 

The Academy of Natural Sciences of Philadelphia 

1900 Benjamin Franklin Parkway 

Philadelphia, Pennsylvania 19103-1195, U.S.A. 



EUGENE COAN 

California Academy of Sciences 

San Francisco, CA 



Co-Editors: 



Assistant Managing Editor: 

CARYL HESTERMAN 

Associate Editors: 



CAROL JONES 
Denver, CO 



JOHN B. BURCH 
University of Michigan 
Ann Arbor 



ANNE GISMANN 

Maadi 

Egypt 



MALACOLOGIA is published by the INSTITUTE OF MALACOLOGY, the Sponsor Members 
of which (also serving as editors) are: 



KENNETH J. BOSS 

Museum of Comparative Zoology 

Cambridge, Massachusetts 

JOHN BURCH, President-Eiect 

MELBOURNE R. CARRIKER 
University of Delaware, Lewes 

GEORGE M. DAVIS 
Secretary and Treasurer 

CAROLE S. HICKMAN, Vice-President 
University of California, Berkeley 



JAMES NYBAKKEN, President 
Moss Landing Marine Laboratory 
California 

CLYDE F. E. ROPER 
Smithsonian Institution 
Washington, D.C. 

W. D. RUSSELL-HUNTER 
Syracuse University, New York 

SHI-KUEI WU 

University of Colorado Museum, Boulder 



Participating Members 

EDMUND GITTENBERGER JACKIE L VAN GOETHEM 

Secretary, UNITAS MALACOLOGICA Treasurer, UNITAS MALACOLOGICA 

Rijksmuseum van Natuurlijke Koninklijk Belgisch Instituut 

Historie voor Natuurwetenschappen 

Leiden, Netherlands Brüssel, Belgium 



J. FRANCIS ALLEN, Emérita 
Environmental Protection Agency 
Washington, D.C. 

ELMER G. BERRY, 
Germantown, Maryland 



Emeritus Members 

ROBERT ROBERTSON 

The Academy of Natural Sciences 

Philadelphia, Pennsylvania 

NORMAN F. SOHL 
U.S. Geological Survey 
Reston, Virginia 



Copyright © 1992 by the Institute of Malacology 



1992 
EDITORIAL BOARD 



J. A. ALLEN 

Marine Biological Station 

Millport, United Kingdom 

R. BIELER 
Field Museum 
Chicago, U.S.A. 

E. E. BINDER 

Muséum d'Histoire Naturelle 

Genève, Switzerland 

A. J. CAIN 

University of Liverpool 
United Kingdom 

P. CALOW 

University of Sheffield 
United Kingdom 

J. G. CARTER 

University of North Carolina 

Chapel Hill, U.S.A. 

R. COWIE 
Bishop Museum 
Honolulu, HL, U.S.A. 

A. H. CLARKE, Jr. 
Portland, Texas, U.S.A. 

B. С CLARKE 
University of Nottingham 
United Kingdom 

R. DILLON 

College of Charleston 

SC, U.S.A. 

C. J. DUNCAN 
University of Liverpool 
United Kingdom 

D. J. EERNISSE 
University of Michigan 
Ann Arbor, U.S.A. 

V. FRETTER 
University of Reading 
United Kingdom 



E. GITTENBERGER 
Rijksmuseum van Natuurlijke Historie 
Leiden, Netherlands 

F. GIUSTI 
Université di Siena, Italy 

A. N. GOLIKOV 
Zoological Institute 
Leningrad, U.S.S.R. 

S. J. GOULD 
Harvard University 
Cambridge, Mass., U.S.A. 

A. V. GROSSU 
Universitatea Bucuresti 
Romania 

T HABE 
Tokai University 
Shimizu, Japan 

R. HANLON 

Marine Biomedical Institute 

Galveston, Texas, U.S.A. 

J. A. HENDRICKSON, Jr. 
Academy of Natural Sciences 
Philadelphia, PA, U.S.A. 

D. M. HILLIS 
University of Texas 
Austin, U.S.A. 

K. E. HOAGLAND 

Association of Systematics Collections 

Washington, DC, U.S.A. 

B. HUBENDICK 
Naturhistoriska Museet 
Göteborg, Sweden 

S. HUNT 
Lancashire 
United Kingdom 

R. JANSSEN 

Forschungsinstitut Senckenberg, 
Frankfurt am Main, Germany 

R. N. KILBURN 
Natal Museum 
Pietermaritzburg, South Africa 

M. A. KLAPPENBACH 

Museo Nacional de Historia Natural 

Montevideo, Uruguay 



J. KNUDSEN 

Zoologisk Institut & Museum 

Kobenhavn, Denmark 

A. J. KOHN 

University of Washington 

Seattle, U.S.A. 

A. LUCAS 

Faculté des Sciences 

Brest, France 

C. MEIER-BROOK 
Tropenmedizinisches Institut 
Tübingen, Germany 

H. К. MIENIS 

Hebrew University of Jerusalem 

Israel 

J. E. MORTON 
The University 
Auckland, New Zealand 

J. J. MURRAY, Jr. 
University of Virginia 
Charlottesville, U.S.A. 

R. NATARAJAN 
Marine Biological Station 
Porto Novo, India 

J. 0KLAND 
University of Oslo 
Norway 

T. OKUTANI 
University of Fisheries 
Tokyo, Japan 

W. L. PARAENSE 

Instituto Oswaldo Cruz, Rio de Janeiro 

Brazil 

J. J. PARODIZ 
Carnegie Museum 
Pittsburgh, U.S.A. 

J. P. POINTER 

Ecole Pratique des Hautes Etudes 

Perpignan Cedex, France 

W. F. PONDER 
Australian Museum 
Sydney 

R. D. PURCHON 

Chelsea College of Science & Technology 

London, United Kingdom 

Ql Z. Y. 

Academia Sínica 

Qingdao, People's Republic of China 



D. G. REID 

The Natural History Museum 

London, United Kingdom 

N. W. RUNHAM 

University College of North Wales 

Bangor, United Kingdom 

S. G. SEGERSTRÂLE 
Institute of Marine Research 
Helsinki, Finland 

A. STAÑCZYKOWSKA 
Siedlce, Poland 

F. STARMÜHLNER 

Zoologisches Institut der Universität 

Wien, Austria 

Y. I. STAROBOGATOV 
Zoological Institute 
Leningrad, U.S.S.R. 

W. STREIFF 
Université de Caen 
France 

J. STUARDO 
Universidad de Chile 
Valparaiso 

S. TILLIER 

Muséum National d'Histoire Naturelle 

Paris, France 

R. D. TURNER 
Harvard University 
Cambridge, Mass., U.S.A. 

J.A.M. van den BIGGELAAR 
University of Utrecht 
The Netherlands 

J. A. van EEDEN 
Potchefstroom University 
South Africa 

N. H. VERDONK 
Rijksuniversiteit 
Utrecht, Netherlands 

B. R. WILSON 

Dept. Conservation and Land Management 

Netherlands, Western Australia 

H. ZEISSLER 
Leipzig, Germany 

A. ZILCH 

Forschungsinstitut Senckenberg 

Frankfurt am Main, Germany 



MALACOLOG I A, 1992,34(1-2): 1-24 

BIOLOGY AND COMPARATIVE ANATOMY OF THREE NEW SPECIES OF 
COMMENSAL GALEOMMATIDAE, WITH A POSSIBLE CASE OF MATING 

BEHAVIOR IN BIVALVES 

Paula M. Mikkelsen 1 & Rüdiger Bieler 2 



ABSTRACT 

Three new galeommatid bivalves, Divariscintilla octotentaculata, D. luteocrinita, and D. cordi- 
formis, are described as commensals occupying the burrows of the mantis shrimp Lysiosquilla 
scabricauda from central eastern Florida. Morphological comparisons are made with all other 
known members of the genus, comprising two previously described species from the same 
burrow system (D. yoyo, D. troglodytes) and the type species, D. maoria, from New Zealand. 
Key characters defining this genus (hinge morphology, flower-like organs, "hanging foot" struc- 
ture) are discussed, especially with regard to their presence in other galeommatoidean genera. 
Intraspecific interaction resembling mating behavior is noted and discussed as one of the few 
possible examples in the Bivalvia. 

Key words: Divariscintilla, Galeommatoidea, Bivalvia, systematics, anatomy, mating behavior, 
commensalism, Stomatopoda. 



MATERIAL AND METHODS 

Lysiosquilla burrows in shallow-water sand 
flats at several locations on the central eastern 
Florida coast were sampled using a stainless 
steel bait pump ("yabby pump") and sieves of 
1-2 mm mesh. Sampling depths during ex- 
treme low water ranged from less than 0.5 m 
to supratidal, when the water level lay several 
centimeters below the level of the sand. 

Living clams were maintained in finger 
bowls of seawater at ambient laboratory con- 
ditions (22-25°C), with variable lighting. Water 
was changed every 1 -2 days, and an irregu- 
larly-supplied, unmeasured diet of mixed uni- 
cellular algae (e.g., Isochrysis, Chlorella, 
Chaetoceras) was provided. Behavioral stud- 
ies were aided by video recordings taken of 
the living animals in aquaria using a standard 
commercial 1/2-inch-format video camera 
equipped with a macro lens. 

Transfer of specimens between laboratory 
bowls was best accomplished using small 
spoons. The spoon could be applied against 
the glass from below the specimen to gently 
break the byssus threads while also cradling 
the clam. Handling these kinds of animals 
with forceps is awkward and frequently 
causes damage to fragile shells and tissue. 

Relaxation prior to dissection or preserva- 
tion was most effectively accomplished with 

1 Harbor Branch Océanographie Museum, Harbor Branch Océanographie Institution, 5600 Old Dixie Highway, Ft. Pierce, 

Florida 34946 U.S.A., and Department of Biological Sciences, Florida Institute of Technology, Melbourne, Florida 32901 

U.S.A. 

department of Zoology, Field Museum of Natural History, Roosevelt Road at Lake Shore Drive, Chicago, Illinois 60605 

U.S.A. 



INTRODUCTION 

Investigation of the organisms associated 
with the sand-burrowing mantis shrimp 
Lysiosquilla scabricauda (Lamarck, 1818) 
(Crustacea: Stomatopoda: Lysiosquillidae) in 
shallow waters of eastern Florida has re- 
vealed a community of seven molluscan spe- 
cies that appear highly dependent on this 
specialized habitat. Remarkably, all of these 
species were found to be either poorly known 
or undescribed. Accounts of the two species 
of vitrinellid gastropods — Cyclostremiscus 
beauii (Fischer, 1857) and Circulus texanus 
(Moore, 1965); Bieler & Mikkelsen, 1988— 
and two of the five species of bivalves — Di- 
variscintilla yoyo Mikkelsen & Bieler, 1989, 
and D. troglodytes Mikkelsen & Bieler, 
1989 — have appeared elsewhere. This re- 
port deals with the remaining three species of 
galeommatoidean bivalves. Although superfi- 
cially different from the two species previously 
described, and preliminarily treated as mem- 
bers of another genus (as Scintilla spp.; Mik- 
kelsen & Bieler, 1989: 192; Eckelbarger et al., 
1990), detailed study has revealed their 
proper placement in Divariscintilla, and has 
shown them to be in fact more similar to the 
New Zealand type species, D. maoria Powell, 
1932, than were the two species previously 
described from Florida stomatopod burrows. 



1 



MIKKELSEN & BIELER 



crystalline magnesium sulfate, added directly 
to the finger-bowl water in unquantified small 
amounts. Methylene-blue/basic-fuchsin and 
neutral red were used to delineate tissues and 
organs in gross dissections. 

For histological serial sections, animals 
were fixed in 5% buffered formalin (Humason, 
1 962: 1 4). Shells were decalcified using either 
dilute (approximately 0.5%) hydrochloric acid 
(complete decalcification within minutes) or a 
1% solution of ethylenediamene tetraacetic 
acid (EDTA, adjusted to pH 7.2; decalcification 
complete in 5-6 days). Specimens were em- 
bedded in paraplast, sectioned at 8 jxm and 
stained with Gomori's green trichrome (mod- 
ified from Vacca, 1 985). Staining reactions de- 
scribed in the text refer to this method. Colors 
referred to in the text are supplied for future 
use, that is, to infer homologies of the various 
glands. Photomicrographs of sections were 
taken with an Olympus BH-2 stereomicro- 
scope fitted with an Olympus OM-2 camera 
with Kodak Panatomic-X (ASA 32) film. 

For scanning electron microscopy (SEM), 
preserved specimens were passed through 
an ethanol-to-acetone series and critical-point 
dried. These and air-dried shells were coated 
with gold/palladium and examined using a 
Zeiss Novascan-30 scanning electron micro- 
scope. 

All cited anatomical measurements were 
taken from specimens of average size (see 
under descriptions). Throughout the text, 
"relaxed" refers to the condition of an animal 
in normal crawling posture and does not refer 
to any chemical treatment. 

Cited institutions are (* indicates location of 
type and other voucher material): 

AMNH — American Museum of Natural His- 
tory, New York, New York 

DMNH — Delaware Museum of Natural His- 
tory, Wilmington 

*FMNH — Field Museum of Natural History, 
Chicago, Illinois 

HBOI — Harbor Branch Océanographie In- 
stitution, Ft. Pierce, Florida 

*HBOM — Harbor Branch Océanographie 
Museum [formerly Indian River 
Coastal Zone Museum], HBOI 

*MCZ — Museum of Comparative Zoology, 
Harvard University, Cambridge, 
Massachusetts 

*SBMNH — Santa Barbara Museum of Natural 
History, California 

SMSLP — Smithsonian Marine Station at 
Link Port, Ft. Pierce, Florida 



k USNM — National Museum of Natural 
History, Smithsonian Institution, 
Washington, D. С 



TAXONOMIC DESCRIPTIONS 

Family GALEOMMATIDAE Gray, 1840 

Genus Divariscintilla Powell, 1932 

Type species: Divariscintilla maoria Powell, 
1932: 66; by original designation. Recent, 
New Zealand. 

Remarks: Redescribed by Mikkelsen & 
Bieler (1989: 193). See remarks concerning 
generic placement under Discussion. 

Divariscintilla octotentaculata n. sp. 
(Figs. 1,4, 7-13,23, 33) 

Material examined 

Holotype: 5.3 mm [shell length], FMNH 
223401. Paratypes (31): 4.2, 4.2 mm, FMNH 
223402; 3.2, 3.2, 3.2, 3.3, 3.3, 3.6, 3.8 mm, 
HBOM 064:01866; 4.2, 4.2, 3.6, 2.9 mm (pre- 
served soft-bodies + shells coated for SEM), 
HBOM 064:01867; 3.9, 4.3, 4.3 mm, HBOM 
064:01865; 3.0, 3.1, 3.2, 3.4, 3.6 mm, USNM 
859443; 3.2, 3.2, 3.2, 3.4, 4.1 mm, MCZ 
302510; 1.7, 3.4, 3.4, 3.8, 3.8 mm, SBMNH 
35167. Total material: 262 specimens: FLOR- 
IDA: Ft. Pierce Inlet: 10 March 1987, 3; 2-3 
May 1987, 73; 24 June 1987, 91 ; 03 August 
1 987, 2; 1 4 August 1 987, 1 0; 31 August 1 987, 
1; 28 December 1987, 7; 11 March 1988, 7; 
12 April 1988, 38; 16 October 1990, 2; 03 
February 1991; 13. -Sebastian Inlet: 30 De- 
cember 1987, 26. -St. Lucie Inlet: 18 February 
1982,2. 

Type locality 

Ft. Pierce Inlet, Indian River Lagoon, St. 
Lucie County, eastern Florida, 27°28.3'N, 
80°17.9'W, occupying Lysiosquilla scabri- 
cauda burrows on intertidal sand flats with 
patches of the seagrass Halodule wrightii As- 
cherson. Paratypes all from type locality. 

Diagnosis 

Animal translucent white. Mantle thin, with 
retractable, papillose folds covering anterior 
and posterior thirds of shell. Tentacles origi- 
nating at dorsal shell margin, two pairs ante- 
riorly, two pairs posteriorly. Posterior foot- 
extension relatively short. Shell roundly 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 




FIGS. 1 -3. Photographs of living animals. 1 . Divariscintilla octotentaculata. 2. D. luteocrinita. 3. D. cordi- 
formis. Scale bars = 1 .0 mm. 



MIKKELSEN & BIELER 




FIGS. 4-7. External appearance and internal shell morphology. Tentacle pairs numbered from anterior to 
posterior (for text reference only; identical numbers do not imply homology). 4. Divariscintilla octotentaculata, 
in crawling position, from left side. 5. D. luteocrinita, same as Fig 4. 6. D. cordiformis, same as Fig. 4. I.D. 
octotentaculata, internal surface of right valve, showing approximate locations of muscle insertions. Scale 
bars = 1 .0 mm. (aam, anterior adductor muscle scar; apr, anterior pedal retractor muscle scar; bag, byssus 
adhesive gland; by, location of byssus gland; c, cowl; est, club-shaped tentacles; exc, excurrent "siphon"; 
ft, foot; inc, incurrent "siphon"; ind, shell indentation; m, mantle fold; mf, point of mantle fusion; mp, mantle 
pouch; mpt, median palliai tentacle; pam, posterior adductor muscle scar; pdc, prodissoconch; pfe, posterior 
foot-extension; ppm, pedal protractor muscle scar; ppr, posterior pedal retractor muscle scar; sh, shell; vgr, 
ventral pedal groove). 



triangular, with umbo slightly anterior, smooth 
exteriorly, with weak radial ribs interiorly; 
length approximately 70% of extended mantle 
length. No "flower-like organ" on anterior sur- 
face of visceral mass. 



Description 

External Features and Mantle: Living ex- 
tended animal (Figs. 1 , 4) generally 5-7 mm in 
length, translucent white except for dark di- 
gestive gland within visceral mass. Shell 
largely external, with only anterior and poste- 
rior thirds of shell covered in life by mantle 
folds (Fig. 4, m), not meeting at lateral mid- 
line. Mantle folds fully retractable, finely pap- 
illose, with scattered larger papillae. Mantle 
thin, extending beyond shell edges; entire 
surface finely papillose, with fringe of longer 



papillae along ventral margin of shell. Two 
palliai openings: (1) anteropedal opening, 
from a point anterior to umbones to a point 
posterior to foot (Fig. 4, mf), forming exten- 
sive anterior cowl (Fig. 4, c), the edges of 
which are held together to form an effective 
incurrent "siphon" (Fig. 4, inc); and (2) pos- 
terodorsal excurrent opening (Fig. 4, exc) be- 
tween posterior tentacle pairs, either as sim- 
ple opening or on rounded protuberance 
forming distinct siphon; appearance depen- 
dent on degree of palliai expansion. Posterior 
mantle fusion forming a rounded, protruding 
pouch (Fig. 4, mp) containing the gills. Two 
pairs (one long, one short) retractable, ante- 
rior tentacles arising from near shell edge; 
dorsalmost pair longest. Two pairs (one long, 
one short) retractable tentacles in vicinity of 
excurrent siphon, also arising from near shell 
edge; posteriormost pair longest. A single, 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



short, mid-dorsal tentacle (Fig. 4, mpt), just 
posterior to umbo. An additional 1 -2 pairs of 
short tentacles ( = enlarged papillae?) in 
larger specimens (approximately 5 mm) at 
ventral shell edge at anterior and posterior 
ends of ventral papillose fringe. Each tentacle 
with papillose surface and central core of lon- 
gitudinal muscle and nerve fibers, visible as 
an inner "thread" under low magnification. In- 
ner palliai fold of non-shell areas (e.g., cowl, 
ventral mantle margin, posterior pouch) highly 
muscular. 

Preserved specimens completely (or nearly 
so) retracted into shell, however, shell usually 
gaping, with tentacles (especially anterodor- 
sal) slightly protruding beyond shell edges. 

Shell (Figs. 7-1 1 , 13): Shell generally 3-5 mm 
in length, roundly triangular, longer posteri- 
orly, equivalve, rather compressed, glossy, ir- 
idescent, transparent, smooth except for fine 
concentric growth lines and weak radial ribs 
most evident interiorly at ventral margin (Fig. 
9). Size large relative to mantle, comprising 
approximately 70% of extended mantle 
length. Valves held open at a 50-60x angle 
while crawling, capable of complete closure 
ventrally but gaping slightly anteriorly and 
posteriorly. Adductor muscle scars faint, 
subequal (Figs. 7, 9). Palliai line entire, indis- 
tinct. Periostracum colorless, most evident at 
ventral shell edge (Fig. 9, per). 

Hinge line short (Fig. 10). One small, 
rounded cardinal tooth (Fig. 10, car) in each 
valve, that of left valve sometimes slightly bi- 
fid; lateral teeth absent. Cardinal teeth abut- 
ting, not interlocking. External ligament (Fig. 
10, lig) weak, amphidetic, supported by 
nymph. Internal ligament (resilium; Fig. 10, 
res) stronger, opisthodetic. 

Prodissoconch (Fig. 11) brownish-yellow, 
approximately 360 fm in length. Prodisso- 
conch I corresponding in size to shell of newly 
released larva, approximately 32% of length 
of prodissoconch II; sculpture not discernible 
(surface abraded in adult shell). Prodisso- 
conch II sculptured only with coarse and fine 
concentric growth lines. Prodissoconch I and 
II stages distinct, demarcated more by 
change in convexity than by sculpture or 
growth discontinuity (Fig. 11, single arrow). 
Demarcation between prodissoconch II and 
dissoconch abrupt (Figs. 8; 1 1 , double arrow). 

Shell microstructure (Fig. 13) cross- 
lamellar centrally, with thin prismatic layer 
covering each side, as in Divariscintilla yoyo 
(see Mikkelsen & Bieler, 1989). 



Organs of the Palliai Cavity: Foot (Fig. 4, ft) as 
previously described for Divariscintilla yoyo 
(see Mikkelsen & Bieler, 1989), including 
hatchet-shaped anterior portion, narrowed 
posterior extension (Fig. 4, pfe), anterior bys- 
sus gland (Fig. 4, by), ciliated ventral groove 
(Fig. 4, vgr) supplied with numerous mucous 
glands, and terminal byssus adhesive gland 
(Fig. 4, bag). Byssus gland of undefined 
structure, staining turquoise in histological 
sections; byssus adhesive gland of branching 
lamellar folds, staining purplish-red. One to 
four byssus threads produced, emanating 
from extreme posterior tip of foot. Opaque 
white pigment band (of unknown function) 
along anterodorsal tip of foot, staining dark 
purplish-red in sections. 

Anterior and posterior adductor muscles 
subequal, of moderate diameter. Anterior and 
posterior pedal retractor muscles smaller in 
diameter, inserting on shell just dorsal and 
medial to their respective adductor muscle 
scars. Very small pedal protractor muscle 
merging with anteroventral edge of anterior 
adductor muscle just before both attach to 
shell; inserting into anterior visceral mass just 
dorsal of labial palps. Muscles leaving very 
faint attachment scars on shell (Figs. 7, 9). 

Overall morphologies of visceral mass, la- 
bial palps and ctenidia as in Divariscintilla 
yoyo (see Mikkelsen & Bieler, 1989). Palps 
with 6-8 lamellae each side. Ctenidia smooth, 
unpleated (appearing posteriorly loosely 
pleated in preserved specimens due to con- 
traction). Outer demibranch approximately 
50% smaller than inner; both demibranchs 
ventrally rounded and with both interfilamen- 
tal and interlamellar connectives. Cilial cur- 
rents on palps and ctenidia not verified. 

Flower-like organ absent (see Discussion 
below). 

Digestive System: Structure of digestive sys- 
tem (mouth, esophagus, stomach, midgut, 
hindgut, rectum) of same organization as that 
in Divariscintilla yoyo (see Mikkelsen & Bieler, 
1989: fig. 28). 

Suprabranchial Chamber. Arrangement of 
openings and presence of glandular patches 
(?hypobranchial glands; Fig. 23, hyp) adja- 
cent to rectum as in Divariscintilla yoyo (see 
Mikkelsen & Bieler, 1989: fig. 29). 

Nervous System: Arrangement of ganglia, 
statocysts, and major nerves as described for 
Divariscintilla yoyo (see Mikkelsen & Bieler, 
1989: fig. 31). Left and right posterior tenta- 



MIKKELSEN & BIELER 



des innervated by branches from the palliai 
nerve, adjacent to its junction with the visceral 
ganglion. Anterior tentacles similarly inner- 
vated but both from a common branch of the 
palliai nerve, adjacent to its junction with the 
cerebro-pleural ganglion. 

Reproductive System: Simultaneous her- 
maphrodite. Ovotestis white, encompassing 
most of volume of visceral mass, as in Di- 
variscintilla yoyo (see Mikkelsen & Bieler, 
1989). 

Mature spermatozoa morphologically indis- 
tinguishable from that of D. yoyo (see Eckel- 
barger et al., 1990: figs. 28-29, table 1; D. 
octotentaculata as Scintilla sp.), except 
smaller in relative size. Spermatogenesis fully 
described by Eckelbarger et al. (1990; as 
Scintilla sp.). 

Brooding large number of small larvae for 
variable period; brooding time for individuals 
collected with larvae, 9-15 days (n = 4); total 
brooding time from set to release in labora- 
tory, 9, 10, and 12 days (n = 3). Larvae held 
within both demibranchs and in suprabran- 
chial chamber, where they are circulated via 
palliai expansions and contractions. During 
brooding, excurrent siphonal opening con- 
stricted by sphincter-like muscle, sometimes 
noted around free end of rectum, allowing di- 
gestive processes to continue. Larvae initially 
white, turning pink with shell development on 
day 5-7 (n = 2); released as straight-hinged 
"D" larvae with apical flagella, 1 15-123 |xm in 
shell length (x = 119 цт, n = 40; Fig. 12). 
Larvae expelled through excurrent siphon via 
strong contractions of shell and palliai mus- 
cles. Adults brooding larvae collected in May, 
June, December 1987, and March, April 
1988; additional adults setting larvae in labo- 
ratory in February, May, June 1982 and No- 
vember 1990; one specimen setting two 
broods in laboratory, four months apart, with 
second brood approximately 20% quantity of 
first. No apparent seasonality. 

Circulatory and Excretory Systems: As de- 
scribed for Divariscintilla yoyo (see Mikkelsen 
& Bieler, 1989). 

Distribution and Abundance 

Known from three locations, all on intertidal 
and shallow subtidal sand flats within the In- 
dian River Lagoon, eastern Florida: the type 
locality, Ft. Pierce Inlet (St. Lucie County, 
27°28.3'N, 80°17.9'W), just north of St. Lucie 
Inlet (Martin County, 27 11.4'N, 80°11.1'W), 



and Sebastian Inlet (Brevard County, 
27°51.6'N, 80°27.0'W). May be quite numer- 
ous; largest number per burrow sample = 74. 

Etymology 

An adjective, octotentaculatus, -a, -urn, 
from the Latin octo (eight) and the late Latin 
tentaculum (a "feeler"), referring to the eight 
long mantle tentacles, a diagnostic feature. 

Remarks 

This is the most common of the five Divaris- 
cintilla species in the Lysiosquilla burrows 
(see Ecology and Behavior). 

Prior to the beginning of this study in March 
1987, two individuals of this species were en- 
countered, and are presently the only known 
specimens of any of the Floridian burrow ga- 
leommatoideans from previous collections. 
These were collected in a shovel-and-sieve 
sample from a sand bar in the Indian River 
Lagoon, north of St. Lucie Inlet, Martin 
County, Florida, 27°11.4*N, 80°11.1'W, on 18 
February 1982. These individuals were main- 
tained in the laboratory for approximately four 
months, providing material for notes and pho- 
tographs on behavior, reproduction, and de- 
velopment. 

Divariscintilla luteocrinita n. sp. 

(Figs. 2, 5, 14-22, 24-25) 

Material examined 

Holotype: 5.0 mm (shell length), FMNH 
223403. Paratypes (12): 4.6 mm, FMNH 
223407; 4.4 mm (shell only), FMNH 223404; 
3.2, 3.9 mm, HBOM 064:01863; 4.9, 4.5, 4.1, 
2.9 mm (shells only, coated for SEM), HBOM 
064:01864; 4.8 mm, USNM 860195; 5.5 mm 
(shell only), USNM 859444; 3.4 mm, MCZ 
302517; 3.5 mm, SBMNH 35168. Total mate- 
rial: 16 specimens: FLORIDA: Ft. Pierce Inlet: 
10 March 1987, 1 ; 2-3 May 1987, 5; 24 June 
1 987, 1 ; 03 August 1 987, 1 ; 1 4 August 1 987, 
2; 31 August 1987, 1; 12 April 1988, 1; 03 
February 1990; 4. 

Type locality 

Ft. Pierce Inlet, Indian River Lagoon, St. 
Lucie County, eastern Florida, 27°28.3'N, 
80°17.9'W, occupying Lysiosquilla scabri- 
cauda burrows on intertidal sand flats with 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 




Figs. 8-13. Shell of Divariscintilla octotentaculata (SEM). 8. Left valve, external view, 2.9 mm length, para- 
type, HBOM 064:01867. 9. Right valve, internal view, 3.6 mm length, paratype, HBOM 064:01867. lO.Hinge, 
anterior to left, paratype, HBOM 064:01867. Scale bar = 100 (xm. 1 1 . Prodissoconch, 360 ^m length. Single 
arrow = prodissoconch l-ll boundary. Double arrow = prodissoconch ll-dissoconch boundary. 12. Newly 
released larval shell, 1 19 |xm length. 13. Microstructure, with internal surface at top. Scale bar = 5 (ati. (car, 
cardinal tooth; lig, external ligament; nym, nymph; per, periostracum; res, resilium). 



MIKKELSEN & BIELER 




Figs. 14-20. Shell of Divariscintilla luteocrinita (SEM). 14. Left valve, external view, 2.9 mm length, paratype, 
HBOM 064:01864. 15. Right valve, internal view, 4.9 mm length, paratype, HBOM 064:01864. 16. Hinge, 
anterior to left, paratype, HBOM 064:01864. Scale bar = 100 p.m. 17. Prodissoconch, 390 |xm length. Single 
arrow = prodissoconch l-ll boundary. Double arrow = prodissoconch ll-dissoconch boundary. 18. Internal 
surface, showing smooth adductor muscle scar (left) and adjacent region of opaque thickening (right). Scale 
bar = 10 ^m. 19. Microstructure in region of adductor muscle scar; internal surface at top. Scale bar = 5 
цль 20. Microstructure in region of opaque thickening, showing additional layer covering internal prismatic 
layer (pr); internal surface at top. Scale bar = 5 ц.т. (pr, internal prismatic layer). 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



patches of the seagrass Halodule wrightii As- 
cherson. Paratypes all from type locality. 

Diagnosis 

Animal translucent yellow. Mantle thin, with 
extensive, retractable, papillose folds com- 
pletely covering shell, meeting at mid-line. 
Tentacles originating at shell edge, three 
pairs anteriorly, two singles and 9-11 pairs 
posteriorly, plus one pair club-shaped tenta- 
cles adjacent to excurrent opening. Posterior 
foot-extension relatively short. Shell roundly 
triangular, with umbo slightly anterior, smooth 
exteriorly, with opaque thickenings interiorly; 
length approximately 70% of extended mantle 
length. Single "flower-like organ" on anterior 
surface of visceral mass. 

Description 

External Features and Mantle: Living ex- 
tended animal (Figs. 2, 5) generally 6-7 mm in 
length. Mantle and tentacles translucent pale 
yellow; foot white. Upper portion of digestive 
gland showing through mantle and shell as 
dark elongate-oval spot. Shell entirely cov- 
ered in life by anterior and posterior mantle 
folds (Fig. 5, m), meeting at lateral dorso-ven- 
tral midline on each side. Mantle folds thin, 
incompletely retractable, entirely finely papil- 
lose, with scattered, elongated papillae espe- 
cially posteroventrally. Mantle edge extending 
widely beyond shell edges; entire surface 
finely papillose. Palliai openings, muscula- 
ture, and posterior pouch as in Divariscintilla 
octotentaculata. Numerous, long, retractile 
tentacles originating at shell edge: three pairs 
anterior to umbo; two singles plus five pairs 
posterior to umbo ( = two singles on midline 
+ two pairs + [excurrent siphon] + three 
pairs). Shorter accessory pairs posterior to 
umbo originating from mantle fold near, but 
ventrad of, shell edge: one to three pairs an- 
terior, two to three pairs posterior to excurrent 
siphon. Structure of these tentacles as in D. 
octotentaculata; those on shell edge adjacent 
to cowl and posterior pouch (third and sixth, 
see numbers, Fig. 5) longest. One prominent 
pair of thicker, whitish, club-shaped tentacles 
(Fig. 5, est) immediately posterior to excurrent 
siphon originating just inside shell edge; inter- 
nal structure differing slightly from that of 
other tentacles (see Remarks). 

Preserved specimens incompletely re- 
tracted into gaping shell; mantle folds con- 
tracted to narrow rim along shell edge, expos- 



ing most of shell surface; tentacles, cowl, 
posterior pouch, and foot contracted but still 
usually extending beyond shell edges. 

Shell (Figs. 14-20): Shell generally 4-5 mm in 
length, roundly triangular to oval, longer pos- 
teriorly, equivalve, rather inflated, glossy, 
transparent to translucent white, smooth ex- 
cept for fine concentric growth lines and irreg- 
ular opaque thickening interiorly, imparting a 
white-blotched pattern (Fig. 15). Size large rel- 
ative to mantle, comprising approximately 
70% of extended mantle length. Valves held 
open at approximately 40° angle while crawl- 
ing, incapable of complete closure. Adductor 
muscle scars subequal, distinct due to pres- 
ence of surrounding shell thickening (Fig. 15). 
Periostracum as in Divariscintilla octotenta- 
culata. 

Hinge line short (Fig. 16), similar to that of 
Divariscintilla octotentaculata. Both cardinal 
teeth rounded. 

Prodissoconch (Fig. 17) brownish-yellow, 
approximately 390 (xm in length. Prodisso- 
conch I approximately 145 (xm in length, ap- 
proximately 37% of length of prodissoconch 
II; sculpture not discernible (surface abraded 
in adult shell). Prodissoconch II sculptured 
with coarse concentric growth lines. Demar- 
cation between prodissoconch I and II stages 
(Fig. 17, single arrow), and between prodis- 
soconch II and dissoconch (Figs. 14; 17, dou- 
ble arrow) as in Divariscintilla octotentaculata. 

Shell microstructure as in Divariscintilla oc- 
totentaculata, except with additional layer of 
parallel crystals covering internal perpendic- 
ular prismatic layer, forming regions of 
opaque thickening (Figs. 18-20). 

Organs of the Palliai Cavity. Foot (Fig. 5, ft) as 
in Divariscintilla octotentaculata. Adductor, 
pedal retractor, and pedal protractor muscles 
(including relative positions) as in D. octoten- 
taculata. Muscles leaving distinct attachment 
scars on shell (Fig. 15). 

Visceral mass, labial palps and ctenidia as 
in Divariscintilla octotentaculata. Palps with 
approximately 8 lamellae each side. Outer 
demibranch approximately 40% smaller than 
inner. Cilial currents on palps and ctenidia un- 
known. 

A single "flower-like organ" (Figs. 21-22; 
see Mikkelsen & Bieler, 1989, for explanation 
of term) on anterior surface of visceral mass 
just ventral to labial palps. Size variable, not 
correlated with shell length. 
Digestive System: As in Divariscintilla oc- 
totentaculata. 



10 



MIKKELSEN & BIELER 



Suprabranchial Chamber. As in Divariscintilla 
octotentaculata, except that the whitish glan- 
dular patches (?hypobranchial glands) are 
much more extensive (Fig. 24). 

Nervous System: Arrangement of ganglia, 
statocysts, and major nerves as described for 
Divariscintilla yoyo (see Mikkelsen & Bieler, 
1989: fig. 31). Additional tentacular nerves 
arising (independently) from palliai nerve. 

Reproductive System: Overall gross morphol- 
ogy as in Divariscintilla octotentaculata. Re- 
productive mode could not be determined 
from sectioned specimen, which showed no 
recognizable developed gametes. Adults 
brooding larvae have not been collected. 

Circulatory and Excretory Systems: As in Di- 
variscintilla octotentaculata. 

Distribution and Abundance 

Known only from the type locality, Ft. 
Pierce Inlet, St. Lucie County, Florida, 
27°28.3'N, 80°17.9'W, on intertidal and shal- 
low subtidal sand flats. Uncommon; only 16 
specimens known. 

Etymology 

An adjective, luteocrinitus, -a, -urn, from the 
Latin luteus (yellow) and the Latin crinitus 
(hairy), referring to the numerous, long, yel- 
lowish tentacles, imparting an overall "hairy" 
appearance. 

Remarks 

The single posterior pair of club-shaped 
tentacles is distinct and consistent in both liv- 
ing and preserved specimens. Normal tenta- 
cles (Fig. 25, nt; similar morphology in all five 
Floridian Divariscintilla spp.) show usually 
four haemocoelic compartments in cross sec- 
tion, each of these supplied with a longitudinal 
muscle bundle and nerve fiber, and separated 
by connective tissue septa. The muscle and 
nerve fibers form a more-or-less concentrated 
central core, appearing as a central thread 
under low magnification. Judd (1971: fig. 6) 
figured a similar internal morphology for ten- 
tacles of Divariscintilla maoria, but the individ- 
ual compartmental muscle-plus-nerve bun- 
dles are not as concentrated at the core of the 
tentacle; this difference may not be real, but 
rather implied by Judd's interpretation of the 
histological sections. An additional difference 



is found in the outer surfaces of the tentacles 
of D. luteocrinita, which are highly papillose 
and convoluted rather than smooth as in D. 
maoria as shown by Judd (1971 : fig. 6). Inter- 
nally, the club-shaped tentacles of D. luteo- 
crinita (Figs. 5, 25, est) are identical in struc- 
ture to normal tentacles except that the 
muscle fibers (Fig. 25, arrow) are dispersed 
over the surfaces of the septa instead of be- 
ing concentrated at the core of the tentacle. 
This difference may allow this particular pair 
of tentacles to undergo stronger contraction, 
resulting in the club-like shape. Under full ex- 
tension, the club shape and whitish coloration 
of these Tentacles disappears, indicating that 
these features are products of the normally 
contracted state. 

The function of the club-shaped tentacles is 
unknown. They are probably not homologous 
to the posterior "defensive appendages" of D. 
maoria (see Judd, 1971 : fig. 7) or Galeomma 
takii (Kuroda, 1945) (see B. Morton, 1973a: 
fig. 5), which can be autotomized and pos- 
sess only a single, central haemocoelic tube. 
No tentacles in D. luteocrinita have this mor- 
phology nor were they ever seen to autoto- 
mize. However, these tentacles were often 
observed to hyperelongate when the animals 
were disturbed, for example during specimen 
transfer between laboratory dishes. While 
most other tentacles contracted, the club- 
shaped tentacles elongated immediately to 2- 
3 times the shell length just as the transfer 
spoon made contact. This bears resemblance 
to the dymantic ("threatening") display of ten- 
tacles in Galeomma polita Deshayes, 1856 
(see B. Morton, 1975), and Ephippodonta oe- 
dipus Morton, 1976 (see B. Morton, 1976), 
when disturbed, and suggests defensive 
function. Unlike the dymantic tentacles of the 
latter two species, however, the club-shaped 
tentacles of D. luteocrinita do not fully retract 
into the mantle at rest. 

The large "?hypobranchial glands" of this 
species are located at sites similar to those of 
the smaller glands of other Divariscintilla spe- 
cies, i.e., adjacent to the rectum and the bran- 
chial nerves as they join the visceral ganglia. 
In histological sections, they bear striking re- 
semblance to "seminal receptacles" de- 
scribed and figured for Aligena elevata 
(Stimpson, 1851) by Fox (1979: 101, 103, fig. 
32), although in the single sectioned speci- 
men of D. luteocrinita, they contained no 
sperm. The montacutid Aligena elevata is a 
protandrous hermaphrodite, and its seminal 
receptacles are present only during the fe- 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



11 




Figs. 21-22. Flower-like organ of Divariscintilla luteocrinita (SEM). Anterior tip of foot has been severed to 
enhance visibility. 21 . Scale bar = 100 |xm. 22. Scale bar = 50 |xm. (ct, ctenidium; fl, flower-like organ; ft, 
foot; Ip, labial palp; m, mantle). 



male stage. The occurrence of such struc- 
tures in Galeommatoidea was summarized by 
Fox (1979: table 8, as Leptonacea, 15 spe- 
cies), who noted that sperm storage organs 
were unknown outside of Montacutidae. How- 
ever, protandry is known in members of other 
families (e.g. Lasaeidae: Arthritica crassi- 
formis Powell, 1933 (see B. Morton, 1973b); 
see also Fox, 1979: table 11). The reproduc- 
tive mode of D. luteocrinita is presently un- 
known, but seminal receptacles and/or 
protandry would be no surprise within the 
context of this reproductively complex super- 
family. 

Divariscintilla cordiformis n. sp. 

(Figs. 3, 6, 26-32) 

Material Examined 

Holotype: 5.6 mm (shell length), FMNH 
223405. Paratypes (4): 6.4 mm (sectioned on 
29 microslides), FMNH 223406; 5.4 mm, 
HBOM 064:1861 ; 4.9 mm (partially dissected; 
prodissoconch coated for SEM), HBOM 064: 
01862; 6.4 mm (shell only, coated for SEM), 
USNM 859445. Total material: 6 specimens: 
FLORIDA: Ft. Pierce Inlet: 24 June 1987, 2; 
-Peanut Island: 10 August 1987, 4. 

Type locality 

Peanut Island, near Lake Worth Inlet, Palm 
Beach County, eastern Florida, 26°46.6'N, 
80°02.7'W, occupying Lysiosquilla scabri- 



cauda burrows on intertidal sand flats with 
patches of the seagrass Halodule wrightii As- 
cherson. Paratypes from type locality (FMNH, 
USNM, HBOM 064:01861) or Ft. Pierce Inlet 
(HBOM 064:01862). 

Diagnosis 

Animal translucent white. Mantle thin, with 
extensive, retractable papillose folds covering 
shell completely, meeting at midline. Tenta- 
cles originating at shell edge, six pairs ante- 
riorly, eight pairs posteriorly. Posterior foot- 
extension relatively short. Shell oval, with 
umbo slightly anterior; length approximately 
65% of extended mantle length. Small, ven- 
tral, anteriorly directed indentation in each 
valve. Coarse growth lines and slightly 
beaded radial ribs restricted to edges of oth- 
erwise-smooth shell. Single "flower-like 
organ" on anterior surface of visceral mass. 

Description 

External Features and Mantle: Living ex- 
tended animal (Figs. 3, 6) generally 7-10 mm 
in length, translucent white except for dark 
digestive gland within visceral mass. Shell en- 
tirely covered in life by anterior and posterior 
mantle folds (Fig. 6, m) meeting at lateral 
dorso-ventral mid-line. Mantle folds thin, in- 
completely retractable, entirely finely papil- 
lose, with scattered larger papillae, which are 
longest at posteroventral section. Mantle 
edge extending widely beyond shell edges; 



12 



MIKKELSEN & BIELER 
24 sh 




mus 



est 




*5f ¿g 



Z :; lé¿¥*„ 



Figs. 23-25. Histological sections. 23. ?Hypobranchial gland adjacent to rectum in D. octotentaculata. 24. 
?Hypobranchial gland adjacent to rectum in D. luteocrinita. 25. Cross-sections of normal and club-shaped 
tentacles in D. luteocrinita. Arrow = muscularized septum. Scale bars = 100 \x,m. (est, club-shaped tentacle; 
ct, ctenidium; hyp, ?hypobranchial gland; mus, muscle fibers; n, nerve fiber; nt, normal tentacle; pam, 
posterior adductor muscle; r, rectum; sbc, suprabranchial chamber; sep, connective tissue septum; sh, shell; 
ten, tentacle). 



entire surface finely papillose, with additional 
enlarged papillae near shell edge. Ventral 
mantle edge entire, not cleft in vicinity of ven- 
tral shell indentation. Palliai openings, mus- 
culature, and posterior pouch as in Divariscin- 
tilla octotentaculata. Paired, retractable, 
palliai tentacles numerous, originating near 
shell edge. Six anterior pairs, those adjacent 
to cowl area (third and fourth, see numbers, 
Fig. 6) longest. Eight posterior pairs, those 
adjacent to excurrent siphon (tenth) longest, 
but shorter than longest anterior tentacles. A 
single median tentacle (Fig. 6, mpt) between 
first and second posterior pairs (Fig. 6, nos. 
7, 8). Structure of individual tentacles as in D. 



octotentaculata. Excurrent opening (Fig. 6, 
exc) between ninth and tenth tentacle pairs. 
Preserved animals not fully retracted into 
gaping shell; mantle folds contracted to nar- 
row rim along shell edge, exposing most of 
shell surface; tentacles, cowl, and posterior 
pouch contracted but still usually extending 
beyond shell edges; foot usually completely 
withdrawn into palliai cavity. 

Shell (Figs. 26-30, 32): Shell generally 5-7 
mm in length, nearly evenly oval, longer pos- 
teriorly, equivalve, compressed, glossy, trans- 
parent to translucent white. Small indentation 
(Figs. 6, ind; 27, arrow; 28) at mid-ventral 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



13 




Figs. 26-32. Shell of Divariscintilla cordiformis (SEM). 26. Right valve, external view, 6.4 mm length, para- 
type, USNM 859445. 27. Right valve, internal view, 6.4 mm length, paratype, USNM 859445. Arrow = 
ventral indentation. 28. Close-up of ventral indentation, interior view. Scale bar = 100 ^m. 29. Hinge, 
anterior to left. Scale bar = 200 ц.т. 30.Prodissoconch, 360 jxm length, paratype, HBOM 064:01862. Single 
arrow = prodissoconch l-ll boundary. Double arrow = prodissoconch ll-dissoconch boundary. 31. Newly 
released larval shell, 146 |xm length. 32. Microstructure, with internal surface at top. Scale bar = 5 |xm. (per, 
periostracum). 



14 



MIKKELSEN & BIELER 



margin, slanting anteriorly toward umbo, evi- 
dent only on distal 1 mm or so of shell growth 
(as evidenced by growth lines). Exterior 
sculpture smooth except for fine concentric 
growth lines, heavier at anteroventral margin, 
anterior to indentation. Beaded radial ribs re- 
stricted to distal 1 mm or so of shell edge, 
most prevalent interiorly at posteroventral 
margin (Fig. 27) and exteriorly at antero- and 
posterodorsal margins (Fig. 26), forming dis- 
tinct, fine crenulation at shell edge, absent 
only at ventral margin immediately anterior to 
indentation. Size large relative to mantle, 
comprising approximately 65% of extended 
mantle length. Valves held open at 20-30° an- 
gle while crawling, incapable of complete clo- 
sure. Adductor muscle scars subequal, faint. 
Periostracum as in Divariscintilla octotentac- 
ulata; also evident covering ventral indenta- 
tion (Fig. 28, per). 

Hinge line (Fig. 29) short, as in Divariscin- 
tilla octotentaculata, with two small rounded 
cardinal teeth. 

Prodissoconch (Fig. 30) brownish-yellow, 
approximately 360 |xm in length. Prodisso- 
conch I corresponding in size to shell of newly 
released larva, approximately 45% of length 
of prodissoconch II; sculpture not discernible 
(surface abraded in adult shell). Prodisso- 
conch II sculptured with coarse concentric 
growth lines. Demarcation between prodisso- 
conch I and II stages (Fig. 30, single arrow), 
and between prodicsoconch II and disso- 
conch (Fig. 30, double arrow) as in Divaris- 
cintilla octotentaculata. 



confirmation of structure through gross dis- 
section. 

Suprabranchial Chamber. As in Divariscintilla 
octotentaculata. 

Nervous System: Arrangement of ganglia, 
statocysts, and major nerves as described for 
Divariscintilla yoyo (see Mikkelsen & Bieler, 
1989: fig. 31). Additional tentacular nerves 
arising (independently) from palliai nerve. 

Reproductive System: Simultaneous her- 
maphrodite. Overall gross morphology as in 
Divariscintilla octotentaculata. 

One specimen brooding larvae collected in 
August 1987. Larvae released one day after 
collection. Newly-released larval shells 136- 
148 (¿m in length (x = 143 |xm, n = 40; Fig. 
31). Adult preserved and subsequently sec- 
tioned; apparently intact larvae found in su- 
prabranchial chamber and throughout diges- 
tive system, including intestine and rectum 
(therefore not being digested). 

Circulatory and Excretory Systems: As in Di- 
variscintilla octotentaculata. 

Distribution and Abundance 

Known from the type locality at Peanut Is- 
land, Palm Beach County, and from sand flats 
at Ft. Pierce Inlet, St. Lucie County, Florida, 
27°28.3'N, 80°17.9'W. Rare; only 6 speci- 
mens known. 

Etymology 



Organs of the Palliai Cavity: Foot and shell 
muscles (adductors, pedal retractors, pedal 
protractors, including relative positions) as in 
Divariscintilla octotentaculata. Muscles leav- 
ing very faint attachment scars on shell (Fig. 
27). 

Visceral mass, labial palps and ctenidia as 
in Divariscintilla octotentaculata. Palps with 
approximately seven lamellae each side. 
Outer demibranch approximately 35% smaller 
than inner. Cilial currents on palps and ctenidia 
unknown. 

Single "flower-like organ" (see Mikkelsen & 
Bieler, 1989) on anterior surface of visceral 
mass just ventral to labial palps. 

Digestive System: Similar to that of Divaris- 
cintilla octotentaculata (relative positions of 
gastric shield, style sac, digestive diverticula, 
midgut, etc.), based on histological sections. 
Limited number of specimens did not permit 



An adjective, cordiformis, -e, from the Latin 
cordis (heart) and the Latin forma (shape), 
referring to the ventrally indented shell out- 
line. 

Remarks 

As in Divariscintilla maoria (see Judd, 
1971), the ventral indentation in the shell of 
this species does not seem to be functionally 
important. In both species, it is present in both 
valves, anteriorly inclined, not developed in 
juveniles, and not reflected by soft anatomy. 



ECOLOGY AND BEHAVIOR 

As in the two previously described com- 
mensal galeommatids (Mikkelsen & Bieler, 
1989), no specimens of the newly described 
species were ever found physically attached 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



15 



to a mantis shrimp, either in the field or in 
museum specimens (HBOM). They are as- 
sumed to be free-living within the vertical por- 
tions of the U-shaped burrow, although spec- 
imens were never visible at the opening prior 
to pumping. Again as with the previous spe- 
cies (Mikkelsen & Bieler, 1989), these clams 
were never found free-living outside of the 
burrows or associated with any other burrow- 
ing invertebrate in the area (e.g. other mantis 
shrimps, callianassid shrimps, polychaetes, 
sipunculans), nor were any empty shells lo- 
cated in dry collections (AMNH, DMNH, 
FMNH, HBOM, USNM), probably because of 
their fragile nature. 

Divariscintilla octotentaculata was the most 
frequently encountered commensal mollusk in 
the Lysiosquilla burrows; of the 35 burrows 
containing mollusks, 31 contained D. octoten- 
taculata, 20 D. yoyo, 19 D. troglodytes, 8 D. 
luteocrinita, 6 Cyclostremiscus beauii, 7 Cir- 
culus texanus, and only 2 D. cordiformis. Di- 
variscintilla octotentaculata was usually col- 
lected with other commensals, occurring alone 
in only 6 of the 31 samples. Divariscintilla lu- 
teocrinita was always collected with other 
commensals. Divariscintilla cordiformis was 
collected once with D. octotentaculata, and 
once alone. Densities of D. octotentaculata 
varied greatly, ranging from 1-74 per sample 
(x = 8.3, n = 31). Divariscintilla luteocrinita 
was usually present in numbers of only one or 
two specimens per burrow; one sample con- 
tained four specimens. Divariscintilla cordi- 
formis was encountered only twice, once as 
two specimens, once as four. Only a small 
number of burrows sampled contained com- 
mensals. And, as previously reported (Mik- 
kelsen & Bieler, 1989), it must be emphasized 
that in no case could an entire burrow be sam- 
pled using the yabby pump, which only effec- 
tively samples its own length (0.5-1 .0 m) of the 
vertical parts of the U-shaped burrow. Esti- 
mates of occurrence and/or density of any 
clams living in the deeper horizontal section of 
the burrow was thus not possible. 

Animals of all three species spent most of 
their time in the laboratory attached to the 
glass surface of laboratory bowls by up to four 
byssus threads, and it is assumed that this is 
also their habit on the smooth walls of the 
Lysiosquilla burrow. When dislodged, they 
actively crawl about using an even, gliding 
motion produced by ciliary action on the cen- 
tral surface of the foot. This was equally ef- 
fective on the underside of the water surface 
as on glass. They made no attempts to bur- 



row when offered a substrate of loose sand in 
the laboratory. 

All three Divariscintilla species previously 
described (D. maoria, Judd, 1971: fig. 4; D. 
yoyo and D. troglodytes, Mikkelsen & Bieler, 
1989: fig. 32) are known to "hang" from a 
vertical substrate by the posterior foot-exten- 
sion, which in these species is extremely long 
and elastic; byssus threads secreted by the 
anteriorly located byssus gland are laid down 
within the ventral groove of the foot and 
emerge from the terminus to attach to the 
substrate. The threads are secured within the 
groove by secretions of the posteriorly lo- 
cated byssus adhesive gland. Byssus and 
byssus adhesive glands of similar morpholo- 
gies are present in each of the three new spe- 
cies described here and are assumed to func- 
tion in the same way. This has been 
confirmed for D. octotentaculata and D. luteo- 
crinita, in which secretion of byssus threads, 
accompanied by distinct pulsing of the byssus 
gland area, was observed as described for D. 
yoyo and D. troglodytes (Mikkelsen & Bieler, 
1989). Following this activity, the clam hangs 
from the posterior foot-extension, with the 
byssus threads emerging from the posterior 
terminus of the foot in the vicinity of the bys- 
sus adhesive gland. The posterior foot-exten- 
sion of these species, as well as of D. cordi- 
formis, is not as elongated and extensible as 
in those previously described, therefore the 
distinctive "hanging" posture, wherein the 
clam "dangles" from an elongated foot, is not 
as pronounced. 3 

A peculiar interaction between pairs of Di- 
variscintilla species was observed in the lab- 
oratory on three occasions. In two instances 
involving D. octotentaculata, one animal of a 
pair was noted reaching its foot into the man- 
tle cavity of the second specimen, either from 
in front of or behind its partner. On one of 
these occasions, the two individuals per- 
formed this activity simultaneously, "facing" 
one another, with each one reaching around 
the visceral mass of the other to contact the 
posterior surface with the tip of its foot (Fig. 
33). Two specimens of D. yoyo have been 



3 Careful notes were not recorded on the behavior of D. 
cordiformis in the early phase of the study, and the inavail- 
ability of additional living specimens prevented confirma- 
tion of the presence or absence of "hanging" behavior. 
Morphology suggests that this behavior does occur, how- 
ever, the shorter posterior extension and the absence of 
any mention in our preliminary written observations indi- 
cates that the "hanging" posture was inconspicuous, as in 
D. octotentaculata and D. luteocrinita. 



16 



MIKKELSEN & BIELER 




Fig. 33. Mating (?) behavior between two specimens of Divariscintilla octotentaculata, as observed in 
laboratory. 



noted performing this same type of activity. In 
all three instances, the interaction was initi- 
ated by the smaller of the two specimens of 
the pair, and the activity was sustained for 3-7 
minutes. Although there is no direct evidence, 
it seems likely that this observed interaction is 
part of some sort of reproductive activity, per- 
haps stimulation of sperm transfer (external 
gonadal openings are located on the poster- 
odorsal surface of the visceral mass). Each of 
the two specimens involved in the simulta- 
neous behavior described above had larvae 
in their suprabranchial chambers within three 
weeks of the noted activity. (If this behavior is 
in fact copulatory in function, then this con- 
firms simultaneous hermaphroditism in these 
species.) 

Observed animals of these Divariscintilla 
species do not respond to changes in light 
intensity (e.g. photographic strobes) and did 
not seek darkness when offered "artificial bur- 
rows" (in the form of black plastic tubes) in the 
laboratory. 



DISCUSSION 
Generic Placement 

The genus Divariscintilla was redescribed 
in a previous part of this study on Lysiosquilla- 
associated mollusks (Mikkelsen & Bieler, 
1989). Major emphasis was placed on two 
features, the presence of flower-like organs 
and a bipartite foot with both byssus and bys- 
sus-adhesive glands, shared between these 
species and the type species, D. maoria Pow- 
ell, 1932 (the latter described by Judd, 1971). 



We regarded the presence of an apparently 
functionless (Judd, 1971) shell notch in D. 
maoria, as well as the difference in degree of 
shell coverage by the mantle, as species- 
level rather than generic characters. At the 
time, we also considered placement of D. 
yoyo and D. troglodytes in the monotypic 
genus Phlyctaenachlamys Popham, 1939, 
based on P. lysiosquillina Popham, 1939. 
However, there are a number of anatomical 
characters that distinguish the Divariscintilla 
species (flower organs, hindgut typhlosole, 
interlamellar ctenidial junctions, relative size 
of adductor muscles, hinge teeth; compare 
data of Popham, 1939, and Mikkelsen & 
Bieler, 1989). 

Divariscintilla luteocrinita and D. cordi- 
formis fit well within Divariscintilla, as rede- 
scribed. In fact, these new species actually 
bridge several morphological gaps between 
D. maoria and D. yoyolD. troglodytes, that is 
shell notch, posterior shell prolongation, less 
shell internalization, and more numerous pos- 
terior tentacles. Divariscintilla octotentaculata 
does not possess the flower-like organs; how- 
ever, in all other taxonomic characters (hinge, 
mantle, ctenidia, etc.), it does conform to the 
redefined genus. The hinge teeth of this spe- 
cies, comprised (as in other Divariscintilla 
species) of only one small cardinal tooth in 
each valve, was an important element in de- 
ciding generic placement. Hinge structure is 
presently considered taxonomically important 
at the generic level in Galeommatoidea (P. H. 
Scott, in litt., October 1990). Most members of 
the superfamily possess more than one car- 
dinal tooth in at least one valve; some show 
distinct lateral teeth as well. The genus Sein- 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



17 



TABLE 1 . Distinguishing characteristics of all described species of Divahscintilla: an expansion of table 1 
from Mikkelsen & Bieler (1989). 





D. maoria 














(from Judd, 




D. 


D. 


D. 


D. 




1971) 


D. yoyo 


troglodytes 


octotentaculata 


luteocrinita 


cordiformis 


Shell: 














General shape 


oval 


elongate- 
pointed 


oval 


roundly 
triangular 


roundly 
triangular 


oval 


Ventral indentation 


present 


absent 


absent 


absent 


absent 


present 


Prolongation 


posterior 


anterior 


anterior 


posterior 


posterior 


posterior 


Internal sculpture 


unribbed 


unribbed 


radially 


radially 


unribbed, with 


radially ribbed 








ribbed 


ribbed 


internal 


(marginally) 








(marginally) 


(marginally) 


thickening 




Length relative to 














extended 














mantle length 


68% 


40% 


50% 


70% 


70% 


65% 


Prodissoconch 














length (ц.т) 


"small" 


360 


380 


360 


390 


360 


Mean newly-released 














larval shell 














length (ц.т) 


unknown 


132 


126 


119 


unknown 


143 


Mantle: 














Color, thickness 


(not 


whitish, 


yellowish, 


whitish, thin 


yellowish, 


whitish, thin 




given) 


thick 


thin 




thin 




Extent covering 


margins 


entire, 


entire, 


anterior and 


entire, 


entire, 


shell 


only 


umbonal 
foramen 


anterior slit 


posterior thirds 


midline 
overlap 


midline 
overlap 


Papillae 


very 


sparse, 


numerous, 


numerous, 


numerous, 


numerous, 




small 


very small 


small, 


small, 


small, 


small, 








evenly- 


longer at 


evenly- 


evenly- 








distributed 


ventral 
edge 


distributed 


distributed 


Anterior tentacles 


2 pairs 


1 pair 


2 pairs 


2 pairs 


3 pairs 


6 pairs 


Posterior tentacles 


1 single 


1 single 


1 single, 


1 single, 


2 singles, 


1 single, 








1 pair 


2-3 pairs 


5 pairs, 

+ 3-6 pairs 

accessory, 

+ 1 pair 

club-shaped 


8 pairs 



Defensive 










appendages 


6-8 
present 


absent 


absent 


absent 


Pedal protractor 










muscle 










insertion relative to 










anterior adductor 










muscle 


(not 
given) 


dorsal 


dorsal 


ventral 


Flower-like organs: 










number 


1 


3-7 
(usually 5) 


1 





Labial palps: 










Lamellae per palp 


approx. 9 


10-14 


14-20 


6-8 


Ctenidia: 


smooth 


pleated 


pleated 


smooth 


Geographical range: 


New 


eastern 


eastern 


eastern 




Zealand 


Florida 


Florida 


Florida 



absent 



ventral 



absent 



ventral 



approx. 8 


approx. 7 


smooth 


smooth? 


eastern 


eastern 


Florida 


Florida 



Шопа Finlay, 1927, is the only other genus 
known to us In which members have a single 
cardinal tooth in each valve. However, mem- 



bers of the latter genus are all attached ecto- 
commensals on echinoderms, and in addi- 
tion, are distinguished by a highly specialized, 



18 



MIKKELSEN & BIELER 



laterally compressed and furrowed foot. Both 
of these considerations exclude the new 
species described here. In view of the taxo- 
nomic confusion present in this group, we 
prefer to use an existing genus until a generic 
revision of this superfamily based on both 
shell and anatomical characters can be ac- 
complished. 

Coney (1990), in a review of "ventrally 
notched galeommatid genera," assigned Di- 
variscintilla yoyo and D. troglodytes to the ge- 
nus Phlyctaenachlamys, and reinstated 
Divariscintilla as a monotypic genus. His rea- 
soning revolved around (1) the shell notch, (2) 
hinge teeth and ligament morphology, (3) 
shell ultrastructure, (4) mantle tentacles, (5) 
degree of shell coverage by the mantle, and 
(6) ctenidial morphology. We disagree with 
his taxonomic decisions and address these 
points as follows: 

(1) Shell notch: The newly described spe- 
cies Divariscintilla cordiformis has an appar- 
ently functionless ventral shell notch very sim- 
ilar to that of the type species D. maoria, and 
agrees well anatomically with the "un- 
notched" species assigned to Divariscintilla. 
Importantly, the notch does not influence 
mantle or other soft-part morphology. 

(2) Hinge and ligament: Coney (1990: 131, 
135) offered conflicting statements regarding 
hinge teeth. His table of generic characters 
and description of Divariscintilla maoria 
stated that there was one cardinal tooth in the 
right valve, whereas his generic description 
indicated two. Although his scanning photo- 
micrograph of the right hinge (Coney, 1990: 
fig. 1 1 ) seemed on first examination to reveal 
two cardinal teeth, comparison with our pho- 
tomicrographs as well as Coney's subse- 
quent text indicated that the posteriormost 
"knob" was actually the resilium, not a sec- 
ond cardinal tooth. Coney (1990) upheld the 
original interpretation by Powell (1932) that 
the left valve was edentulous, although his 
scanning photomicrograph (Coney, 1990: fig. 
12) featured a small structure that may corre- 
spond to Powell's (1932: 67) "minute and 
shapeless vestige" of a left cardinal. Addition- 
ally, Coney (1990) noted a ridge in the left 
valve that he called a posterior lateral tooth. 
All Divariscintilla species (by our definition) 
possess strengthened or thickened hinge 
lines to varying degrees, but without develop- 
ment into distinct and/or interlocking lateral 
teeth/lamellae. As we lack relevant ontoge- 
netic data on shell development, we do not 
wish to infer homology in this instance and 



prefer not to call these structures "lateral 
teeth." 

According to Popham (1939: 66, text-fig. 6), 
the hinge of Phlyctaenachlamys lysiosquillina 
has two discrete cardinal teeth in the right 
valve, one cardinal in the left, and an inter- 
locking set of posterior laterals. This is dis- 
tinctly different from the situation in Divaris- 
cintilla yoyo, D. troglodytes, and D. maoria. 

Ligament morphology in all Divariscintilla 
species (by our definition) consists of an ex- 
ternal amphidetic ligament (inappropriately 
called "periostracal webbing" at one point in 
our earlier paper; Mikkelsen & Bieler, 1989: 
178) supported by a nymph, and an internal 
opisthodetic resilium. The resilium is also ap- 
parently present in Phlyctaenachlamys lysios- 
quillina, but the external ligament was not 
mentioned by Popham (1939). 

In total, these findings do not agree with 
Coney's (1990: 142) statement that "the 
hinge teeth and ligament are remarkably sim- 
ilar between the three species [lysiosquillina, 
yoyo, troglodytes], but are quite different than 
those of Divariscintilla maoria. ..." 

(3) Shell ultrastructure: This feature was 
also utilized by Coney (1990) in his discus- 
sion of distinguishing characteristics of Di- 
variscintilla and Phlyctaenachlamys species, 
although he also (1990: 142) admitted that 
the "shell ultrastructure of Phlyctaenach- 
lamys lysiosquillina is unknown". Studies of 
the three new species described here (Figs. 
13, 19-20, 32), as well as a réévaluation of 
this character in D. yoyo and D. troglodytes 
(see Mikkelsen & Bieler, 1989: figs. 11, 15), 
indicate that both the inner and outer non- 
cross-lamellar layers may be prismatic rather 
than "homogenous" as previously labelled by 
us. Variability in thickness and/or presence of 
the various layers among individuals and 
among different locations on a single valve 
(e.g. extra internal layer in D. luteocrinita, see 
above) has also been noted. A more exacting 
study using more appropriate methodology 
(e.g. sections rather than fractions, see Taylor 
et al., 1973) is necessary before differences 
at this level should be employed in taxonomic 
decisions. 

(4) Mantle tentacles: Coney (1990: 142) 
maintained that "number and placement of 
mantle tentacles and defensive appendages 
is strongly similar between Phlyctaenach- 
lamys] lysiosquillina and those of P. yoyo and 
P. troglodytes," noting that none of these spe- 
cies possess the numerous posterior defen- 
sive tentacles seen in Divariscintilla maoria. 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



19 



The three new species described here in Di- 
variscintilla all possess a number of posterior 
tentacles, albeit none "defensive." The two 
primary anterior tentacles, also mentioned 
specifically by Coney (1990: 142) to combine 
P. lysiosquillina, D. yoyo, and D. troglodytes, 
are in fact also present in Divariscintilla mao- 
ria. 

(5) Shell coverage: Although there is a dis- 
tinct difference in the degree of shell cover- 
age by the mantle between the type species 
and Divariscintilla yoyo and D. troglodytes, 
the three newly described species show inter- 
mediate conditions. All Divariscintilla species 
(by our definition) show at least some degree 
of shell exposure, thus differing from the con- 
dition in Phlyctaenachlamys lysiosquillina 
("the shell is completely embedded"; Pop- 
ham, 1939: 65). A similar range of variability 
was described for the genus Ephippodonta 
Tate, 1889, by Arakawa (1960: 57), and can 
also be found in Entovalva Völtzkow, 1890, 
sensu lato, wherein the genus Devonia was 
distinguished by Winckworth (1930: 14) for a 
species with incomplete shell coverage. 

(6) Ctenidial morphology: Coney (1990: 
142) noted that the ctenidia in Divariscintilla 
maoria are smooth, whereas those of Phlyc- 
taenachlamys lysiosquillina, D. yoyo and D. 
troglodytes have been described as pleated. 
However, unlike pleated (= plicate) gills in 
other bivalve groups (e.g. Pectén; see Rice, 
1897), pleating in these species is not based 
on structural differences in the filaments 
(pers. obs.; Popham, 1939: 71). Some degree 
of "pleating" caused by contraction in pre- 
served specimens has been noted during the 
present study. An additional ctenidial charac- 
ter separates P. lysiosquillina and the five Flo- 
ridian species of Divariscintilla, in that the lat- 
ter have interlamellar junctions (unknown for 
New Zealand D. maoria). 

Comparative characteristics for all six 
known species of Divariscintilla are presented 
in Table 1 . The three new species described 
here more closely resemble the type species, 
D. maoria (see Judd, 1971 : fig. 1), in general 
morphology than do the other species previ- 
ously described during this study (D. yoyo 
and D. troglodytes, see Mikkelsen & Bieler, 
1 989: figs. 1 , 2). Like D. maoria, the three new 
species have posteriorly prolonged, relatively 
large shells and smooth ctenidia (differences 
in percent reduction of the outer demibranch 
as cited in the text may not be reliable, as they 
were taken from preserved specimens and 
were affected by contraction). One species, 



D. octotentaculata, has a shell that is similarly 
incompletely covered by the mantle. Another 
species, D. cordiformis, shows a similar ven- 
tral indentation that is likewise apparently 
functionless. These similarities effectively re- 
move most of the dissimilarity between the 
New Zealand type species and the included 
eastern Florida species that existed at the 
completion of the previous paper (Mikkelsen 
& Bieler, 1989). Divariscintilla maoria is the 
only species in the genus for which detach- 
able defensive papillae have been described. 

Distribution 

The peculiar pattern of geographic distri- 
bution of Divariscintilla species, with now 
five members in the western Atlantic and one 
in New Zealand, is most likely a result of in- 
sufficient sampling of burrow fauna. No eco- 
logical niche separation between the five 
sympatric species was recognized, leaving in- 
teresting questions for future research. 

Comparison with Other Genera 

Divariscintilla was formerly treated as a 
subgenus of Vasconiella Dall, 1899, by Cha- 
van (1969), apparently on the basis of 
notched shells. However, members of V. jef- 
freysiana (P. Fischer, 1873), type and sole 
species of the genus, possess a deep inden- 
tation only in the comparatively smaller right 
valve, and importantly, there are modifica- 
tions in the right mantle and ctenidial tissues 
corresponding to the notch (Cornet, 1982). 
This species is also the only other galeomma- 
toidean with a published account of a "flower- 
like organ" (Table 2). Unfortunately, although 
recognizably illustrated by Cornet (1982: fig. 
5), no details on structure or possible function 
were provided for the briefly mentioned 
"rounded tubercle just under the labial palps" 
(Cornet, 1982:39). Vasconiella jeffreysiana is 
probably also commensally associated with a 
mantis shrimp, Lysiosquilla eusebia (Risso, 
1816) (Table 2; Cornet, 1982). Differences of 
Vasconiella from Divariscintilla (i.e. two cardi- 
nal teeth in the left valve, lack of a posterior 
foot-extension, ?lack of ctenidial interlamellar 
junctions) prevent synonymy of the two gen- 
era as currently defined, but clearly, their re- 
lationship should be investigated further. 

Mention has been made several times 
above to the relationships of galeommatoide- 
ans with certain phyla of host invertebrates 
(e.g. echinoderms versus stomatopods). Evi- 



20 



MIKKELSEN & BIELER 



TABLE 2. Galeommatoidean 
structure (x = presence; - = 



species, hosts, and occurrences of flower-like organs and hanging foot 
absence). 







Flower- 


Hanging 








like 


foot 






Host 


organs 


structure 


References(s) 


Divariscintilla maoria Powell, 


Heterosquilla tricarinata 


X 


X 


Judd, 1971 


1932 


(Claus) 








D. yoyo Mikkelsen & Bieler, 


Lysiosquilla scabricauda 


X 


X 


Mikkelsen & Bieler, 


1989 


(Lamarck) 






1989 


D. troglodytes Mikkelsen & 


Lysiosquilla scabricauda 


X 


X 


Mikkelsen & Bieler, 


Bieler, 1989 


(Lamarck) 






1989 


D. octotentaculata n. sp. 


Lysiosquilla scabricauda 
(Lamarck) 




X 


This study 


D. luteocrinita n. sp. 


Lysiosquilla scabricauda 
(Lamarck) 


X 


X 


This study 


D. cordiformis n. sp. 


Lysiosquilla scabricauda 
(Lamarck) 


X 


X 


This study 


Parabornia squillina 


Lysiosquilla scabricauda 


X 


X 


Boss, 1965a; this study 


Boss, 1965 


(Lamarck) 








Vasconiella jeffreysiana 


?L. eusebia (Risso) 


X 


- 


Cornet, 1982 


(P. Fischer, 1873) 










Phlyctaenachlamys 


L. maculata (Fabricius) 


- 


X 


Popham, 1939 


lysiosquillina Popham, 










1939 










Ceratobornia longipes 


Callianassa major Say or 


- 


X 


Jeffreys, 1863; Dall, 


(Stimpson, 1855) 


Upogebia affinis (Say) 






1899; Norman, 1891; 
Jenner & McCrary, 1968 


С. сета Narchi, 1966 


Callianassa major Say 


— 


X 


Narchi, 1966 



dence suggests that host specificity applies 
mainly to those cases where modifications for 
locomotion and attachment have occurred. 
Most "generalist" species (sensu В. Morton & 
Scott, 1989; e.g. Mysella bidentata (Montagu, 
1803), associated with a wide variety of inver- 
tebrates [Boss, 1965b]) possess a simple foot 
structure consisting of a strong crawling por- 
tion (similar to the anterior foot of Divariscin- 
tilla spp.) ending in a bluntly rounded "heel" 
from which byssus threads emanate (B. Mor- 
ton & Scott, 1989: fig. 1 [Lasaea], figs. 9-10 
[Pseudopythina]). This is probably the plesio- 
morphic condition in the superfamily, com- 
pared to the more derived states seen in 
some of the host specialists. For example, the 
flattened foot of Santulona spp. (J. E. Morton, 
1 957) may be a modification for laterally ap- 
plied locomotion among the vertical spines 
and papillae of echinoderms to which they at- 
tach (Dali et al., 1938: 145; Yamamoto & 
Habe, 1974; Ó Foighil & Gibson, 1984: 75). 
Also, the sucker-like anterior foot of Entovalva 
( = Devonia) perrieri (Malard, 1903) (see An- 
thony, 1916) allows attachment to the 
smooth, outer body walls and/or inner cloacal 



walls (Bruun, 1938) of burrowing holothuri- 
ans. 

An elongate posterior foot-extension for at- 
tachment to a smooth vertical substrate is 
found in three specialist genera: Divariscin- 
tilla, Phlyctaenachlamys Popham, 1939, and 
Ceratobornia Dall, 1899 (Table 2). All species 
involved are associated with crustaceans that 
produce smooth-walled burrows in sand 
(Table 2). Most are believed to live attached 
to the crustacean burrow walls (Ceratobornia. 
cema may also "temporarily" attach to its host 
(Narchi, 1966); the biology of C. longipes is 
unknown). Although insufficiently studied in 
Phlyctaenachlamys and Ceratobornia, all 
species apparently possess the same 
"hanging apparatus" comprised of an anterior 
byssus gland, ventral groove, and posterior 
byssus adhesive gland (see Popham, 1939: 
fig. 2, P. lysiosquillina, hanging behavior not 
specifically described; Narchi, 1966: figs. 1,5, 
С cema; Dall, 1899: pi. 88, figs. 10-11,13, C. 
longipes). They also all possess similar gen- 
eral morphologies of the palliai, ctenidial, di- 
gestive, circulatory, excretory, and nervous 
systems (C. longipes incompletely known). 



NEW SPECIES OF COMMENSAL GALEOMMATIDAE 



21 



TABLE 3. Distinguishing characteristics for the three galeommatoidean genera possessing the "hanging 
foot." See text for included species and sources of data. [L, left; R, right] 





Divariscintilla 


Phlyctaenachlamys 


Ceratobornia 


Shell internalization 


incomplete 


complete 


incomplete 


Hinge: 








Cardinal teeth 


1 R, 1 L 


2 R, 1 L 


2R.2L 


Lateral teeth 


absent 


1 posterior, 


1 posterior, 






reduced 


reduced 


Retraction into shell 


yes (1 sp.) 
no (5 spp.) 


no 


yes 


Adductor muscles 


subequal 


posterior reduced 


subequal 


Flower-like organs 


present (5 spp.) 
absent (1 sp.) 


absent 


absent 


Interlamellar ctenidial 


present 


absent 


absent 


junctions: 








Hindgut typhlosole 


present 


absent 


absent 


Hypobranchial gland 


present? 


absent 


absent 


Supportive chondroid 


absent 


absent 


present 


edge in foot 









The anatomical differences among the three 
genera that presently prevent their synonymy 
are listed in Table 3. Until the importance of 
each of these characters can be reassessed, 
it is uncertain whether possession of a 
"hanging foot" reflects convergence or phylo- 
genetic relationship, and the three genera are 
best treated separately. 

Another possible difference between Di- 
variscintilla on one hand and Phlyctaenach- 
lamys and Ceratobornia on the other lies in 
the mode of reproduction. Members of Di- 
variscintilla for which such data are available 
(D. yoyo, D. troglodytes, D. octotentaculata, 
D. cordiformis) are known to be simultaneous 
hermophrodites. From literature data, it 
seems that Phlyctaenachlamys lysiosquillina 
and Ceratobornia cema are forms with sepa- 
rate sexes (Popham, 1939, p. 80: "specimen 
sectioned was a male"; and Narchi, 1966, p. 
521: "sectioned specimen was a female"). 
However, while simultaneous hermaphrodit- 
ism can be documented from individuals with- 
out observations over extended periods of 
time, "males" or "females" could belong to 
forms with consecutive hermaphroditism. 
Protandrous and protogynous hermaphrodit- 
ism have both been reported for the super- 
family (summarized by Fox, 1979: tab. 1 1 , as 
Leptonacea). 

Rhamphidonta Bernard, 1975, represented 
by the single species R. retifera (Dall, 1899), 
also possesses a bipartite foot but one which 
is different from that discussed above, both 
morphologically and functionally. According 
to Bernard (1975), the foot of R. retifera is 



anteriorly elongated, with the main enlarged 
crawling portion located posteriorly. Members 
of this species are not known to hang; they 
burrow into sand to avoid illumination, and are 
apparently free-living. The hinge of R. retifera 
(see Bernard, 1975: fig. 1) is distinctly "mon- 
tacutid." Close relationship with the three 
genera discussed above is unlikely. 

The mantis shrimp Lysiosquilla scabri- 
cauda also serves as host to Parabornia 
squillina Boss, 1965, a galeommatoidean that 
attaches to the inner surface of the abdominal 
sclera of the shrimp (Table 2). Parabornia 
squillina has been collected in the region of 
this study (Sebastian Inlet, Peanut Island), 
but has not been collected in burrows contain- 
ing Divariscintilla species. Interestingly, the 
animal of P. squillina also possesses a single 
"flower-like organ" (pers. obs.), although this 
was not mentioned in the original species de- 
scription based on preserved material. (Boss, 
1965a: fig. 3 shows an unexplained five-part 
zig-zag outline for the visceral mass below 
the gills and labial palps.) Its foot (Boss, 
1965a: fig. 3) appears morphologically similar 
to the "hanging type" described above; a 
deep ventral groove ends with an opaque 
white area at the end of a very short posterior 
extension (pers. obs.). Boss (1965a: 4) inter- 
preted the white area at the end of the foot as 
the byssus gland (as did Judd [1971] for D. 
maoria, and Narchi [1966] for Ceratobornia 
cema). In view of the byssus adhesive gland 
described here and previously (Mikkelsen & 
Bieler, 1989), this organ (which was depicted 
with internal lamellae by Boss, 1965a: fig. 3) 



22 



MIKKELSEN & BIELER 



warrants reinvestigation. If this structure is the 
primary byssus gland, what is the function of 
the ventral groove? (Histological study of the 
structure of the entire foot, and observations 
of living animals producing byssus threads 
would resolve this question.) Boss (1965a) 
placed Parabornia in the Erycinidae (Erycini- 
nae), in part, because its hinge has two car- 
dinal teeth in each valve and a central resil- 
ium. If hinge structure is eventually confirmed 
as a reliable taxonomic character based on 
phylogeny of the group, the "hanging foot" 
and "flower-like organs" would need to be ex- 
plained as results of evolutionary conver- 
gence and/or symplesiomorphies. 

Mating Behavior in Bivalves 

The observed "mating" behavior (Fig. 33) 
between individuals of Divariscintilla octoten- 
taculata as well as of D. yoyo is of special 
interest. Mating behavior has not been con- 
firmed for bivalves (Mackie, 1984: 362). 
Mackie (1984: 363) summarized the reported 
cases of associations between females and 
dwarf males in bivalves and concluded that 
those interactions would qualify as mating be- 
havior if it can be shown that the associations 
are confined to a breeding period. 

True copulatory behavior is absent in bi- 
valves because they lack copulatory organs. 
However, several authors (Clapp, 1951: 7; 
Townsley et al., 1966: 49) have observed 
male teredinids {Bankia spp.) inserting their 
exhalent siphons into the inhalent siphons of 
females during sperm release. Purchon 
(1977: 295), and subsequently Mackie (1984: 
362), interpreted this as the use of an intromit- 
tant organ, the development of which would 
be of "great survival value to species ... living 
in isolated pieces of drift wood (Purchon, 
1977: 295). Similarly, quasi-copulatory be- 
havior in Divariscintilla would be advanta- 
geous for these small bivalves with limited ga- 
mete production, living in restricted, relatively 
isolated populations within the burrows of 
their hosts. Selective mating in burrows that 
contain several other congeneric species 
would guarantee successful fertilization and 
make conservative use of male gametes. In 
our previous discussion of flower-like organs, 
now known from five species of Divariscintilla, 
we (Mikkelsen & Bieler, 1989: 191) specu- 
lated that these organs might emit a phero- 
mone for attracting reproductive partners 
(another possibility, if they are indeed phero- 



mone-emitting organs, would be the attraction 
of larvae to settlement sites). 



ACKNOWLEDGMENTS 

The following are gratefully acknowledged: 
Dr. Raymond B. Manning (USNM), for bring- 
ing this interesting fauna to our attention; 
William D. ("Woody") Lee (SMSLP), for in- 
valuable field help; Paul H. Scott (SBMNH), 
for comments on generic placement; Patricia 
A. Linley and Pamela Blades-Eckelbarger 
(both H80I) for SEM assistance; Tom 
Smoyer (HBOI) for photographic work; Dr. 
Kerry B. Clark (Florida Institute of Technol- 
ogy, Melbourne) and John E. Miller (formerly 
HBOI) for the use of and help with video re- 
cording equipment; Paul S. Mikkelsen (for- 
merly HBOI), for behavioral and developmen- 
tal notes and the photograph in Fig. 1 ; Chris 
Bauman (Vero Beach, Florida) for the illustra- 
tion in Fig. 33; Paul H. Scott (SBMNH), Dr. 
Kenneth J. Boss (MCZ), and two anonymous 
reviewers for valuable comments on the 
manuscript. 

This research was supported in part by the 
Smithsonian Marine Station at Link Port; the 
cooperation of Dr. Mary E. Rice and her staff 
is gratefully acknowledged. Funding for this 
project was derived in part from a National 
Capital Shell Club scholarship to P. M., and a 
NATO Postdoctoral Research Fellowship at 
SMSLP to R. B. This is Harbor Branch 
Océanographie Institution Contribution no. 
862 and Smithsonian Marine Station Contri- 
bution no. 273. 



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ARAKAWA, K. Y., 1960, Ecological observations 
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BERNARD, F. R., 1975, Rhamphidonta gen. n. 
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BIELER, R. & P. M. MIKKELSEN, 1988, Anatomy 
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BOSS, K. J., 1965a, A new mollusk (Bivalvia, Ery- 
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BOSS, K.J., 1965b, Symbiotic erycinacean bi- 
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BRUUN, A. F., 1938, A new entocommensalistic 
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167. 

CHAVAN, A., 1969, Superfamily Leptonacea Gray, 
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CLAPP, W. F., 1951, Observations on living Tere- 
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CONEY, С. О, 1990, Bellascintilla parmaleeana 
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CORNET, M., 1982, Anatomical description of Vas- 
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DALL, W. H., 1899, Synopsis of the Recent and 
Tertiary Leptonacea of North America and the 
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DALL, W. H., P. BARTSCH & H. A. REHDER, 
1938, A manual of the Recent and fossil marine 
pelecypod mollusks of the Hawaiian Islands. Ber- 
nice P. Bishop Museum Bulletin, 153: ¡v + 233 
pp., 58 pis. 

ECKELBARGER, K. J., R. BIELER & P. M. MIK- 
KELSEN, 1990, Ultrastructure of sperm develop- 
ment and mature sperm morphology in three 
species of commensal bivalves (Mollusca: Ga- 
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75. 

FOX, T. H., 1979, Reproductive adaptations and life 
histories of the commensal leptonacean bivalves. 
Ph.D. dissertation, University of North Carolina, 
Chapel Hill, ix + 207 pp. 

HUMASON, G. L, 1962, Animal Tissue Tech- 
niques. W. H. Freeman & Co., San Francisco & 
London, xv + 468 pp. 

JEFFREYS, J. G., 1863, British Conchology, oran 
account of the Mollusca which now inhabit the 
British Isles and the surrounding seas. Vol. II. 
Marine Shells, comprising the Brachiopoda, and 
Conchífera from the family of Anomiidae to that of 
Mactridae. London, John Van Voorst, xiv + 465 
pp., 8 pis. 

JENNER, С. E. & A. B. McCRARY, 1968, A "gas- 



tropod" bivalve, commensal on Squilla empusa 
(Abstract). Annual Report for 1968, American 
Malacological Union: pp. 20-21 . 

JUDD, W., 1971, The structure and habits of Di- 
variscintilla maoria Powell (Bivalvia: Galeomma- 
tidae). Proceedings of the Malacological Society 
of London, 39: 343-354. 

MACKIE, G. L, 1984, Bivalves. Pp. 351-418, 
in: A. S. TOMPA, et al., The Mollusca, Vol. 7, Re- 
production. Academic Press, Orlando, xix + 486 
PP- 

MIKKELSEN, P. M. & R. BIELER, 1989, Biology 
and comparative anatomy of Divariscintilla yoyo 
and D. troglodytes, two new species of Galeom- 
matidae (Bivalvia) from stomatopod burrows in 
eastern Florida. Malacologia, 31(1): 1-21. 

MORTON, В., 1973a, The biology and functional 
morphology of Galeomma (Paralepida) takii (Bi- 
valvia: Leptonacea). Journal of Zoology, 169(2): 
133-150. 

MORTON, В., 1973b, Some factors affecting the 
location of Arthritica crassiformis (Bivalvia: Lep- 
tonacea) commensal upon Anchomasa similis 
(Bivalvia: Pholadidae). Journal of Zoology, 170: 
463-473. 

MORTON, В., 1975, Dymantic display in Ga- 
leomma polita Deshayes (Bivalvia: Leptonacea). 
Journal of Conchology, 28: 365-369. 

MORTON, В., 1976, Secondary brooding of tem- 
porary dwarf males in Ephippodonta (Ephipp- 
odontina) oedipus sp. nov. (Bivalvia: Leptona- 
cea). Journal of Conchology, 29: 31-39. 

MORTON, B. & P. H. SCOTT, 1989, The Hong 
Kong Galeommatacea (Mollusca: Bivalvia) and 
their hosts, with descriptions of new species. 
Asian Marine Biology, 6: 129-160. 

MORTON, J. E., 1957, The habits of Santulona ze- 
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N ARCH I, W., 1966, The functional morphology of 
Ceratobornia cema, new species of the Erycina- 
cea (Mollusca, Eulamellibranchiata). Anais da 
Academia Brasileira de Ciencias, 38(3-4): 513- 
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NORMAN, A. M., 1891, Lepton squamosum (Mon- 
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Ó FOIGHIL, D. & A. GIBSON, 1984, The morphol- 
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POPHAM, M. L, 1939, On Phlyctaenachlamys 
lysiosquillina gen. and sp. nov., a lamellibranch 
commensal in the burrows of Lysiosquilla macu- 
late. British Museum (Natural History), Great Bar- 
rier Reef Expedition 1928-29, Scientific Reports, 
6(2): 549-587. 

PURCHON, R. D., 1977, The Biology of the Mol- 
lusca, 2nd ed. Pergamon Press, Oxford, xxv + 
560 pp. 

RICE, E. L, 1897, Die systematische Verwert- 
barkeit der Kiemen bei den Lamellibranchiaten. 



24 



MIKKELSEN & BIELER 



Jenaische Zeitschrift für Naturwissenschaft, 31 
(N.F. 24): 29-89, pis. 3-4. 

TAYLOR, J. D., W. J. KENNEDY & A. HALL, 1973, 
The shell structure and mineralogy of the B¡- 
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sions. Bulletin of the British Museum (Natural 
History), 22(9): 253-294, pis. 1-15. 

TOWNSLEY, P. M., R. A. RICHY & P. С TRUS- 
SELL, 1 966, The laboratory rearing of the ship- 
worm, Bankia setacea (Tryon). Proceedings of 
the National Shellfisheries Association, 56: 49-52. 



VACCA, L. L, 1985, Laboratory manual of his- 
tochemistry. Raven Press, New York, xvii + 578 
pp. 

WINCKWORTH, R., 1930, Notes on nomenclature. 
Proceedings of the Malacological Society of Lon- 
don, 19(1): 14-16. 

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matica (Pilsbry) new to Japan. Venus, Japanese 
Journal of Malacology, 33(3): 116. 

Revised Ms accepted 1 August 1991 



MALACOLOGIA, 1992, 34(1-2): 25-32 

EMBRYONIC DEVELOPMENT OF BIOMPHALARIA GLABRATA 

(SAY, 1818) (MOLLUSCA, GASTROPODA, PLANORBIDAE): 

A PRACTICAL GUIDE TO THE MAIN STAGES 

Toshie Kawano (Carney), 1 Kayo Okazaki 2 & Liliane Ré 1 

ABSTRACT 

The morphology of the gastropod mollusk Biomphalaria glabrata (Say, 1818) from the stage 
proceeding the first egg cleavage to the hippo stage is presented. The morphogenetic changes 
along its development were followed by in vivo observation and by cell lineage studies, with 
description of the origin and function of the main structures that characterize the embryonic and 
larval phases. 

Key Words: Biomphalaria glabrata, Planorbidae, cleavage, embryonic, development. 



INTRODUCTION 

The studies of cell lineages in mollusks are 
of great interest for research in experimental 
embryology. The first ones date back to the 
end of the last century, when different species 
were investigated by many authors, such as 
Neritina fluviatilis by Blochmann (1882), Um- 
brella mediterránea by Heymons (1893), 
Ischnochiton sp. by Heath (1899), Planorbis 
trivolvis by Holmes (1 900), Physa fontinalis by 
Wierzejski (1905), Littorina obtusata by Dels- 
man (1912, 1914), Limnaea stagnalis by 
Raven (1946, 1958) and Verdonk (1965) and 
Dentalium sp. by Dongen (1976). Carney & 
Verdonk (1970) carried out studies on cell lin- 
eage of the gastropod Biomphalaria glabrata 
(Say, 1818); they analyzed its embryonic 
stages, with emphasis on the cephalic region, 
from the first egg cleavages to the veliger 
stage. 

This guide was prepared in order to de- 
scribe the various stages of the embryonic 
and larval development of B. glabrata and 
was also especially planned for those initiat- 
ing research activities or attending or giving 
lectures on embryology. It covers the stages 
between the first egg cleavages, which are 
characterized by the spiral (helicoidal) type, 
and the trochophore and veliger stages, all of 
them occurring within the egg capsule. 

MATERIAL AND METHODS 

Egg masses of Biomphalaria glabrata (Say, 
1818) were picked up in Belo Horizonte, State 



of Minas Gerais, Brazil, and maintained for 
several years in the laboratory, at 25°C, in a 
climatic chamber. 

The developmental stages were followed 
and photographed in vivo by using a Zeiss 
photomicroscope. From the first to the fourth 
cleavages, the stages were photographed at 
intervals of 1 to 30 minutes. From the fourth 
cleavage on, the time intervals were of 5 
hours. From the early trochophore stage on, 
the larvae were immobilized with ethyl ether 
for photography. These time intervals were 
decided upon based on data obtained in pre- 
vious observations of the embryonic develop- 
ment of this species. 

Stage 1 (Fig. 1B) was considered as the 
starting point for the determination of the em- 
bryo age due to the variation of the time in- 
tervals elapsing between egg laying and first 
egg cleavage, as pointed out by Raven (1946). 
Due to the asynchrony of egg division in the 
same egg-mass, zero time was considered to 
be the time when 50% of the eggs were in 
stage 1. 

For the cell lineage study, the embryos were 
prepared according to the technique de- 
scribed by Verdonk (1965), which basically 
consists of the following steps: the embryos 
were decapsulated, fixed with 0.75% AgN0 3 
for a few seconds until the cell contours be- 
came sharp, dehydrated in an alcohol series, 
cleared with xylene, and mounted with Per- 
mount between a slide and a coverslip. Em- 
bryos and larvae were drawn using a Zeiss 
camera lucida, and the nomenclature adopted 
was that of Conklin (1897). 



1 Serviço de Zoonoses Parasitarias e Parasitología, Instituto Butantan Cx. Postal 065, CEP 01000, Säo Paulo, SP, BRAZIL. 
2 Divisáo de Radiobiología, Departamento de Aplicaçoes em Ciencias Biológicas, Instituto de Pesquisas Energéticas e 
Nucleares, Comissáo Nacional de Energía Nuclear/SP, Sao Paulo, SP, BRAZIL 



25 



26 



f P-b. 




KAWANO, OKAZAKI & RÉ 
a. P. 






50jir 



c.c. 



mm m m # 

ч fi 7 ft a 



• 





13 






17 МЦ£ 19 j 20 

FIG. 1. Early development of Biomphalaria glabrata. (1A) Undivided egg with two polar bodies. (1B) Begin- 
ning of first cleavage, with appearance of the cleavage furrow in the animal pole (0 hr). (2) Appearance of 
the cleavage furrow, after 3 min. (3) After 4 min. (4) Two blastomeres attached to each other by a small 
cytoplasmic area, after 10 min. (5), (6) and (7) the blastomeres present a gradually larger contact surface: 
after 21 , 26 and 38 min., respectively, (8) Emergence of the cleavage cavity after 45 min. (9) Cleavage cavity 
appearing more clearly, after 75 min. (10) Second cleavage, after 80 min. (11) Four blastomere stage, after 
91 min. (12) Beginning of the appearence of the cleavage cavity, at 98 min. (13) After 103 min. (14) After 
123 min. (15) Almost rounded surface of the blastomeres, at 134 min. (16) Third cleavage with formation of 
the first quartette of micromeres, at 160 min. (17) At 165 min. (18) At 173 min. (19) At 181 min. (20) Fourth 
cleavage, at 230 min. p.b. = polar body, a.p. = animal pole, v. p. = vegetative pole, c.c. = cleavage cavity, 
m = micromere, M = macromere. 



RESULTS 

Table 1 shows the main stages of the em- 
bryonic development of B. glabrata, from the 
beginning of the first cleavage to the hippo 
stage. 

The morphological features of B. glabrata 
during the main stages of its embryonic and 
larval development are illustrated in Figures 
1-3. The embryonic and larval cells are sche- 
matically presented in Figure 4. 

Figure 1 presents a sequence of morpho- 
genetic alterations covering the period of time 
between the zygote (or fertilized egg) and the 



fourth cleavage stage. Stage 1A shows the 
zygote before the first cleavage; two polar 
bodies (p.b.) can be seen at the upper apical 
region, exactly in the animal pole. The diam- 
eter of a viable egg is approximately 100 цт 
long. Stages from 1 to 19 are equivalent to 
those described for Limnaea stagnalis by 
Raven (1946). The first cleavage furrow ap- 
pears during stage 1B in the animal pole. This 
phase corresponds to the beginning of the 
first cleavage, which is total and passes 
through the animal and vegetative poles. 
Stages 2 and 3 occur approximately 3 or 4 
minutes after stage 1B, respectively, with 



EMBRYONIC DEVELOPMENT OF BIOMPHALARIA 



27 



TABLE 1 . Main stages of the embryonic development of B. glabrata at 25°C. 





Time interval 


Number of stage 


Embryonic stage 


between observations 


in figures 


Beginning of the 1st cleavage 





1B 


2nd cleavage 


80 min. 


10 


3rd cleavage 


160 min. 


16 


4th cleavage 


230 min. 


20 


blástula 


15hrs. 


21 


gastrula 


26 hrs. 


23 


early trochophore 


43 hrs. 


25 


late trochophore 


66 hrs. 


27 


early veliger 


96 hrs. 


28 


late veliger 


120 hrs. 


29 


hippo stage 


144 hrs. 


30 



cleavage furrow becoming gradually more 
predominant; the cleavage of the animal pole 
region is more rapid than that of the vegeta- 
tive one. At stage 4 the cleavage furrow al- 
most completely divides the zygote into two 
blastomeres, both of them presenting a well- 
rounded surface and linked to each other by 
only a small cytoplasmic bridge. This phase 
occurs approximately 10 minutes after stage 
1B (Fig. 1, stage 4; Fig. 4, stage 31). After 
stage 5, at approximately 21 minutes from the 
beginning of zero time, the two blastomeres 
approach each other, increasing their contact 
surface, that is, they flatten against each 
other, with the formation of a separating blas- 
tomeric membrane (Fig. 1, stage 5). During 
stages 6 and 7 (Fig. 1), the boundaries of the 
blastomeres can hardly be defined; these 
stages occur approximately 26 and 38 min- 
utes after stage 1 B, respectively. 

The cleavage cavity, the function of which is 
osmotic regulation, can be seen between the 
two blastomeres at stage 8, approximately 45 
minutes after stage 1B (Fig. 1). This cavity 
increases in size, and both blastomeres have 
an egg shape (Fig. 1, stages 8 and 9). 

The beginning of the second cleavage (Fig. 
1, stage 10) occurs 80 minutes after the first 
cleavage. Because the blastomeres do not di- 
vide synchronously, the cleavage furrow of 
one of the blastomeres appears before the 
other, so that this stage presents an asym- 
metrical shape (Fig. 1, stage 10). This cleav- 
age is meridional and total. 

During stage 11 (Fig. 1), after approxi- 
mately 91 minutes, the blastomeres show a 
rounded shape but are not on the same plane 
when observed obliquely; that is, blastomeres 
A and С (Fig. 4, stage 32) are linked to each 
other by the furrow in the animal pole, and 



blastomeres В and D by the furrow in the veg- 
etative pole. 

At stage 12 (Fig. 1), after approximately 98 
minutes, the cleavage furrow linking the alter- 
nate blastomeres in the animal and vegeta- 
tive poles of the egg can be seen more clearly 
(Fig. 1, stage 12; Fig. 4, stage 32). At stages 
13 (after 103 min) and 14 (after 123 min) (Fig. 
1), the blastomeres are already more closely 
joined, and the cleavage cavity between them 
begins to appear. Material from the cleavage 
cavity then starts coming out, with contraction 
of the whole egg causing a visible reduction in 
diameter (Fig. 1, stage 15). 

The third cleavage is laeotropic and occurs 
in the subequatorial plane of the egg, with the 
formation of the first micromere quartette (1a 
to 1 d), approximately 1 60 minutes after the be- 
ginning of the first cleavage (Fig. 1, stage 16; 
Fig. 4, stage 33). A gradual increase of the 
cleavage cavity is observed during stages 17 
(after 165 min), 18 (after 173 min) and 19 
(Fig. 1). 

Stage 20 (Fig. 1 ) illustrates the egg at the 
fourth cleavage stage, which is dexiotropic. 
The fourth cleavage is subequatorial and oc- 
curs approximately 230 minutes after stage 
1B (Fig. 4, stage 34), originating the second 
micromere quartette (2a to 2d). 

The embryo reaches the blástula stage ap- 
proximately 1 to 23 hours after the 1 st cleav- 
age (Figs. 21, 22). 

Stages 35 and 36 (Fig. 4) show the embryo 
with about 64 and 130 cells, respectively, and 
presenting a set of cells forming a cross-like 
figure (1a 11 to 1d 11 , 1a 12 to 1d 12 and 2a 11 to 
2d 11 cell lineages) in the animal pole, which 
gives origin to almost the whole head region. 

Gastrulation occurs about 24 to 39 hours 
after the first cleavage (Fig. 2, stages 23, 24A, 



28 



KAWANO, OKAZAKI & RE 



• 





21 



22 



23 



\ 

50 urn 





v.p. 



24A 



24B 




FIG. 2. Different stages of development of Biomphalaria glabrata. (21) Blástula, 15 hrs. after 1st cleavage. 
(22) Blástula, 21 hrs. and 30 min. after 1st cleavage (23) Gastrula, 26 hrs. after 1st cleavage. (24A) Late 
gastrula as seen from the vegetative pole after 34 hrs. (24B) Lateral view of the 34 hrs. gastrula stage. (25) 
Early trochophore, after 43 hrs. a. p. = animal pole, v.p. = vegetative pole. 



24B; Fig. 4, stages 37 and 38). Flattening of 
the vegetative pole region towards the animal 
pole then occurs, followed by the invagination 
of this region, with the formation of a spherical 
opening, the blastopore, which then becomes 
gradually smaller. 

The early trochophore stage appears ap- 
proximately 40 to 65 hours after the first cleav- 
age, and is characterized by the first larval 
movements (Fig. 2, stage 25; Fig. 3, stage 26) 
by means of the ciliated cells of the prototroch, 
which divides the larva in the pretrochal re- 
gion, characterized by the presence of an api- 
cal plate, head vesicle and cephalic plates, 
and the posttrochal region, characterized by 
the presence of a shell gland, stomodeum and 
foot (Fig. 4, stage 39). The prototroch shows 
a double row of cells, an upper one formed by 
2d 111 , 2b 112 , 1a 21 , 1a 22 , 1b 21 and 1b 22 cells, 
and a lower ciliated one consisting of 2b 211 
and 2b 121 cells and of cells descending from 
2b 122 and 2b 212 cells (Fig. 4, stage 39). 

The late trochophore stage occurs approx- 
imately 65 to 80 hours after the first cleavage, 
with the larva having a slightly elongated, kid- 
ney-like shape. In this phase, cells responsi- 
ble for the formation of the head and foot can 
be seen in its anterior region, and in the dor- 



sal region one can see the shell gland (Fig. 3, 
stage 27). Stage 40 (Fig. 4) shows a greater 
development of the prototroch, leading to 
more active larval movements. The pretrochal 
region, located above the prototroch, is 
formed by a set of cells that gives rise to such 
larval structures as the apical plate (1a 111 to 
1d 11 \ 1a 1121 , 1b 1121 and 1b 1211 ), cephalic 
plates (cells located laterally to the apical 
plate), and the head vesicle (1c 21 , 1d 21 , 1c 22 , 

1(j 22 1a 122 1c 122 1d 122 -,¿1211 -,¿12^ 

2a 11 , 2c 11 , 2d 11 ), which in turn will form the 
head. In the posttrochal region, below the pro- 
totroch, are the stomodeum (M), which will 
give origin to the mouth, and cells that will 
form the foot and the shell gland. 

The time interval between 80 and 100 
hours after the first cleavage corresponds to 
the early veliger stage. The shell and foot are 
then more developed (Fig. 3, stage 28). The 
prototroch has evolved into the velum (Fig. 4, 
stage 41). In this stage, the shell gland shifts 
towards the right side. 

The veliger stage occurs about 1 20 hours 
after the first egg cleavage. The eyes become 
more visible, and the elevation of the tentacle 
regions can be observed, as well as the 
mouth, foot and shell (Fig. 3, stage 29). Stage 



EMBRYONIC DEVELOPMENT OF BIOMPHALARIA 



29 




62 pm 



H^ 



F^ 




29 




30 M 



FIG. 3. Different stages of development of Biomphalaria glabrata. (26) Early trochophore, 55 hrs. after 1st 
cleavage. (27) Trochophore, after 66 hrs. (28) Early veliger, after 96 hrs. (29) Veliger, after 1 20 hrs. (30) 
Hippo stage, 144 hrs. after first cleavage. E = eyes, F = foot, H = head, M = mouth, S = shell, Pr = 
prototroch. 



42 (Fig. 4) shows that the apical plates and 
head do not undergo any changes, while the 
cephalic plates go on dividing to form the ten- 
tacles and eyes. 

The hippo stage is a phase of larval devel- 
opment that occurs within the egg capsule ap- 
proximately 144 hours after the first cleavage. 
At this stage, the eyes and tentacles in the 
pretrochal region are already well developed, 
and in the posttrochal region the foot has 
grown in size and is much more differentiated. 
The shell starts coiling and covers almost the 
whole body (Fig. 3, stage 30). At 25°C, the 
young snail hatches from the egg capsule be- 
tween the sixth and ninth day after the first 
cleavage. 



DISCUSSION 

Due to their semi-transparence and easy 
maintenance in the laboratory, the eggs of B. 
glabrata represent a suitable material for 
studies on embryology, allowing a good in 
vivo observation. At egg-laying, its egg- 
masses already contain fertilized eggs and, 
after approximately 30 minutes (at 25°C), the 
emission of the first polar body occurs, as a 
result of the first meiotic division. Approxi- 
mately 60 minutes later, the second polar 
body emerges. Both polar bodies remain in 
the animal pole during the first cleavages. 
Within 60 minutes after the emission of the 
second polar body, the fusion of the male and 



30 



KAWANO, OKAZAKI & RÉ 




'b n¿+ ~l^ 



35 



36 



FIG 4 Outline of cell boundaries on developing eggs of Biomphalaria glabrata. (31) 1st cleavage with 2 
blastomeres. (32) 2nd cleavage with 4 blastomeres, 80 min. after 1st cleavage. (33) 3rd cleavage with 1st 
quartette of micromeres (1a to 1d) and with 8 blastomeres, after 160 min (34) 4th Ravage with 2nd 
quartette of micromeres (2a to 2d) and 12 blastomeres, after 230 min. (35 Blastu a i stage with 64 blas- 
tomeres, showing the cross figure at the animal pole after 15 hrs. (36). Blástula with 130 cells (animal pole) 
after 21 hrs. and 30 min. (37). Gastrula, after 26 hrs. (38) Gastrula after 34 hrs. (39) Early trochophore after 
34 hrs (40) Trochophora after 66 hrs. (41) Early veliger after 96 hrs. (42) Veliger stage after 120 hrs. M - 
mouth! A.P. = apical plate, H.V. = head vesicle, C.P. = cephalic plate, V = velum, T = Tentacle, E - 
ey eSi F = foot, Pr. = prototroch. 



EMBRYONIC DEVELOPMENT OF BIOMPHALARIA 



31 




122 



32 



KAWANO, OKAZAKI & RÉ 



female pronuclei occurs, the egg now being 
ready for the first cleavage (Carney, 1968). 

Except for cephalopods, the cleavage pat- 
tern of mollusks is of the spiral type; that is, 
blastomere cleavage is oblique in relation to 
its axis and is of the determinative type, be- 
cause the different egg regions are going to 
form the future organs (Raven, 1958). 

The developmental stages described here 
are similar to those of Limnaea stagnalis, with 
some differences. Cleavage is laeotropic or 
reverse in B. glabrata (Carney & Verdonk, 
1970) and dexiotropic in L. stagnalis (Ver- 
donk, 1965). The differentiation of the type of 
cleavage occurs according to the orientation 
of the division spindle. When this is oblique in 
relation to the axis of the egg in clockwise 
direction, cleavage is dexiotropic, and when 
the spindle is oblique but in a counterclock- 
wise direction, cleavage is laeotropic. The 
first indication of the orientation of the cleav- 
age spindle in B. glabrata occurs during the 
third cleavage (Carney & Verdonk, 1970). 

In the trochophore stage, the shell gland is 
shifted to the right side in B. glabrata (Carney, 
1 968) and to the left side in L. stagnalis (Ver- 
donk, 1965). 

When the helicoidal shell of an adult of L. 
stagnalis is placed with the apex turned up, it 
can be seen that the opening is towards the 
right side. In B. glabrata, with a planispiral 
shell, it is difficult to determine the direction of 
the shell opening, except for its shift in a di- 
rection opposite to that of L. stagnalis during 
larval development. It may be concluded, 
therefore, that the cleavage pattern and shell 
opening of B. glabrata are laeotropic. 



ACKNOWLEDGEMENTS 

Thanks are due to Dr. Kaoru Hiroki for the 
critical reading of this paper and to Miss. Vera 
Helena Monezi for typing this manuscript. 



LITERATURE CITED 

BLOCHMANN, F., 1 882, Ueber die Entwicklung der 
Neritina fluviatilis. Mull. Zeitschrift für Wissen- 
shaftliche Zoologie. Leipzig. 36: 125-174. 

CAMEY, T, 1968, Estágios iniciáis do desenvolvi- 
mento embrionario da Biomphalaria glabrata 
(Say, 1818). Tese de Doutoramento. Belo Hori- 
zonte, MG, Brasil. 

CAMEY, T. & N. H. VERDONK, 1970, The early 
development of the snail Biomphalaria glabrata 
(Say) and the origin of the head organs. Nether- 
lands Journal of Zoology, Leiden, 20(1 ): 93-1 21 . 

CONKLIN, E. G., 1897, The embryology of Crepi- 
dula. Journal of Morphology, 13: 1-226. 

DELSMAN, H. C., 1912, Ontwikkelingsgeschiede- 
nis van Littorina obtusata. Thesis, Univ. of Am- 
sterdam, Amsterdam, The Netherlands. 

DELSMAN, H. C., 1914, Cf. Entwicklungsge- 
schichte von Littorina obtusata. Tijdschrift Neth- 
erlandsche Dierkundige Vereeniging. Leiden, 
13(2): 170-340. 

DONGEN, С. A. M. van, 1976, The development of 
Dentalium with special reference to the signifi- 
cance of the polar lobe. Utrecht, Rijksuniversiteit, 
92 pp. PhD. Thesis. 

HEATH, H., 1899, The development of Ischnochi- 
ton. Zoologische Jahrbucher, Anatomie, 12: 567- 
656. 

HEYMONS, R., 1893, Zur Entwicklungsgeschichte 
von Umbrella mediterránea Lam. Zeitschrift für 
Wissenschaftliche Zoologie, 36: 245-298. 

HOLMES, S. J., 1900, The early development of 
Planorbis. Journal of Morphology. 16: 369-458. 

RAVEN, С. P., 1 946, The development of the egg of 
Lymnaea stagnalis L. from the first cleavage till 
the trochophore stage, with special reference to 
its "chemical embhology". Archives Néerlan- 
daises de Zoologie, Leiden, 7(3-4): 353-434. 

RAVEN, С. P., 1958, Morphogenesis: the analysis 
of molluscan development. London, Pergamon 
Press. 31 1 pp. (International series of mono- 
graphs on pure and applied biology. Division Zo- 
ology, v. 2) 

VERDONK, N. H., 1965, Morphogenesis of the 
head region in Lymnaea stagnalis L. Utrecht, 1 33 
pp. PhD. Thesis. 

WIERZEJSKI, A., 1905, Embryologie von Physa 
fontinalis L. Zeitschrift für Wissenchaftliche Zool- 
ogie, 83: 502-706. 

Revised Ms. accepted 31 August 1991 



MALACOLOGIA, 1992, 34(1-2): 33-61 

ALLOZYME FREQUENCIES IN ALBINARIA (GASTROPODA: PULMONATA: 
CLAUSILIIDAE) FROM THE IONIAN ISLANDS OF KEPHALLINIA AND ITHAKA 

Th. С M. Kemperman & G. H. Degenaars 1 

Systematic Zoology Section, Population Biology Department, Rijksuniversiteit Leiden, 
c/o National Museum of Natural History, P.O. Box 9517, 2300 RA Leiden, The Netherlands 



ABSTRACT 

In 42 samples comprising 11 of the 1 2 subspecies of the four species of Albinaria occurring 
on the islands of Kephallinia and adjoining Ithaka, Greece, allozyme variation was studied at 28 
gene loci by starch gel electrophoresis. To serve as outgroups, single samples of two Cretan 
Albinaria species and both an Albinaria and an Isabellaria species from the Péloponnèse were 
included. The various taxa varied at 0.0% to 35.7% of the loci, with an average of 12.2%. In 71% 
of the populations, a deficiency of heterozygosity was found. Taxa differ in the number of 
populations that must be analysed to reach a point where further addition of populations does 
not increase the genetic variation. The results show relatively low genetic distances between the 
taxa from Kephallinia; between the subspecies of the two non-endemic species hardly any 
allozyme differentiation is found. In the two endemic species, however, the subspecies are less 
similar to each other and show conspicuous genetic resemblances with different non-endemic 
species. The taxon from Ithaka differs markedly from all Kephallinian taxa and thus geographical 
isolation seems to be an important factor with respect to genetic differentiation. There is evi- 
dence for introgressive hybridization. In at least one case, we found indications for hybrizymes. 
A Hennigian analysis did not result in a resolved phylogeny. The present classification of species 
and subspecies, based on morphological and biogeographical data, is maintained only with 
some doubt. 

Key words: systematics, allozymes, genetic variation, hybridization, electrophoresis, Albina- 
ria, Pulmonata. 



INTRODUCTION 

General Notes 

Within the context of a study on the sys- 
tematics and evolutionary history of the pul- 
monate genus Albinaria Vest, 1867 (Gitten- 
berger, in press), special attention has been 
given to the Albinaria species of the neighbor- 
ing Ionian islands Kephallinia and Ithaka. The 
aims of this subproject are (1) to unravel the 
pattern of taxonomic relationships among the 
various taxa and (2) to get some insight into 
the population genetics and the evolutionary 
history of these pulmonates. This goal is ap- 
proached by using shell morphology (Kem- 
perman & Gittenberger, 1988), radula mor- 
phology (Kemperman et al., in prep.), genital 
morphology, distributional patterns, and allo- 
zyme variation (Degenaars, in press). In ad- 
dition to the latter preliminary report, the 



present paper deals with the biochemical data 
in more detail. 

The value of electrophoretic techniques for 
systematic study is widely recognized (Avise, 
1974; Buth, 1984; Richardson et al., 1986; 
Ferguson, 1988; Nevo, 1988; Murphy et al., 
1990). Allozyme variation among samples of 
different populations provides a data-set that 
may be considered virtually independent of 
the morphological data (Avise, 1974; Ember- 
ton, 1988). Albinaria has been the subject of 
electrophoretic studies before. Ayoutanti et al. 
(1 988) reported on an allozyme analysis of 23 
Albinaria populations from 12 Aegean is- 
lands, including Crete. Using 27 loci, these 
authors tried to unravel the genetic affinities 
among a group of Albinaria species from 
many islands with generally only one or two 
Albinaria taxa each. We limited our study to 
the rich Albinaria fauna of only two, neighbor- 
ing islands. The studies show some overlap in 



'Present address: Hugo de Vries Lab., University of Amsterdam, Kruislaan 318, 1098 SM Amsterdam, The Netherlands. 

33 



34 



KEMPERMAN & DEGENAARS 



that we used, for example, Cretan material as 
outgroup and partly analyzed the same en- 
zyme systems. 

Geography of Kephallinia and Ithaka 

The largest island, Kephallinia (Fig. 1), 
measures about 50 km from north to south 
and somewhat more than 20 km across the 
middle of the main part of the island (that is, 
without the Paliki Peninsula), which is domi- 
nated by a mountain ridge, reaching 1 ,628 m 
altitude. All over the island are areas with 
limestone rocks and/or rock faces inhabited 
by Albinaria. Ithaka is only 23 km long and up 
to 6 km wide. It consists of two large penin- 
sulas joined by a narrow, high isthmus. 

The present 1 30 m isobath can probably be 
looked upon as the Pleistocene shore line of 
approximately 18,000 B.P., when sea level 
reached its lowest point (Andel & Shackleton, 
1982: 447-448) during the Wurm. Judging 
from the isobaths on Admiralty Chart no. 203 
(1988) and data given by Bird (1984: 81, fig. 
41), we may assume that between approxi- 
mately 29,000 and 12,000 years ago there 
was a minimally 1 km-broad land connection 
between Kephallinia and Ithaka, running from 
Cape Dichalia, northeast of Sami on Kephal- 
linia, and the south-southwest coast of Ithaka. 
At present, this is a stretch of approximately 4 
km sea water. North of this area, there have 
not been relatively recent land bridges, be- 
cause there the sea between Kephallinia and 
Ithaka is over 1 45 m deep, with a depth of 1 82 
m near Fiskardo, northeastern Kephallinia. 
Thus, Ithaka and Kephallinia have been sep- 
arate islands for at least 12,000 years. 

Because the sea between Lixouri and Ar- 
gostoli is at present only 25 m deep, we have 
to accept that during glacial sea-level lower- 
ings the entire Gulf of Argostoli between the 
Paliki Peninsula and the main island was 
land. 

Distribution of Albinaria on Kephallinia 
and Ithaka 

In Figure 1 , the ranges of the Albinaria taxa 
from Kephallinia and Ithaka are indicated, 
based on our data from approximately 800 
localities (Kemperman, in prep. A). Different 
shadings indicate different species, whereas 
broken thick lines separate the ranges of sub- 
species. Based on classical conchological 
characters, four species are distinguished, 
each with at least two subspecies. 



On Kephallinia, two species are endemic, 
both with limited ranges, namely A. jónica and 
A. adrianae. On Ithaka no endemic Albinaria 
species have been found. 

Albinaria contaminata ranges on the Ionian 
islands from Lefkas to Zakynthos and on the 
mainland from the southern Epirus to the 
province of Phocis. It is the only Albinaria 
known from Ithaka. On Kephallinia it is the 
most widely distributed species, occurring al- 
most everywhere, except for places along the 
west coast and the Paliki Peninsula. The sec- 
ond widespread species is A. senilis, occur- 
ring on all the Ionic islands, except for Za- 
kynthos, and from the Epirus southward to the 
mainland opposite Lefkas. 

MATERIALS AND METHODS 
Species Sampled 

In total, 46 populations are examined elec- 
trophoretically, 42 of which were from Kephal- 
linia and Ithaka, representing the following 
taxa (between square brackets, the number of 
investigated populations: individuals is given): 
Albinaria contaminata contaminata (Ross- 
mässler, 1835) [7: 128], A. с incommoda 
(Boettger, 1 878) [4: 36], A. с odysseus (Boett- 
ger, 1878) [4: 53], A. с liebetruti (Charpentier, 
1852) [1: 18], A. senilis senilis (Rossmässler, 
1836) [12: 140], A. s. flavescens (Boettger, 
1878) [2: 27], A. s. kolpomyrtensis Kemper- 
man & Gittenberger, 1990 [1 : 15], A. adrianae 
adrianae Gittenberger, 1979 [4: 52], A. a. du- 
bia Gittenberger, 1979 [1 : 19], A. jónica assi- 
cola Kemperman & Gittenberger, 1 990 [2: 36], 
and A. j. jónica (Pfeiffer, 1866) [2: 16]. 

All specimens from Kephallinia and Ithaka 
were collected by the first author in March 
1988. See Appendix for detailed information 
on the localities indicated in Figure 2. 

As outgroups we used populations of A. 
teres nordsiecki Zilch, 1977 [1 : 8] and A. re- 
bel! Wagner, 1924 [1:13] from Crete, and Al- 
binaria spec. [1 : 20] and Isabellaria edmundi 
Gittenberger, 1987 [1: 20] from the Pélo- 
ponnèse (Parnon) (see Appendix). The out- 
group material was collected by E. Gitten- 
berger in April and June 1988. 

In total 613 individuals are examined, an 
average of 13 specimens per population. 

Collecting and Transport 

Populations were sampled at a location lim- 
ited to 1 to 20 m 2 . The snails could be col- 



ALLOZYME FREQUENCIES IN ALBINARIA 



35 



Ithaka 




Albinaria species and subspecies 

I I contaminate 
С = с. contaminate 

I = с incommoda 

О = с. odysseus 

L = с. liebctruti 

M = с. muraria 



senilis RsS$^ jónica 

S = s. senilis JA = j. assicola 

F = s. flavescens JJ =j. jónica 

К = s. kolpomyrtensis 



%4 adrianae 
A = a. adrianae 

D = a. dubia 



10 



km 



FIG. 1. Map of Kephallinia and Ithaka, with distributions of Albinaria taxa. 



36 



KEMPERMAN & DEGENAARS 



- A. 

- A. 

- A. 

- A. 

- A. 

- A. 



contaminata contaminate 

с incommoda 

с. odysseus 

с. liebetruti 

senilis senilis 

s. flavescens 

- A. s. kolpomyrtensis 

- A. jónica jónica 
-A. j. assicola 

- A. adrianae adrianae 

- A. a. dubia 

- Albinaria hybrids 




FIG. 2. Map of Kephallinia and Ithaka, with collection sites of material used in allozyme studies. 



lected easily, without any special device. 
Forced into aestivation, they are not very de- 
manding during transport. Plastic sandwich- 
bags with some dry paper wipes proved to be 
adequate for storage of several weeks. 

Electrophoresis 

Complete animals, including shells, were 
ground in two drops of distilled water in ice- 



cooled porcelain cups. Wicks (Whatman no. 3 
filter paper; 2x5 mm) were used to apply the 
homogenate on the gels. After 15 initial run- 
ning minutes at approximately 16 V/cm, the 
wicks were removed and the electric field was 
raised to approximately 19 V/cm. Running 
times depended on the buffer system that 
was used (Table 1). The gel/buffer system 
was ice-cooled. The complete set of enzyme 
systems of a specimen was tested, usually in 



ALLOZYME FREQUENCIES IN ALBINARIA 



37 



TABLE 1 . Enzymatic systems and loci scored for electrophoretic analysis, with buffer systems employed 
and stain references. 



Enzymatic system (E.C.) 



Loci 



Migration 



Buffer* 



Stain reference 



Adenylate kinase (2.7.4.3) 
Aldehyde oxidase (1.2.3.1) 
Alkaline phosphatase (3.1.3.1.) 
Aspartate aminotransferase (2.6.1.1) 
a-Esterase (3.1.1.1) 
Glucose-6-phosphate dehydr. (1.1.1.49) 

Glucophosphate isomerase (5.3.1.9) 
a-Glycerophosphate dehydrog. (1.1.1.8) 
Hexokinase (2.7.1.1) 
L-lditol dehydrogenase (1.1.1.14) 
Isocitrate dehydrogenase (1.1.1.42) 
Lactate dehydrogenase (1.1.1.27) 
Leucine aminopeptidase (3.4.11.1) 
Malate dehydrogenase (1.1.1.37) 
Mannose phosphate isomerase (5.3.1.8) 
NADH dehydrogenase (1.6.99.3) 
Phosphoglucomutase (5.4.2.2) 
Phosphogluconate dehydrog. (1.1.1.43) 
Superoxide dismutase (1.15.1.1) 
Xanthine dehydrogenase (1.2.1.37) 



Ak 


anodic 


TC7 


Ao 


anodic 


Poulik 


Aph 


anodic 


TC7 


Aat-1,2 


cathodic/anodic 


TEB9 


Est-1,2,3 


cathodic/anodic 


TC7 


G6pd 


anodic 


TEB9 


Gpi 


anodic 


Poulik 


a-Gpdh-1,2 


anodic 


TEB8 


Hk 


anodic 


TEB8 


Idh 


anodic 


TEB9 


lsdh-1,2 


anodic 


TC7 


Ldh 


anodic 


TEB8 


Lap 


anodic 


TEB8 


Mdh-1,2 


cathodic/anodic 


TC7 


Mpi 


anodic 


TEB9 


Nadd-1,2 


anodic 


Poulik 


Pgm 


anodic 


Poulik 


Pgd 


anodic 


TEB9 


Sod-1,2 


anodic 


TEB8 


Xdh 


anodic 


TEB8 



Shaw& Prasad (1970) 
Ayalaetal. (1972) 
Shaw& Prasad (1970) 
Selander et al. (1971) 
Shaw& Prasad (1970) 
Shaw & Prasad, mod. 

Brewer (1970) 
Shaw& Prasad (1970) 
Shaw& Prasad (1970) 
Shaw& Prasad (1970) 
Shaw& Prasad (1970) 
Shaw& Prasad (1970) 
Shaw& Prasad (1970) 
Shaw & Prasad (1970) 
Shaw& Prasad (1970) 
Nichols et al. (1973) 
Menken (1980) 
Shaw & Prasad (1970) 
Shaw& Prasad (1970) 
Shaw & Prasad (1970) 
Shaw& Prasad (1970) 



*) TC 7 = Tris— Citrate, pH 7, running time (rt) 2 h (Shaw & Prasad, 1970); Poulik = Poulik, gel pH 8.75-8.9 & electrode 
pH 7.6-8.0, rt 3 h (Poulik, 1957); TEB 8 = Tris— EDT A— borate, pH 8, rt 3.5 h (Shaw & Prasad, 1970); TEB 9 = 
Tris— EDTA— borate, pH 9, rt 4.5 h (Ayala et al., 1973). All at 350 V or 35 mA. 



two days, for which the homogenate had to be 
stored overnight at -80°C. For references 
concerning the histochemical staining proce- 
dures, see Table 1 . All samples were scanned 
for 28 loci from 20 enzyme systems. In Table 
1 a list of the enzyme systems is given, with 
the abbreviations used in the present article. 
Because of the risk of a substantial decline in 
detectability of the weaker enzyme systems, 
we analyzed freshly prepared homogenates 
only. 

Data Analyses 

Scoring of the loci resulted in a "single-in- 
dividual" genotype dataset. Doubtful pairs of 
bands were by convention scored as single 
bands, and in case of a doubtful position of a 
single band the alternative with the highest 
frequency was accepted. Data analyses were 
performed with the BIOSYS-1 computer pro- 
gram, release 1.7 (Swofford & Selander, 
1989). To assess the degree of genetic vari- 
ability, we calculated per population and per 
subspecies the mean observed heterozygos- 
ity per locus (Ho), the mean heterozygosity 
per locus based on Hardy-Weinberg expecta- 
tions (He), the accompanying standard devi- 
ations, as well as the percentage of polymor- 



phic loci (Pp and Pt). Also allelic frequencies 
were estimated per population and per sub- 
species. In the latter case, populations with 
morphologically determined hybrids have not 
been taken into account. On the basis of the 
allele frequencies, the genetic similarity coef- 
ficients were computed for each pair of pop- 
ulations or taxa, according to Nei's (1972) ge- 
netic distance (D) and Rogers' (1972) genetic 
distance (R). On the resulting genetic dis- 
tance coefficient matrices, a UPGMA cluster 
analyses (Sneath & Sokal, 1973) was per- 
formed. 

To achieve a phylogeny, we also carried 
out a Hennigian analysis following Richard- 
son et al. (1986: 337), Mooi (1989) and Em- 
berton (1990). We used the computerpro- 
gram HENNIG86 (Farris, 1988). 

The ingroup is not polyphyletic, but consists 
of a representative sample of subspecies of 
the wide-spread species A. contaminata and 
A. senilis and all the known subspecies of 
the endemic species A. jónica and A. adrianae. 
It is hypothesized that these species form a 
monophyletic group and that both A. adrianae 
and A. jónica have evolved on Kephallinia, 
sharing a common ancestor with one of the 
other species. Although our ingroup — all 
the Albinaria species and subspecies from 



38 



KEMPERMAN & DEGENAARS 



Kephallinia and Ithaka, except for A. contam- 
inata muraría — does not consist of all the 
known members of the clade, this does not 
invalidate its use as a monophyletic group. In 
principle one can never be sure whether all its 
extinct and extant members are known. Many 
valuable phylogenetic revisions have been 
published in monographs dealing with incom- 
plete ingroups due to geographical or practi- 
cal restrictions. Our outgroup is composed of 
not too closely related congeneric species 
from relatively distant localities (Péloponnèse 
and Crete) and, in the case of Isabellaria ed- 
mundi, a species of a genus of doubtful status 
that is at least very closely related to Albinaria 
(Gittenberger, 1987). 

Classification on Shell Morphology 
and Distribution 

It should be emphasized that we primarily 
classified the various taxa according to shell 
morphology and distribution, following Nord- 
sieck (1977) and Kemperman & Gittenberger 
(1990). From the shell characters we mention 
(1) the sculpture of the protoconch, (2) the 
sculpture of the teleoconch (without cervix), 
(3) the cervical sculpture, (4) the cervical 
structures, (5) the microsculpture, (6) the 
general shell shape, (7) the shell size, (8) the 
apertural shape, (9) the shape of the apertural 
lip, (10) the apertural attachment to the teleo- 
conch, (1 1 ) the shape of the columellaris, (1 2) 
the length of the parietalis, (13) the length of 
the spiralis, (14) the presence of a paralellis, 
(1 5) the shape of the palatalis, (16) the shape 
of the lunella, and (17) the shape of the clau- 
silium. These characters, which proved to 
vary independently among the species and 
subspecies, will be discussed in detail in 
Kemperman (in prep. A). It turned out that A. 
contaminata differs in eight independent char- 
acters from A. senilis, in ten from A. jónica 
and in eleven from A. adrianae. A. senilis dif- 
fers in nine characters from A. jónica and in 
four from A. adrianae, whereas A. jónica dif- 
fers in seven characters from A. adrianae. 
These morphological data, in combination 
with the distributional patterns and the fact 
that intermediate forms are an extreme minor- 
ity found only in a few contact zones lead us 
to consider these forms separate species. 

RESULTS 

Among all taxa, including the outgroups, 
variation was found at 1 6 of the 28 loci (57%), 



namely, Aat-1, Ak, Aph, Est-3, Gpi, Hk, Idh, 
Lap, Ldh, Mdh-1, Mdh-2, Mpi, Nadd-1, Nadd- 
2, Pgd, and Pgm. All the taxa, except for A. a. 
dubia, of which only a single population could 
be analysed, are polymorphic to some extent. 
At up to 35.7% of the loci, with an average of 
12.2% (± 10.4%), variation has been demon- 
strated (Table 2). The observed percentage 
polymorphic loci per taxon (Pt) depends to 
some extent on the number of scanned pop- 
ulations. In a single population, only a part of 
the specific allozyme variation is found. 
Therefore, to estimate the amount of polymor- 
phic loci in a certain taxon, a minimum num- 
ber of populations must be studied. For A. c. 
contaminata and A. s. senilis, we computed 
the percentage polymorphic loci for one pop- 
ulation, for two combined populations, and so 
on, each time adding randomly specimens of 
one more population. This procedure was re- 
peated three times, starting and continuing 
with different populations. The averaged val- 
ues for the different numbers of populations 
are indicated in Figure 3. From this figure it 
becomes obvious that in A. s. senilis from four 
populations on, the growth in variation is very 
low. In A. с contaminata, however, there is 
still no important reduction in increase of vari- 
ation after the addition of six populations. 

The polymorphic loci per population (Pp) 
varies between 0.0% and 1 7.9%, with an av- 
erage of 7.5% (± 4.9%). Two of sixteen pop- 
ulations of A. contaminata and four of five 
populations of A. adrianae proved to be in- 
variable. The averaged values of Pp per 
taxon vary between 0.0% for A. a. dubia and 
10.7% for A. j. jónica. 

In Figure 4, we plotted the Pp of the anal- 
ysed populations of the various species 
against the number of analysed individuals. It 
turned out that there is only a limited relation 
between the number of analysed individuals 
and the Pp of that population (Table 2). 

In all taxa that could be investigated, the 
level of heterozygosity was low (Table 2). 
With Ho = 0.030, A. s. senilis was the most 
variable subspecies. No genetic variation was 
found in the population of A. a. dubia, and 
only very little was found in A. с liebetruti (Ho 
= 0.003), A. с odysseus (Ho = 0.005) and 
A. a. adrianae (Ho = 0.001). In all subspe- 
cies, except for A. s. kolpomyrtensis, the 
mean observed heterozygosity was below the 
mean expected heterozygosity. The differ- 
ences were often considerable, as in A. с 
liebetruti (Ho = 0.003 versus He = 0.017) 
and A. с odysseus (Ho = 0.005 versus He = 



ALLOZYME FREQUENCIES IN ALBINARIA 



39 



TABLE 2. Percentages polymorphic loci per taxon and per population and the mean heterozygosity. For 
abbreviations, see Results; "s" in an abbreviation means standard deviation. *: based on 702, 706-708 
710 and 712 only. 



Taxon/ l ос. nr. 


P, 


Pp 




Pp 


*Pp 


Ho 


sH„ 


He 


sH, 


contaminate 


32.1 




9 


.2 


5.8 


0.013 


0.006 


0.026 


0.013 


713 




14.3 








0.020 


0.012 


0.032 


0.020 


715 




17.9 








0.024 


0.013 


0.039 


0.021 


716 




7.1 








0.005 


0.004 


0.018 


0.015 


721 




10.7 








0.037 


0.024 


0.031 


0.019 


735 




7.1 








0.000 


0.000 


0.019 


0.013 


749 




0.0 








0.000 


0.000 


0.000 


0.000 


759 




7.1 








0.012 


0.008 


0.012 


0.008 


incommoda 


14.3 




4 


.4 


3.4 


0.009 


0.006 


0.014 


0.009 


723 




7.1 








0.022 


0.018 


0.019 


0.015 


727 




7.1 








0.008 


0.006 


0.015 


0.012 


772 




0.0 








0.000 


0.000 


0.000 


0.000 


773 




3.6 








0.009 


0.009 


0.019 


0.019 


liebet rut i 


3.6 


3.6 








0.003 


0.003 


0.017 


0.017 


odysseus 


17.9 




8 


.0 


3.4 


0.005 


0.003 


0.044 


0.021 


764 




3.6 








0.000 


0.000 


0.013 


0.013 


765 




7.1 








0.003 


0.003 


0.012 


0.010 


766 




10.7 








0.015 


0.009 


0.023 


0.014 


770 




10.7 








0.000 


0.000 


0.029 


0.019 


sen i lis 


32.1* 




9 


.9 


4.9 


0.030 


0.010 


0.050 


0.016 


702 




7.1 








0.020 


0.018 


0.020 


0.015 


706 




14.3 








0.036 


0.023 


0.045 


0.025 


707 




14.3 








0.018 


0.011 


0.030 


0.017 


708 




7.1 








0.023 


0.018 


0.033 


0.024 


709 




3.6 








0.018 


0.018 


0.018 


0.018 


710 




3.6 








0.000 


0.000 


0.019 


0.019 


712 




3.6 








0.030 


0.030 


0.023 


0.023 


719 




14.3 








0.022 


0.011 


0.031 


0.018 


732 




14.3 








0.042 


0.022 


0.044 


0.023 


733 




10.7 








0.009 


0.007 


0.034 


0.021 


752 




10.7 








0.029 


0.018 


0.037 


0.021 


756 




17.9 








0.065 


0.030 


0.061 


0.028 


f lavescens 


14.3 




7. 


2 


5.0 


0.010 


0.005 


0.028 


0.016 


734 




3.6 








0.004 


0.004 


0.004 


0.004 


751 




10.7 








0.016 


0.011 


0.039 


0.026 


kolpomyrtensis 


10.7 




10. 


7 




0.022 


0.013 


0.020 


0.012 


jónica 


17.9 




10. 


7 


0.0 


0.028 


0.015 


0.035 


0.016 


707 




10.7 








0.021 


0.013 


0.027 


0.016 


708 




10.7 








0.048 


0.027 


0.057 


0.032 


ass i со I a 


10.7 




8. 


9 


2.6 


0.011 


0.009 


0.018 


0.014 


742 




10.7 








0.010 


0.008 


0.019 


0.014 


744 




7.1 








0.013 


0.010 


0.018 


0.015 


adrianae 


3.6 




0. 


9 


1.8 


0.001 


0.001 


0.001 


0.001 


725 




0.0 








0.000 


0.000 


0.000 


0.000 


728 




0.0 








0.000 


0.000 


0.000 


0.000 


729 




0.0 








0.000 


0.000 


0.000 


0.000 


730 




3.6 








0.002 


0.002 


0.002 


0.002 


dubia 


0.0 










0.000 


0.000 


0.000 


0.000 


teres 


3.6 










0.000 


0.000 


0.015 


0.015 


rebel i 


7.1 










0.005 


0.005 


0.024 


0.017 


Albinaria sd. 


7.1 










0.002 


0.002 


0.016 


0.014 


Isabel laria 


3.6 










0.000 


0.000 


0.018 


0.018 



0.044). From the individual population sam- 
ples in which Ho and He are not equal to zero 
(Table 2), 13% have Ho > He, whereas 16% 
have Ho = He. So, in 71% of the populations 
there is a deficiency of heterozygosity. 

Among the various taxa there is only little 
allozymic differentiation (Table 3). All taxa 



show the same allele at highest frequency for 
all but nine loci, namely, Aat-1, Hk, Idh, Lap, 
Ldh, Mdh-1, Mdh-2, Mpi and Pgm. Of these 
loci, only Aat-1, Mdh-1 and Mdh-2 concern 
Albinarias from Kephallinia or Ithaka. Although 
Aat-1, Mdh-2 and Lap are highly polymorphic, 
there is always a most common allele. 



40 KEMPERMAN & DEGENAARS 

percentage polymorphic loci 




T 1 1 г 

3 4 5 6 7 

number of populations 



10 



c. contaminate 



s. senilis 



FIG. 3. Allozyme variation in Albinaria subspecies at increasing number of populations. For explanation, see 
Discussion. 



Albinaria a. adrianae is monomorphic at the 
Mdh-1 locus for an allele that is rare or absent 
in the other taxa. Albinaria a. dubia is mono- 
morphic at the Mdh-2 locus for an allele that is 
not found in any other Kephallinian taxon, but 
occurs in the Ithakian A. с odysseus and in 
the outgroup species, namely, Albinaria sp. 
and Isabellaria edmundi from the Pélopon- 
nèse and A. teres nordsiecki and A. rebel! 
from Crete. No other Kephallinian or Ithakian 
taxa are monomorphic for rare alleles. The 
outgroup species, however, have at the loci 
Aat-1, Lap, Ldh, Mdh-1, Mdh-2, Mpi, Nadd-1, 
Idh either unique alleles or alleles that they 
share with other outgroup species. 

Based on the estimated allelic frequencies, 
coefficients of genetic distances (D & R) were 
calculated for all pair-wise combinations of 
populations as well as taxa (Table 4). The 
results of the UPGMA cluster analyses, which 
are based on the various matrices are given 
in Figures 5 and 6, the population dendro- 
grams, and Figures 7 and 8, the taxon den- 
drograms. Intraspecific genetic distances in 
the non-endemic species A. contaminata and 
A. senilis on Kephallinia are very low, 
whereas the differentiation among both the 



subspecies of the endemic A. adrianae and A. 
jónica is large enough to cluster them sepa- 
rately with different groups of the non- 
endemic species. The interspecific variation 
of the non-endemic species is limited, but 
large enough to enable a complete partition. 
The genetic distance between the endemic 
species is larger than that between the en- 
demic and the non-endemic species. 

There may be a correlation between ge- 
netic distance and geographical isolation, 
which is demonstrated by the genetic dis- 
tance between the fully isolated subspecies of 
both A. adrianae and A. jónica, as well as by 
the allelic composition of the Ithakian A. с 
odysseus. The latter is genetically more dif- 
ferentiated from the combined other Kephal- 
linian taxa than these are from each other. 
The distance between the Kephallinian- 
Ithakian taxa and the Cretan-Peloponnesian 
outgroup species also reflects the geograph- 
ical isolation. However, among the outgroup 
species, the differentiation between taxa from 
Crete on the one hand and the Péloponnèse 
on the other hand does not strictly conform to 
the geographical distances. 

To test the goodness of fit of the trees we 



20 



ALLOZYME FREQUENCIES IN ALBINARIA 
percentage polymorphic loci / population 



41 



15 - 



10 - 



- 


+ + + 




• / 


- 


1— 4— — — .i л / 








-^ + y*\ / / + \ 




/ Ш 








1 1 



10 15 20 25 

number of individuals / population 



30 



35 



contaminate 



senilis 



-Ж- 



jonica 



"Э - adrianae 



FIG. 4. Allozyme variation in Albinaria species within samples of different size. For explanation, see Dis- 
cussion. 



repeatedly calculated Nei's D and ran a 
UPGMA cluster analysis while each time a 
different subspecies was removed from the 
data set. This procedure resulted in two types 
of trees that differ only in the position of A. с 
odysseus and A. a. adrianae. When either A. 
с contaminata, A. с incommoda or A. a. du- 
bia was removed, a tree was obtained with A. 
с odysseus as the sister group of the com- 
bined A. contaminata subspecies and A. j. 
jónica. In case of the removal of A. с contam- 
inata and A. с incommoda, A. a. adrianae 
switched from the A. contaminata cluster to 
the A. senilis cluster. However, when either A. 
с liebetruti, A. s. senilis, A. s. flavescens, A. 
s. kolpomyrtensis, A. j. jónica, A. j. assicola or 
A. a. dubia were removed the dendrogram 
was identical to the one obtained when all 
subspecies are included (Fig. 7). It can be 
concluded from this "jackknife" procedure 
that the complete dendrogram is very stable 
except for the position of A. с odysseus and 
A. a. adrianae. 

To be used in a Hennigian analysis, Table 5 
gives the characters of the various taxa, as 
they are derived from the allele frequencies 



(Table 3). There are 17 informative alleles. 
The apomorphic character states are deter- 
mined by outgroup comparison. Table 5 
shows that a plesiomorphic state or states 
have been proposed for 1 5 of the 1 6 polymor- 
phic loci. Mdh-2 defines a large group to 
which only A. a. dubia is not linked. Albinaria 
a. dubia has no synapomorphies with any of 
the other taxa. The goal of separating the taxa 
into groups that are characterized by derived 
character states turned out to be impractica- 
ble. This means that a Hennigian analysis of 
the entire group does not produce a resolved 
phylogeny. The low number of synapomor- 
phies distributed among the loci even troubles 
partial analyses. 

Some alleles are very locally distributed in 
a certain species, sometimes exclusively 
where its range comes close to that of an- 
other species. This shared presence of alleles 
may be considered then the result of intro- 
gression. This applies to (1) the Nadd-1 c- 
and d-allele in A. j. jónica at loo 708, and in 
the adjoining populations of A. s. senilis at 
Iocs. 702 and 706; (2) Gpi alleles in A. s. se- 
nilis at Iocs. 756 and 706, and A. с contam- 



42 



KEMPERMAN & DEGENAARS 



TABLE 3. Allele frequencies in populations for 16 polymorphic loci. N 
at the locus. For taxon abbreviations, see Figure 5. 



number of individuals sampled 



















Populan 


on 
















sen 


sen 


sen 


Jon 


Jon 


sen 


sxj 


sen 


sen 


sen 


con 


con 


con 


sen 


cxs 


Locus 


702 


706 


707 


707 


708 


708 


709 


709 


710 


712 


713 


715 


716 


719 


721 


Aat-1 


6 


9 


12 


5 


3 


4 


3 


2 


10 


16 


20 


24 


17 


12 


5 


a 


1.000 


1.000 


0.917 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.975 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.083 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.025 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Ak 
CN) 


6 


9 


12 


5 


2 


5 


3 


1 


1 


6 


9 


24 


5 


11 


5 


a 


1.000 


1.000 


0.708 


1.000 


0.750 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.909 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.292 


0.000 


0.250 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.091 


0.000 


Aph 
(N) 


8 


6 


12 


5 


2 


5 


3 


1 


8 


10 


21 


21 


22 


12 


5 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Est-3 
































(N) 


8 


6 


12 


8 


3 


5 


3 


1 


8 


10 


17 


21 


22 


12 


5 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Gpi 
(N) 


1 


6 


12 


5 


2 


5 


3 


1 


8 


10 


17 


18 


14 


12 


5 


a 


1.000 


0.833 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.676 


0.667 


0.750 


0.917 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.029 


0.028 


0.179 


0.000 


0.000 


с 


0.000 


0.083 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.059 


0.056 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.056 


0.000 


0.000 


0.000 


e 


0.000 


0.083 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.083 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.059 


0.056 


0.000 


0.000 


0.000 


9 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.176 


0.139 


0.071 


0.000 


0.000 


Hk 
(N) 


6 


3 


1 


3 


1 


1 


1 


1 


10 


16 


9 


7 


13 


1 


1 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Idh 
(N) 


5 


9 


14 


6 


2 


6 


4 


2 


11 


15 


23 


23 


17 


17 


7 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.913 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.087 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


lap 
































(N) 


1 


1 


12 


6 


2 


4 


3 


1 


10 


10 


21 


16 


13 


12 


5 


a 


1.000 


1.000 


0.917 


0.833 


0.750 


0.500 


0.500 


0.000 


0.500 


1.000 


1.000 


0.906 


0.962 


0.958 


1.000 


b 


0.000 


0.000 


0.000 


0.167 


0.250 


0.000 


0.500 


1.000 


0.500 


0.000 


0.000 


0.000 


0.038 


0.042 


0.000 


с 


0.000 


0.000 


0.083 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.500 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.094 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


9 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Ldh 
(N) 


6 


9 


12 


9 


3 


4 


3 


2 


10 


6 


24 


25 


10 


12 


5 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 






























{continued) 



¡nata at Iocs. 713, 715 and 716; (3) the c- 
allele of the Ak locus in A. s. senilis at Iocs. 
756 and 707 and in A. j. jónica at loo 708; (4) 
the b-allele of Aat-1 in А. с liebetruti at loc. 
746, in A. с incommoda at loc. 773 and in all 
Ithakian populations (Table 5). Therefore, a 
Hennigian phylogenetic analysis has also 
been performed on data in which the alleles 
probably acquired by introgression were re- 
moved. This, however, did not result in an 
obvious improvement of the analyses. 



DISCUSSION 



General 



There is still much debate on the relevance 
of various algorithms determining genetic dis- 
tance coefficients and the combination of 
these coefficients with clustering methods 
(Nei et al., 1983; Buth, 1984; Richardson et 
al., 1986; Emberton, 1988; Ferguson, 1988). 
For a phenetic comparison of the allozyme 



ALLOZYME FREQUENCIES IN ALBINARIA 



43 



TABLE 3. (Continued) 



Mdh-1 
































(N) 


6 


9 


14 


8 


3 


7 


4 


1 


9 


6 


19 


29 


1 


17 


7 


a 


0.000 


0.000 


0.000 


1.000 


1.000 


0.000 


1.000 


0.000 


0.000 


0.167 


1.000 


0.931 


1.000 


0.000 


0.857 


Ь 


0.750 


1.000 


1.000 


0.000 


0.000 


1.000 


0.000 


1.000 


1.000 


0.583 


0.000 


0.017 


0.000 


1.000 


0.143 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.017 


0.000 


0.000 


0.000 


e 


0.250 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.083 


0.000 


0.000 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.167 


0.000 


0.034 


0.000 


0.000 


0.000 


Mdh-2 
































(N) 


6 


9 


14 


8 


3 


7 


4 


1 


9 


6 


20 


29 


2 


17 


7 


a 


1.000 


0.889 


1.000 


0.063 


0.000 


0.786 


0.250 


1.000 


1.000 


1.000 


1.000 


0.897 


1.000 


0.676 


1.000 


b 


0.000 


0.111 


0.000 


0.000 


0.000 


0.214 


0.000 


0.000 


0.000 


0.000 


0.000 


0.103 


0.000 


0.324 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.938 


1.000 


0.000 


0.750 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Mpi 
































(N) 


1 


6 


12 


6 


2 


4 


3 


2 


10 


1 


23 


15 


14 


12 


5 


a 


1.000 


1.000 


0.958 


1.000 


1.000 


1.000 


1.000 


0.750 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.700 


b 


0.000 


0.000 


0.042 


0.000 


0.000 


0.000 


0.000 


0.250 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.300 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Nadd-1 
































(N) 


14 


9 


14 


8 


3 


7 


4 


1 


9 


16 


18 


26 


22 


17 


7 


a 


0.929 


0.611 


1.000 


1.000 


0.667 


1.000 


1.000 


1.000 


1.000 


1.000 


0.917 


1.000 


1.000 


1.000 


0.929 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.036 


0.167 


0.000 


0.000 


0.167 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.036 


0.111 


0.000 


0.000 


0.167 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.083 


0.000 


0.000 


0.000 


0.000 


f 


0.000 


0.111 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Э 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.071 


Nadd-2 
































(N) 


14 


9 


14 


8 


3 


7 


4 


1 


9 


15 


15 


23 


22 


17 


7 


a 


1.000 


1.000 


1.000 


0.813 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.188 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Pqd 
































(N) 


6 


9 


14 


9 


3 


6 


4 


2 


11 


16 


24 


23 


17 


4 


2 


a 


1.000 


0.944 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.056 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Pgm 
































(N) 


6 


9 


14 


3 


3 


2 


4 


1 


9 


16 


20 


17 


14 


5 


2 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.971 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.029 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 
















Population 


















i ne 


adr 


ine 


adr 


adr 


adr 


dub 


sxd 


sxd 


fia 


con 


kol 


ass 


ass 


lie 


Locus 


723 


725 


727 


728 


729 


730 


731 


732 


733 


734 


735 


736 


742 


744 


746 



Aat-1 
(N) 



9 

1.000 
0.000 
0.000 
0.000 
0.000 



6 

1.000 
0.000 
0.000 
0.000 
0.000 



9 

1.000 
0.000 
0.000 
0.000 
0.000 



9 

1.000 
0.000 
0.000 
0.000 
0.000 



12 
1.000 
0.000 
0.000 
0.000 
0.000 



16 

1.000 
0.000 
0.000 
0.000 
0.000 



19 
1.000 
0.000 
0.000 
0.000 
0.000 



6 

1.000 
0.000 
0.000 
0.000 
0.000 



12 
1.000 
0.000 
0.000 
0.000 
0.000 



1.000 
0.000 
0.000 
0.000 
0.000 



13 
0.846 
0.000 
0.000 
0.000 
0.154 



15 

1.000 
0.000 
0.000 
0.000 
0.000 



18 
0.944 
0.000 
0.056 
0.000 
0.000 



17 

1.000 
0.000 
0.000 
0.000 
0.000 



13 
0.692 
0.269 
0.000 
0.000 
0.038 



Aph 
(N) 



1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



6 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



9 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



3 
1.000 
0.000 
0.000 
0.000 



5 

1.000 
0.000 



3 

1.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 



1.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 



1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



11 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



8 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



8 

1.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 



6 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



1 

1.000 
0.000 
0.000 
0.000 



9 
1.000 
0.000 



1 

1.000 
0.000 
0.000 
0.000 



10 
1.000 



16 

1.000 
0.000 
0.000 
0.000 



10 
1.000 



0.000 0.000 

{continued) 



data, we have chosen the well known Nei's 
Genetic Distance (D). This index measures 
the mean number of electrophoretically de- 
tectable substitutions per locus, which accu- 
mulated because the two populations di- 
verged from a common ancestor (Buth, 
1984). Nei's D might also enable assumptions 



concerning the period of time after the taxa 
evolved from their most recent common an- 
cestor, presuming that constant rates of evo- 
lution occur within the lineages. No agree- 
ment exists, however, on the translation of 
genetic distances into time-spans (Rich- 
ardson et al., 1986). After a comparison of 



44 



KEMPERMAN & DEGENAARS 



TABLE 3. (Continued) 



Est-3 
































(N) 


9 


8 


9 


1 


10 


10 


6 


4 


11 


1 


10 


8 


16 


18 


18 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Gpi 
(N) 


6 


6 


9 


3 


13 


16 


8 


4 


11 


1 


10 


8 


10 


10 


10 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


0.969 


1.000 


1.000 


0.909 


1.000 


1.000 


0.938 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.031 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.091 


0.000 


0.000 


0.063 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


g 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Hk 
(N) 


6 


6 


9 


3 


12 


16 


2 


1 


1 


8 


13 


15 


18 


8 


12 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Idh 
(N) 


9 


1 


10 


6 


9 


10 


17 


8 


14 


10 


13 


15 


9 


10 


16 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.950 


1.000 


0.867 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.050 


0.000 


0.133 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Lap 
(N) 


6 


1 


8 


1 


9 


10 


2 


6 


1 


1 


10 


9 


10 


10 


10 


a 


0.750 


1.000 


0.813 


1.000 


1.000 


1.000 


1.000 


0.917 


1.000 


1.000 


1.000 


0.889 


1.000 


1.000 


1.000 


b 


0.250 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.083 


0.000 


0.000 


0.000 


0.111 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


g 


0.000 


0.000 


0.188 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Ldh 
(N) 


9 


6 


9 


9 


12 


6 


19 


6 


12 


1 


13 


9 


17 


8 


18 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Mdh-1 
(N) 


9 


6 


10 


8 


13 


6 


16 


6 


13 


10 


13 


14 


9 


8 


8 


a 


0.944 


0.000 


1.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


1.000 


0.000 


0.000 


0.000 


1.000 


b 


0.056 


0.000 


0.000 


0.000 


0.000 


0.000 


1.000 


0.917 


1.000 


1.000 


0.000 


1.000 


1.000 


1.000 


0.000 




0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


1.000 


0.000 


1.000 


1.000 


1.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.083 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Mdh-2 
































(Ю 


9 


6 


10 


8 


13 


6 


16 


6 


13 


10 


13 


14 


9 


8 


8 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.000 


0.750 


0.846 


1.000 


0.846 


1.000 


0.222 


0.250 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.250 


0.154 


0.000 


0.154 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


1.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.778 


0.750 


0.000 


Mpi 
































(N) 


3 


6 


9 


9 


12 


6 


19 


6 


12 


1 


10 


9 


9 


10 


10 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.333 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.667 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Nadd-1 
































(N) 


9 


14 


10 


8 


13 


16 


16 


6 


13 


10 


13 


6 


18 


18 


18 


a 


1.000 


1.000 


0.950 


1.000 


1.000 


1.000 


1.000 


1.000 


0.500 


1.000 


1.000 


1.000 


0.972 


0.944 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.028 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.500 


0.000 


0.000 


0.000 


0.000 


0.056 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


g 


0.000 


0.000 


0.050 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 



{continued) 



various combinations of genetic coefficients 
and clustering methods, Nei et al. (1983) con- 
cluded that Nei's D in combination with an 
UPGMA is the best choice for phylogenetic 
analysis. However, a phylogenetic interpreta- 
tion of the Nei's D dendrograms is seriously 
affected by (a) the failure of Nei's D to satisfy 



the triangle of equality, which may result 
in biologically impossible negative branch- 
lengths, and (b) the fact that the dendrograms 
only show phenetic similarity or dissimilarity. 
Rogers' R (1972) meets the metrical require- 
ments, which is therefore also applied in com- 
bination with UPGMA cluster analyses. 



ALLOZYME FREQUENCIES IN ALBINARIA 



45 



TABLE 3. (Continued) 



Nadd-2 
































(N) 


3 


14 


10 


8 


13 


16 


16 


6 


13 


10 


13 


14 


18 


18 


6 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Pgd 
































(N) 


9 


1 


10 


6 


7 


10 


17 


2 


u 


2 


13 


9 


17 


18 


18 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Рят 
































(N) 


6 


1 


10 


5 


10 


10 


H 


2 


13 


2 


13 


8 


18 


18 


12 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


















Population 
















con 


fia 


sen 


sen 


con 


ody 


ody 


ody 


ody 


inc 


inc 


ter 


reb 


Isa 


par 


Locus 


749 


751 


752 


756 


759 


764 


765 


766 


770 


772 


773 











Aat-1 
(N) 



8 

1.000 
0.000 
0.000 
0.000 
0.000 



17 
0.735 
0.000 
0.265 
0.000 
0.000 



11 
0.227 
0.773 

0.000 
0.000 
0.000 



15 
1.000 
0.000 
0.000 
0.000 
0.000 



8 
0.938 
0.000 
0.000 
0.000 
0.063 



12 
0.000 
1.000 
0.000 
0.000 
0.000 



13 

0.000 
0.962 
0.000 
0.038 
0.000 



13 

0.000 
1.000 
0.000 
0.000 
0.000 



12 
0.583 
0.417 
0.000 
0.000 
0.000 



1 

1.000 
0.000 
0.000 
0.000 
0.000 



4 
0.625 
0.375 
0.000 
0.000 
0.000 



0.000 
1.000 
0.000 
0.000 
0.000 



12 
0.750 
0.250 
0.000 
0.000 
0.000 



20 
0.000 
1.000 
0.000 
0.000 
0.000 



20 
1.000 
0.000 
0.000 
0.000 
0.000 



Aph 
(N) 



Est-3 
CM) 



SEI 



12 

1.000 
0.000 
0.000 
0.000 



12 

1.000 
0.000 



16 

1.000 
0.000 



12 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



1.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 



1.000 
0.000 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



3 

1.000 
0.000 
0.000 
0.000 



10 

1.000 
0.000 



10 

1.000 
0.000 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



15 

0.767 
0.000 
0.033 
0.200 



12 

1.000 
0.000 



12 
1.000 
0.000 



12 
0.833 
0.083 
0.042 
0.042 
0.000 
0.000 
0.000 



3 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



3 

1.000 
0.000 



1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



11 

1.000 
0.000 
0.000 
0.000 



6 

1.000 
0.000 



12 

1.000 
0.000 



1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



13 
1.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 



13 

1.000 
0.000 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



13 

1.000 
0.000 
0.000 
0.000 



10 
0.000 
1.000 



13 
1.000 
0.000 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



11 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



15 

1.000 
0.000 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



1 

1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



1 

1.000 
0.000 



1 

1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



11 

1.000 
0.000 
0.000 
0.000 



13 

1.000 
0.000 



1.000 
0.000 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



4 
1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



1.000 
0.000 



1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



5 

1.000 
0.000 
0.000 
0.000 



13 

1.000 
0.000 



13 
0.154 
0.846 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



12 
1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



13 

1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



12 
1.000 
0.000 
0.000 
0.000 



1.000 
0.000 



20 20 
0.400 0.750 
0.600 0.250 



12 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



Idh 
(N) 



Lap 
(N) 



1.000 
0.000 
0.000 



1.000 
0.000 
0.000 



11 

1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



17 

1.000 
0.000 
0.000 



1.000 
0.000 
0.000 



10 
0.950 
0.000 
0.000 
0.000 
0.000 
0.050 
0.000 



12 

1.000 
0.000 
0.000 



12 
1.000 
0.000 
0.000 



9 
0.833 
0.167 
0.000 
0.000 
0.000 
0.000 
0.000 



3 

1.000 
0.000 
0.000 



15 
1.000 
0.000 
0.000 



12 
0.750 
0.167 
0.000 
0.083 
0.000 
0.000 
0.000 



8 

1.000 
0.000 
0.000 



1.000 
0.000 
0.000 



5 

1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



7 
1.000 
0.000 
0.000 



1.000 
0.000 
0.000 



1 

1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



1.000 
0.000 
0.000 



13 

1.000 
0.000 
0.000 



10 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 
0.000 



13 

1.000 
0.000 
0.000 



10 
0.950 
0.050 
0.000 
0.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 
0.000 



15 
1.000 
0.000 
0.000 



10 
0.900 
0.100 
0.000 
0.000 
0.000 
0.000 
0.000 



1.000 
0.000 
0.000 



1.000 
0.000 
0.000 



1 

1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



1.000 
0.000 
0.000 



1.000 
0.000 
0.000 



10 

1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



0.000 
0.750 
0.250 



1.000 
0.000 
0.000 



1.000 
0.0G0 
0.000 
0.000 
0.000 
0.000 
0.000 



10 
1.000 
0.000 
0.000 



10 
0.000 
1.000 
0.000 



10 
0.000 
0.000 
0.000 
1.000 
0.000 
0.000 
0.000 



1 1 

1.000 1.000 

0.000 0.000 

0.000 0.000 



12 12 

1.000 0.000 

0.000 0.000 

0.000 1.000 



12 

0.000 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 



12 
1.000 
0.000 
0.000 
0.000 
0.000 
0.000 
0.000 



(continued) 



The percentages of polymorphic loci per 
population (Pp) (Table 2) are generally below 
the values given by Nevo (1978: 126) for 
other stylommatophoran gastropods. The av- 
erage Pp of Kephallinian and Ithakian Albina- 



ría taxa is 7.5% (± 4.9%). Nevo listed mean 
estimates of polymorphism which range from 
6% to 100% with an average of 31.5% (± 
26.6%) (excluding Rumina decollata). How- 
ever, it is not clear from his data what is 



46 

TABLE 3. (Continued) 



KEMPERMAN & DEGENAARS 



Ldh 
<N) 


15 


10 


12 


15 


5 


11 


13 


13 


15 


1 


13 


8 


13 


20 


20 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.885 


0.933 


1.000 


1.000 


0.000 


0.000 


1.000 


1.000 


Ь 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


1.000 


1.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.115 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.067 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Mdh-1 
(N) 


16 


7 


13 


15 


6 


5 


13 


13 


5 


4 


13 


8 


3 


20 


18 


а 


1.000 


0.000 


0.000 


0.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.000 


0.000 


0.000 


0.000 


b 


0.000 


1.000 


1.000 


1.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


1.000 


1.000 


1.000 


1.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Mdh-2 
(N) 


16 


7 


13 


15 


6 


5 


13 


13 


5 


4 


13 


8 


3 


20 


20 


а 


1.000 


0.500 


1.000 


1.000 


1.000 


0.800 


0.154 


0.192 


0.000 


1.000 


1.000 


0.000 


0.000 


0.000 


0.000 


b 


0.000 


0.429 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.200 


0.846 


0.808 


1.000 


0.000 


0.000 


1.000 


1.000 


1.000 


1.000 


d 


0.000 


0.071 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Mpi 
(N) 


11 


10 


9 


12 


5 


6 


10 


10 


10 


1 


11 


4 


10 


12 


12 


а 


1.000 


1.000 


1.000 


0.958 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.000 


Ь 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


1.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.042 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Nadd-1 
































(N) 


16 


17 


3 


15 


7 


11 


13 


13 


15 


4 


13 


4 


13 


20 


20 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


1.000 


с 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


d 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


e 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


f 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


9 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Nadd-2 
































(N) 


16 


17 


13 


15 


7 


11 


13 


13 


5 


4 


13 


4 


13 


20 


20 


a 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Pgd 
(N) 


15 


10 


12 


15 


5 


11 


13 


13 


15 


4 


13 


8 


13 


20 


20 


a 


1.000 


1.000 


1.000 


0.500 


0.900 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


b 


0.000 


0.000 


0.000 


0.500 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


с 


0.000 


0.000 


0.000 


0.000 


0.100 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


Pgm 
(N) 


16 


10 


13 


3 


4 


6 


10 


10 


10 


4 


13 


8 


13 


20 


20 


a 


1.000 


1.000 


0.769 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


1.000 


0.025 


b 


0.000 


0.000 


0.192 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.975 


с 


0.000 


0.000 


0.038 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 



meant by a population. According to our find- 
ings, the genetic variability depends on the 
extent of the area which is considered to be 
inhabited by a single population (Fig. 3). It is 
uncertain, therefore, whether Nevo's P values 
are comparable with our averaged Pp values 
for each taxon. 

Detailed results on mobility, dispersion and 
homing in Albinaria are not yet available. 
However, we have observed individuals mov- 
ing on rockfaces for at least one meter during 
a single rainy night. Baur (1988) found in bed- 
rock-inhabiting Chondrina dienta (Wester- 
lund), on the island of Öland, Sweden, an av- 
erage distance of 291 cm was bridged during 
six months, with a maximum of 814 cm. Ex- 
cept for the more temporate climate to which 
Chondrina dienta is confined, this species is 
very well comparable with Albinaria species in 



habitat preferences (Gittenberger, pers. 
com.). In view of these data, and because our 
samples are collected from an area limited to 
1 to 20 m 2 of substratum, each sample is con- 
sidered to belong to a panmictic population. 
Genetic differences within a subspecies 
proved to be present within distances less 
than 200 m, for example in A. j. assicola at 
Iocs. 744 and 742, in A. s. senilis and in A. j. 
jónica at Iocs. 707 and 708, as well as in A. с 
contaminata at Iocs. 713, 715 and 716. 
Nevo's (1978: 126) values for populations are 
approximately of the same magnitude as 
what we find within some whole subspecies 
(Pt = of all populations together), namely, 
32.1% for A. с contaminata, 17.9% for A. с 
odysseus, but only 3.6% for A. с liebetruti 
and A. a. adrianae. In A. s. senilis, we looked 
separately at the central west coast range 



ALLOZYME FREQUENCIES IN ALBINARIA 47 

TABLE 4. Matrix of genetic distance coefficients: above diagonal Roger's R, below diagonal Nei's D. 



Subspec i es 


1 


2 


3 


4 


5 


6 


7 


8 


9 


10 


11 


12 


13 


14 


15 


1 senilis 


..... 


0.028 


0.026 


0.058 


0.053 


0.062 


0.115 


0.087 


0.046 


0.058 


0.056 


0.187 


0.220 


0.165 


0.235 


2 flavescens 


0.003 


..... 


0.023 


0.054 


0.052 


0.054 


0.102 


0.089 


0.033 


0.052 


0.041 


0.171 


0.212 


0.158 


0.227 


3 kolpomyrtensis 


0.002 


0.004 


..... 


0.051 


0.047 


0.057 


0.114 


0.090 


0.041 


0.046 


0.047 


0.186 


0.213 


0.167 


0.226 


4 contaminata 


0.037 


0.039 


0.038 


..... 


0.015 


0.021 


0.077 


0.058 


0.071 


0.046 


0.079 


0.181 


0.221 


0.169 


0.228 


5 incommoda 


0.036 


0.039 


0.037 


0.001 


..... 


0.013 


0.070 


0.051 


0.069 


0.042 


0.077 


0.177 


0.214 


0.159 


0.228 


6 liebet rut i 


0.039 


0.041 


0.042 


0.004 


0.002 


..... 


0.061 


0.061 


0.074 


0.047 


0.082 


0.165 


0.211 


0.154 


0.233 


7 odysseus 


0.085 


0.083 


0.093 


0.051 


0.049 


0.037 


..... 


0.088 


0.108 


0.107 


0.088 


0.128 


0.211 


0.116 


0.239 


8 iónica 


0.071 


0.069 


0.075 


0.035 


0.036 


0.040 


0.062 


..... 


0.059 


0.087 


0.088 


0.191 


0.224 


0.167 


0.234 


9 assicola 


0.022 


0.017 


0.023 


0.058 


0.059 


0.062 


0.091 


0.041 


..... 


0.066 


0.035 


0.172 


0.215 


0.162 


0.219 


10 adrianae 


0.038 


0.040 


0.038 


0.037 


0.037 


0.040 


0.091 


0.074 


0.059 


..... 


0.072 


0.176 


0.218 


0.165 


0.223 


11 dubia 


0.036 


0.032 


0.038 


0.073 


0.074 


0.078 


0.070 


0.076 


0.030 


0.074 


..... 


0.139 


0.182 


0.129 


0.187 


12 teres 


0.184 


0.179 


0.194 


0.189 


0.186 


0.170 


0.114 


0.194 


0.183 


0.190 


0.147 


..... 


0.161 


0.125 


0.255 


13 rebel i 


0.225 


0.226 


0.221 


0.232 


0.229 


0.234 


0.210 


0.231 


0.228 


0.234 


0.189 


0.161 


..... 


0.143 


0.244 


14 Isabellaria sp. 


0.151 


0.158 


0.165 


0.168 


0.159 


0.151 


0.093 


0.159 


0.164 


0.171 


0.129 


0.122 


0.142 


..... 


0.226 


15 Albinaria sp. 


0.242 


0.241 


0.242 


0.243 


0.245 


0.250 


0.244 


0.245 


0.236 


0.244 


0.199 


0.284 


0.264 


0.249 


..... 



populations (Iocs. 702, 706-708, 710 and 
712), which resulted in a Pt of 32.1%. Taking 
all A. s. senilis material into account (thus in- 
cluding 71 9, 752 and 756) leads to only a very 
slight increase of Pt to 35%. The wide range 
of the Pt values for the various taxa is to a 
certain extent due to differences in the num- 
ber of populations that could be investigated 
(Table 2). A percentage of 3.6 in A. с liebe- 
truti is based on a single population. In fact, 
this value cannot be compared with, for ex- 
ample, 32.1% in A. s. senilis, because the 
latter number is based on six populations. As 
is obvious from Table 2, the range of variation 
of species and subspecies is not reached in a 
single population. This cannot be a conse- 
quence of simply a larger number of speci- 
mens that is studied when more populations 
are investigated. Figure 4 shows that there is, 
except for A. contaminata, only a small trend 
towards an increasing P at populations with 
higher numbers of analysed individuals. Ac- 
cording to the results presented in Figure 3, a 
minimal number of populations is needed to 
reach a point where the addition of more pop- 
ulations contributes relatively little to the ge- 
netic variability. In case of A. s. senilis, this 
number is five populations, whereas in case 
of A. с contaminata it is at least more than six 
populations. This indicates that for A. с con- 
taminata the total variation has not yet been 
established. It is concluded that there can be 
important differences in genetic variability 



throughout the range of Albinaria subspecies, 
which recommends, at least in the case of 
subspecies with large ranges, the use of min- 
imally six populations. Because of the lack of 
a sufficient amount of living material, we were 
not able to investigate equal numbers of indi- 
viduals per population and six or more popu- 
lations for all taxa. 

Nevo (1978) investigated the correlation 
between several biological parameters and 
the genetic variability observed in over 200 
species of plants and animals. Some of his 
parameters are briefly discussed here. 

One might expect that the genetic variabil- 
ity depends on the mode of reproduction. In 
our material, there is a considerable defi- 
ciency of heterozygosity in most populations 
(Table 2). This pattern has also been found in 
other hermaphroditic Mollusca (Seiander & 
Kaufman, 1973; Nevo, 1978; Hillis et al., 
1987; Boato, 1988). A deficiency of heterozy- 
gosity and a low proportion of polymorphic 
loci are considered indicative of a breeding 
system with incomplete panmixia. It has been 
reported that a complete or near absence of 
genetic variability within populations may oc- 
cur in facultative self-fertilizing breeding sys- 
tems, as is known, for example, for Rumina 
decollata (see Selander & Kaufman, 1973), 
Partula gibba (see Johnson et al., 1977) and 
some Arion species (see McCracken & Se- 
lander, 1980). The fact that most populations 
of A. adrianae proved to be invariable (Table 



48 



KEMPERMAN & DEGENAARS 



.40 



.36 



.28 



.24 



Distance 
.20 



.16 



.04 



.00 



Legend 






con - 


A. 


contaminata contaminate 


inc - 


A. 


с incommonda 


ody - 


A. 


с odysseus 


lie - 


A. 


c. liebetruti 


sen - 


A. 


senilis senilis 


fla - 


A. 


s. flavescens 


kol - 


A. 


s. kolpomyrtensis 


jon - 


A. 


jónica jónica 


ass - 


A. 


jónica assicola 


adr - 


A. 


adrianae adrianae 


dub - 


A. 


a. dubia 


ter - 


A. 


teres nordsiecki 


reb - 


A. 


rebel i 


Alb - 


Albinaria sp. 


Isa - 


Isabellaria sp. 




.36 



.28 



.20 



.04 



FIG. 5. Dendrogram of UPGMA cluster analysis on Neis (1972) genetic distance coefficients. Cophenetic 
correlation = 0.947. 



ALLOZYME FREQUENCIES IN ALBINARIA 



49 



.40 



.28 



.24 



Distance 
.20 



.16 



.08 



.00 



-C 



sen 702 
sen 712 
fla 734 
kol 736 
sen 710 
sen 707A 
sen 708 
sen 719 
fla 751 
sen 706 
sen x dub 733 
dub 731 
ass 742 
ass 744 
sen x dub 732 
sen 756 
sen752 
sen 709 
con 713 
con 715 
con 716 

inc 727 
odk 749 

inc 772 

con759 

inc 723 

con 735 

lie 746 

inc 773 

con x sen 721 

adr 725 

adr 728 

adr 729 

adr 730 

jon 707 

sen x jon 709 

jon 708 

ody 764 

ody 765 

ody 770 

ody 766 

ter 

Isa 

reb 

Alb 



.40 



.36 



.32 



.28 



.24 



.20 



.16 



.12 



.00 



FIG. 6. Dendrogram of UPGMA cluster analysis on Rogers' (1972) genetic distance coefficients. Cophenetic 
correlation = 0.941 . For legend, see Figure 5. 



50 



.40 

+ + . 



KEMPERMAN & DEGENAARS 
Distance 



33 



.27 

.+ +. 



.20 

.+ +. 



.13 

.+ +. 



.07 
.+ +. 



,00 



s. senilis 
s. kolpomyrt. 
s. flavescens 
j. assicola 
a. dubia 
с. contaminata 
с. incommoda 
с. liebetruti 
j. jónica 
a. adrianae 
с. odysseus 
teres 

Isabellaria sp. 
rebel i 
Albinaria sp. 



+ + + + + + + + + + + + + 

.40 .33 .27 .20 .13 .07 .00 

FIG. 7. Dendrogram of UPGMA cluster analysis on Nei's (1972) genetic distance coefficients. Cophenetic 
correlation = 0.960. 



2) might be an indication for apomixis. How- 
ever, there is no additional information that 
could support this. The observations within, 
for example, A. с contaminata (namely, P = 
17.9 in loc. 715 and P = 0.0 in loc. 749) show 
that populations of a single Albinaria subspe- 
cies may be strikingly different in genetic vari- 
ability. Whether this is caused by multiple re- 
productive systems or by one or more other 
reasons, remains unknown. Because no in- 
formation on the breeding system of Kephal- 
linian and Ithakian Albinaria is present, it also 



remains unclear whether the observed defi- 
ciency of heterozygosity in the majority of the 
sampled populations is caused by facultative 
apomixis or by inbreeding. As has been 
stated before, a sample is considered to be- 
long to a panmictic population. Hence, it is 
concluded that the Wahlund effect (Rich- 
ardson et al., 1986) is probably not partly re- 
sponsible for the paucity of the mean het- 
erozygosity. It can also be suggested that the 
convention of scoring doubtful pairs of bands 
as single bands might have contributed to the 







ALLOZYME FREQUENCIES IN ALBINARIA 






Distance 


.40 


.33 


.27 .20 .13 .07 


+ + __ 


- + + 


+ __+ + +__ _+ _ + + _+_ _ 



.00 

+ 



51 



- s. seni 1 is 

- s. flavescens 

- s. kolpomyrt. 

- j. assicola 

- a. dubia 

- с. contaminata 

- с. incommoda 

1 — cl iebetruti 

- a. adrianae 

- j. jónica 

- с. odysseus 

- teres 

- Isabel laria sp, 

- rebel i 

- Albinaria sp. 



+ + + + + + + + + + + + + 

.40 .33 .27 .20 .13 .07 .00 

FIG. 8. Dendrogram of UPGMA cluster analysis on Rogers' (1972) genetic distance coefficients. Cophenetic 
correlation = 0.963. 



observed deficiency of heterozygotes. How- 
ever, this cannot have influenced our values 
substantially. Moreover, the effect is also re- 
duced by the acceptance of the most frequent 
alternative in case of a doubtful position of a 
single homozygous band. 

Most Albinaria habitats on Kephallinia and 
Ithaka are seasonally subject to heat and 
drought. Therefore, in summer at least, most 
snails are in aestivation. Relatively long peri- 
ods of severe drought and/or very high tem- 
peratures might cause a high mortality, its ex- 



tent depending on the local microclimate. 
Especially in small populations, a high mor- 
tality could result then in a bottleneck effect 
and, therefore, a low genetic variability. Be- 
cause we could not make reliable estimations 
of population size, correlations between this 
factor and allozyme variation cannot be inves- 
tigated. 

It is suggested (Nevo, 1978: 162) that a 
relatively low genetic variability (low P value) 
is characteristic for geographically restricted 
habitat specialists, whereas a high variability 



52 



KEMPERMAN & DEGENAARS 



TABLE 5. Character states for 16 polymorphic loci, showing the plesiomorphic states for the in- 
group taxa. Synapomorphies are underlined, autapomorphies are in italics. For taxon abbreviations, see 
Figure 5. 



Ingroup: 



Outgroup: 



Ak A£h 



Gp1 



Locus as taxonomic character 
jdh_ La£ Ldh_ Mdh-1 



f la 


ac_ 


kol 


a 


con 


ade 


inc 


ab 


lie 


abe 


ody 


abd 


jon 


a 


ass 


ac 



3 les1omorph1c state: 



Mp1 Nadd-1 Nadd-2 Pgd 



£2£ 



is associated with more wide-spread habitat 
generalists. This rule can be checked in our 
material, consisting of two endemic habitat 
specialists and two non-endemic, much more 
wide-spread, less stenoecious species. In 
populations of A. contaminata and A. senilis, 
the mean P values vary between 3.6% and 
10.7%, with an average of 8.0% for A. con- 
taminata and 9.6% For A. senilis (Table 2). In 
the endemic habitat specialist A. adhanae, we 
found no polymorphic loci in one population of 
A. a. dubia and three populations of A. a. adri- 
anae; in a fourth population of the latter sub- 
species, a P of 3.6% was found. Although 
these differences are small, the data support 
Nevo's rule. However, P values in the second 
endemic species A. jónica contradict it. The 
habitat specialist A. j. jónica, known only from 
barren rock faces, has a mean P of 8.9%, and 
the more catholic A. j. assicola has a mean P 
of 10.7%, both moderately high values. Also, 
the low P value of 3.6% for A. с liebetruti, 
which is considered a habitat generalist, does 
not support Nevo's suggestion. We do not 
consider Nevo's rule falsified by our results, 
however. It could equally well be argued that 
there are only minor ecological differences in 



the habitats of the various subspecies; they 
are all represented in relatively warm, dry 
limestone areas. 

Apparently a certain genetic distance does 
not reflect a divergence on either population, 
subspecies, species or genus level. Gener- 
ally, electrophoretic data alone are not suffi- 
cient for final decisions on this matter (Gould 
& Woodruff, 1978: 407; Gould & Woodruff, 
1 986: 464; Richardson et al., 1 986: 308; Men- 
ken & Ulenberg, 1987: 318). It may be impos- 
sible to distinguish subspecies or even repro- 
ductively isolated species on the basis of 
biochemical characters (Avise, 1975; John- 
son et al., 1977). Several authors list ranges 
of Nei's D corresponding with various taxo- 
nomic ranks (e.g., Davis et al., 1981; Menken 
& Ulenberg, 1987; Thorpe, 1982). It becomes 
obvious from these publications that the rank 
indications for a certain group of taxa do not 
necessarily hold for another group (Davis, 
1984). The genetic distances found in Albina- 
ria are very low, but, on the whole, the group- 
ing of populations by UPGMA cluster analy- 
ses of Nei's D and Rogers' R is in congruence 
with the results of the classical, morphological 
approach. 



ALLOZYME FREQUENCIES IN ALBINARIA 



53 



The amount of genetic divergence among 
the Kephallinian and Ithakian Albinaria taxa 
can be considered indicative of a recent radi- 
ation of the group, assuming that a clear ge- 
netic differentiation is secondary to the speci- 
ation process. The occurrence of different 
fixed alleles in several species, which are de- 
fined on conchological and biogeographical 
data, demonstrates that these taxa also have 
individuality with regard to allozyme composi- 
tion. 

A Hennigian analysis of the allozyme data 
did not result in a resolved phylogeny. The 
results are contradictory and often inconsis- 
tent with the conchological data (Kemperman, 
in prep. A) and biogeographical patterns. It is 
considered premature to discuss these differ- 
ences in detail here. 

There are several possible reasons for the 
poor outcome of the Hennigian analyses. 
Generally, the number of apomorphic charac- 
ters states is too low to validate dichotomies 
(Table 5). This number is even lowered when 
alleles are removed from which the shared 
presence is suspected to result from intro- 
gression, rather than being based on apomor- 
phy. Although, the resulting phylogeny should 
become more realistic when the "pseudo"- 
apomorphic characters based on introgres- 
sion are removed, the results did not improve. 
Another source of discrepancies might be the 
use of characters based on very low allele 
frequencies that are given the same weight as 
characters based on high frequencies. Char- 
acters based on low allele frequencies are 
considered the least supportive, mostly be- 
cause they are more error prone. 



Albinaria spp. on Kephallinia and Ithaka 

Albinaria contaminata: The subspecies A. с 
contamínala, А. с. incommoda, and А. с. lie- 
betruti, cannot be recognized as separate en- 
tities in the UPGMA cluster dendrograms of 
Nei's D and Rogers' R (Figs. 5, 6). Material 
from Iocs. 727 (A. с incommoda), 735, 749 
and 759 (A. с contaminata) and 772 (A. с 
incommoda), situated in various quarters of 
the species range, is even identical according 
to Nei's D dendrogram. 

All phenetic analyses illustrate the relatively 
high genetic distance between populations 
from Ithaka on the one hand and those from 
Kephallinia on the other hand. The cause of 
this divergence is difficult to find out. Ithaka is 
geologically older than Kephallinia (Hagn et 



al., 1962: 185), but it has been unified with 
Kephallinia several times in the past due to 
sea-level oscillations. Nothing is known yet 
about the early history of Albinaria in this re- 
gion. The present situation on Kephallinia and 
Ithaka is certainly affected by a series of sep- 
arations and secondary contacts. It may be 
questioned whether each geographical sepa- 
ration resulted in genetic divergence and to 
what extent the secondary contacts reestab- 
lished a fully mixed gene-pool. The present 
genetic divergence is at least the result of the 
postglacial separation of minimally 12,000 
years. It cannot be excluded that it is the ef- 
fect of an accumulation of smaller diver- 
gences caused by the repetitive splitting and 
fusion of ranges. 

It remains a puzzling fact that, according to 
both Nei's and Rogers' genetic distance co- 
efficients, A. с odysseus, from Ithaka, gets a 
position in the UPGMA dendrograms (Figs. 
5-8) opposite all Kephallinian Albinaria taxa, 
thus including the not conspecific ones. This 
implies that the amount of allozyme diver- 
gence cannot be correlated with taxonomic 
status, that is, with species boundaries. It 
must be stressed here, however, that the po- 
sition of A. с odysseus is relatively unstable. 
This is demonstrated by the jackknife proce- 
dure, resulting in dendrograms in which A. с 
odysseus is grouped either among the other 
A. contaminata subspecies or opposite the 
combined Kephallinian taxa. Except for A. a. 
adrianae in two cases, the subspecies remain 
always in the same position. 

An explanation for the large genetic dis- 
tance between Ithakian and Kephallinian taxa 
might be that, due to, for example, extensive 
introgressive hybridization, a certain amount 
of gene flow continues between all Kephallin- 
ian taxa, opposing the genetic differentiation 
among them. Although introgression among 
Kephallinian taxa occurs, our biochemical 
data are insufficient to demonstrate the level 
of gene flow. 

A population of A. contaminata sampled 
near Fiskardo, northern Kephallinia (loc. 
749), was identified as A. с odysseus by 
Rähle (1979: 215). Although the shells show 
some resemblance to Ithakian A. с odys- 
seus, the allozyme data indicate no genetic 
difference with other Kephallinian A. contam- 
inata populations. The conchological similar- 
ity might result from convergent evolution 
only. However, because of the location of loc. 
749, it can also be hypothesized that ancient 
shipments between the harbour of Fiskardo 



54 



KEMPERMAN & DEGENAARS 



and the opposite Mycenean harbour in the 
Ormos Poleos on Ithaka caused human 
transport of A. с odysseus from Ithaka to 
Kephallinia, followed by hybridization with the 
local A. с contaminata. If that is the case, 
only some shell characters still witness the 
introgression. 

The range of A. с liebetruti on Kephallinia 
is very small, comprising probably less than 
one km 2 along the coast east of Sami. This 
area is situated close to the former land 
bridge to Ithaka. Nevertheless, A. с liebetruti 
is not found on Ithaka. Maybe this is because 
it could not penetrate an area occupied by 
another conspecific taxon. It is also possible 
that this subspecies is only a relatively recent 
human introduction on Kephallinia. If so, A. с 
liebetruti did not occur close to the Pleis- 
tocene land bridge until recently. Most con- 
chological characters of A. с liebetruti from 
Kephallinia are very similar to those of this 
subspecies from Zakynthos, where it is widely 
distributed (Rähle, 1979; Kemperman, in 
prep. A). This supports the second view. 
Meanwhile, the results of the allozyme anal- 
yses, especially with regard to the popula- 
tions of Iocs. 746 and 773, and conchological 
data suggest that there is introgressive hy- 
bridization between A. с liebetruti and A. с 
incommoda. Unfortunately, there are no allo- 
zyme data for populations of the former sub- 
species from Zakynthos. 

There are no synapomorphies defining A. 
contaminata as a monophyletic group (Table 
5). Within A. contaminata, A. с contaminata 
shares a synapomorphic allele (d) of the 
Aat-1 locus with A. с odysseus and a syn- 
apomorphic allele (e) of the same locus with 
A. с liebetruti; the shared presence of the 
b-allele of the Aat-1 locus in A с incommoda, 
А. с. liebetruti and A. с odysseus is consid- 
ered to be based on introgression, as dis- 
cussed below. No other synapomorphies 
were found within A. contaminata. With other 
species, however, A. contaminata has sev- 
eral apomorphies in common. The apomor- 
phic b-allele of the Gpi locus is shared with A. 
s. senilis and A. a. adrianae, whereas the 
apomorphic alleles с and d of Gpi are exclu- 
sively found in A. с contaminata and A. s. 
senilis. The presence of the Gpi alleles b, с 
and d in A. с contaminata and A. s. senilis is 
further discussed below. The b-allele at Mdh- 
2 is the synapomorphic character state for A. 
с contaminata, A. s. senilis and A. s. fla- 
vescens. Because these synapomorphies re- 
sult in conflicting conclusions, they do not al- 



low decisions on phylogenetic relations with 
respect to A. contaminata. 

Albinaria senilis: According to all distance co- 
efficients, only minor genetic variation occurs 
among the various populations of A. senilis. 
There is no differentiation between the sub- 
species. In both population dendrograms 
(Figs. 5, 6), A. s. senilis from loc. 709 is the 
sister group of the combined other A. senilis 
samples. At Iocs. 708 and 709, A. s. senilis 
lives truly sympatrically with A. j. jónica (Iocs. 
707, 708). At those sites, some specimens 
have been found that are morphologically in- 
termediate between A s. senilis and A /. jón- 
ica; allozyme analyses, however, group these 
intermediate specimens with A j. jónica (Fig. 
5). The position in the UPGMA-dendrograms 
of A s. senilis of loc. 709, of which only two 
specimens could be analysed, results from 
the fixation of both the Mdh-1 b-allele and the 
Lap b-allele. For Mdh-1 b, most other A s. 
senilis populations are also fixed; this allele is 
absent in A y. jónica. The Lap b-allele occurs 
in some other populations of A. s. senilis, A. с 
contaminata and A. j. jónica. Although ac- 
cording to the shell morphology this material 
was convincingly identified as A s. senilis, the 
analysed enzyme systems indicate that the 
specimens were hybrids between A s. senilis 
and A /. jónica. 

Within A s. senilis two geographically iso- 
lated populations, namely, loc. 752 on the Pa- 
liki-Peninsula and loc. 756 from Kastro, 
southeast of Argostoli, exhibit relatively high 
genetic distances from the other A senilis 
populations. Both populations are not clearly 
differentiated morphologically. 

The results of the allozyme frequency anal- 
yses did not support the taxonomic distinction 
of A s. kolpomyrtensis Kemperman & Gitten- 
berger, 1990, which was based on concho- 
logical and distributional data. In the dendro- 
grams, this subspecies is closely linked to A 
s. flavescens (Figs. 5-8), which is the geo- 
graphically adjoining subspecies. 

There are no apomorphic character states 
that point to A senilis, s.l., as a monophyletic 
group. This is not affected by limiting the anal- 
yses to a subset of the outgroup species. The 
apomorphic allele e of the Gpi locus is the 
only derived character state defining a sub- 
group within A senilis, namely, the combined 
A s. senilis and A s. kolpomyrtensis. All other 
apomorphies are shared with one or more 
other species, which results in conflicting phy- 
logenetic trees. The apomorphic c-allele of 



ALLOZYME FREQUENCIES IN ALBINARIA 



55 



Aat-1 defines A. s. senilis, A. s. flavescens 
and A. j. assicola as a monophyletic group. 
The c-allele of Ak groups A. s. senilis and A. 
j. jónica together; however, the presence of 
this allele in both taxa is considered to be the 
result from introgression, as discussed below. 
Both apomorphic alleles с and d of Nadd-1 
would point to the combined A. s. senilis, A. j. 
jónica and A. j. assicola as a monophyletic 
group, but the presence of these alleles in A. 
s. senilis is regarded too as being due to in- 
trogression. The cause of the distribution of 
the "synapomorphic" alleles b, с and d of Gpi 
in both A. s. senilis and A. с contaminata is 
further discussed below. The status of the b- 
allele of Gpi with respect to A. a. adrianae and 
A. s. senilis is unclear. The d-allele of Mdh-2 
is shared by A. s. flavescens, A. j. jónica and 
A. j. assicola, whereas the b-allele of Mdh-2 
could be a synapomorphy for A. s. senilis, A. 
s. flavescens and A. с contaminata. 

Albinaria jónica: The endemic A. jónica is di- 
vided into two widely disjunct subspecies. Al- 
binaria j. assicola occurs along the Kephallin- 
ian northwest coast, including the Assos 
Peninsula. Its range overlaps partly with that 
of A. с contaminata (Fig. 1), but the two spe- 
cies remain separate in habitat. Albinaria j. 
jónica is known from 3 to 4 km north of Ar- 
gostoli, along the west coast of the main is- 
land. It is found on the rocks along about 500 
m of the coastal road, where it occurs in a 
mixed population with A. s. senilis. Morpho- 
logically intermediate specimens have occa- 
sionally been found between A. j. assicola 
and A. с contaminata, as well as between A. 
j. jónica and A. s. senilis. 

With respect to both subspecies of A. jón- 
ica, there is a striking contrast between the 
results of morphological studies and those of 
allozyme analyses. The taxa have been rec- 
ognized as conspecific on the basis of con- 
chological features (Kemperman & Gitten- 
berger, 1990). Quite different from what is 
found among subspecies of A. senilis and A. 
contaminata, all UPGMA analyses demon- 
strate a relatively large genetic distance be- 
tween A. j. assicola (Iocs. 742 and 744) and 
A. j. jónica (Iocs. 707 and 708). Moreover, the 
affinities between these and other taxa, as 
indicated by the results of the UPGMA cluster 
analyses, suggest that A. jónica is not a 
monophyletic entity. Albinaria j. assicola is 
grouped among A. senilis subgroups, al- 
though it is partly sympatric not with this spe- 
cies, but with A. contaminata. The position of 



A. j. jónica is less constant. Nei's D (Figs. 5, 7) 
suggests that A. j. jónica and A. с contami- 
nata are sister groups; combined they are the 
sister group of A. a. adrianae. After Rogers' R 
(Figs. 6, 8), A. j. jónica is the sister group of 
the combined Kephallinian taxa. 

Contrary to the results of the UPGMA anal- 
yses is the outcome of a Hennigian analysis 
defining A. jónica, s.l., as a monophyletic 
group. At the Nadd-1 locus, с (present in Iocs. 
708 and 742) and d (present in Iocs. 708 and 
744) are synapomorphic alleles (Table 5); it is 
assumed that the occurrence of both alleles in 
A. s. senilis is the result of introgressive hy- 
bridization. Another synapomorphy for A. jón- 
ica would be the d-allele of Mdh-2, if one con- 
siders its occurrence in A. s. flavescens (loc. 
751 only) due to introgression. This could 
then date back to times when the present 
floor of the Gulf of Argostoli was dry land. The 
c-allele of Ak in A. j. jónica, shared with A. s. 
senilis (Table 5), will be discussed below. The 
presence of the apomorphic c-allele of Aat-1 
in A. j. assicola as well as in A. s. senilis and 
A. s. flavescens, might result from convergent 
evolution. 

Albinaria adrianae: The endemic A. adrianae 
is subdivided into two subspecies. This sub- 
division, primarily based on shell characters, 
is also strongly supported by the allozyme 
data. The four analyzed samples of A. a. adri- 
anae (Iocs. 725, 728, 729 and 730), proved to 
be genetically very similar. According to 
UPGMA population dendrograms based on 
Nei's D and Rogers' R (Figs. 5, 6), these pop- 
ulations are even identical. Material from loc. 
730, with a P value of 3.6%, differs only 
slightly from the other three localities, which 
were found to be monomorphic. Albinaria a. 
adrianae has not a stable position within the 
various dendrograms. According to Rogers' 
R, A. a. adrianae and A. contaminata are sis- 
ter groups. Corresponding to Nei's D, A. a. 
adrianae is the sister group of the combined 
A. contaminata and A. j. jónica. When Nei's D 
is calculated while either A. с contaminata or 
A. с incommoda are removed from the data- 
set in a jackknife procedure, A. a. adrianae 
switches from this position and becomes the 
sister group of the combined A. senilis, A. j. 
jónica and A. a. dubia. The second subspe- 
cies, A. a. dubia, is clustered either as the 
sister group of the combined A. senilis sub- 
species and A. j. assicola (Nei's D) or as the 
sister group of A. j. assicola alone (Rogers' 
R). This results from the fixation of the Mdh-1 



56 



KEMPERMAN & DEGENAARS 



allele b in A. a. dubia, as well as in A. j. assi- 
cola, A. s. flavescens and A. s. kolpomyrten- 
sis. However, no phylogenetic conclusions 
can be drawn on this allele distribution as no 
plesiomorphic character state can be deter- 
mined for this locus (Table 5). 

The allozyme analyses did not reveal one 
or more synapomorphic character states 
characterizing A. adrianae as a monophyletic 
group (Table 5). Moreover, the absence of the 
a-allele of MDH-2, which is a synapomorphy 
for all subspecies but A. a. dubia, points to the 
paraphyletic origin of A. a. adrianae and A. a. 
dubia. With the b-allele of the Gpi locus, A. a. 
adrianae has an apomorphic character state 
in common with both A. s. senilis and A. с 
contaminata. Because the presence of this al- 
lele in both A. s. senilis and A. с contaminata 
is considered to be the result of introgression 
from A. с contaminata to A. s. senilis, it is a 
synapomorphic character state for A. с con- 
taminata and A. a. adrianae. Because in our 
analyses A. a. dubia has no apomorphic char- 
acter states at all, its phylogenetic position 
remains puzzling. 



Hybridization and Introgression: Among Albi- 
naria species both hybridization and intro- 
gression are known to occur (e.g. Fuchs & 
Käufei, 1936; Nordsieck, 1983; Gittenberger, 
1991). For Kephallinian taxa, these phenom- 
ena are reported by Rähle (1979), Gitten- 
berger (1979) and Nordsieck (1979), who 
based their conclusions on morphological 
characters. We found some biochemical data 
pointing to introgression (Table 3). 

(1) As mentioned above, the c- and d-al- 
leles of the Nadd-1 locus are characteristic for 
A. jónica, s.l. ; in A. j. jónica they are present in 
loc. 708. These alleles occur also in the ad- 
joining A. s. senilis populations 702 and 706. 
It is assumed, therefore, that their very local 
presence in A. s. senilis results from intro- 
gression. 

(2) The o-allele of the Ak locus is only 
known from A. s. senilis at Iocs. 756 and 707, 
as well as from A. j. jónica at loc. 708. As 
localities 707 and 708 are adjoining, intro- 
gression of the Ak c-allele is hypothesized be- 
tween A. s. senilis and A. j. jónica. Although 
the distribution of this allele in A. s. senilis is 
less local than it is in A. j. jónica, the direction 
of the introgression is unclear. 

(3) The b-allele of Aat-1 occurs in all Ith- 
akian populations as well in A. с liebetruti of 
loc. 746 and A. с incommoda of population 



773. This is in roughly an area constituting 
Ithaka and the central eastern part of Kephal- 
linia. During the late Wurm sea-level lowering, 
Kephallinia and Ithaka were connected by a 
land bridge located close to this area. There- 
fore, introgression is a possible explanation. 
Albinaria с liebetruti might have received the 
allele either during the late Wurm period di- 
rectly from the same Ithakian source, or sec- 
ondarily from local Kephallinian A. с incom- 
moda. Thus, whether or not A. с liebetruti is a 
recent invader in Kephallinia cannot be de- 
cided on the basis of these data. For the pres- 
ence of the Aat-1 b-allele in A. s. senilis of loc. 
752, introgression cannot be hypothesized. 
We have to accept either convergent evolu- 
tion or indistinguishable, yet different electro- 
morphs. 

(4) The b-, c- and d-alleles of the Gpi locus 
are found in the adjoining A. с contaminata 
populations at Iocs. 713 (alleles b and c), 715 
(alleles b, с and d) and 716 (allele b). These 
alleles are also found in the nearby A. s. se- 
nilis populations at Iocs. 756 (alleles b, c, and 
d) and 706 (allele b). Their local presence in 
both species can be seen as the result of in- 
trogression, but it can also be hypothesized 
that the b-, c- and d-alleles of Gpi are exam- 
ples of what Woodruff (1989: 282) called hy- 
brizymes, "unexpected allelic electromorphs 
associated with hybrid zones." We found mor- 
phologically intermediate specimens, indica- 
tive of hybridization, in this area. Only the 
presence of the Gp/'-b allele in A. a. adrianae 
of loc. 730 is not in agreement with the hy- 
pothesis that hybrizymes are involved. 

(5) Locally, both in A. с contaminata pop- 
ulation 715 and in A. s. senilis population 752, 
the b-allele of the Pgm locus occurs. This can 
hardly be the result of introgression, unless 
ancestral populations once met during a gla- 
cial sea-level lowering. 

(6) At loc. 732, specimens morphologically 
intermediate between A. s. senilis and A. a. 
dubia are found, whereas at the nearby loc. 
721 , we collected specimens intermediate be- 
tween A. s. senilis and A. с contaminata. 
Only at these two localities we found the c- 
allele of the Mpi locus. Because loc. 732 is 
situated in a hybrid zone, the Mpi c-allele 
might be a hybrizyme (Woodruff, 1989). With 
respect to the material of loc. 721 , one might 
postulate introgression between loc. 732 and 
loc. 721 . 

(7) Some specimens of loc. 709 are con- 
chologically intermediate between A. s. seni- 
lis and A. j. jónica; the shells have the sien- 



ALLOZYME FREQUENCIES IN ALBINARIA 



57 



derness of A. j. jónica, the apertural shape of 
A. s. senilis, and an intermediate sculpture. In 
contrast, the allozyme analyses point to a ge- 
netic similarity with A. j. jónica (Figs. 5, 6). 
This is, for example, indicated by the pres- 
ence of the Mdh-2 d-allele, which might be 
considered typical for A. j. jónica. 

Apparently there are several localities on 
Kephallinia and Ithaka where introgression 
might explain the distribution of rare alleles 
(1-4). As an alternative for introgression, the 
presence of hybrizymes is suggested (4, 6). 
Again, in all cases, one must remember the 
possibility of convergent or parallel evolution 
or the presence of electromorphs that are in- 
distinguishable by our methods. 

The Outgroup Species 

All dendrograms (Figs. 5-8) show the 
same topology for the species serving as out- 
groups, namely, Albinaria sp. and Isabellaria 
edmundi irom the Peloponnesos, and A. teres 
nordsiecki and A. rebeli from Crete. They are 
linked outside the cluster formed by the spe- 
cies from Kephallinia and Ithaka, with a ge- 
netic distance varying between 0.18 and 0.41 . 
Albinaria t. nordsiecki is the sister group of 
Isabellaria edmundi; the combination of A. t. 
nordsiecki and /. edmundi is the sister group 
of A. rebeli. This cluster is the sister group of 
the combined Kephallinian and Ithakian Albi- 
naria taxa. The Albinaria sp. is grouped as the 
sister group of all other taxa. 

The grouping of Isabellaria edmundi within 
a cluster of Albinaria species, with a genetic 
distance far below the distance between a 
Cretan and a Peloponnesian Albinaria spe- 
cies, supports the view that Isabellaria sensu 
auctt. might be of polyphyletic origin and 
partly synonymous with Albinaria (Gitten- 
berger, 1987: 79). 



Electrophoretic Data, Morphology 
and Classification 



In this paper, we do not discuss in detail the 
results of morphological and morphomethcal 
analyses; these will be presented in a forth- 
coming article. Nevertheless, some remarks 
concerning electrophoretic data in relation to 
morphology can be made already. 

The genetic distance (D = 0.05 for Kephal- 
linian and Ithakian taxa, and D < 0.284 when 
including the outgroup-species) are very low 



with regard to their status as separate species 
(Menken & Ulenberg, 1987; Thorpe, 1982). 
Apparently we are dealing with most closely 
related "young" species, clearly character- 
ized by their distribution and a series of inde- 
pendent conchological characters, among 
which some localized introgression still oc- 
curs. This reminds one of what is described 
for five Partula species by Johnson et al. 
(1977: 122, 125), who found an D of 0.09. 
Davis et al. (1981) mentioned 0.01 < D < 0.1 
for the Elliptio complánala species group (Bi- 
valvia: Unionidae). For Cerion species, the 
following values are reported in the literature: 
D = 0.015 (D < 0.056) (Gould & Woodruff, 
1978: 407); D < 0.06 (Woodruff & Gould, 
1980: 397); D ± 0.05 (Gould & Woodruff, 
1986: 464); and D < 0.01 (Gould & Woodruff, 
1987: 343). Evidently, "taxonomic decisions 
do not follow directly from the estimation of D" 
(Gould & Woodruff, 1986: 466). 

Preliminary results of allozyme analyses of 
several Albinaria taxa from Crete are in line 
with this statement. Although both the distri- 
butional patterns and the conchological diver- 
sification strongly remind one of what is re- 
ported for the Kephallinian and Ithakian taxa 
here, genetic distances between the Cretan 
taxa were seven times larger (D = 0.35). This 
is considered indicative of speciation events 
that occurred further back in geological time 
(Schilthuizen, pers. com., 1991). 

For the Albinaria under study, the taxo- 
nomic relationships, derived from morpholog- 
ical and biogeographical data, are not in close 
congruence with the patterns found in allo- 
zyme variation. This becomes obvious sev- 
eral times independently. Illustrative are the 
widely separate positions of the subspecies of 
both A. jónica and A. adrianae among differ- 
ent clusters of A. contamínala and A. senilis, 
according to the allozyme data. Despite the 
fact that A. с odysseus from Ithaka is only 
slightly different morphologically from the 
Kephallinian A. с contamínala, the Ithakian 
subspecies has to be placed opposite the to- 
tal group of four polytypic Albinaria species 
represented in Kephallinia according to its al- 
lozymes. In some cases, forms that are con- 
sidered hybrid origin by morphological and 
distributional data, cannot be recognized as 
such electrophoretically. 

Despite the puzzling facts that remain, the 
electrophoretic analyses were an important 
tool in the multidisciplinary approach towards 
an understanding of the differentiation of 
Kephallinian and Ithakian Albinaria. 



58 



KEMPERMAN & DEGENAARS 



ACKNOWLEDGMENTS 

We are indebted to Prof. Dr. E. Gitten- 
berger, who initiated and supervised this 
project and gave critical comments on the 
various versions of this text. The second au- 
thor wishes to express her gratitude to Dr. G. 
M. Davis for teaching her the ins and outs of 
allozyme analyses on molluscs at his labora- 
tory at the Academy of Natural Sciences of 
Philadelphia, USA; many thanks are due also 
to C. Hesterman, who gave the daily technical 
support there. At Amsterdam, we were guests 
of Prof. Dr. S. B. J. Menken, who helped us 
adapting and continuing our runs and also 
commented on this paper. At Amsterdam, 
daily technical support was given by W. van 
Ginkel, Dr. J. W. Arntzen and Dr. W. M. 
Scheepmaker, for which we are very grateful. 
Dr. J. W. Arntzen also helped us with debug- 
ging the data-set and introduced us to 
BIOSYS. Dr. T. Backeljau of Brussels, Bel- 
gium, is kindly acknowledged for his introduc- 
tion to biochemical molluscan systematics. 



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ology, 9: 445-454. 

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Revised Ms. received 18 September 1991 



APPENDIX: LIST OF COLLECTION 
LOCALITIES 

Unless otherwise stated, the locality is on 
Kephallinia island. Locality numbers corre- 
spond with those on the map of Figure 2. 

319 A. senilis senilis (Rossmässler) — Near junc- 
tion to Moni Theotokou Sision (= 17.4 km 
SE of Argostoli), on rock, 210 m alt., 
DH701 7; 24-04-1987 

702 A. senilis senilis (Rossmässler) — 1 km E of 
Argostoli, W of junction Angon — Sami/Poros, 
on walls along road, 10 m alt., DH5626; 
16-03-1988 

706 A. senilis senilis (Rossmässler) — 400 m N of 
Ag. Konstantinos ( = 1 .5 km NE of Argostoli) 
on rocks along road, 10 m alt., DH5627; 17- 
03-1988 

707 A. jónica jónica (L. Pfeiffer) — Akros Kokkinos 
Vrachos, most western rocks along road, 
(= 3.5 km N of Argostoli), 50 m alt., 
DH5429; 17-03-1988 

707 A. senilis senilis (Rossmässler) — Akros Kok- 
kinos Vrachos (= 3.5 km N of Argostoli), 
most western rocks along road, 50 m alt., 
DH5429; 17-03-1988 

708 A. senilis senilis (Rossmässler) — 200 m N of 
Akros Kokkinos Vrachos ( = 3.7 km N of Ar- 
gostoli); on rocks along road, 50 m alt., DH 
5429; 17-03-1988 

708 A. jónica jónica (L. Pfeiffer) — 200 m N of Ak- 
ros Kokkinos Vrachos ( = 3.7 km N of Argos- 
toli), on rocks along road, 50 m alt., DH5429; 
17-03-1988 

709 A. senilis senilis (Rossmässler) — 500 m N of 
Akrcs Kokkinos Vrachos (= 4 km N of Ar- 
gostoli), along road on rocks, 55 m alt., 
DH5429; 17-03-1988 

710 A. senilis senilis (Rossmässler) — Spilia, S 
exit village ( = 1 .7 km S of Argostoli), along 



road on rocks covered with vegetation, 120 
m alt., DH5524; 18-03-1988 

712 A. senilis senilis (Rossmässler) — Lakythra, 
50 m N of northern entrance ( = 6.2 km SSE 
of Argostoli), on rocks in field and on low 
rock-face, W side of road, 230 m alt., 
DH5720; 18-03-1988 

713 A. contaminata contamínala (Rossmäss- 
ler) — Lakythra, 50 m N of northern entrance 
( = 6.2 km SSE of Argostoli), on rocks in field 
and on low rock-face, E side of road, 230 m 
alt., DH5720; 18-03-1988 

715 A. contaminata contaminata (Rossmäss- 
ler) — 0.5 km NW of Lakythra, in road-cleft on 
hill-top (= 5.3 km SSE of Argostoli), W side 
of road on rock-face, 240 m alt., DH5721; 
18-03-1988 

716 A. contaminata contaminata (Rossmäss- 
ler) — 0.5 km NW of Lakythra, in road-cleft on 
hill-top (= 5.3 km SSE of Argostoli), E side 
of road on rock-face, 240 m alt., DH5721 ; 
18-03-1988 

719 A. senilis senilis (Rossmässler) — Atsou- 
pades, 200 m NW of junction for Arginia 
(= 22.1 km ESE of Argostoli), on rocks 
along road, 290 m alt., DH7416; 21-03-1988 

722 A. adrianae dubia Gittenberger — W exit of 
Markopoulon (= 24.1 km ESE of Argostoli), 
on rocks along road, 285 m alt., DH7615; 
21-03-1988 

723 A. contaminata incommoda (Boettger) — 300 
m N of Ag. Georgios (= 24.3 km ESE of 
Argostoli), on wet soft sand-'stone', 220 m 
alt., DH781 8; 21-03-1988 

725 A. adrianae adrianae Gittenberger — Poros, 
rock-faces N side of bridge in cleft (25.1 km 
E of Argostoli), type loa, 10 m alt., DH8023; 
21-03-1988 

727 A. contaminata incommoda (Boettger) — 180 
m N of Poros-town bridge (= 25 km E of 
Argostoli), on isolated rocks between 
houses along coastal road, 1 m alt., 
DH8023; 21-03-1988 

728 A. adrianae adrianae Gittenberger — 200 m 
N of Poros-town bridge ( = 25 km E of Ar- 
gostoli), on isolated rocks between houses 
along coastal road, 1 m alt., DH8023; 21- 
03-1988 

729 A. adrianae adrianae Gittenberger — Gorge 
3.8 km SE of Poros ( = 28.3 km ESE of Ar- 
gostoli) on lower and higher rocks of gorge. 
2-10 m alt., DH8220; 21-03-1988 

730 A. adrianae adrianae Gittenberger — Poros, 
N rocks along river E from bridge in cleft 
(= 25.1 km E of Argostoli), 5-15 m alt., 
DH8023; 21-03-1988 

731 A. adrianae dubia Gittenberger — 2.5 km NW 
of Ag. Georgios, type loc. ( = 22.8 km ESE of 
Argostoli) on high rock faces, 400 m alt., 
DH761 8; 21-03-1988 

734 A. senilis flavescens (Boettger) — 1 km W of 
Kourouklata (= 6.4 km N of Argostoli), on 



ALLOZYME FREQUENCIES IN ALBINARIA 



61 



rocks along road, 140 m alt., DH5432; 22- 
03-1988 

735 A. contaminata contaminata (Rossmäss- 
ler) — 1 km NE of N exit of Angon ( = 14 km 
NNE of Argostoli); on rocks along road, 250 
m alt., DH5641; 22-03-1988 

736 A. s. kolpomyrtensis Kemperman & Gitten- 
berger — 2 km NE of N exit of Angon ( = 1 5.3 
km N of Argostoli), on rocks along road, 260 
m alt., DH5641; 22-03-1988 

742 A. j. assicola Kemperman & Gittenberger — 
Assos Peninsula, rocks along lower part of 
road to main entrance fortress ( = 22.5 km 
NNE of Argostoli), 2-5 m alt., DH5948; 22- 
03-1988 

744 A. j. assicola Kemperman & Gittenberger — 
Assos Peninsula ( = 22.5 km NNE of Argos- 
toli), fortress and southern rocks, 100 m alt., 
DH5948; 22-03-1988 

746 A. contaminata liebetruti (Boettger) — 1 km 
NE of Sami (= 17.2 km ENE of Argostoli), 
on rocks W of Tomb-cave, 35 m alt., 
DH7035; 22-03-1988 

749 A. contaminata (Rossmässler) — 1 km S of 
Fiskardo ( = 30.8 km NNE of Argostoli), on 
rocks along bay, 10 m alt., DH6356; 24-03- 
1988 

751 A. senilis flavescens (Boettger) — 1 .5 km S of 
Livadi, = 1 km N of Ag. Dimithos (= 8.5 km 
NW of Argostoli), on sand-stone wall along 
road, 5 m alt., DH5033; 23-03-1988 

752 A. senilis senilis (Rossmässler) — Lepada, 
2.5 km S of Lixouri ( = 4.5 km W of Argos- 
toli), 100 m N of beach entrance on low 
rocks along coast, 2 m alt., DH5125; 23-03- 
1988 

756 A. senilis senilis (Rossmässler) — Kastro Ag. 
Georgios, inside fortress on stone walls 
(= 7.0 km SE of Argostoli), 290 m alt., 
DH6021; 24-03-1988 

759 A. contaminata contaminata (Rossmäss- 
ler) — 4 km NW of broadcasting plant on Me- 
gas Oros ( = Oros Aenos)( = 1 4.3 km E of 
Argostoli), on rocks along road, 1260 m alt., 
DH6723; 25-03-1988 



764 A. contaminate odysseus (Boettger) — Ithaka 
Isl., Perachori, in/on ruin Venetian Church in 
lower part village ( = 1 .7 km S of Vathi), 1 70 
m alt., DH7545; 26-03-1988 

765 A. contaminata odysseus (Boettger) — Ithaka 
Isl., 600 m S of Agros, on rock face along 
road-curve (= 4.5 km W of Vathi), 160 m 
alt., DH7147; 26-03-1988 

766 A. contaminata odysseus (Boettger) — Ithaka 
Isl., W side of Poly-bay (= 11.2 km NW of 
Vathi), on rocks and walls, 1-5 m alt., 
DH6855; 26-03-1988 

770 A. contaminata odysseus (Boettger) — Ithaka 
Isl., Kioni, most eastern port of town ( = 9 km 
N of Vathi), along coastal road on wall, 5 m 
alt., DH7355; 26-03-1988 

772 A. contaminata incommoda (Boettger) — 2 
km NW of Tzanata ( = 21 .8 km E of Argos- 
toli), near bridge on sediment wall along old 
road to Mon. Atrou, 130 m alt., DH7623; 27- 
03-1988 

773 A. contaminata incommoda (Boettger) — 1.1 
km E of Koulourata ( = 16.9 km E of Argos- 
toli), on high rock-faces along road, 420 m 
alt., DH71 28; 27-03-1988 

Outgroup species, locality-numbers are not 
on Figure 2. 

1 A. teres nordsiecki Zilch — Greece, Crete, 
Province of Lasithi, Kavousi ravine, 50 m alt; 
28-04-1988 

2 A. rebel! Wagner — Greece, Crete, Province 
of Lasithi, NW of Orino, 620 m alt.; 6-04- 
1988 

3 A. nov. spec. — Greece, Peloponnesos, 
Province of Arkadhia, E slope Parnon Moun- 
tain at 8.5 km SW of Astros, shadowed rocks 
in coniferious forest, 400 m alt., UTM FG43; 
27-07-1988 

4 Isabellaria edmundi Gittenberger — Greece, 
Peloponnesos, Province of Arkadhia, high 
rock-face along the forest road, 5 km (6 km 
along the road) N of Kosmas, 975 m alt., 
UTM FG51; 27-07-1988 



MALACOLOGIA, 1992, 34(1-2): 63-73 

EFFECT OF STARVATION AND HIBERNATION ON THE 

FINE STRUCTURAL MORPHOLOGY OF DIGESTIVE GLAND CELLS 

OF THE SNAIL HELIX LUCORUM 

Vasilis K. Dimitriadis & Dimitris Hondros 

School of Biology, Faculty of Sciences, Aristotle University of Thessaloniki, 
Thessaloniki 540 06, Greece 

ABSTRACT 

Thirty seven days hibernation resulted in significant changes in the morphology of the diges- 
tive gland epithelium of Helix lucorum compared to control animals: the number of the digestive 
cells significantly decreased, but the number of the other cell types of digestive gland, calcium 
and excretory cells significantly increased. The apical granules and the cisternae with dense 
cores of the digestive cells significantly increased in number and size, while the calcium granules 
of the calcium cells increased in number and showed more intense concentric rings compared 
to those of the control animals. In the excretory cells, the large vacuoles significantly increased 
in size, while in all the digestive gland cells the lipid inclusions decreased in number as hiber- 
nation proceeded. In some cases, cytoplasmic material was extruded into the lumen, and in- 
tercellular spaces appeared to be very dilated. 

Forty days of starvation induced similar but less intense phenomena in the digestive gland of 
Helix lucorum. The possible physiological function of the crop epithelial cells and the digestive 
gland cells of Helix lucorum is discussed. 

Key words: Helix lucorum, snail, digestive gland, fine structure, starvation, hibernation. 



INTRODUCTION 

Snails exposed to starvation or hibernation 
respond by minimizing their energy require- 
ments (Prosser, 1973). Carbohydrate metab- 
olism is the principal energy source during hi- 
bernation, whereas during starvation oxygen 
uptake decreases sharply during the first day 
and more slowly later (von Brand, 1931; von 
Brand et al., 1948). However, little is known 
about the biochemistry and the physiology, as 
well as the fine structural morphology of 
snails exposed to such conditions as hiberna- 
tion and starvation. There is little information 
about the fine structure of the gut and diges- 
tive gland epithelium and other visceral tis- 
sues of snails exposed to starvation and hi- 
bernation (Sumner, 1965; Oxford & Fish, 
1979; Janssen, 1985; Roldan, 1987). The 
majority of the data available deals with the 
fine structure and the physiology of the diges- 
tive gland mainly under normal conditions 
(see, for example, Sumner, 1965, 1966; 
Owen, 1966; Walker, 1970; Bowen & Davies, 
1971; Oxford & Fish, 1979; Pipe, 1986; 
Roldan, 1987). 

In the present study, the cells of the diges- 
tive gland of Helix lucorum were studied with 
the light and electron microscope under nor- 



mal conditions. In addition, the morphology of 
the digestive gland was studied in starving 
and hibernating animals and was compared 
with the morphology existing under normal 
conditions. 



MATERIALS AND METHODS 

Adult snails of Helix lucorum (Gastropoda, 
Pulmonata, Helicidae) were collected from 
Edessa, northern Greece. The shell diameter 
of the snails used in the present study varied 
from 41 to 43 mm and the body weight from 
about 19 to 21.5 g. 

Snails, deprived of food, were kept in a cold 
room at 4± 1 °C under a 9 h light (L): 1 5 h dark 
(D) photopehod for 8, 12, 22 and 37 days, in 
order to be exposed to hibernating conditions 
(Lazahdou-Dimitriadou & Saunders, 1986). 
Control feeding snails were kept at 19±1°C 
and 13L:11D photopehod. Starved snails 
were kept at 19±1°C and 13L:11D photope- 
hod for 1 5, 25 and 40 days. Four animals from 
each experimental group were dissected late 
morning, and the digestive gland was fixed in 
Karnovsky's fixative (Karnovsky, 1965), post- 
fixed in 2% osmium tetroxide, dehydrated and 
embedded in Spurr's resin. Sections were cut 



63 



64 



DIMITRIADIS & HONDROS 




FIG. 1. Drawing of the various cell types observed in the digestive gland epithelium of H. lucorum. CC, 
calcium cell; DC, digestive cell; EC, excretory cell. 



using a Reichert Om U3 ultramicrotome. The 
sections were pcststained with lead citrate 
and uranyl acetate and examined under a Ja- 
pan Electron Optics Laboratory Co. 100B 
electron microscope operating at 80 KV. For 
light microscopic observations, thick sections 
were stained with 1 % toluidine blue. 

Morphometric evaluation was performed 
according to Weibel (1979) and Steer (1981). 
Samples of five cells were counted from every 
digestive gland section of four different snails. 
The volume densities of certain morphometric 
parameters of crop and digestive gland or- 
ganelles were determined from point counting 
stereology, using a test square lattice with a 
period d = 10 mm, equivalent to 1 |xm on the 
specimen. A minimum of 480 points were 
counted per cell. The mean absolute volume 
of the various cells were estimated using the 
mean absolute volumes of the nuclei (using, 
depending of the cell type, the formula for a 
spheroid of prolate spheroid) and the ratio of 
nuclear/cytoplasmic volume. The mean total 
volume per cell of the various cell compo- 
nents were determined by their volume den- 
sities and the absolute volumes of the cor- 



responding cells. The percentages of the var- 
ious epithelial cell types within the digestive 
gland epithelium were determined from mea- 
surements on light microscopic micrographs 
at a final magnification of x 500. Cells were 
identified on the basis of morphologic criteria 
given in the Results section. Mean values and 
standard deviations of the morphometric pa- 
rameters were calculated and statistically 
compared using Student's t-test, significant 
level P < 0.05. 



RESULTS 

Morphology of the Digestive 
Gland Epithelium 

The digestive gland epithelium of Helix lu- 
corum consisted of three cell types: digestive 
cells, calcium and excretory cells (Fig. 1). In 
cross sections of the gland tubules, the cells 
were located around a central lumen, and the 
whole structure was surrounded by connec- 
tive tissue, muscular layers and a sys- 



STARVATION IN HELIX LUCORUM 



65 



tern of haemocoelic spaces surrounded by 
amoebocytes. 

Digestive cells were the most frequent cell 
type found in the digestive gland (Fig. 6). 
They usually appeared in a columnar shape, 
varied in size, and usually prossessed mi- 
crovilli (Fig. 2). Digestive cells were charac- 
terized by numerous granules and cisternae 
of varying size and electron density. In the 
apical portion of these cells, there was usually 
a number of granules filled with a homoge- 
neous material of a low electron density (Figs. 
2, 3). In the middle region of the cells, smaller 
cisternae were also observed containing two 
or more intracisternal cores (Figs. 2, 3). At the 
base of the digestive cells, a few lipid inclu- 
sions were concentrated in clusters (Fig. 4). 
The nuclei of the digestive cells were usually 
located in the basal or in the middle regions of 
the cell and were usually well defined. 

Calcium cells were the second cell type 
found in the digestive gland epithelium of H. 
lucorum. Their number was smaller than that 
of the digestive cells (Fig. 6). They were usu- 
ally pyramidal in shape and did not usually 
protrude into the lumen, but they were located 
at the base of the epithelium. Their height was 
less than that of the digestive cells (Fig. 6). 
Calcium cells were usually filled with calcium 
granules which were located mainly in the 
basal region of the cells. Calcium granules 
were spherical in shape (Fig. 5) and were of a 
similar size throughout each cell. In high mag- 
nification, the calcium granules showed inner 
concentric rings. The nuclei of the calcium 
cells were usually located in their basal re- 
gion, where lipid inclusions were often ob- 
served. 

Excretory cells were the third cell type 
found in the digestive gland epithelium of H. 
lucorum (Figs. 12, 14). Their size was larger 
than that of digestive cells, their shape varied 
between ovoid and columnar, and their cyto- 
plasm was characterized by one or more 
large cisternae containing cores or amor- 
phous mass of high electron density. These 
cisternae occupied a large portion of the cell 
volume (Fig. 1 2). The nucleus was usually po- 
sitioned at the basal region of the excretory 
cells and in their apical portion small vacuoles 
of low electron density were observed. 

Morphology of the Digestive Gland 
Epithelium under Starving Conditions 

Snails exposed to starving conditions 
showed modified morphology of the digestive 



cells compared to the control animals. After 
40 days of starvation, a significant decrease 
in the number of these cells were noticed (Fig. 
6). A significant increase in the total volume 
per cell of the apical granules located in the 
digestive cells compared to the control ani- 
mals was also noticed in the same period 
(Figs. 7, 8). In contrast, the cisternae with the 
electron dense cores did not change signifi- 
cantly their total volume per cell after 40 days 
of starvation compared to the control animals 
(Fig. 7). Lipid inclusions decreased in number 
and almost disappeared after 40 days of star- 
vation compared to control animals, while 
calcium granules increased their number and 
consequently their total volume per cell (Figs. 
6, 7). In high magnification, the calcium gran- 
ules also showed more concentric rings in 
starved snails (Fig. 11). However, the diame- 
ter of the calcium granules was similar in both 
starved and control animals. 

Excretory cells significantly increased in 
number and volume after 40 days of starva- 
tion compared to the control animals (Figs. 6, 
9). Moreover, the large cisternae of these 
cells significantly increased in size (Figs. 7, 9) 
so that they occupied over than half of the cell 
volume. Generally, the Golgi complexes and 
the rough endoplasmic reticulum of these 
cells were quite distinguishable and well or- 
ganised in this stage. 

After 25 and 40 days of starvation, extru- 
sions of cytoplasm of whole cells into the 
gland lumen were apparent. Degeneration 
phenomena were rarely observed, while no of 
autophagic vacuoles were visible. The mem- 
brane infoldings appeared more intense, 
mainly in the basal region, and the intercellu- 
lar spaces were more dilated than that of the 
control animals. 

Morphology of the Digestive Gland 
Epithelium under Hibernating Conditions 

After eight days of dormancy, the digestive 
gland cells did not show significant morpho- 
logical changes. The only change noticed 
concerned the calcium cells, which showed 
more electron-dense concentric rings com- 
pared to those in control animals. 

Thirty seven days exposure of H. lucorum 
in hibernating conditions induced fine struc- 
tural modification of digestive gland cells. The 
digestive cells significantly increased in size, 
while their number decreased by about 25% 
(Fig. 6) compared to the control animals. The 
total volume per digestive cell of the apical 



66 



DIMITRIADIS & HONDROS 




' 



Mv 




L~- 



DV, 




¥Mdv 2 



s 



#* 



PM 



œ/1 



3 




*** 




Li 



BM. 



1»***лЕ 




H 



61 



FIG. 2. Digestive cells contain apical granules with homogenous material (DV,) and smaller cisternae with 

electron dense cores (DV 2 ). Mv, microvilli. Bar = 4 p.m. 

FIG. 3. Higher magnification view of the apical granules (DV,) and of the cisternae with dense cores (DV 2 ) 

located in the apical portion of digestive cells. Mv, microvilli; Pm, plasma membranes. Bar = 6 ц.т. 

FIG. 4. Numerous lipid inclusions (Li) appear in clusters at the base of digestive cells. BM, basement 

membrane, IS, intercellular space; MS, muscles. Bar = 3 |xm. 

FIG. 5. Calcium cells contain numerous calcium granules (CGs) with inner concentric rings. Bl, basal 

infoldings; N, nucleus. Bar = 3 цт. 



STARVATION IN HELIX LUCORUM 



67 



OJO 

•>— > 
С 
<D 
О 

i-H 




Control 
gg Starved 
Щ Hibernated 



E 



< 



10000 -, 




Digestive cells Calcium cells Excretory cells 

FIG. 6. Distribution (%) (A) and absolute volumes (B) of the various cell of control, starved (40 days) or 
hibernated animals (37 days) observed in the digestive gland of H. lucorum. Significantly different values (P 
< 0.05) among control and starved or control and hibernated animals are indicated by asterisks. 



granules, as well as of the cisternae with the 
electron dense cores, significantly increased 
compared to the control animals (Fig. 7). Cal- 
cium cells significantly increased in number 
and slightly in size at that period (Fig. 6). As 
hibernation proceeded, calcium granules in- 
creased in number (Fig. 7) and appeared in 
clusters. The concentric rings in the calcium 
granules also had a higher electron density 



and increased in number in relation to the 
control animals (Fig. 11). 

Another morphological effect of hibernation 
was a significant increase in the absolute vol- 
umes of the excretory cells compared to the 
control animals (Fig. 6). After 37 days of hi- 
bernation, the large cisternae of the latter 
cells also greatly increased in size and often 
occupied more than half of their cell volume 



68 



DIMITRIADIS & HONDROS 



n 

E 



Q. 

S 

о 
> 



£ 



10000 -, 



8000 - 



6000- 



4000. 



2000. 



Control 
Starved 
Hibernated 




Apical granules Cistemae with Granules of Large cisternae 
of digestive cells dense cores of calcium cells of excretory 
digestive cells cells 

FIG. 7. Total volume per cell of cell components of digestive, calcium and excretory cells of control, starved 
(40 days) or hibernated cells (37 days) of digestive gland of H. lucorum. Significantly different values (P < 
0.05) among control and starved or control and hibernated animals are indicated by asterisks. 



(Figs. 7, 12, 13, 15). In all the cells studied, 
the number of lipid inclusions observed at the 
basal region greatly decreased 37 days after 
the exposure of the animals to hibernation, 
while the extracellular spaces formed by the 
infoldings of the basal plasma membrane and 
the intercellular spaces appeared very dilated 
in relation to the control animals. Cytoplasmic 
regions often extruded into the lumen. In ad- 
dition, whole cells appeared to have degen- 
erated and been released into the lumen (Fig. 
10). 



DISCUSSION 

There are numerous studies on the mor- 
phology and physiology of epithelial cells in 
the digestive gland of the Pulmonata (Sum- 
ner, 1965, 1966; Walker, 1970; Bowen & Dav- 
ies, 1971 ; Oxford & Fish, 1979) that are con- 
sistent with the fact that this, the largest 
organ, participates in digestion, absorption 
and storage of food material previously di- 
gested in the crop and stomach lumen. How- 
ever, there are conflicting reports about the 
cell types that constitute the digestive gland 
epithelium of Pulmonata. Thus, Krijgsman 
(1925, 1928) considered digestive and excre- 
tory cells as different cell types. David & 
Götze (1963) distinguished one category of 
digestive calcium cells, whereas Billett & Mc- 



Gee-Russell (1955) considered calcium and 
excretory cells as distinct forms of digestive 
cells. Other studies supported the existence 
of three types of digestive gland cells in slugs: 
digestive, calcium and thin cells (Abolins-Kro- 
gis, 1961; Morton, 1979), considering excre- 
tory cells as digestive cells. 

After 40 days of starvation or 37 days of 
hibernation, there was an increase in the vol- 
ume of the digestive cells, as well as an in- 
crease in the total volume per cell of the api- 
cal granules in the digestive cells. This fact is 
consistent with the results of Sumner (1965), 
who reported that starvation resulted in vac- 
uolisation of the digestive cells of Helix as- 
persa, as well as in an increase in size of the 
green granules compared to the control ani- 
mals. Similar granules to the apical granules 
of digestive cells of H. lucorum were reported 
in the digestive glands of Arion hortensis, Lit- 
torina littorea and Cepaea nemoralis (Bowen 
& Davies, 1971; Oxford & Fish, 1979; Pipe, 
1986) and were characterized as hetero- 
phagic, phagolysosomal or heterolysosomal 
cisternae. On the contrary, Krijgsman (1925, 
1928), Walker (1970) and Sumner (1965), in 
Helix pomatia and Agriolimax reticulatus, 
referred to similar granules as green vacuoles 
due to their staining with light green. There 
are also histochemical results in other snails 
indicating that these cisternae display a pos- 
itive PAS (Walker, 1969), acid phosphatase, 



STARVATION IN HELIX LUCORUM 



69 



1 ^i». /^' 



« 



DV, 



8 



% 



Mv 



Mv 




Ж UN 

10 ? ï 




CRs *•» 

/ 



11 




FIG. 8. 40 days' starvation. Apical portion of digestive cells possessing large number of apical granules 
(DVJ. DV 2 , cisternae with dense cores; Lu, lumen; Mv, microvilli; PM, plasma membrane. Bar = 3 \im. 
FIG. 9. 40 days' starvation. Excretory cells display many large cisternae (EV) with large electron dense 
cores. Mv, microvilli; PM, plasma membrane; CC, calcium cell. Bar = 5 p.m. 

FIG. 10. 22 days' hibernation. Part of the cytoplasm of a digestive gland cell shows degenerating phenom- 
ena. Mi, mitochondria; G, granules. Bar = 3 |xm. 

FIG. 1 1 . 37 days' hibernation. Calcium granules containing two or more inner concentric rings (CRs). Bar = 
1 |i.m. 



70 



DIMITRIADIS & HONDROS 



Mv 



DVo 



DV, 



CGs | 




«f 




2 во 



Li 



CGs 



13 




FIGS. 12, 13. 37 days' hibernation. Part of large cisternae (EV) with large electron-dense cores occupying 
great cytoplasmic areas of excretory cells. CGs, calcium granules; DV,, apical granules; DV 2 , cisternae with 
dense cores; Li, lipid inclusions; Mv, microvilli. Fig. 12 Bar = 5 |xm. Fig. 13 Bar = 6 p.m. 
FIGS. 14, 15. Light microscopic view of digestive gland epithelium of control (Fig. 14) and hibernating 
animals (Fig. 15). Many excretory cells (EC) and calcium cells (CC) are observed in the epithelium of the 
hibernating snails, compared to the control ones. DC, digestive cells; Lu, lumen. Fig. 14 Bar = 20 |xm. Fig. 
15 Bar = 20 ц -m. 



STARVATION IN HELIX LUCORUM 



71 



esterase (Bowen, 1970; Bowen & Davies, 
1971) and B-glucuronidase reactions (Billett 
& McGee-Russell, 1955). This fact probably 
indicates that elaboration of material with cer- 
tain enzymes inside these cisternae occurs. 

After 37 days of hibernation, the total vol- 
ume per digestive cell of the cisternae with 
dense cores, which probably correspond to 
the green granules of Helix aspersa, was 
found to have increased. Similar cisternae 
have been reported in the equivalent cells of 
H. aspersa, where they were characterized as 
yellow granules and exhibited positive reac- 
tions for calcium and lipofuscin, as well as 
positive PAS activity (Sumner, 1965). Also, 
Pipe (1986) observed similar cisternae in the 
digestive cells of Littorina littorea and sug- 
gested that they probably were cut off from 
heterolysosomal vesicles and filled with 
waste material. Sumner (1965) also sup- 
ported the view that similar cisternae could 
contain remnants of intracellular digestion 
that must be extruded to the central lumen. 

From the above-mentioned reports it could 
be concluded that the apical granules and cis- 
ternae with dense cores in digestive gland 
cells are structures involved in endocytotic 
processes. There are reports supporting the 
existence of phagocytosis in the digestive 
glands of most Gastropoda (Owen, 1966; Ox- 
ford & Fish, 1979). Owen (1966) claimed that 
the existence of phagocytosis in certain spe- 
cies of Mollusca may be an adaptation to 
functional needs. In the present study, the 
fine structural characteristics of digestive cells 
of H. lucorum support the view that these cells 
are responsible for absorption and digestion 
of food material, as well as production and 
secretion of digestive enzymes. However, 
phagocytosis of solid food material by the 
cells examined was not observed. It is possi- 
ble that nutrients are absorbed by small pi- 
nocytotic vesicles forming the apical granules 
and consequently are degraded by lysosomal 
action, producing the cisternae with dense 
cores. Thus, the increase of the total volume 
per cell of the apical granules and of the cis- 
ternae with dense cores could probably be 
attributed to the accumulation of residual in- 
digestible material inside the digestive cell, 
reflecting decreased digestive and excretory 
functions of these cells due to hibernation. 

The absolute volume of the excretory cells, 
and the volume of their large cisternae, in- 
creased during hibernation. There is a con- 
troversy about the origin and the function of 
excretory cells. Thiele (1953) and Sumner 



(1965) suggested that they are degenerating 
calcium cells, as they contained only a small 
quantity of DNA in their nuclei, little cytoplasm 
and a small number of mitochondria. Fretter 
(1952), Billett & McGee-Russell (1955) and 
Walker (1970) proposed that they are differ- 
entiated forms of digestive cells. Abolins- 
Krogis (1961) found mucopolysaccharides, 
proteins, lipids and small quantities of RNA in 
similar vacuoles of Helix aspersa and sug- 
gested that this material is used for shell re- 
pair. In H. lucorum, the increase in size of the 
large cisternae, is probably a result of accu- 
mulation of excretory material inside the ex- 
cretory cells, which probably reflects the min- 
imizing of their excretory processes during 
hibernation. 

In the present study, the electron micro- 
scopic examination of H. lucorum digestive 
glands revealed the presence of three cell 
types: digestive cells, calcium cells and ex- 
cretory cells. The morphological features sug- 
gest that excretory cells constitute a distinct 
cell type. Morton (1975, 1979) supported the 
view that excretory cells of Pulmonata are di- 
gestive cells in different stages of a sequence 
of cytological changes over the course of 24 
hours. Also Walker (1970) supported the view 
that excretory cells of Agriolimax reticulatus 
digestive glands are degenerating cells full of 
lipofuscin, which are formed from digestive 
cells. If this hypothesis can be supported, the 
excretory cells should be the final step in the 
development of the digestive cells. In that 
case, the excretory cells contain the digestive 
remnants of ingested material, which are de- 
graded by lysosomal action and are finally 
discharged into the gland lumen. Arising from 
this hypothesis are the following questions: 
(1) Why is the population density of the ex- 
cretory cells in H. lucorum digestive glands 
apparently smaller than that of the digestive 
cell? (2) Why are there no cell types interme- 
diate between digestive and excretory cells in 
the examined digestive glands? 

Another characteristic feature that 37 days 
hibernation induced was an increase in the 
number of calcium cells. This is evidently due 
to the participation of these cells in the forma- 
tion of the calcareous epiphragm that covers 
the peristome of the snail during hibernation. 
The role of calcium in gastropods and other 
molluscs is multiple: it is regarded as a com- 
ponent being implicated in pH homeostasis 
(Sminia et al., 1977; Akberalli et al., 1977; 
Jokumsen & Fyhn, 1982), in reproduction 
(Tompa & Wilbur, 1977; Fournie & Chetail, 



72 



DIMITRIADIS & HONDROS 



1982), in regulation of freezing tolerance 
(Murphy, 1977), as a component of mucus 
(Barr, 1928), in metabolic waste production 
(Kniprath, 1975), and in shell regeneration 
and formation of the epiphragm during hiber- 
nation (Abolins-Krogis, 1961, 1965; Wagge, 
1951). In pulmonates, calcium appears as 
carbonate (Wagge, 1951) or phosphate 
(Khjgsman, 1928; Oxford & Fish, 1979), in a 
crystalline form or as an amorphous mass 
(Wagge, 1951). In the hepatopancreas of He- 
lix aspersa (Howard et al., 1981), calcium 
granules contain CaMgP 2 7 . These granules 
are sites for accumulation of a wide variety of 
cations, acting as a detoxification mechanism 
that traps a number of dietary metals. How- 
ever, the wide functions of these granules are 
largely unknown (Taylor et al., 1988). 

Autolysis and extrusions of cytoplasmic 
material into the digestive gland lumen of H. 
lucorum were increased significantly as star- 
vation and hibernation proceeded. A similar 
cell response is also referred to by Bowen & 
Davies (1971) and Oxford & Fish (1979) in 
Arion hortensis and Cepaea nemoralis, where 
release of hydrolytic enzymes from the cyto- 
plasm of digestive cells was related to an in- 
crease in the rate of cell autolysis. In H. luco- 
rum, the extrusion of cytoplasmic pieces into 
the lumen was possibly related to the reduc- 
tion of energy requirements of the epithelium 
and the maintenance of low function activity 
of the digestive gland cells. 

By using light and electron microscopic 
observations and stereological evaluation, 
the present study provides fine structural in- 
formation about how hibernation and starva- 
tion affects the cells of the digestive gland of 
H. lucorum. However, more information, es- 
pecially from the use of cytochemical meth- 
ods, is needed for a better understanding of 
the physiology of the phenomenon of hiber- 
nation in snails. 



ACKNOWLEDGEMENTS 

This work was supported financially by the 
Greek Ministry of Agriculture. We are grateful 
to Dr. M. Lazaridou-Dimithadou, who pro- 
vided the snails and kept them under normal, 
starvation and hibernation conditions. 



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Revised Ms. accepted 19 September 1991 



MALACOLOGIA, 1992, 34(1-2): 75-86 

SUPRASPECIFIC NAMES OF MOLLUSCS: A QUANTITATIVE REVIEW 

Philippe Bouchet & Jean-Pierre Rocroi 
Muséum National d'Histoire Naturelle, 55 Rue de Buffon, Paris 75005, France 

ABSTRACT 

The number of nomenclaturally available genus-group taxa described since 1758 for Recent 
and fossil molluscs has been estimated by two methods. One result stands at 28,400 names; 
another at 24,900 names, of which 12,700 are gastropods, 6,000 are cephalopods, 5,100 are 
bivalves and 1,100 are in the smaller classes. The yearly increment appears to have remained 
relatively stable since the late 1 9th century. It is presently at an average of 224 new genus-group 
names per year, with Cephalopoda representing precisely one-third of the names. At least 20% 
of the recently introduced taxa escape the Zoological Record, with the Soviet and paleontológ- 
ica! literatures especially underrepresented. In the last 30 years, journals and books published 
in USSR, USA, and China contain altogether 50% of the new names. The total number of 
nomenclaturally available genus-group names of Recent molluscs is on the order of 12,000. It 
is estimated that the number of family-group names for molluscs is over 5,000. 

Key words: genus-group names, family-group names, numbers, nomenclature, nomenclators, 
trends, literature coverage. 



INTRODUCTION 

Although estimates of molluscan species 
diversity (Nicol, 1969; Boss, 1971; for a criti- 
cism, see Solem, 1978) exist, genus-group 
names, as defined by Article 42 of the Inter- 
national Code of Zoological Nomenclature, 
have not received similar attention for many 
years. In molluscs, as in other large groups of 
Recent and fossil animals, taxonomic work is 
hampered by the lack of adequate, up-to- 
date, comprehensive manuals, and by the 
vast number of journals and books containing 
descriptions of new taxa. The fate of every 
family- or genus-level taxonomic treatise is to 
be incomplete or outdated the very year it is 
published. In malacology, the currently avail- 
able taxonomic academic treatises are 20-50 
years old (Wenz, 1938-1944; Zilch, 1959- 
1960; Moore, 1960-1971). In 1987, as a re- 
sult of this frustration, we started to compile a 
loose-leaf index to the new supraspecific 
names proposed since 1960 for Recent and 
fossil molluscs, exclusive of cephalopods. As 
this work is now being extended to encom- 
pass the older literature, we have found it in- 
teresting to quantify the abundance of names 
involved, to identify trends in current taxo- 
nomic literature, and to evaluate the efficiency 
of the supposedly most complete indexing 
system, the Zoological Record. The present 
paper documents these findings and dis- 
cusses their implications. 



Taxonomic research in malacology demon- 
strates a strong division between cephalopod 
and non-cephalopod literature. In particular, 
ammonoid research is almost totally sepa- 
rated from other fields of molluscan research: 
malacological journals only exceptionally 
contain papers on ammonites; ammonite and 
other cephalopod specialists rarely interact 
with other malacologists in national and inter- 
national congresses. Our index, which re- 
flects our own research interests, excludes 
cephalopods and is itself an illustration of this 
dichotomy. As a consequence, while some of 
our results are concerned with Mollusca in 
general, others deal only with Mollusca exclu- 
sive of cephalopods. 



METHODS 

The number of genus-group taxa in Mol- 
lusca is based on an evaluation of the number 
of names contained in various catalogues, 
nomenclators and checklists. Names enu- 
merated in the sources listed in Table 1 were 
counted. For the Nomenclátor Zoologicus, we 
counted the molluscan names in 50 pages 
(chosen from five consecutive pages in ten 
random samples) per volume. The total of 
250 pages sampled represents about 5% of 
4,816 pages included in the Nomenclátor. 
Only original spellings were considered. 



75 



76 



BOUCHET & ROCROI 



To index the supraspecific names pro- 
posed after 1960, we first scanned the Mol- 
lusca volumes of the Zoological Record for 
the relevant years. For Cephalopoda, only ge- 
nus-group names were considered: we sim- 
ply counted them without further checking. 
For classes other than cephalopods, genus- 
group and family-group names were consid- 
ered separately. The original publications 
were consulted, and all newly introduced 
names were checked and recorded. 

In a second phase, we used a combination 
of approaches to find names (of non-Ceph- 
alopods) omitted in the Zoological Record: (a) 
We scanned biographical compilations of ma- 
lacologists containing their lists of papers and 
taxa (e.g. Powell by Cernohorsky, 1988; 
Habe by Inaba & Oyama, 1977), and yearly or 
cumulative indices for malacological journals 
(e.g. Anonymous, 1979); (b) We searched 
malacological and regional journals not cov- 
ered by the Zoological Record that we knew 
contained new malacological taxa (e.g. Noti- 
ziario del CISMA, Roma; Schriften zur Mala- 
kozoologie, Cismar); (с) We used library facil- 
ities in Paris (Muséum National d'Histoire 
Naturelle, Société Géologique de France, 
Centre National de la Recherche Scientifique, 
Université Pierre et Marie Curie and several 
personal libraries), Leningrad (Zoological In- 
stitute of the Academy of Sciences and All- 
Union Geological Institute of the Ministry of 
Geology), London (The Natural History Mu- 
seum), Stockholm (Naturhistoriska Riksmu- 
seet and University Library) and Frankfurt 
(Senckenberg Library), including browsing 
through collections of reprints in departmental 
libraries; (d) We reviewed available papers for 
secondary uses of supraspecific names pub- 
lished elsewhere in unrecorded places; this 
method proved particularly useful with the So- 
viet literature; (e) The recent (since 1985) 
Chinese paleontological literature is covered 
by the quarterly Gushengwuxue Wenzhai 
(English subtitle: Paleontological Abstracts), 
and we discovered many names in this ab- 
stracting journal; (f) Recently published cata- 
logues of names (such as Vokes, 1980, 1990; 
Vaught, 1989) were also scanned for omis- 
sions, and several colleagues provided refer- 
ences to obscure names. 

Our data base is therefore far more com- 
plete for non-cephalopods than for cephalo- 
pods. Even for non-cephalopods, it is admit- 
tedly still a little incomplete but provides the 
best available source of supraspecific names 
for the years 1960-1989. 



RESULTS 

Number of Genus-Group Taxa in Mollusca 

We estimated the total number of genus- 
group names by two indépendant methods. 
One method relies primarily on a statistical 
analysis of the names listed in Nomenclátor 
Zoologicus. The other is based on the various 
catalogues and checklists available for se- 
lected classes of the phylum. 

Evaluation Based on 
Nomenclátor Zoologicus 

(a) 1758-1965: We counted molluscan ge- 
nus-group names in a set of 250 pages of the 
Nomenclátor Zoologicus, selected as de- 
scribed under "Methods." Excluding incorrect 
subsequent spellings, there is an average 
4.77 molluscan names per page, and the total 
number of nomenclaturally available genus- 
group names can be estimated at 23,000 for 
the period 1758-1965. This number is a min- 
imum because some names certainly have 
escaped the Nomenclátor Zoologicus. It is our 
experience, however, that while omissions 
appear to become more numerous with re- 
cent volumes, as we document below, the 
coverage of the Nomenclátor Zoologicus is 
fairly good for the years 1758-1935. We 
therefore believe 23,000 names to be a reli- 
able estimate. 

(b) 1966-1989: Using the combination of 
methods outlined above, our index of non- 
cephalopod names for the period 1966-1989 
contains 3,644 genus-group taxa. For the 
same period, the Zoological Record lists 
1 ,448 genus-group names of Cephalopoda. If 
it is conservatively assumed that 20% of the 
cephalopod names have escaped the Zoolog- 
ical Record (see below), then the total num- 
ber of cephalopod names introduced during 
that period would be 1,810. The total number 
of molluscan genus-group names introduced 
in the years 1966-1989 is therefore approxi- 
mately 5,400. 

The total number of genus-group names in 
Mollusca therefore amounts to circa 28,400 
names. 

Evaluation Based on Catalogues 

A number of catalogues or counts are avail- 
able for selected classes or subclasses of 
Mollusca. We have counted the number of 
genus-group names in comprehensive works. 



SUPRASPECIFIC NAMES OF MOLLUSCS 



77 



TABLE 1. Number of genus-group names of 
molluscs, partitioned by class 



Total 



Recent 



Aplacophora 

Monoplacophora 

Polyplacophora 

Bivalvia 

Scaphopoda 

Gastropoda 

Prosobranchia 

Opisthobranchia 

Pulmonata 
Hyolitha, 

Rostroconchia, etc. 
Cephalopoda 

Total 



80(a) 

267 (b) 

282 (c) 

5090 (d) 

63(e) 

12721 

7149(f) 

982 (h) 
4590 (i) 

362 
6000 (k) 

24865 



80(a) 
9 
205 (a,c) 
2043 (d) 
57(a) 
9004 (a,g), 
10451 (a) 
4112(g), 
5559 (a) 
817(a) 
4075 (a) 


305 (I) 

11703-13150 



Sources: 

(a) after Vaught (1989); (b) after Knight & Yochelson, and 
Knight, Batten & Yochelson, in Moore (1960), plus incre- 
ment; (c) after van Belle (1975-78), plus increment; (d) 
after Vokes (1980), plus increment; (e) after Ludbrook, in 
Moore (1960), plus increment; (f) after Schilder (1947) for 
the period 1758-1932, an estimated increment of 45.5 
names/yr. for the period 1933-1959 (see Table 4), and our 
own counts for 1960-1989; (g) same as (f), but estimated 
increment 24.3 names/yr; (h) after Zilch (1959-60), Rus- 
sell (1971, 1986), and increment; (i) after Zilch (1959-60), 
plus increment; (j) Hyolitha and Tentaculitida after Fisher, 
in Moore (1962), plus increment; Rostroconchia after 
Vokes (1980), plus increment; (k) Ammonoida and Nauti- 
lida after Hewitt (1989), modified; (I) after Nesis (1987). 



handbooks and catalogues, as listed in Table 
1. Because none of these sources is com- 
plete to 1989, we have added an increment, 
based on our own index for the more recent 
years. When these independent subcounts 
are added, the result is 24,900 names (Figs. 
1,3). 

Number of Genus-Group Names of 
Recent Mollusca 

In this analysis, a name is classified as Re- 
cent if it is based on a Recent type species. 
Therefore, Recent and fossil constitute two 
mutually exclusive categories: we are fully 
aware that this is only an approximation of the 
actual situation. 

From the sources listed in Table 1 , we con- 
clude the number of genus-group names of 
Recent Mollusca (Fig. 2; see also Fig. 4) to be 
approximately 12,000, or 42% of all mollus- 
can genus-group names. As a comparison, 
Vaught (1989), who also listed incorrect sub- 
sequent spellings, chresonyms (one name 



may be tabulated several times), and some 
genus-group names with fossil type species, 
ennumerated approximately 1 5,000 names. 

Discussion 

The two estimates, 28,400 and 24,900, are 
consistent with each other and differ only by 
12%. We believe that the discrepancy be- 
tween the two figures arises mainly from our 
use of Schilder (1947) for an evaluation of the 
number of names in the Prosobranchia. 
Schilder's data were based on Wenz (1938- 
44), which is fairly complete only for names 
published before 1933. We do not know to 
what extent Schilder corrected his counts with 
the literature published in 1933-1947, and 
this fact alone could explain a "loss" of sev- 
eral hundred names. 

Our results are intermediate between two 
previously published estimates: (1) Schilder 
(1949) estimated that there were 1 1 ,260 gen- 
era established for "living and fossil shells." 
Because he did not explain how he obtained 
his results, or what he meant with "shells," it 
is difficult to comment. We consider that this 
very low figure is a gross underestimation of 
the actual number of names involved, even 
when one appreciates that Schilder made his 
statement more than 40 years ago. 

(2) Vokes (1967) stated he had a card file 
containing 40,000 names of Mollusca ex- 
tracted from volumes 1-5 of Nomenclátor 
Zoologicus, a much higher figure than our 
own results indicates. We corresponded with 
Dr. Vokes to discuss this difference, and we 
have had access to the card-index that was 
the basis for his 1967 and 1980 catalogues, 
which he generously donated in 1991. 

It appears that Vokes listed incorrect sub- 
sequent spellings as well as original spellings: 
on average, 22.5% of the names enumerated 
in Vokes (1980) are incorrect subsequent 
spellings, and as many as 37.9% of the 8,200 
cards in his bivalve card-index are for nomen- 
claturally unavailable names (incorrect sub- 
sequent spellings, chresonyms, etc.). From 
this evidence, we conclude that 40,000 was a 
gross overestimate, and that it should not be 
regarded as an accurate number of mollus- 
can genus-group names. 

Naming Activity in the Last 30 Years 

In the period 1960-1989, 6,720 new mol- 
luscan genus-group names have been intro- 
duced. Numbers for classes other than ceph- 



78 BOUCHET & ROCROI 

Partition by class, 1758-1989 

33.9 
Cephalopoda 




Bivalvia 



Gastropoda 



■ Aplacophora 

S3 Monoplacophora 

Щ Polyplacophora 

■ Bivalvia 

D Scaphopoda 

П Gastropoda 

Щ Hyolitha, etc... 

Щ Cephalopoda 



44.71% 



FIG. 1. Number of genus-group names of Mollusca introduced since 1758, partitioned by class. 
Recent Mollusca, Partition by class 

Cephalopoda 



17.46% 



Gastropoda 




Bivalvia 



■ 


Aplacophora 


в 


Monoplacophora 


H 


Polyplacophora 


■ 


Bivalvia 


D 


Scaphopoda 


D 


Gastropoda 


m 


Hyolitha, etc.. 


ш 


Cephalopoda 



76.94% 



FIG. 2. Number of genus-group names of Recent Mollusca introduced since 1758, partitioned by class. 



alopods represent genus-group names ac- 
tually found by us. For the Cephalopoda, we 
use the count of names recorded by the Zoo- 
logical Record, corrected on the basis of an 
estimated 20% omission rate by the Zoologi- 
cal Record. 
The breakdown by class, and Recent vs. 



fossil, is presented in Table 2 and Figure 5. 
For simplicity, Caudofoveata and Solenogas- 
tra have been grouped under "Aplacophora," 
and the minor fossil classes are grouped un- 
der a single entry. For the controversial con- 
tents of the Monoplacophora, we have fol- 
lowed Runnegar & Pojeta (1985). 



SUPRASPECIFIC NAMES OF MOLLUSCS 



79 



Gastropod breakdown by subclass 
1758-1989 




Prosobranchi: 
56 2% 



FIG. 3. Number of genus-group names of Gas- 
tropoda introduced since 1758, partitioned by sub- 
class. 



Recent Gastropoda 
breakdown by subclass 




FIG. 4. Number of genus-group names of Recent 
Gastropoda introduced since 1758, partitioned by 
subclass. 



These results lead to several observations: 

1 . Cephalopod names can be estimated at 
33% of the total on the basis of volume 
7 of Nomenclátor Zoologicus, the only 
one that indicates class level. Counts for 
the entire period 1960-1989 give 
33.3%. It is remarkable that 98.2% of all 
cephalopod genus-group names are 
based on fossils. 

2. Within the Gastropoda, which com- 
prises 37.5% of the total, the number of 
taxa can be further subdivided by sub- 
class as presented in Table 3 and Fig- 
ure 6. 

Of all gastropod names, 62.6% are 
based on Recent species, 37.4% on fos- 
sil species. It is interesting to note that 
61.7% of Archaeogastropoda names, 
which date back to the early Paleozoic, 
are based on fossils, whereas only 



40.9% of Neogastropoda, which date 
back to the Cretaceous, are based on 
fossils. Considering that there are many 
more taxonomists working on Recent 
gastropods than on Paleozoic ones, this 
would tend to indicate that name counts 
do reflect some pattern of overall diver- 
sity, rather than the activity of taxono- 
mists. 
3. Bivalves are, to a considerable extent, 
dominated by paleontological research: 
79.6% of all bivalve genus-group names 
are based on fossil species. 

Zoological Record Coverage 

We have estimated the Zoological Record 
coverage by two alternative methods. 

1. Volume 7 (1956-1965) of the Nomen- 
clátor Zoologicus is largely based on the 
Zoological Record (the same typograph- 
ical errors that crept in the Zoological 
Record also appear in the Nomenclátor, 
e.g. Apollonia misspelled Appollonia). 
We searched for 311 names (all the 
names beginning with А, В or C) known 
to us, published during this span of 
years — 56 names (18.0%) were found 
missing in that volume. 

2. For the period 1966-1989, the Zoolog- 
ical Record listed some 2,848 genus- 
group names of Mollusca exclusive of 
Cephalopoda. (Names published in the 
1950s but recorded by the ZR in the 
1960s were not counted by us in this 
total. Names published in 1989 and re- 
corded by the ZR in the 1989-90 vol- 
ume were counted.) Our own index con- 
tains 3,644 names for the same period. 
This indicates a coverage by the Zoo- 
logical Record of 78.2%, or an omission 
rate of 21 .8%. 

This is admittedly an optimistic esti- 
mate, because our own index has also 
certainly missed a number of names. 
For instance, we know of another 85 
names with incomplete references or no 
references at all, which we have not yet 
been able to trace to their primary 
source. If these 85 names are consid- 
ered when calculating the omission rate, 
it then jumps to 23.7%. 

Therefore, it seems fair to conclude that at 
least 20% of new genus-group names have 
been omitted in the last 35 years by a nomen- 



80 



BOUCHET & ROCROI 



TABLE 2. Breakdown, by class, of new genus-group names introduced in 1960-1989 

Total Increment 

Paleozoic Mesozoic Cenozoic Fossil Recent Total (names/yr.) 



Aplacophora 
Monoplacophora 
Polyplacophora 
Bivalvia 


173 

30 

301 


1 

2 

525 


1 
275 


174 

33 

1101 


32 

7 

15 

282 


32 

181 

48 

1383 


1.1 

6.0 

1.6 

46.1 


Scaphopoda 

Gastropoda 

Hyolitha, Rostroconchia, 


7 
164 
305 


1 

293 

2 


1 
477 

1 


9 
934 
308 


24 
1566 


33 

2500 

308 


1.1 
83.3 
10.3 


etc. 
Cephalopoda 








2197 


38 


2235 


74.5 


Grand Total 








4756 


1964 


6720 


224 



y\ 



3000 



2000 - 



1000 - 



Mollusca genus-group names, 1960-1989 




D Recent 
LJ Cenozoic 
flesozoic 
Paleozoic 



FIG. 5. Number of genus-group names of Mollusca introduced in the period 1 960-1 989, partitioned by class. 
All fossil Cephalopoda have been plotted under Mesozoic, which is admittedly an oversimplification. 



clator considered to be the most complete in 
terms of coverage of the taxonomic literature. 
It is important to emphasize that omission 
by the Zoological Record is not random. The 
most imperfect coverage is of the Soviet liter- 
ature, in particular the coverage of paleonto- 
lógica! monographs. Altogether, probably as 



much as one-third of names proposed in the 
Soviet literature escapes the Zoological Rec- 
ord. Other imperfectly covered literatures are 
those from China, Japan, South America and 
southern Europe. We confirm the generally 
accepted belief that books are much more 
poorly recorded than journals. 



SUPRASPECIFIC NAMES OF MOLLUSCS 



81 



Gastropoda genus-group names 
1960-1989 



D Recent 
LJ Cenozoic 
Щ tlesozoic 
I Paleozoic 




aaaaamsa 



•• 



FIG. 6. Number of genus-group names of Gas- 
tropoda introduced in the period 1960-1989, parti- 
tioned by subclass. 



Temporal Variation in Names Output 

The year of publication of genus-group 
names has been extracted from the samples 
of the Nomenclátor Zoologicus described 
above, and grouped by periods of 30 years. 
Table 4 shows the total number of genus- 
group names proposed for each period of 30 
years, and the yearly rate within each of these 
eight periods. 

An examination of the number of supraspe- 
cific names contained per volume of Zoolog- 
ical Record for the period 1 960-1 989 reveals 
considerable variability in the yearly output 
and indicates that no single randomly se- 
lected year can be considered representative 
for the period. 

Table 4 and Figure 7 show a regular growth 
of the yearly increment during the first 100 
years, with a faster growth in the latter part of 
the 19th century. The yearly output has sub- 
sequently remained remarkably stable at 
170-200 genus-group names per year since 
1880, with a slight minimum in the period 
1936-1965 (also noted by Solem, 1978). 

Hewitt (1989) stated that the recent de- 
cades are characterized by important "mono- 
graphic bursts" in nautiloid and ammonoid 
taxonomy. Such bursts have certainly oc- 
curred elsewhere in selected groups and fau- 



nas, such as the Aplacophora, for which 40% 
of the names have been introduced in the last 
30 years. However, contrary to a rather gen- 
eral belief among many molluscan taxono- 
mists, Table 4 and the graph suggest only a 
moderate increase, not an overall explosion, 
in the output of names over this 30-year pe- 
riod. 



Which Country Produces the 
Most Molluscan Taxa? 



To answer this question we have plotted for 
1960-1989 the number of new genus-group 
names (non Cephalopods only) by country of 
origin of the author (Fig. 8), and separately by 
country of publication. When multiple author- 
ship is involved, each author is counted for 
0.5 (two authors), 0.33 (three authors), etc. 

In the period 1960-1989, authors from 53 
countries are involved, with those from 
USSR, USA and China accounting for a little 
over half (50.3%) of the names. Below the 6th 
rank (New Zealand, 4.1%), the percentage is 
already below 5%, and the 1% mark is 
reached at the 17th rank (Cuba). 

National output should not be used as a 
key to overall molluscan biological diversity of 
the different parts of the world. While it is true 
that, to some extent, the vast majority of the 
Japanese naming activity focuses on the Jap- 
anese (paelo)fauna, the naming activity of 
many countries involves faunas from several 
oceans or continents: a marine snail from the 
Caribbean is just as likely to be named by an 
American or a European author. In other 
words, there is no immediate biological expla- 
nation for the differences in numbers of mol- 
lusc genus-group names described in each 
country. 

It is remarkable that New Zealand, Austra- 
lia and Czechoslovakia, with national popula- 
tions of respectively 3.4, 16.8 and 15.6 mil- 
lion, rank among the 10 countries with the 
highest output. New Zealand is then not only 
the country with the highest number of sheep 
per capita, but also the country with by far the 
highest output of mollusc genus-group names 
per capita! 

Ranking by country of publication (Table 6) 
is not significantly different from the ranking 
by country of origin of authors. This means 
that authors do not, as a rule, "expatriate" 
their papers in journals of other countries. 
However, Germany and, to a lesser extent, 
Great Britain rank higher as publishers than 



82 



BOUCHET & ROCROI 



TABLE 3. Breakdown, by subclass, of new gastropod genus-group names introduced in 1960-1989 











Total 








Paleozoic 


Mesozoic 


Cenozoic 


Fossil 


Recent 


Total 


Prosobranchia 


163 


245 


364 


772 


916 


1688 


Archaeogastropoda 

Mesogastropoda 

Neogastropoda 

Opisthobranchia 

Pulmonata 


148 
15 

1 


61 
167 
17 
24 
24 


26 

156 

182 

21 

92 


235 
338 
199 
45 
117 


146 
482 
288 
168 
482 


381 
820 
487 
213 
599 


Total 


164 


293 


477 


934 


1566 


2500 



TABLE 4. Estimates of number of genus-group names introduced during 30-year periods 



period 


1758- 
1787 


1788- 
1817 


1818- 
1847 


1848- 
1877 


1878- 
1907 


1908- 
1935 


1936- 
1965 


1966- 
(1) 


-1989 
(2) 


time (yr) 

names 

increment 


30 
171 
5.7 


30 
874 
29.1 


30 
2755 
91.8 


30 
3002 
100.1 


30 
5643 
188.1 


28 
5947 
212.4 


30 
5115 
170.5 


24 
4333 
180.5 


5454 
227.3 



(1) Names actually recorded by the Zoological Record (2) Names found by us (non-cephalopods) + cephalopod names 
estimated on the basis of a 20% omission rate. 



300 



200 



100 



Mean yearly increment 



1 1 1 1 1 

1750 1850 1950 ( yea r) 

FIG. 7. Mean yearly increment of new genus-group names of Mollusca, calculated over 30-year periods. 



as authors, whereas the opposite is true for 
New Zealand. 

Family-Group Names 

No estimate of the number of family-group 
names, as defined by Article 35 of the Code, 



available for molluscs has ever been pub- 
lished. A fact well known to taxonomists is that 
a new family may be erected without even 
mentioning that a new name is being intro- 
duced. For that reason, we believe that our 
data base is slightly less complete for family- 
group names than for genus-group names. 



SUPRASPECIFIC NAMES OF MOLLUSCS 



83 



Ranking by country of author 




FIG. 8. Ranking of number of genus-group names of Mollusca (exclusive of Cephalopoda) introduced in 
1960-1989, partitioned by country of origin of author. 



TABLE 5. Ranking of country of origin of author 
as a function of total number of genus-group 



TABLE 6. Ranking of number of genus-group 
names arranged by place of publication (1960- 
1989) 



1 
Country 



no. of names 



4 
Rank 



Rank 



Country 



no. of names 



% 



USSR 

USA 

China 

Japan 

Germany 

New Zealand 

France 

Australia 

Great Britain 

Czechoslovakia 

Total 



970 
831 
451 
374 
314 
184 
154 
143 
108 
96 

3625 



21.6 
18.5 
10.1 
8.3 
7.0 
4.1 
3.4 
3.2 
2.4 
2.1 

80.7 



1 
2 
3 
4 
5 
6 
7 
8 
9 
10 



1 
2 
3 
4 
5 
6 
7 
8 
9 
10 

1-10 



USSR 

USA 

China 

Germany 

Japan 

Australia 

France 

Great Britain 

New Zealand 

Italy 



955 


21.3 


901 


20.1 


446 


9.9 


415 


9.3 


390 


8.7 


154 


3.4 


146 


3.3 


143 


3.2 


133 


3.0 


99 


2.2 



3782 



84.4 



The only partial checklist of family-group 
names is that by Ponder & Waren (1988) for 
the Caenogastropoda and Heterostropha. It 
lists 833 names, of which 187 were intro- 
duced in the period 1960-1987. 



Our own index lists 1 ,102 new family-group 
names proposed for molluscs exclusive of 
cephalopods in the period 1960-1989. If we 
assume that the ratio of names published be- 
fore and after 1960 is the same for Caeno- 



84 



BOUCHET & ROCROI 



gastropoda + Heterostropha as for the rest of 
the molluscs, then the total number of fam- 
ily-group names available for molluscs ex- 
clusive of cephalopods can be estimated 
at 4,900. No similar figure is available for 
Cephalopoda, but Hewitt (1989) counted 409 
nautiloid and ammonoid families. 

In our view, this demonstrates the need for a 
new nomenclatural tool that would be to fam- 
ily-group names what the Index Animalium 
(Sherborn, 1902; 1922-1932) and the No- 
menclátor Zoologicus are to species-group 
and genus-group names respectively. 

Epilogue 

The sheer magnitude of the numbers dis- 
cussed in this paper will certainly draw con- 
trasting opinions among malacologists. Two 
extreme views can be expected. In one view, 
the newly named taxa are deemed to repre- 
sent taxonomically valid units, and the current 
naming activity simply demonstrates the 
gross incompleteness of the knowledge on 
the diversity of this phylum. In the opposite 
view, most of the newly created names are 
synonyms and the current naming activity is 
the symptom of a system gone crazy. 

Superfluity in molluscan nomenclature ap- 
pears to be a recurrent concern among pro- 
fessional taxonomists (e.g. Schilder, 1949; 
Nicol, 1958; Boss, 1971, 1978). Schilder 
(1 949) estimated that 34% of the names avail- 
able to classify prosobranchs were synonyms. 
However, whereas 49% of the names estab- 
lished in 1 808-1 857 were synonyms, the syn- 
onymy ratio decreased to 37%, 34% and 21% 
respectively for the next consecutive 25-year 
periods (Schilder, 1949). This can be inter- 
preted in two ways: either taxonomists have 
been doing better work since the early 19th 
century, or it takes a long period of time (in 
excess of 75 years) before the value of a ge- 
nus-group name can be properly assessed. It 
is likely that both elements reflect the actual 
situation, an opinion already expressed by 
Schilder & Schilder (1947). Schilder (1947) 
had also suggested that genus-group names 
of Recent and fossil prosobranchs, described 
and undescribed, would amount to about ap- 
proximately 20,000, of which 5,000 would still 
be extant. This prediction may not be as un- 
realistic as it may first seem: in bivalves, fossils 
outnumber Recent genus-group names in the 
proportion of 2-3 to 1 ; and there are already 
in the order of 5,000 Recent prosobranch ge- 
nus-group names. If Schilder was correct, this 



would mean that more than 10,000 fossil 
prosobranch genera still await naming, a 
daunting perspective! 

Naming activity strongly reflects national 
traditions. There used to be a time when ma- 
lacologists from Australia and New Zealand 
did not expect that their fauna might already 
have been described by workers in other 
parts of the world, and consequently engaged 
in overnaming what they considered to be en- 
tirely endemic faunas. The expression "an- 
other creation" was even used for the Austra- 
lian biota. It is clear that this scientific isolation 
has now ended, and that the high level of mol- 
luscan naming activity in New Zealand and 
Australia is the result of a healthy descriptive 
malacology there in a worldwide context. 

Notwithstanding our unfamiliarity with Pa- 
leozoic and Mesozoic faunas, we remain 
greatly concerned by the introduction of new 
taxa at the genus and family levels based on 
poorly preserved fossils or molds. Certain 
branches of malacology are also undoubtedly 
suffering from "inbreeding," a case in point 
being the immense literature on the Neogene 
Ponto-Caspian basins. 

Despite these reservations, we do not 
share the view that overnaming is the single 
most important factor explaining yearly incre- 
ments of 224 molluscan genus-group names, 
and 40+ family-group names. In the last 30 
years, whole new faunas have been discov- 
ered, either Recent (e.g. oceanic hydrother- 
mal vents) or fossil (e.g. lower Cambrian of 
China); old faunas have been readressed us- 
ing new techniques (e.g. scanning electron 
microscopy, SCUBA diving) and new charac- 
ters (e.g. anatomy of minute species). And, 
perhaps most importantly, the phase of intel- 
lectual stagnation that followed Thiele's ep- 
och-making Handbuch is giving way to stim- 
ulating and provocative ideas. In this process, 
superfluous names undoubtedly become in- 
troduced, but we are convinced that these do 
not minimize the considerable amount of gen- 
uine discoveries being made every year. 

The unexpectedly high omission rate of 
Zoological Record should cause concern to 
all taxonomists. Because this nomenclátor is 
the main bibliographical source of many (pa- 
leo)zoologists, this factor alone poses an im- 
portant threat to nomenclatural stability. We 
believe that this justifies the establishment 
of new criteria of availability of zoological 
names, whereby a published name would 
have to be registered by the International 
Commission of Zoological Nomenclature be- 



SUPRASPECIFIC NAMES OF MOLLUSCS 



85 



fore it is declared nomenclaturally available 
(Bouchet, in press). 



ACKNOWLEDGMENTS 

For assistance with literature and/or calling 
our attention to names not listed in the Zoo- 
logical Record, we thank A. Bogan (Academy 
of Natural Sciences, Philadelphia), S. Freneix 
(Institut de Paléontologie, Paris), R. Janssen 
(Senckenberg Museum, Frankfurt), A. Kabat 
(National Museum of Natural History, Wash- 
ington, D.C.), T. Kase (National Science Mu- 
seum, Tokyo), I. Loch (Australian Museum, 
Sydney), A. Lum (Natural History Museum, 
London), Z.-G. Ma (Institute of Geology and 
Paleontology, Academia Sinica, Nanjing), A. 
Matsukuma (National Science Museum, To- 
kyo), R. Petit (North Myrtle Beach, South 
Carolina), Yu. Starobogatov and M. Dolgo- 
lenko (Zoological Institute, Leningrad), H. 
Vokes (Tulane University, New Orleans) and 
A. Waren (Naturhistoriska Riksmuseet, 
Stockholm). For assistance in English, we 
thank A. Kabat. A. Foubert helped generating 
computer-produced figures. 



LITERATURE CITED 

ANONYMOUS, 1979, Venus: Japanese Journal of 
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Malacological Society of Japan, Tokyo. 

BOSS, K. J., 1971, Critical estimate of the number 
of Recent Mollusca. Occasional Papers on Mol- 
lusks, 3(40) :81 -135. 

BOSS, K. J., 1978, Taxonomic concepts and su- 
perfluity in bivalve nomenclature. Philosophical 
Transactions of the Royal Society of London, 
(B)284:41 7-424. 

BOUCHET, P. (in press), Proposal that a zoological 
name becomes nomenclaturally available from 
the date when the work containing its introduction 
is received by the Secretariat of ICZN. Bulletin of 
Zoological Nomenclature 

CERNOHORSKY, W. O., 1988, Arthur William Ba- 
den Powell (1901-1987). A brief biography and 
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EDWARDS, M. A. & A. T. HOPWOOD, 1966, No- 
menclátor Zoologicus, vol. 6. The Zoological So- 
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EDWARDS, M. A. & H. G. VEVERS, 1975, Nomen- 
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GUSHENGWUXUE WENZHAI [= Paleontológica! 



Abstracts], 1986-1991. Quarterly. Institute of 
Geology and Paleontology, Nanjing. 

HEWITT, R. A., 1989, Recent growth of nautiloid 
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INABA, T. & К. OYAMA, 1977, Catalogue of mol- 
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MOORE, R. O (ed). Treatise on invertebrate pale- 
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lusca 6 (vol. 1, 1969; vol. 2, 1969; vol. 3, 1971); 
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NEAVE, S. A., 1939-1940, Nomenclátor Zoologi- 
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don. 

NESIS, K. N., 1987, Cephalopods of the world. TFH 
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NICOL, D., 1958, Trends and problems in pelecy- 
pod classification (the genus and subgenus). 
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NICOL, D., 1969, The number of living species of 
molluscs. Systematic Zoology, 18:251-254. 

NOMENCLÁTOR ZOOLOGICUS. See Neave; Ed- 
wards & Hopwood; Edwards & Vevers. 

PONDER, W. & A. WAREN, 1988, Classification of 
the Caenogastropoda and Heterostropha. A list 
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RUNNEGAR, B. & J. POJETA, 1985, Origin and 
diversification of the Mollusca. In: E. R. TRUE- 
MAN & M. R. CLARKE, eds; The Mollusca, vol. 
10, Evolution, 1-58. Academic Press, Orlando. 

RUSSELL, H. D., 1971, Index Nudibranchia (1554- 
1965). Delaware Museum of Natural History, 
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RUSSELL, H. D., 1986, Index Nudibranchia. Sup- 
plement I, 1966-1975. Department of Mollusks, 
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SCHILDER, F. A., 1947, Die Zahl der Prosobran- 
chier in Vergangenheit und Gegenwart. Archiv für 
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SCHILDER, F. A., 1949, Statistical notes on mala- 
cology. Proceedings of the Malacological Society 
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SCHILDER, M. & F. A. SCHILDER, 1947, Zur 
Entwicklung der Molluskenkunde. Archiv für Mol- 
luskenkunde, 76:163-166. 

SHERBORN, О D., 1902, 1922-1932, Index Ani- 
malium. Sectio I (1758-1800), 1902, Cambridge; 
Sectio II (1801-1850), 1922-1932 (29 parts). 
British Museum, London. 

SOLEM, A., 1978, Classification of the land mol- 
lusca. In: V. Fretter & J. Peake, eds, Pulmo- 
nates, vol. 2A. Systematics, evolution and ecol- 
ogy, 49-97. Academic Press, London. 

VAN BELLE, R. A., 1975-78, Sur la classification 
des Polyplacophora. Informations de la Société 
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(4)6:135-152 (1975); 5(2):15-42 (1977); 6(1): 
3-28 (1978); 6(2):35-44 (1978). 

VAUGHT, K. C, 1989, A classification of the living 
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VOKES, H. E., 1967, Genera of the Bivalvia: a sys- 
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VOKES, H. E., 1980, Genera of the Bivalvia: a sys- 
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and errata. Tulane Studies in Geology and Pale- 
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WENZ, W., 1938-1944, Handbuch der Paläozoo- 
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ZILCH, A., 1959-1960, Handbuch der Paläozoo- 
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ZOOLOGICAL RECORD for the years 1960-1989, 
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London (1981-1990). 

Revised Ms. accepted 2 January 1992 



MALACOLOGIA, 1992,34(1-2): 87-97 

SHELL PATTERN POLYMORPHISM IN A 13-YEAR STUDY OF THE LAND SNAIL 
THEBA PISANA (MÜLLER) (PULMONATA: HELICIDAE) 

Robert H. Cowie 
Bishop Museum, P.O. Box 19000 A, Honolulu, Hawaii 96817-0916, U.S.A. 

ABSTRACT 

Shell pattern gene frequencies in the helicid Theba pisana at Tenby, South Wales, did not 
change over a 13-year period at a range of sites differing in frequency. This apparent stability 
and the differences among sites may be maintained by selection, although the selective agent(s) 
is(are) unknown. Alternatively, selection may be absent (or too weak to be detected), and the 
differences among sites may be random due to the founder effect; in this case, stochastic 
fluctuations in gene frequency would be small in these large populations and directional changes 
would not be expected. The need for long-term ecological studies in order to address such 
questions is emphasized. 

Key words: Pulmonata, Theba pisana, genetics, polymorphism, selection, founder effect. 



INTRODUCTION 

Long-term field studies are essential in ev- 
olutionary biology yet are rarely possible 
(Cain, 1983; Cowie, 1989). Subtle but impor- 
tant changes in the composition of communi- 
ties, the relative distributions of species or the 
morph frequencies in polymorphic popula- 
tions, or indeed the absence of such changes, 
can only be demonstrated by studies pursued 
over many years. 

In the extensively studied European land 
snails Cepaea nemoralis (L.) and С hortensis 
(Müller), stability in morph frequencies over 
periods of 5-50 years, as well as significant 
changes over such periods, have been found 
and attributed to natural selection (Cain, 
1983; Murray & Clarke, 1978; Wall et al., 
1980). Other studies have failed to detect se- 
lection (Cain & Cook, 1989; Cain et al., 1990). 
Striking changes in the relative distributions of 
the two species have also been reported 
(Cowie & Jones, 1987). Longer-term studies 
also have demonstrated significant changes 
in morph frequencies, attributable to both nat- 
ural selection and historical factors (Cain, 
1983; Cameron & Dillon, 1984). 

The rather strict criteria for the successful 
study of gene frequencies over time (Murray 
& Clarke, 1978; and see also Endler, 1986, 
pp. 73-75) are that (i) accurately dated and 
localised data be available; (ii) the genetic 
control of the polymorphism be understood 
sufficiently to allow accurate scoring of geno- 
types; (iii) there be genetic continuity over the 
period of the study; and (iv) the selective sig- 



nificance of the characters be understood. 
The present study of morph frequencies in the 
land snail Theba pisana (Müller) fulfills all 
these criteria: the samples were all taken by 
the author from the same group of sites on 
specific dates and using the same collecting 
techniques (Cowie, 1984a, b); the genetics of 
the polymorphism have been elucidated by 
breeding experiments (Cowie, 1984a); sam- 
ples were taken from continuously extant 
populations with overlapping generations ev- 
ery year from 1977 to 1989 (except 1982); 
and both visual selection by predators and 
climatic selection appear to influence both 
this polymorphism (Hazel & Johnson, 1990; 
Heller, 1981; Heller & Gadot, 1984; Johnson, 
1980, 1981) and other aspects of visually de- 
tectable variation in T. pisana (Cowie, 1990). 
Theba pisana is a coastal Mediterranean 
land snail, the distribution of which extends 
along the Atlantic coasts of north Africa and 
western Europe as far north as the south- 
western parts of England and Wales and the 
east coast of Ireland. It has been introduced 
to Australia, South Africa and California. A 
five-year study of morph frequencies at six 
sites at Tenby, South Wales (Cowie, 1984a), 
failed to demonstrate any changes in morph 
frequencies over this period despite a hint 
that selection was acting against the un- 
banded (i.e. pale) morphs. The aims of the 
present study were to confirm or discount the 
possible selection and to assess the stability 
or otherwise of the morph frequencies over a 
longer period. All shells are deposited in the 
Mollusca Section of The Natural History Mu- 



87 



88 



COWIE 



seum (London) (Mollusca Section accession 
number: 2355). 



METHODS 

Samples were collected from up to six sites 
between 24 May and 31 August of each year 
from 1977 to 1989 (except 1982) following the 
precise protocols of Cowie (1984a, b). All 
snails in a sample area were collected, so no 
bias in morph frequency, due to differential 
collection of certain morphs, was introduced. 
A map of the site locations is given by Cowie 
(1984a). Theba pisana is biennial at Tenby, 
breeding only once at the end of its second 
year and then dying. Summer samples taken 
prior to the start of the late summer/early au- 
tumn breeding season therefore have dis- 
tinctly bimodal size frequency distributions, 
allowing easy separation of two-year-old 
adults from one-year-old juveniles by simple 
graphical analysis (Cowie, 1984b). Morph 
scores of the two age classes could, there- 
fore, be analysed separately. 

The data of Cowie (1984a) have been in- 
corporated in the present analysis. The regu- 
lar taking of more than one sample from some 
of the six sites, sometimes on more than one 
occasion during the 24 May-31 August pe- 
riod, was discontinued from 1983 onwards as 
being redundant: one sample only was taken 
from each site, and on only one occasion (be- 
tween 24 May and 31 August) in each year. 
From 1986 onwards, a single sample only 
was taken from each of three sites, represent- 
ing the three broad sampling areas at Tenby 
(see Cowie, 1984a). 

Morphs were scored according to Cowie 
(1984a). The discussion over whether T. 
pisana is pentataeniate or tetrataeniate 
(Cowie, 1984a; Gittenberger & Ripken, 1987; 
Heller, 1981) does not affect the scoring of 
the polymorphism. The polymorphism in the 
Tenby populations is controlled by three loci 
with two alleles showing dominance at each. 
Epistasy allows only four main phenotypes to 
appear: plain unbanded, dotty unhanded 
(with a row of dots along the mid-line of the 
shell), dark five-banded and yellow five- 
banded; these are illustrated by Cowie 
(1984a). There is some blurring of the bound- 
aries between the dark and yellow five- 
banded morphs such that it has been neces- 
sary to include an "intermediate" five-banded 
category (Cowie, 1984a). The blurred distinc- 
tions between the five-banded categories, 



combined with the variable shell size at which 
the dark banding lineolations appear, allow 
only limited conclusions to be drawn regard- 
ing this aspect of the variation, and indeed 
juveniles were scored simply as banded with- 
out distinction into dark, intermediate and 
yellow. However, the distinction between 
banded and unbanded shells, in both adults 
and juveniles, is clear since, in these popula- 
tions, the apex (in fact the protoconch) of 
banded shells is always darker than that of 
unbanded shells. This association of banding 
pattern and apex colour is due to the very 
close linkage of the banded/unbanded and 
apex colour loci, combined with strong link- 
age disequilibrium, due presumably to selec- 
tion (Cain, 1984; Cowie, 1984a). Mistakes in 
scoring are considered, then, to be insignifi- 
cant. 



RESULTS 

Morph scores for all samples are given in 
Table 1 . The differences between sites in the 
proportion of unbandeds, indicated by Cowie 
(1984a), have been maintained. Sites 2, 3 
and 4, which are less than 100 m apart and 
more or less connected by suitable habitat 
with T. pisana present, show similar frequen- 
cies, most samples having 25-45% unband- 
eds (average about 34%). Sites 1, 5 and 6, 
the last two of which are close together but 
separated from site 1 by about 0.5 km of cliffs 
with no T. pisana, also have similar frequen- 
cies, most samples having 10-25% unband- 
eds (average about 17%). 

Percentages of unbandeds through the 
course of the study at each site, for adults and 
juveniles separately, are shown in Figures 1- 
6. Regression analyses (Feldman et al., 
1987) of these percentage data (arcsinh 
square root transformed) on year reveal no 
significant change (p>0.05 in all cases) in fre- 
quencies of unbandeds in either adults or ju- 
veniles at any site throughout the course of 
the study, either when all samples or only 
those of 30 or more individuals are included 
(Table 2). Morph frequencies have not 
changed significantly for 13 years. Two-way 
log-likelihood G-statistic analyses (Sokal & 
Rohlf, 1969) of the actual frequencies of un- 
bandeds by year for each site separately 
(Table 3) indicate some heterogeneity among 
years (as detected by Cowie (1984a)) but no 
obvious trend, and do not contradict the over- 
all absence of change. (Because of the lack of 



POLYMORPHISM IN THEBA PISANA 
TABLE 1 . Morph scores for all samples. 



89 



Site Year 












Total 




- Juveni les- 




Total 


Total 














— unbanded — 




--banded- - 




adults 


— unbanded — 


banded 


juveni I 


es 




plain 


dotty 


dark 


intermed 


ye I low 




plain 


dotty 








1 1977 


48 


26 


258 


81 


20 


433 


45 


16 


360 


421 


854 


1978 


27 


9 


150 


36 


4 


226 


42 


22 


294 


358 


584 


1979 


5 


2 


37 


7 


2 


53 


4 


4 


65 


73 


126 


1980 


7 


4 


43 


8 


5 


67 


6 


6 


66 


78 


145 


1981 


3 


3 


31 


10 


3 


50 


17 


4 


72 


93 


КЗ 


1983 


8 


2 


29 


1 


2 


42 


5 





37 


42 


84 


1984 


5 


3 


24 


1 


1 


34 


7 


2 


55 


64 


98 


1985 


4 


1 


22 


4 


3 


34 


2 


1 


24 


27 


61 


1986 


3 


1 


22 


1 


2 


29 


8 


4 


91 


103 


132 


1988 








9 


1 





10 


2 





23 


25 


35 


1989 


3 


1 


12 


3 


4 


23 


2 





13 


15 


38 


2 1977 


203 


42 


269 


63 


18 


609* 


245 


43 


372 


660 


1269 


1978 


133 


51 


204 


67 


21 


476 


90 


22 


164 


276 


752 


1979 


21 


12 


52 


9 


4 


98 


32 


11 


89 


132 


230 


1981 


81 


39 


159 


59 


20 


358 


47 


16 


88 


151 


509 


1983 


24 


8 


36 


3 


11 


82 


32 


11 


57 


100 


182 


1984 


14 


7 


35 


4 


4 


64 


36 


20 


129 


185 


249 


1985 


77 


28 


115 


10 


13 


243 


17 


3 


51 


71 


314 


1986 


18 


5 


35 


5 


4 


67 


43 


6 


91 


140 


207 


1987 


43 


11 


60 


9 


17 


140 


22 





63 


85 


225 


1988 


39 


15 


57 


12 


12 


135 


25 


11 


50 


86 


221 


1989 


28 


10 


54 


9 


6 


107 


18 


7 


38 


63 


170 


3 1977 


24 


12 


62 


20 


4 


122 


33 


12 


79 


124 


246 


1978 


38 


6 


70 


16 


6 


136 


37 


18 


113 


168 


304 


1979 


1 





1 


1 





3 


23 


15 


88 


126 


129 


1980 


9 


5 


23 


7 


2 


46 


20 


10 


65 


95 


141 


1981 


5 


2 


14 


4 


1 


26 


1 





5 


6 


32 


1983 


19 


5 


26 


8 


10 


68 


5 


5 


35 


45 


113 


1984 


18 


6 


50 


8 


13 


95 


30 


17 


90 


137 


232 


1985 


32 


14 


81 


14 


10 


151 


14 





30 


44 


195 


4 1977 


51 


23 


104 


29 


8 


215 


66 


6 


106 


178 


393 


1978 


55 


35 


143 


33 


7 


273 


106 


65 


297 


468 


741 


1979 


12 


5 


30 


5 


3 


55 


52 


15 


132 


199 


254 


1980 


34 


17 


44 


13 


2 


110 


42 


15 


110 


167 


277 


1981 


26 


21 


83 


21 


11 


162 


16 


10 


43 


69 


231 


1983 


17 


4 


39 


8 


10 


78 


17 


12 


77 


106 


184 


1984 


59 


9 


146 


9 


9 


232 


23 


15 


59 


97 


329 


1985 


65 


15 


115 


9 


10 


214 


8 





32 


40 


254 


5 1977 


24 


7 


119 


38 


15 


203 


13 


2 


53 


68 


271 


1978 


14 


5 


78 


17 


9 


123 


20 


14 


157 


191 


314 


1979 


1 





10 


2 


2 


15 


3 


2 


6 


11 


26 


1980 


4 


3 


35 


8 


3 


53 


1 


2 


11 


14 


67 


1981 


4 


1 


16 


1 


3 


25 


1 


1 


2 


4 


29 


1983 


4 


1 


11 


2 


2 


20 


1 





11 


12 


32 


6 1977 


23 


10 


139 


39 


10 


221 


16 


4 


104 


124 


345 


1978 


19 


2 


87 


18 


7 


133 


20 


7 


219 


246 


379 


1979 








16 


12 


1 


29 


3 


2 


29 


34 


63 


1981 


10 


2 


57 


21 


7 


97 


21 


5 


94 


120 


217 


1983 


11 


1 


103 


14 


13 


142 


7 


1 


57 


65 


207 


1984 


13 


8 


86 


15 


17 


139 


7 


1 


88 


96 


235 


1985 


22 


5 


82 


5 


9 


123 


10 


3 


130 


143 


266 


1986 


11 


3 


61 


7 


4 


86 


12 


2 


89 


103 


189 


1987 


9 


4 


49 


7 


12 


81 


28 


2 


111 


141 


222 


1988 


5 


1 


57 


15 


8 


86 


29 


12 


165 


206 


292 


1989 


10 


5 


57 


18 


11 


101 


33 


10 


253 


296 


397 


Total 


1443 


517 


3807 


847 


415 


7043 


1465 


494 


5232 


7191 


14234 



In certain years, samples were not collected as follows: 1980— sites 2, 6; 1982— all sites; 1984-1989— site 5; 1986- 
1989— sites 3, 4; 1987— sites 1, 3, 4, 5. 

"There were 14 banded adults that were not scored as dark, intermediate or yellow. These have been included in analyses 
of banded/unbanded but excluded from analyses within the banded class. 



90 



COWIE 



40- 


ADS 












SITE 1 




30- 


















20- 












\ 








10- 












































i 
1977 




8 


i i i 
83 




i i 
86 


8 



40 -i 


JUVS 












SITE 1 


"" 


30- 
























20- 






















10- 
0- 


I 






1 1 








i 


i 



FIG. 1. Percent unbandeds in adults (ADS) and juveniles (JUVS) at site 1 for 1977-1989, with 95% 
confidence limits derived using the tables of Sokal & Rohlf (1973) supplemented by Goldstein (1964). 



60 -i 
50- 
40- 
30- 
20- 




ADS 



SITE 2 




Í 



i — i — i — г 
1977 80 



~ i i i i i i i 
83 86 89 



60-1 
50- 
40- 
30- 
20- 




JUVS 



SITE 2 




г 



i — i — i — i — г 
1977 80 



83 



FIG. 2. Percent unbandeds at site 2; details as in Fig. 1. 



60 n 


ADS 












SITE 3 


50- 












40- 
















30- 


















20- 










10- 






0- 


i i i i i i i i i i i i i 



60-1 
50- 
40- 
30- 
20- 
10- 
0- 



JUVS 



1 — I — I — I — I — I 



SITE 3 



1977 



83 



i i i i i i i i i i i i i 
1977 80 83 86 89 



FIG. 3. Percent unbandeds at site 3; details as in Fig. 1 . (The 1979 adult value is omitted as the sample was 
too small to allow calculation of confidence limits.) 



samples from some sites in some years, an ing against unbandeds, at least during the 

overall three-way G-statistic analysis of fre- second year of life (Cowie, 1984a), is re- 

quency by year by site was not performed). jected. Comparisons of unbanded frequen- 

The suggestion that selection was operat- cies in adults with those in juveniles of the 



POLYMORPHISM IN THEBA PISANA 



91 



% 



60-1 ADS 
50- 
40 
30- 
20- 
10- 




SITE 4 




1977 



i — i — i — i — г 

80 83 



60 -i 


JUVS 












SITE 4 


50- 






T 


40- 
















30- 






















- 1 - - 1 - j 








20- 








10- 






















l 






l l 1 



FIG. 4. Percent unbandeds at site 4; details as in Fig. 1 . 



% 



80- 


ADS 










SITE 5 


70- 




60- 




50- 






40- 








30- 










20- 
















10- 
















0- 


! ! 


1 I I I I 1 1 1 ! 1 



% 



80n 
70- 
60- 
50- 
40- 
30- 
20- 
10- 
0- 



JUVS 



SITE 5 




1977 



83 



— i — i — i — i — i — i — i — i — i — i i i i 
1977 80 83 86 89 



FIG. 5. Percent unbandeds at site 5; details as in Fig. 1. (The 1981 juvenile value is omitted as the sample 
was too small to allow calculation of confidence limits.) 



SITE 6 




1977 



40 -i 
30- 
20- 
10 




JUVS 



SITE 6 




— i — i — i — i — i — i — i — i — i — i i i i 
1977 80 83 86 89 



FIG. 6. Percent unbandeds at site 6; details as in Fig. 1. 



92 



COWIE 



TABLE 2. Values of r 2 and p from the regression analyses of morph frequency (transformed) on year. If 
any sample(s) or sub-sample(s) (e.g. unbanded adults when considering percent plain among adults) 
contained fewer than 30 individuals, the regression was also computed omitting these samples and the 
results are given in parentheses. 



X unbanded 
2 



X plain 
2 



% yel tow 
2 



X unbanded 
2 



X plain 
2 



0.160 0.222 0.109 0.351 0.012 0.747 0.006 0.828 0.286 0.090 0.351 0.055 

(0.124) (0.392) (0.305) (0.256) (0.234) (0.332) (0.103) (0.439) 



0.001 0.930 0.021 0.674 0.009 0.785 0.356 0.053 0.095 0.356 0.001 0.934 

(0.015) (0.753) (0.133) (0.375) 



0.086 0.481 0.035 0.657 0.011 0.807 0.264 0.193 0.044 0.620 0.025 0.711 

(0.029) (0.749) (0.064) (0.838) (0.047) (0.680) (0.588) (0.075) (0.117) (0.453) (0.267) (0.373) 



0.033 0.667 0.430 0.077 0.174 0.304 0.175 0.303 0.411 0.087 0.006 0.859 

(0.418) (0.165) (0.414) (0.241) 



0.281 0.279 0.000 0.972 0.228 0.338 0.219 0.349 0.101 0.540 0.006 0.888 

(0.961) (0.127) (0.708) (0.364) (0.893) (0.212) (-)(-)* 



0.096 0.354 0.071 0.458 0.033 0.593 0.469 0.020 0.012 0.750 0.092 0.364 

(0.017) (0.716) (0.015) (0.732) (0.403) (0.049) (0.472) (0.518) 



'There were only two samples with 30 or more juveniles at site 5, so a regression based on just these two points has not 
been calculated. 



previous year (i.e. the population from which 
those adults were derived) at each site (40 
possible comparisons) do not show a signifi- 
cant trend; the points in Figure 7 fall more or 
less evently on either side of the line of equal 
percent unbanded, and a Wilcoxon's signed 
ranks test detected no significant difference 
(p>0.1) in the frequencies of unbandeds be- 
tween these adult and juvenile samples. (On 
checking the data on which Table 3 of Cowie 
(1984a) is based, the percent unbanded in 
that table for juveniles in 1979 at site 1 was 
found to be incorrect; the correct value is 
1 1 .0, not 32.6. Recalculation of the Wilcox- 
on's tests of Cowie (1984a) but incorporating 
this correction, in fact, indicates no significant 
difference for the comparison of adults with 
juveniles of the previous year — p>0.05; al- 
though the less biologically meaningful com- 
parison of adults and juveniles of the same 



year remains significant, but less strongly 
so— p<0.05.) 

Within the unbandeds, there are no appar- 
ent differences among sites in the proportion 
of plain as opposed to dotty shells, most sam- 
ples having 60-85% plain (average about 
74%). As above, regression analysis (Table 
2) detected no significant trends, while G-sta- 
tistic analysis (Table 3) indicated some heter- 
ogeneity among years but without suggesting 
any trends. Again, morph frequencies show 
no consistent change. 

Within the bandeds, there are no apparent 
differences among sites in the proportions of 
dark, intermediate and yellow shells. Most 
samples had 65-85% dark and 0-20% yel- 
low (average about 75% and 9%, respec- 
tively), with the remainder made up of inter- 
mediates. Regression analysis revealed a 
single significant change in morph frequen- 



POLYMORPHISM IN THEBA PISANA 93 

TABLE 3. G-statistic analysis of morph frequencies by year. 

Site G d.f. p 

adults juveniles adults juveniles adults juveniles 

unbanded vs. banded 



1 


4.714 


9.800 


9* 


10 


n. s. 


n. s. 


2 


9.694 


27.172 


10 


10 


n. s. 


<0.01 


3 


1.215 


4.426 


7 


7 


n. s. 


n. s. 


4 


14.446 


11.010 


7 


7 


<0.05 


n. s. 


5 


2.890 


7.430 


5 


5 


n. s. 


n. s. 


6 


17.646 


23.762 


9* 


10 


<0.05 


<0.01 



plain vs. dotty 



1 3.502 7.340 

2 16.858 18.416 

3 6.164 2.942 

4 24.900 30.184 

5 0.866 4.794 

6 9.730 9.030 



9 


6* 


n. s. 


n. s. 


10 


9* 


n. s. 


<0.05 


6* 


5* 


n. s. 


n. s. 


7 


6* 


<0.001 


<0.001 


4* 


3* 


n. s. 


n. s. 


9 


10 


n. s. 


n. s. 



1 


17.248 


2 


22.400 


3 


5.690 


4 


25.072 


5 


2.180 


6 


24.520 



dark vs. intermediate/yellow 

9* - <0.05 

10 - <0.05 

6* - n.s. 

7 - <0.001 

5 - n.s. 

10 - <0.01 



*Some years pooled to give adequate cell values. 



94 



COWIE 




% UNBANDED - ADULTS 



FIG. 7. Scatter diagram of percent unbanded in adult samples against juvenile samples of the previous year 
(see text for explanation). The diagonal indicates equal percentages. Open circles represent cases in which 
the adult or juvenile sample, or both, contained fewer than 30 individuals. 



cies — an increase in percent yellow at site 
6 — which is only just significant (p = 0.049) 
when those samples of fewer than 30 individ- 
uals are omitted from the analysis (Table 2). 
This one significant result can be attributed to 
chance, given the large number of regression 
analyses performed, and does not indicate a 
real change. G-statistic analysis (Table 3, with 
frequencies of yellow and intermediate shells 
combined) again indicates heterogeneity 
among years (including at site 6, where the 
regression analysis indicated a significant 
trend) but this probably also reflects stochas- 
tic effects associated with multiple testing, 
combined with the difficulty, and consequent 
inconsistency, of scoring morphs within the 
banded class (see above). It is reasonable to 
conclude, once again, that there has been no 
consistent change in morph frequency over 
the course of the study. 

In the absence of any trends, selection co- 
efficients have not been calculated (cf. Mur- 
ray & Clarke, 1978; Wall et al., 1980). 



DISCUSSION 

Theba pisana was probably introduced ar- 
tifically to the U.K. during the eighteenth cen- 
tury (Cowie, 1984a; Turk, 1966, 1972). Cowie 
(1984a) suggested that selection acting since 
that time (over about 100 generations) may 
have given rise to the characteristic appear- 
ance of the morphs at Tenby and nearby lo- 
calities in South Wales. While this may or may 
not be true, the current differences among 
sites in morph frequency, yet the lack of de- 
tectable morph frequency change over the 
13 years of the study, can be explained in a 
number of ways (cf. Endler, 1986: 73-75), in- 
volving both selection and stochastic pro- 
cesses: 

(1) Directional selection, following the initial 
introduction, produced the differences among 
sites; these differences are now being main- 
tained by stabilising selection. While visual 
selection by predators and climatic selection 
are known to influence variation in shell pat- 



POLYMORPHISM IN THEBA PISANA 



95 



tern and colour of soft parts in T. pisaría 
(Cowie, 1990; Hazel & Johnson, 1990; Heller, 
1981; Heller & Gadot, 1984; Johnson, 1980, 
1981 ), as in the better known helicids Cepaea 
nemoralis and C. hortensis (Cain, 1983; 
Cowie & Jones, 1985; Jones et al., 1977), it is 
difficult to see what selective differences exist 
among the three sampling areas (site 1 , sites 
2, 3 and 4, sites 5 and 6) because all occur 
along a short length (about 0.75 km) of cliff 
face in apparently fairly similar habitat; any 
putative selective agent(s) is(are) unknown. 
Nevertheless, subtle and arcane habitat dif- 
ferences may be important, as has been sug- 
gested as one of a number of explanations of 
"area effects" in Cepaea (Cain & Currey, 
1963a, b), and an influence of selection on 
morph frequencies in T. pisana at Tenby can- 
not be ruled out. 

(2) The differences among sites arose from 
founder effects; frequent extinction of local 
populations, followed by recolonisation, at 
this, the climatic edge of the species' range 
(Cowie, 1987), leads to morph frequencies 
being the results of a succession of founding 
events; directional selection operating simi- 
larly at all sites has never been able to bring 
the populations to equilibrium. None of the 
present populations became extinct during 
the course of the study, although numbers 
were low at sites 1 and 5; selection may not 
be sufficiently strong to have been detected 
during the 13 years of the study. The sugges- 
tion that apparently strong directional selec- 
tion was operating over all sites against un- 
bandeds, and perhaps against plain morphs 
within the unbanded class (Cowie 1984a), is 
rejected, with the explanation that the initial 
five-year study was not long enough and the 
apparent selection was a statistical artefact, 
exagerated by a computational error (see 
above). At this cold extreme of the range of T. 
pisana, selection for darker colour (i.e. in- 
creased banding) might be expected, and in- 
deed, at all other localities in this part of South 
Wales (Cowie, 1986), only five-banded (no 
unbanded) snails are found (Cowie, 1984a, 
1987; Fowles & Cowie, 1989). (The absence 
of unbandeds at these other localities may 
also be due to the founder effect if these 
smaller populations are all derived ultimately 
from the much larger and well-established 
Tenby populations via small propagules con- 
sisting only of banded snails — Cowie, 1984a; 
Fowles & Cowie, 1989.) 

(3) The differences among sites are due to 
founder effects; selection is not operating; 



and the differences are being maintained, or 
at least changing only slowly, because large 
population sizes prevent significant genetic 
drift. Cowie (1984c) estimated effective 
neighbourhood numbers as ranging up to 
4130 individuals, large enough for drift to be 
insignificant. 

It is not possible to distinguish between 
these various scenarios. Even though at least 
some aspects of the biology of T. pisana, es- 
pecially at Tenby, are fairly well understood, 
there remain major gaps in our knowledge of 
the selective pressures to which the snails are 
subject, pressures that will differ from locality 
to locality, so that studies of T. pisana in Israel 
(Heller, 1981; Heller & Gadot, 1984), South 
Africa (Hickson, 1972) and Australia (Baker & 
Vogelzang, 1988; Hazel & Johnson, 1990; 
Johnson, 1980, 1981) can only be of broad, 
and not specific, relevance to our understand- 
ing of factors controlling the polymorphism in 
South Wales. Furthermore, Cain & Cook 
(1989) and Cain et al. (1990), studying Ce- 
paea nemoralis over periods of 1 8 years and 
23 years, respectively, have indicated the 
considerable difficulties involved in distin- 
guishing among systematic changes in gene 
frequencies due to selection, maintenance of 
unchanging frequencies by selection, and 
stochastic fluctuations (see also Endler, 
1986, ch. 4); Cain & Cook (1989) suggest that 
studies of 250 years or more may be neces- 
sary. The unanswered questions posed by 
the present study, reinforce the view (Endler, 
1986; Wade & Kalisz, 1990) that only with 
detailed ecological knowledge, necessarily in- 
volving long-term studies, combined with ex- 
perimentation, can the factors influencing 
polymorphisms in natural populations be thor- 
oughly understood. 



ACKNOWLEDGEMENTS 

This work was begun at Liverpool Univer- 
sity and continued, in part, at University Col- 
lege London. Scoring of post-1982 samples, 
new analyses presented here and writing up 
were done at the Bishop Museum. I thank 
these institutions for the opportunities and fa- 
cilities provided. I also thank Professor A. J. 
Cain and Dr. M. S. Johnson for comments on 
the manuscript, Dr. L. A. Mehrhoff for compu- 
tational and statistical assistance and the var- 
ious friends and colleagues, especially С. T. 
French and A. M. Cassin, who assisted with 
field work. 



96 



COWIE 



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CAIN, A. J., 1983, Ecology and ecogenet- 
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pp. 597-647. 

CAIN, A. J., 1984, Genetics of some 
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CAIN, A. J., & L. M. COOK, 1989, Persis- 
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CAIN, A. J., L M. COOK & J. D. CURREY, 
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I 



POLYMORPHISM IN THEBA PISANA 



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SOKAL, R. R. & F. J. ROHLF, 1973, Intro- 
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WADE, M. J. & S. KALISZ, 1990, The 
causes of natural selection. Evolution, 44- 
1947-1955. 

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CLARKE, 1980, Temporal changes of gene 
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Revised Ms. accepted 16 December 1991 



MALACOLOGIA, 1992, 34(1-2): 99-106 

NULL ALLELES AND HETEROZYGOTE DEFICIENCIES AMONG 

MUSSELS (MYTILUS EDULIS AND M. GALLOPROVINCIALIS) 

OF TWO SYMPATRIC POPULATIONS 

J. P. A. Gardner 1 

School of Biological Sciences, University College of Swansea, Swansea, SA2 8PP, 

United Kingdom 

ABSTRACT 

The frequencies of null (enzymatically inactive) alleles estimated from two-banded pheno- 
types at an esterase-D (dimeric enzyme) locus among two sympatric Mytilus edulis/Mytilus 
galloprovincialis populations (Croyde and Whitsand) in southwestern England are 0.00823 and 
0.00751 respectively. These values fall in the range of null allele frequencies predicted for British 
mussel populations. At Croyde, there were significantly more M. galloprovincialis than M. edulis 
null heterozygotes. This is thought to result from linkage of M. edulis null alleles to genes at a 
selective disadvantage compared with M. galloprovincialis, which indirectly reduces null allele 
frequency. Observed and predicted frequencies of null alleles are used to test the hypothesis 
that heterozygote deficiencies (compared to Hardy-Weinberg expectations), which are often 
observed among marine molluscan populations, might be caused by null alleles. 

Key words: null alleles, heterozygote deficiency, Mytilus edulis, Mytilus galloprovincialis. 



INTRODUCTION 

Null (enzymatically inactive) alleles at loci 
encoding enzymes have been reported in 
populations of plants (Allendorf et al., 1982; 
Lafiandra et al., 1987) and animals (Voelker 
et al., 1 980a, b; Langley et al., 1 981 ; Skibinski 
et al., 1983; Foltz, 1986; Katoh & Foltz, 1989). 
Null alleles usually occur only rarely, often 
<0.005 (Ayala et al., 1972; Voelker et al., 
1980a; Langley et al., 1981; Allendorl et al., 
1982), although higher frequencies have 
been reported (Seiander & Yang, 1969; 
Coyne & Felton, 1977; Freeth & Gibson, 
1985). 

In southwestern England, the marine mus- 
sels Mytilus edulis and M. galloprovincialis 
occur sympatrically and hybridize (Skibinski, 
1983; Gardner & Skibinski, 1988). The sys- 
tematic status of M. galloprovincialis has 
been reviewed by Seed (1978), Gosling 
(1984) and Koehn (1991). One of the best 
methods for distinguishing between the two 
mussel types is electrophoresis of an es- 
terase-D {Est-D) locus, which has a genetic 
identity (/ value; Nei, 1972) of 0.039 in com- 
parisons between M. edulis from south Wales 
and M. galloprovincialis from Italy (Skibinski 
et al., 1980). Est-D is a dimeric enzyme 



(Harris & Hopkinson, 1976) that produces a 
three-banded heterozygote phenotype. It is 
a particularly good locus for observing null 
alleles not only because it is the most effec- 
tive locus for detecting differences between 
the two mussel types, but also because two- 
banded null allele phenotypes are easily 
scored. However, estimates of this sort will 
tend to underestimate overall null allele 
frequency if the null allele produces a protein 
that fails to dimerize with the normal protein, 
that is, no heterodimer is produced, or if the 
heterodimer is formed but is not enzymatically 
expressed, that is, it is not visible on the gel. 
Although the frequency of such events can 
be determined by pedigree analysis and 
biochemical methods, these techniques gen- 
erally preclude investigations of the large 
numbers (several thousands) of individuals 
necessary for population studies. 

A phenomenon often observed among ma- 
rine molluscan populations is a significant 
deficit of heterozygotes compared to Hardy- 
Weinberg expectations (Milkman & Beaty, 
1970; Berger, 1983; Skibinski et al., 1983; 
Singh & Green, 1984; Zouros & Foltz, 1984; 
Gaffney et al., 1990). It has been suggested 
that null heterozygotes are one of the possi- 
ble explanations for this deficit (Skibinski et 



1 Present address: Marine Sciences Research Laboratory, Memorial University of Newfoundland, St. John's, Newfoundland, 
A1C5S7, Canada 



99 



100 



GARDNER 



TABLE 1. Allele distributions in pure M. edulis and M. galloprovincialis populations, after Skibinski et al. 
(1980) and Sanjuan et al. (1990). 



Allele 


M edulis^ 


M. edulis 1 


M. galloprovincialis 3 


M. galloprovincialis 4 


Est-D 










76 








0.019 





82 





0.005 


0.048 


0.038 


90 


0.036 


0.024 


0.905 


0.950 


93 





0.001 








100 


0.946 


0.941 


0.016 


0.013 


103 








0.013 





107 


0.004 


0.004 








110 


0.015 


0.020 








118 





0.003 








Odh 










80 







0.012 




100 


0.020 




0.396 




115 


0.930 




0.131 




129 


0.050 




0.459 




140 







0.004 





1 Mean allele frequency of M. edulis from Holland and Denmark (Sanjuan et al., 1990). 

2 Allele frequencies of M. edulis from Swansea, South Wales (Skibinski et al., 1980). 

3 Mean allele frequency of M. galloprovincialis from Spanish Mediterranean (Sanjuan et al., 1990). 

4 Allele frequencies of M. galloprovincialis from Venice, Italy (Skibinski et al., 1980). 



al., 1983; Beaumont et al., 1985; Mallet et al., 
1985; Gaffney et al., 1990). Despite the fact 
that few data exist concerning null allele fre- 
quencies, it is usually assumed that null alle- 
les do not occur at sufficiently high frequen- 
cies to explain the observed heterozygote 
deficits. 

In this paper, genotype-dependent differ- 
ences in the frequencies of two-banded null 
heterozygotes are reported. Observed and 
predicted frequencies of null alleles are ap- 
plied to data from mussel populations to test if 
null alleles can explain heterozygote deficien- 
cies. 



MATERIALS AND METHODS 

During the period from October 1986 to 
June 1989, 3,159 mussels from Croyde Bay, 
north Devon, and 2,929 mussels from Whit- 
sand Bay, south Cornwall, southwestern En- 
gland, were collected from locations as previ- 
ously reported (Skibinski, 1983; Gardner & 
Skibinski, 1988). Mussels were electro- 
phoresed, and stained for the esterase-D 
(EST-D; EC 3.1.1.1) and octopine dehydro- 
genase loci (ODH; EC 1.5.1.11). Some mus- 
sels were also stained for the mannose phos- 
phate isomerase locus (MPI; EC 5.3.1.8). 
Tissue samples were prepared from hepato- 
pancreas (liver) homogenized in an equal vol- 



ume of distilled water. Following centrifuga- 
tion at 0°C and 3000 rpm for 15 min, the 
supernatant was used as the enzyme source. 
EST-D, ODH and MPI were run on a Tris-citric 
acid (pH 6.9) buffered gel (Grant & Cherry, 
1985). The following staining methods were 
employed; that of Ahmad et al. (1977) for 
EST-D, Beaumont et al. (1980) for ODH, and 
Grant & Cherry (1985) for MPI. 

The migratory position (identity) of a null 
allele cannot be determined because it can be 
anodal or cathodal to the two-banded null 
phenotype. Similarly, the identity of the ex- 
pressed allele in the heterozygous state with 
the null allele cannot be determined because 
it is unknown which band of the phenotype is 
the allele and which the heterodimer. It is 
therefore not possible to determine individual 
null allele frequencies, nor to determine the 
frequencies of Est-D alleles in the heterozy- 
gous state with the null alleles. For this rea- 
son, the synthetic compound allele system 
described by Skibinski (1983) was used to 
classify null genotypes. At a given locus, the 
compound E allele is obtained by pooling 
those alleles at highest frequency in M. edu- 
lis, and the compound G allele by pooling 
those alleles at highest frequency in M. gal- 
loprovincialis. For example, the Est-CF allele 
occurs at frequencies of 0.01 and 0.96 at 
Bude and Swansea, "pure" M. galloprovincia- 
lis and M. edulis populations, respectively 



NULL ALLELES AMONG HYBRID MUSSELS 



101 



(Edwards & Skibinski, 1987). The allele dis- 
tributions among the two mussel types (Table 
1) have been established by Skibinski et al. 
(1980) and Sanjuan et al. (1990). Although 
most alleles at a locus occur in both mussel 
types, the frequency differences are quite 
pronounced for the most common alleles. 
Frequency differences of rare alleles are less 
pronounced, but rare alleles contribute less 
often to compound alleles. The compound 
genotype of Est-D null heterozygotes was es- 
timated by reference to the genotype at the 
Odh locus, which is in tight linkage disequilib- 
rium with the Est-D locus. There are signifi- 
cant excesses of the E/E E/E (M. edulis-Wke) 
and G/G G/G (M. galloprovincialis-Wke) dilo- 
cus compound genotypes (Skibinski, 1983; 
Gardner & Skibinski, 1988). Mussels with Est- 
D null alleles were classified as M. edulis-Wke 
if they possessed the Odh E/E compound 
genotype, as intermediate or putative F1 hy- 
brid if they exhibited the Odh E/G compound 
genotype, and as M. galloprovincialis-Wke if 
they possessed the Odh G/G compound geno- 
type. For one null heterozygote, genotype 
was assigned on the basis of the compound 
genotype at the Mpi locus (which is also in 
tight linkage disequilibrium with the Est-D lo- 
cus; Gardner & Skibinski, 1988) because the 
Odh genotype was not easily scored. Analy- 
sis was carried out to determine if null alleles 
occur at significantly greater frequency 
among M. edulis, M. galloprovincialis, or hy- 
brid mussels. 

Finally, null allele frequencies for three loci 
were predicted for the Croyde and Whitsand 
populations from the equation F IT = 2x/ 
(1 + x) (where x is null allele frequency and F IT 
is a measure of deviation from Hardy-Wein- 
berg expectations in the compound popula- 
tion obtained by pooling all subdivisions) if 
Hardy-Weinberg equilibrium (HWE) is as- 
sumed and if null heterozygotes are scored 
as homozygotes and null homozygotes are 
inviable. F IT estimates have been obtained for 
British M. edulis and M. galloprovincialis pop- 
ulations, and for "pure" M. edulis populations 
by Skibinski et al. (1983). The observed Est-D 
two banded null phenotype frequencies and 
the predicted null allele frequencies were both 
applied to estimates of the D statistic (D = 
( H o /H eH - where H is the observed, and H e 
the expected heterozygote frequencies), 
which is commonly used to estimate devia- 
tions from Hardy-Weinberg expectations (a 
negative D value indicates a deficiency of het- 
erozygotes compared with expectations). D 



statistics for the Croyde and Whitsand popula- 
tions are given by Gardner & Skibinski (1 988), 
and have been recalculated to determine if 
heterozygote deficiencies are attributable to 
observed and predicted null allele frequen- 
cies. New estimates of H have been calcu- 
lated by multiplying the original H values of 
Gardner & Skibinski (1988) by 1 +x, where x 
is null allele frequency. Null allele frequencies 
(x values) give a direct estimate of the under- 
estimation of observed heterozygote fre- 
quency and the overestimation of observed 
homozygote frequency. New H e estimates 
should be greater than the original H e esti- 
mates because with the inclusion of null het- 
erozygotes, heterozygosity will increase 
when mussels incorrectly scored as homozy- 
gotes are now correctly scored as heterozy- 
gotes. However, the original estimates of H e 
have been used because the identities of the 
null alleles remain unknown, so new esti- 
mates of H e have not been calculated. This 
then gives a conservative test, in the sense 
that (1) the calculated D values are slightly 
greater than they should be, that is, the het- 
erozygote deficiency at each locus is esti- 
mated to be slightly smaller than it really is, 
and (2) the corresponding X 2 and P (signifi- 
cance) values are slightly less significant than 
they should be. Because the method of esti- 
mating null allele frequency assumes HWE, 
the F IT values of Skibinski et al. (1983) for 
"pure" M. edulis populations (which, on the 
whole, are in HWE), and F, T values for M. 
edulis and M. galloprovincialis populations 
(which are in HWE less often) have both been 
employed (Table 3). 



RESULTS 

At Croyde, two-banded null allele pheno- 
types occurred at an average frequency of 
0.00823. There were 1 .93 times as many M. 
galloprovincialis null heterozygotes as ex- 
pected, based upon an hypothesis of equal 
null allele frequency for all genotypes 
(G- 10.508, df = 2, P = 0.003). The E/E and 
E/G compound genotypes showed null het- 
erozygote frequencies that were 55.1% and 
53.8% of their expected frequencies (Table 
2). At Whitsand, null alleles were observed at 
a mean frequency of 0.00751 . There was no 
significant association between null heterozy- 
gote frequency and compound genotype (G 
= 0.816, df = 2, P = 0.672). There were 
non-significant excesses of E/G and G/G, and 



102 



GARDNER 



TABLE 2. Frequencies of compound genotypes and 
mussels from Croyde and Whitsand. 


null heterozygotes 


at 


an 


esterase-D 


locus for 


Site 






Croyde 








Whitsand 




Compound genotype 


E/E 


E/G 


G/G 




E/E 


E/G 


G/G 



No. of null heterozygotes 6 3 17 7 7 8 

No. of mussels without null alleles 1473 647 1013 1194 831 822 

Observed % frequency of null heterozygotes 0.473 0.462 1 .650 0.583 0.835 0.899 

Expected number of null heterozygotes* 12.104 5.317 8.324 8.968 6.242 6.625 

'assuming equal frequencies of null heterozygotes for all compound genotypes 



a non-significant deficit of E/E null heterozy- 
gotes compared with expectations (Table 2). 
Inter-population comparisons reveal that 
there were no significant differences between 
the two populations in the total number of null 
heterozygotes, or in the frequencies of M. 
edulis null heterozygotes, hybrid {E/G) null 
heterozygotes, and M. galloprovincialis null 
heterozygotes. 

D values calculated by Gardner & Skibinski 
(1988) for the Ap, Est-D and Pgi loci at 
Croyde and Whitsand are shown in Table 3. 
In all cases, there were significant deficien- 
cies of heterozygotes compared with expec- 
tations. Recalculation of D for the Est-D locus 
based upon the observed two-banded null 
heterozygote frequencies does not reduce 
the significance of the deficiencies at either 
site. Changes in the associated X 2 values are 
very small and do not have a significant effect 
upon probability levels (Table 3). With the ex- 
ception of the Ap locus at Whitsand, the same 
is true when predicted null allele frequencies 
(firstly for "pure" M. edulis populations, and 
secondly for M. edulis and M. galloprovincialis 
populations) estimated from the equation F, T 
= 2x/(1 +x) are used to re-calculate D val- 
ues of the Ap, Est-D and Pgi loci. 



DISCUSSION 

Est-D null heterozygotes occur significantly 
more often among M. galloprovincialis than 
among M. edulis at Croyde. This is in line with 
the findings of Skibinski et al. (1983), who 
predicted an Est-D null allele mean frequency 
of 0.018 in M. edulis populations and a mean 
frequency of 0.071 in M. edulis and M. gallo- 
provincialis populations. Null alleles most fre- 
quently result from single point mutations 
(Schwartz & Sofer, 1975; Voelker et al., 



1980b; Scallon et al., 1987), but can also be 
due to small insertions that diminish transcrip- 
tion (Burkhart et al., 1984; Gibson & Wilks, 
1989; but see Jiang et al., 1988). Two sepa- 
rate explanations can account for the lower 
frequency of nulls in M. edulis than in M. gal- 
loprovincialis. First, the M. edulis genome 
might be less susceptible to the mutational 
events that give rise to null alleles. Second, 
tight linkage of M. edulis nulls to other loci in 
the M. edulis genome, which experiences a 
significantly higher age-dependent mortality 
rate than M. galloprovincialis (Gardner & Ski- 
binski, submitted), might indirectly reduce the 
frequency of nulls in M. edulis. At Croyde, 
comparison of the mean shell length of M. 
edulis versus M. galloprovincialis with null al- 
leles (mean ± SD of 21 .2 mm ± 6.20, n = 6, 
versus 39.7 mm ± 4.02, n = 17) indicates 
that M. edulis null alleles occur among 
smaller, younger mussels. No M. edulis larger 
than 30 mm length were found with null al- 
leles, supporting the suggestion that in- 
creased mortality experienced by M. edulis 
compared to M. galloprovincialis acts indi- 
rectly to reduce null allele frequency among 
M. edulis. This second explanation is pre- 
ferred because of supporting data, whereas 
the first explanation requires the invoking of 
differential mutation rates, for which there 
is no direct evidence. Evidence of length- 
dependent change in genotype frequencies 
(Gardner & Skibinski, 1988) indicates that M. 
edulis viability with regard to M. galloprovin- 
cialis is greater at Whitsand than at Croyde, 
providing an explanation for the non-signifi- 
cant differences in genotype-dependent null 
allele frequencies at Whitsand. 

Few estimates of null allele frequencies ex- 
ist for marine molluscan populations. Gaffney 
et al. (1990) estimated mean null allele fre- 
quencies in the coot clam, Mulinia lateralis, to 



NULL ALLELES AMONG HYBRID MUSSELS 



103 



TABLE 3. Recalculation of D values to take into consideration (a) the observed two banded null 
heterozygote frequencies at the esterase-D locus, and the predicted frequencies of null heterozygotes at 
three loci for (b) "pure" M. edulis populations, and (c) for M. edulis and M. galloprovincialis populations. 







CROYDE 




Ap 


WHITSAND 




Ap 


Est-D 


Pgi 


Est-D 


Pgi 


1 Ho 


0.379 


0.196 


0.457 


0.417 


0.277 


0.516 


1 He 


0.462 


0.505 


0.651 


0.493 


0.485 


0.687 


1 D 


-0.179* 


-0.61 Г" 


-0.298*** 


-0.153* 


-0.430*** 


-0.250*** 


1 n 


422 


474 


501 


381 


376 


411 


2 x 


— 


0.00823 


— 





0.00751 


_ 


2 Ho 


— 


0.198 


— 


— 


0.284 





2 He 


— 


0.505 


— 


— 


0.485 





2 D 


— 


-0.608*** 


— 


— 


-0.425*** 





2 n 


— 


474 


— 


— 


376 


— 


3 x 


0.0101 


0.0304 


0.0163 


0.0101 


0.0304 


0.0163 


3 Ho 


0.383 


0.202 


0.465 


0.422 


0.285 


0.524 


3 He 


0.462 


0.505 


0.651 


0.493 


0.485 


0.687 


3 D 


-0.171* 


-0.600*** 


-0.287*** 


-0.145* 


-0.413*** 


-0.237*** 


3 n 


422 


474 


501 


381 


376 


411 


4 x 


0.0225 


0.2903 


0.0482 


0.0225 


0.2903 


0.0482 


4 Ho 


0.388 


0.253 


0.479 


0.427 


0.357 


0.541 


4 He 


0.462 


0.505 


0.651 


0.493 


0.485 


0.687 


4 D 


-0.16Г 


-0.499*** 


-0.264*** 


-0.134 NS 


-0.264*** 


-0.213*** 


4 n 


422 


474 


501 


381 


376 


411 



x null allele frequency 

Ho observed number of heterozygotes 

He expected number of heterozygotes 

D heterozygote deviation compared with Hardy-Weinberg expectations, calculated as D = (Ho/He)-1 

NS not significant 

*P<0.05 

*"P<0.001 

n number of mussels 

1 Data from Gardner & Skibinski (1988). 

2 Data from Gardner & Skibinski (1988) recalculated incorporating the observed frequencies of two banded null heterozygote 

phenotypes. 

3 Data from Gardner and Skibinski (1988) recalculated incorporating frequencies of predicted null alleles for "pure" M. edulis 

populations (Skibinski et al., 1983). 

"Data from Gardner & Skibinski (1988) recalculated incorporating frequencies of predicted null alleles for M. edulis and M. 

galloprovincialis populations (Skibinski et al., 1983). 



be 0.001 at the GPI locus and 0.0003 at the 
AP2 locus. In common with other large stud- 
ies, the null allele frequencies reported by 
Gaffney et al. (1990) are for two-banded phe- 
notypes of dimeric enzymes, and as in the 
present study, must therefore be viewed as 
minimum estimates. Because of the difficulty 
of distinguishing one-banded null heterozy- 
gotes from homozygotes, no null allele 
frequency data are given for monomeric en- 
zymes for the coot clam. No null heterozy- 
gotes were observed at the other dimeric loci. 
Skibinski et al. (1983) predicted that null allele 
frequency in British populations of M. edulis 
and M. galloprovincialis should be in the 
range 0.003 to 0.102 for 1 1 allozyme loci. The 



observed null allele frequencies reported here 
(0.00823 at Croyde and 0.00751 at Whitsand) 
are therefore consistent with predicted values 
over several loci (Skibinski et al., 1983). 

Mean null allele frequency estimated from 
the equation F, T = 2x/(1 +x) for seven Est-D 
alleles is 0.102 for M. edulis and M. gallopro- 
vincialis, and for the pure M. edulis is 0.019 
(Skibinski et al., 1983); that is, both values are 
substantially greater than the observed fre- 
quencies of two-banded null heterozygotes. 
This might be accounted for by the undetect- 
able presence of null alleles that do not pro- 
duce a heterodimer, such individuals being 
scored as homozygotes. This is particularly 
important because it is often suggested that 



104 



GARDNER 



null alleles might be responsible for observed 
heterozygote deficiencies (Boyer, 1974; Ski- 
binski et al., 1983; Beaumont et al., 1985; 
Mallet et al., 1985; Foltz, 1986; Gaffney et al., 
1990). However, estimates from natural pop- 
ulations suggest that null alleles are not com- 
mon enough, often by orders of magnitude, to 
account for homozygote excesses, and recal- 
culation of D statistics (Table 3) seems to con- 
firm this. D values increase (heterozygote de- 
ficiencies decrease) in all cases as expected 
when observed or predicted null allele fre- 
quencies are used to re-estimate observed 
and expected heterozygote frequencies. 
However, changes in significance levels are 
generally small and only effect probability val- 
ues in one case. This indicates that null alle- 
les generally do not occur at sufficient fre- 
quency to significantly effect heterozygote 
deficiencies. At Whitsand, the heterozygote 
deficiency at the Ap locus reported by Gard- 
ner & Skibinski (1988) is small but significant 
(X 2 = 4.389, df = 1, P = 0.0362). Recalcu- 
lation of D, adjusting for predicted null allele 
frequency among "pure" M. edulis popula- 
tions, reduces the X 2 value to 3.909 (df = 1 , 
P = 0.048), that is, to borderline significance. 
Further recalculation adjusting for null allele 
frequency among M. edulis and M. gallopro- 
vincialis populations reduces the X 2 value to 
3.361 (df = 1, P = 0.0668), that is, a non- 
significant deficiency of heterozygotes. It 
seems likely that only in such cases as this, 
where heterozygote deficiencies are small but 
(borderline) significant, can observed or pre- 
dicted null allele frequencies contribute signif- 
icantly to heterozygote deficiencies. 

In the single case among marine bivalves 
where very high frequencies of null alleles 
have been observed (Foltz, 1986), it was sug- 
gested (Gaffney et al., 1990) that the produc- 
ton of aneuploid gametes (i.e., gametes with 
one or more chromosomes, or chromosome 
segments, absent) might better explain the 
high frequency of apparent null alleles. There 
is thus still no evidence from natural popula- 
tions to indicate that heterozygote deficien- 
cies result solely from high frequencies of null 
alleles. Although null alleles appear likely to 
contribute to heterozygote deficiencies, more 
significant causes include selection, the 
Wahlund effect, aneuploidy or imprinting, that 
is, the differential expression of genetic ma- 
terial, at the chromosomal or allelic level, de- 
pending on whether the genetic material was 
derived from the male or female parent 
(Gaffney et al., 1990). In the case of the sym- 



patic Mytilus populations of southwestern 
England, selection and the Wahlund effect 
are the most likely explanations for significant 
heterozygote deficiencies (Skibinski, 1983; 
Gardner & Skibinski, 1988). 



ACKNOWLEDGEMENTS 

Thanks to Eric Roderick for help in the field; 
to Donna Gardner for data collation; to Andy 
Beaumont, David Skibinski and three anony- 
mous reviewers for helpful comments. The re- 
search was supported in part by a NERC Stu- 
dentship to the author. 



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106 GARDNER 

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MALACOLOGIA, 1992, 34(1-2): 107-128 

A NEW PROBLEMATICAL HYGROMIIDAE FROM THE AEOLIAN ISLANDS 
(ITALY) (PULMONATA: HELICOIDEA) 1 

Folco Giusti 2 , Giuseppe Manganelli 2 & Jorge V. Crisci 3 

ABSTRACT 

Helicotricha n. gen. is proposed for a very small hygromiid from the Aeolian Islands, Italy: H. 
carusoi n. sp. The new species has a shell with persistent postembhonal hairs and is charac- 
terized anatomically by: a right ommatophore retractor independent of the genitalia; a penial 
nerve originating from the right cerebral ganglion; a vaginal complex with digitiform glands and 
dart-sac complex consisting of two couples of stylophores disposed on opposite sides of the 
vagina (each couple is formed by a larger dart-bearing outer stylophore and a smaller dartless 
inner stylophore); and a penial complex with a very peculiar penial papilla. Direct anatomical 
comparison of the new genus with other genera of the Hygromiidae suggested that it may be 
very closely related to Microxeromagna. This hypothesis was subsequently found to be sup- 
ported by the results of cladistic analysis. New evidence is thus provided to confirm that the 
distinction between the Hygromiinae and the Trichiinae is artificial. A neotype is designated for 
Helix aetnea Benoit, 1 857, a junior synonym of Xerotricha conspurcata (Draparnaud, 1 801 ), and 
the presence of Helicopsis (s. str.) in western North Africa is confirmed. 

Key words: Hygromiidae, Aeolian Islands, Italy, western Mediterranean, Helicella, Xerotricha, 
Helicopsis, systematics. 



INTRODUCTION 

Recent insular equilibrium theory studies 
based on analytical comparison of the mala- 
cofaunas of the Tuscan and Aeolian archipel- 
agos, Italy (Piantelli et al., 1990), have moti- 
vated new field research and study of the 
material collected from the various islands. As 
happened for the islands of the Tuscan Archi- 
pelago (Giusti & Manganelli, 1989, 1990), 
new data emerged for the Aeolians. A new 
species of the Hygromiidae was identified 
amongst material of Xerotricha conspurcata 
(Draparnaud, 1801) collected in all the Aeo- 
lian Islands. The Hygromiidae are a group of 
helicoids of western Palaearctic distribution 
recently separated from the Helicidae as a 
distinct family and characterized by a bursa 
copulatrix duct free from the diaphragm wall 
and a variable number of stylophores. 

The peculiar structure of the genitalia made 
it difficult to establish relationships and ge- 
neric status of the new species. Although the 
2 + 2 structure of the dart-sac complex as- 
sociated with the vagina, and the right om- 
matophore retractor free of penis and vagina, 
at first glance suggested a relationship with 



the genus Helicopsis, details of the penial 
complex suggested other possibilities. In par- 
ticular, as in the case of Helicella (one large, 
evident and one vestigial, externally invisible, 
stylophore on opposite sides of the vagina) 
and Xerolenta (one normal and one modified 
stylophore on opposite sides of the vagina) 
which appear to form monophyletic groups re- 
spectively with Candidula (one large, evident 
and one vestigial, externally invisible, stylo- 
phore on one side of the vagina) and Xero- 
munda (one normal and one modified stylo- 
phore on one side of the vagina) respectively 
(Hausdorf, 1988, 1990a; Giusti & Manganelli, 
1989; Manganelli & Giusti, 1989), it seems 
highly probable that the new genus (one 
larger outer stylophore and one smaller dart- 
less inner stylophore on opposite sides of the 
vagina) is a member of the monophyletic 
group to which also Microxeromagna (one 
larger outer stylophore and one smaller dart- 
less inner stylophore on one side of the va- 
gina) belongs. Accordingly, in view of the fact 
that Helicella, Xerolenta, Candidula, Xero- 
munda are unanimously regarded as distinct 
generic taxa and in view also of our consid- 
erations on character weighting in establish- 



1 Notulae Malacologicae, LI. 

2 Dipartimento di Biológica Evolutiva, Università degli Studi di Siena, Via Mattioli 4, 1-53100 Siena, Italy 

laboratorio de Sistemática y Biología Evolutiva, Museo de La Plata, Paseo del Bosque 1900, La Plata, Argentina 

107 



108 



GIUSTI, MANGANELLI & CRISCI 



ing systematic rank in the Hygromiidae (Man- 
ganelh & Giusti, 1988), a new genus is intro- 
duced for the new species. 

MATERIALS AND METHODS 

Empty shells and whole specimens were 
collected in the litter or under stones and 
wood on rocky slopes of the islands (locality 
data follows species description). Living spec- 
imens were left in water to drown for 24 h then 
preserved in ethanol 75%. Relaxed material 
was studied by optical microscopy (Wild 
M5A). Bodies were isolated then dissected 
using thin, pointed watchmaker's forceps. Im- 
ages of isolated portions of body and genitalia 
were drawn using a Wild camera lucida. Rad- 
ulae were manually extracted from buccal 
bulbs, then washed in pure 75% ethanol, 
mounted on copper blocks with electron-con- 
ductive glue, sputter-coated with gold and 
photographed using a Philips 505 SEM. 

All shell parameters — shell maximum di- 
ameter, shell height, aperture maximum di- 
ameter, aperture height — were measured in 
variable numbers of shells from the different 
islands using a Wild M5A microscope and a 
millimetric lens. Whole shells were photo- 
graphed under optical microscope. Whole 
shells and shell surface details were photo- 
graphed under optical and scanning electron 
microscopes using the procedure described 
for the radulae. 

Detailed study of the genitalia followed, 
particularly the distal portion (penis and va- 
gina), the external and internal details of 
which proved to be diagnostic characters in 
similar previous studies (see literature cited in 
the Discussion) (Table 2). 

The entire set of character states was used 
for classical evolutionary and cladistic analy- 
sis to define the new genus. 

Cladistic analysis was performed using the 
method of phylogenetic systematics as origi- 
nally developed by Hennig (1966), who main- 
tained that only strictly monophyletic taxa 
may be regarded as historical entities and 
demonstrated that the only logical basis for 
inferring monophyly is by showing synapo- 
morphies. The distribution of the synapomor- 
phies is determined by the parsimony crite- 
rion (minimizing homoplasy). On the basis of 
these synapomorphies, the taxa are ordered 
into a specific pattern represented by a hier- 
archical branching diagram. 

The new genus Helicotricha belongs to the 
family Hygromiidae, which is delimited by 
the following unique combination of features: 



bursa copulatrix free from diaphragm; vari- 
able number of stylophores (2 + 2; + 2; 
1 + 1; + 1; + 0); stylophores, when 
present and not extremely regressed or mod- 
ified, forming dart-sac complex consisting of 
one or two double units with a common base 
and distinct distal sacs lying side by side in 
the same plane; diverticulum of bursa copul- 
atrix absent; digitiform gland tufts variable in 
number inserted on proximal vagina not close 
to where dart sacs, when present, open into 
vagina. Helicidae are characterized by: bursa 
copulatrix inserted into diaphragm; dart-sac 
complex consisting of a single stylophore; di- 
verticulum of bursa copulatrix duct present. 
Helicidae and Hygromiidae share the follow- 
ing characters: dart-sac complex, vaginal dig- 
itiform glands. 

Twenty (Table 1 ) genera are considered to 
be the terminal taxa. The minute details of the 
distal genitalia of some have recently been 
reviewed (Schileyko, 1978a, 1978b; Giusti & 
Manganelli, 1987, 1989; Hausdorf, 1988, 
1990a, 1990b; Manganelli & Giusti, 1988, 
1989). 

In the case of the two sets of genera re- 
viewed by Schileyko and Hausdorf, some 
characters were uncertain or not discussed. A 
question mark sometimes follows or substi- 
tutes the character states in Table 3. A ques- 
tion mark also substitutes the character states 
when one character is known to be present 
with more than two states in the same taxa 
(character 5 in Xeromunda and Cernuella (s. 
str.); character 16 in Xerotricha). Character 
polarity was determined by outgroup compar- 
ison methods (Watrous & Wheeler, 1981; 
Maddison et al., 1984) using the family col- 
lectively Helicidae as outgroup, because 
there is no singie genus that is a clear sister 
group. Three out fifteen characters (5, 10, 15) 
had more than two character states. These 
characters were treated as nonadditive. Two 
characters (11, 14) were autoapomorphies, 
with an additional autoapomorphy in a multi- 
state nonadditive character (10). 

The data was analyzed using a Wagner 
parsimony algorithm from Farris's phyloge- 
netic program HENNIG86, applying the im- 
plicit enumeration option for calculating trees 
(version 1.5; Farris, 1989; see also Platnick, 
1989) run on a IBM AT computer. When cla- 
distic analysis yielded more than one tree, the 
Nelsen consensus method was applied 
(Nelsen, 1979). We also used the successive 
weighting procedure (Farris, 1989), which cal- 
culates weights from the best fits to the most 
parsimonious trees, and applied them in the 



A NEW HYGROMIIDAE FROM ITALY 



109 



TABLE 1. Acronyms, genus-group taxa, type species and bibliographical sources of anatomical data. 
Some of the genera listed have a subgeneric division. For cladistic analysis, only species of 
nominotypical subgenera have been considered. 



Acronyms Genus-group taxa 

CAND Candidula Kobelt, 1871 



CAUC 
CERN 



Caucasigena Lindholm, 1927 
Cernuella Schlüter, 1838 



EDEN Edentiella Polinski, 1929 

HELL Helicella Férussac, 1821 

HELP Helicopsis Fitzinger, 1833 



HELT Helicotricha Giusti & 

Manganelli, 1992 
HYGR Hygromia Risso, 1826 

HYGH Hygrohelicopsis Schileyko, 

1978a 
KOKO Kokotschashvilia Hudec & 

Lezhawa, 1969 
LEUC Leucozonella Lindholm, 1927 

MICR Microxeromagna Ortiz de 

Zarate Lopez, 1950 

NANA Nanaja Schileyko, 1978b 

PLIC Plicuteria Schileyko, 1978a 

PXER Pseudoxerophila Westerlund, 

in Westerlund & Blanc, 1879 

TRIC Trichia, Hartmann, 1840 

XERL Xerolenta Monterosato, 1 892 

XERM Xeromunda Monterosato, 
1892 



XERS Xerosecta Monterosato, 1892 

XERT Xerotricha Monterosato, 1 892 



Type-species 
Glischrus (Helix) candidula, 
Studer, 1820, = Helix 
unifasciata Poiret, 1801 

Helix eichwaldi Pfeiffer, 1 846 

Helix variabilis Draparnaud, 

1801, = Cochlea virgata 

Da Costa, 1778 

Helix edentula Draparnaud, 

1805 

Helix itala Linnaeus, 1758; cf. 

Opinion 431 

Helix striata Müller, 1774 



Helicotricha carusoi Giusti, 

Manganelli & Crista, 1992 

Helix cinctella Draparnaud, 

1801 

Hygrohelicopsis darevskii 

Schileyko, 1978a 

Helix holotricha O. Boettger, 

1884 

Helix rubens von Martens, 1 874 

Helix stolismena Bourguignat, 

in Servain, 1880, = Helix 

armillata Lowe, 1852 

Nanaja cumulate Schileyko, 

1978b 

Helix lubomirskii Slosarski, 1 881 

Helix (Pseudoxerophila) 

bathytera Westerlund, in 

Westerlund & Blanc, 1879 

Helix hispida Linnaeus, 1 758 

Helix obvia Menke, 1 828 

Helix turbinate, sensu 

Monterosato, 1892, non De 

Cristofori & Jan, 1832) (cf. 

Hausdorf, 1988; Manganelli & 

Giusti, 1988; 1989; an 

application to the I. C.Z.N, is in 

progress by Giusti & Manganelli 

Helix explánala Müller, 1 774 

Helix conspurcata Draparnaud, 

1801 



Sources 
Hausdorf, 1988: (С. unifasciata, 
С. gigaxii); personal 
unpublished data on С spadae, 
С intersecta, С unifasciata 
Schileyko, 1978a, 1978b 
Hausdorf, 1988; Manganelli & 
Giusti, 1988 

Schileyko, 1978a, 1978b 

Hausdorf, 1988; Giusti & 
Manganelli, 1989 
Giusti & Manganelli, 1989; 
Schileyko, 1978b; 
Hausdorf, 1990b 
this paper 

Giusti & Manganelli, 1987 

Schileyko, 1978a, 1978b 

Schileyko, 1978a, 1978b 

Schileyko, 1978a, 1978b 
Hausdorf, 1988, 1990c; 
Manganelli & Giusti, 1988 

Schileyko, 1978b 

Schileyko, 1978a, 1978b 
Hausdorf, 1988 



Schileyko, 1978a, 1978b 
Hausdorf, 1988 
Hausdorf, 1988, 1990a; 
Manganelli & Giusti, 1989 



Manganelli & Giusti, 1988 
Hausdorf, 1988; Giusti & 
Manganelli, 1989 



weighting procedure until there were no 
changes in successively produced trees. 

SYSTEMATIC DESCRIPTION 
Helicotricha n. gen 

Diagnosis 

Very small hygromiid with shell having per- 
sistent postembronial hairs; anatomically 
characterized by right ommatophore retractor 
independent of genitalia; penial nerve from 



right cerebral ganglion; vaginal complex with 
digitiform glands and dart-sac complex con- 
sisting of two pairs of stylophores, each cou- 
ple comprising a larger dart-bearing outer sty- 
lophore and a smaller dartless inner 
stylophore; penial complex having a very pe- 
culiar penial papilla. 

Description 

Shell: Small, hairy, opaque-brown in colour, 
with white flecks. Spire consisting of approx- 



1 1 GIUSTI, MANGANELLI & CRISCI 

TABLE 2. List of characters 

1 — Penial nerve. 

— From right cerebral ganglion = 

— From right pedal ganglion = 1 

Remarks: no data for Caucasigena, Edentiella, Hygrohelicopsis, Kokotschashvilia, Leucozonella, Nanaja, 

Plicuteria. In Manganelli & Giusti (1989: 4) was wrongly reported for Xeromunda "from right pedal 

ganglion." Revision of original data indicates that penial nerve comes out of right cerebral ganglion. 

2 — Right ommatophore retractor. 
— Between penis and vagina = 
— Independent of penis and vagina = 1 
Remarks: No data for Psudoxerophila. 

3 — Number of stylophores and/or their dérivâtes forming the dart-sac complex. 
—2 + 2 = 
—0 + 2 = 1 

4 — Shape and position of stylophore groups in relation to vagina. 

— Stylophores Trichia type: each stylophore group (each composed of an outer and an inner stylophore) 

slender and entering vagina through a slender neck (Manganelli & Giusti, 1988: fig. 14 E) = 

— Stylophores not Trichia type: each stylophore group (each composed of an outer and an inner 

stylophore) wide and fused to inner walls of vagina for a long tract (Manganelli & Giusti, 1988: fig. 14 A) 

= 1 

Remarks: Helicopsis: based on the type species only (Giusti & Manganelli, 1989). Schileyko (1978b) and 

Hausdorf (1990b) show drawings of the genitalia of H. striata and of some other species (H. likharevi, H. 

retowskii) in which the situation is slightly different. A slightly different situation also occurs in the species 

studied in the present paper and referred to as Helicopsis sp. 

5 — Shape and dimensions of stylophore groups. 

— Each group formed by an inner and an outer stylophore of similar dimensions (Manganelli & Giusti, 

1988: fig. 14 A) = 

— Each group consisting of large outer stylophore and small externally visible inner stylophore (Giusti & 

Manganelli, 1989: fig. 9 A) = 1 

— Each group consisting of large outer stylophore and externally visible residues of the inner stylophore 

(Manganelli & Giusti, 1989: fig. 1 E) = 2 

— Each group consisting of larger outer stylophore and very small, not externally visible, inner stylophore 

(Schileyko, 1987a: fig. 43) = 3 

— Each group consisting of large outer stylophore and extremely reduced not externally visible inner 

stylophore (Giusti & Manganelli, 1989: fig. 9 C) = 4 

Remarks: the scheme of the dart-sac complex in Helicella reproduced by Hausdorf (1988: fig. 8) is 

incorrect: the inner stylophore, referred to as "Nebensack," appears too large and externally visible. 

6 — Digitiform glands. 
— All around vagina = 
— On one side of vagina = 1 

Remarks: situation not clear enough in drawings by Schileyko (1978a, 1978b) of the genitalia of 
Edentiella, Nanaja and Plicuteria. The situation in Hygrohelicopsis and Leucozonella showed by the 
same author (Schileyko, 1978a, 1978b) seems to indicate digitiform glands all around vagina. Species of 
Cernuella (s. str.) show digitiform glands all around vagina (C. caruanae) or on one side of the vagina 
(C. virgata). 

7 — Basal portion 

— Stylophore groups opening directly into vagina without a wide basal dilated portion (Manganelli & 

Giusti, 1988: fig. 14 E) = 

— Stylophore groups opening in a wide dilated basal portion (Manganelli & Giusti, 1989: fig. 1 E) = 1 

8 — Inner stylophores. 

— With thin muscular walls and large internal cavity (Manganelli & Giusti, 1988: fig. 14 E) = 
— With thick muscular walls and small internal cavity (Manganelli & Giusti, 1988: fig. 14 A) = 1 
Remarks: transverse and longitudinal sections of dart-sac complex in Edentiella, Kokotschashvilia, 
Leucozonella, Nanaja, Plicuteria unknown. Situation as reported by Schileyko (1978a, 1978b) for 
Caucasigena and Hygrohelicopsis not clear enough. 



A NEW HYGROMIIDAE FROM ITALY 1 1 1 

9 — Opening of stylophores. 

— Openings of inner and outer stylophore cavities into vagina close to each other (Manganelli & Giusti, 

1988: fig. 14 E) = 

— Openings of inner and outer stylophore cavities into vagina very far apart (Manganelli & Giusti, 1988: 

fig. 14 A) = 1 

10 — Inner vaginal accessory structures. 

— Opening of stylophores in a groove bordered by folds (Manganelli & Giusti, 1988: fig. 8 A) = 

— Opening of stylophores bordered by rows of papillae (Schileyko, 1987b: fig. 215) = 1 

— Vagina with one tongue-like structure for each stylophore group (two tongue-like structures when 2 

stylophore groups present) (Giusti & Manganelli, 1989: figs. 3, 9A) = 2 

— Vagina with a groove-like structure for each stylophore group (unique tube-like structure when 2 

stylophore groups present) (Giusti & Manganelli, 1989: figs. 7, 9 C) = 3 

—Vagina with dart-gun through which dart is shoot (Manganelli & Giusti, 1989: figs. 4 E, 14 A) = 4 

11 — Joint of penis and vagina 

— Penis joins vagina distally with respect to stylophores (Manganelli & Giusti, 1989: Fig. 5 F) = 

— Penis joins distal vagina level with stylophores (Manganelli & Giusti, 1989: fig. 8 B) = 1 

12 — Proximal penis. 

— Proximal penis Helicopsis type: transverse sections reveal a duct in the lumen; this duct joins the 

epiphallus lumen directly with the ejaculatory canal of the penial papilla (Giusti & Manganelli, 1989: fig. 8 

F, H) = 0. 

— Proximal penis simple: transverse sections only show the penial walls (Manganelli & Giusti, 1988: fig. 

11 F) = 1. 

Remarks: A structure, only apparently similar to those in Helicopsis, seems present in drawings by 

Manganelli & Giusti (1990: figs. 2C, ЗА, 4E for Xeromunda), Schileyko (1978b: Fig. 253 for "Helicella 

candicans)", Schileyko, in Damjanov & Likharev (1975: fig. 274, for "Helicella candicans" and fig. 278 for 

"Helicella spiruloides"). This is due to the fact that the thin external layer of the penis has been detached 

during dissection. 

13 — Glandular area on one side of terminal penis walls. 

— absent = 

— present (this paper: Fig. 3) = 1 

Remarks: no data for Caucasigena, Edentiella, Hygrohelicopsis, Kokotschashvilia, Leucozonella, Nanaja, 

Plicuteria, Pseudoxerophila; for Microxeromagna unpublished personal data. 

14 — Frenula. 

— Penial papilla with no frenula joining it to the distal penis walls (Manganelli & Giusti, 1988: fig. 7 F-H) 

= 

— Penial papilla joined by frenula to the distal penis walls (Manganelli & Giusti, 1988: fig. 6 C-D) = 1 

15 — Sections of penial papilla. 

—Trichia type (Schileyko, 1978b: fig. 221) = 

— Caucasigena type (Schileyko, 1978b: fig. 199) = 1 

—Xerosecta type (Manganelli & Giusti, 1988: fig. 8 C) = 2 

— Helicotricha type (this paper: Fig. 3C) = 3 

— Microxeromagna type (Manganelli & Giusti, 1988: fig. 11 E) = 4 

—Leucozonella type (Schileyko, 1978b: fig. 146) = 5 

—Cernuella type (Manganelli & Giusti, 1988: fig. 7 G) = 6 

Remarks: Species of Xerotricha have sections of penial papilla of Trichia type (X. apicina) and of 

Cernuella type (X. conspurcata). Due to variability, Leucozonella is based only on the type-species. 



¡mately 4 whorls separated by deep sutures, sisting of fine close longitudinal grooves. Ex- 
last whorl angled at periphery. Umbilicus ternal surface of teleoconch with superficially 
open, wide approximately 1/5 of maximum reticulated periostracal layer and transverse 
shell diameter. Aperture oblique, oval, lacking rows of very short hairs, 
internal rib. Peristome not thickened or re- 
flexed. External surface of protoconch with Genitalia: Vaginal complex with relatively long 
few faint growth lines and microsculpture con- distal vagina; dart-sac complex consisting of 



112 



GIUSTI, MANGANELLI & CRISCI 



TABLE 3. 
treated as 


Original data matrix used for cladistic analysis. All characters with 
not additive. 


more 


than two states 


are 






Character 














Taxa 


1 2 3 4 5 6 7 


8 9 


10 


11 


12 


13 


14 


15 



Outgroup 

Candidula 

Caucasigena 

Cemuella 

Edentiella 

Helicella 

Helicopsis 

Helicotricha 

Hygrohelicopsis 

Hygroma 

Kokotschashvilia 

Leucozonella 

Microxeromagna 

Nanaja 

Plicuteria 

Pseudoxerophila 

Trichia 

Xerolenta 

Xeromunda 

Xerosecta 

Xerotricha 



two pairs of stylophores disposed on opposite 
sides of vagina, each pair consisting of a 
smaller inner and a larger outer stylophore; 
outer stylophore containing slightly curved 
darts of circular section near base and oval or 
rhombic section thereafter; each inner stylo- 
phore (called "accessory sacs" by Nordsieck, 
1987, and Hausdorf, 1988; see Giusti & Man- 
ganelli, 1989: 51, for a discussion of homol- 
ogy and terminology of this structure) show- 
ing wide, totally dartless internal cavity; 
cavities of outer and inner stylophore of each 
couple in communication and opening into va- 
gina in a single opening bordered by two large 
anteriorly fused pleats. Digitiform glands 
forming two groups, each of two glands, 
sometimes apically branched, arising from 
opposite sides of distal vagina close to point 
where bursa copulatrix duct arises. Bursa 
copulatrix duct of medium length, with initial 
portion slightly flared. Penial complex with fla- 
gellum almost as long as epiphallus plus pe- 
nis. Epiphallus (from end of vas deferens to 
point of attachment of penial retractor) long, 
almost twice penis length. Penis (from point of 
attachment of penial retractor to genital 
atrium) lacking distinct penial sheath and dis- 
tally enlarged. Wide area yellow in colour 
and covered with glandular tissue, on exter- 
nal side of terminal portion of penis walls. Pe- 



nial papilla cylindrical, formed by a wide tube 
with thin walls. Penial papilla lumen continu- 
ous with that of proximal penis and epiphal- 
lus. T-shaped (in transverse section) pilaster 
running entire length of penial papilla and 
joined to it by a peduncle, so that cross sec- 
tion of papilla plus pilaster resembles card fig- 
ure spades. 

Right ommatophore retractor free of penis 
and vagina. 

Penial nerve apparently arising in the right 
cerebral ganglion (according to Franc, 1968: 
473, even if it comes from cerebral ganglion 
the penial nerve originates in the pedal gan- 
glion). 

Origin of the Name 

Helicotricha, gender feminine. 
The small helicoid shell with hairy perios- 
tracum suggested the name of the new genus. 

Type species 

Helicotricha carusoi n. sp. 

Helicotricha carusoi n. sp. 
[Figs. 1A-C, 2A-D, 3A-D, 4A-D, 5A-D, G, 
6A-C] 

Helicella (Xerotricha) conspurcata, — Giusti, 
1973: 259-260 [partim, non Draparnaud, 
1801]. 



A NEW HYGROMIIDAE FROM ITALY 



113 




FIG. 1. Shells of Helicotricha carusoi n. gen. n. sp. Holotype (A) and one paratype (B) from Alicudi Island: 
Perciato, F. G. leg. 24.10.69. A shell from Salina Island: Pollara, R. Arcidiacono leg. 21.9.66 (C). 



Helicopsis sp., — Piantelli et al., 1990: Table 5 
et passim. 

Diagnosis 

At present the only species of the genus 
Helicotricha known. Specific coincides with 
generic diagnosis. 

Description 

Shell (Figs. 1A-C, 2A-D): Shell small (Figs. 
1A-C), hairy, low conical above, convex be- 
low, opaque brown in colour, with white flecks. 



White flecks widely distributed over shell sur- 
face, concentrated above (near sutures and 
near periphery of last whorl) to form irregularly 
spaced spots of variable shape, below (from 
periphery of last whorl to umbilicus) to form 
white spiral bands of variable width and num- 
ber (2-6). Spire depressed-conical, consisting 
of 4-4V2 convex, regularly increasing whorls 
separated by deep sutures; last whorl angled 
at periphery, dilated, sometimes descending 
slightly near aperture. Umbilicus open and 
wide, approximately 1/5 of maximum shell di- 
ameter. Aperture markedly oblique, oval, lack- 



114 



GIUSTI, MANGANELLI & CRISCI 




FIG. 2. External shell surface of specimens of Helicotricha carusoin. gen. n. sp. collected on Panarea Island, 
D. Caruso & I. Marcellino leg. 27.5.67 (A-B) and Lipari Island: Monte Sant Angelo, F.G. leg. 23.10.69 (C-D). 
A: A view of first whorls. B: Detail of protoconch showing longitudinal grooves. C: Detail of last whorl with one 
hair and reticular microsculpture of teleoconch. D: Detail of reticular microsculpture of teleoconch. 



ing internal rib; peristome not thickened, 
slightly reflexed at its columellar margin. 

External surface of protoconch with few 
faint growth lines near its end (Fig. 2A) and 
microsculpture consisting of fine close spiral 
grooves (Fig. 2B). External surface of teleo- 
conch (Fig. 2A) with many growth lines, more 
marked near sutures. Periostracal layer (Fig. 



2C-D), superficially reticulated (Fig. 2D) to 
form transverse rows of very short, often 
hook-shaped hairs 0.1 mm in length (Fig. 2C). 
Reticulation and hairs of caducous appear- 
ance, absent in large portions of surface. Sur- 
face of mineralized portion of shell (in areas 
devoid of periostracal layer) seems to be 
crossed by fine close spiral grooves. 



A NEW HYGROMIIDAE FROM ITALY 



115 




FIG. 3. Genitalia of Helicotricha carusoi n. gen. n. sp. in specimens collected on Basiluzzo Islet, F. G. leg. 
5.1 1 .69. A-B: Two opposite views of the same genital apparatus (gonad excluded in A; gonad and part of 
ovispermiduct excluded in B). C: Part of penial complex with distal penis opened to show penial papilla, a 
section of proximal penis and two sections of penial papilla. D: Vagina opened to show its inner structure. 
Explanations of the symbols used in Figures 3-5, 8: ag, albumen gland; be, bursa copulatrix (gametolytic 
gland); dbc, duct of bursa copulatrix; dg, digitiform glands; dsc, dart-sac complex; dp, distal penis; e, 
epiphallus; f, flagellum; fc, fertilization chamber; fn, fenestration; fo, free oviduct; fr, frenula; g, penial papilla 
(glans); ga, genital atrium; gp, genital pore; gt, glandular tissue; gtp, gland of the terminal penis; hd, 
hermaphrodite duct; p, penis; po, prostatic portion of ovispermiduct; pp, proximal penis; pr, penial retractor 
muscle; pv, proximal vagina; pw, penial wall; s, stripes; sr, seminal receptacle; t, talon; tsp, t-shaped pilaster 
of the penial papilla; uo, uterine portion of ovispermiduct; v, vagina; vd, vas deferens. 



116 



GIUSTI, MANGANELLI & CRISCI 




FIG. 4. Genitalia of Helicotricha carusoin. gen. n. sp. in specimens collected on Filicudi Island: Stimpagnato, 
F. G. leg. 28.10.69. A: Genital apparatus (gonad and part of ovispermiduct excluded). B-C: Part of two penial 
complexes with distal penis opened to show penial papilla, section of epiphallus, proximal penis and penial 
papilla (B), Section of proximal penis and two sections of penial papilla (C). D: Vagina opened to show its 
inner structure. 



Dimensions (N = 10): shell max. diam.: 
4.5-5.4 mm; shell height: 2.5-3.4; aperture 
max. diam.: 2.0-2.5 mm; aperture height: 
1.7-2.1. 

Dimensions of the holotype: shell max. 



diam.: 5.3 mm; shell height: 2.8; aperture 
max.: diam. 2.1 mm; aperture height: 2.1 

Genitalia (Figs. 3A-D, 4A-D, 5A-D, G): Con- 
voluted first hermaphrodite duct arising from 



A NEW HYGROMIIDAE FROM ITALY 



117 



\J Üb 



4 h 




OG 1mm 



FIG. 5. Outline and scheme of dart-sac complex (darts omitted) (A-B), talon (C-D) and dart (G) of Helicot- 
richa carusoi n. gen. n. sp. in specimens collected on Filicudi Island: Stimpagnato, F. G. leg. 28.10.69, talon 
(E) of Microxeromagna armillata from Corsica: Olmeto, F.G. & G.M. leg. 1 .12.83 and talon (F) of Helicopsis 
striata from Öland, parish Persnäsa (Jordhamn, Sweden). 



118 



GIUSTI, MANGANELLI & CRISCI 




m&WmÊÈSF 



FIG. 6. Radula of Helicothcha carusoi n. gen. n. sp. 
in a specimen collected on Basiluzzo Islet, F. G. 
leg. 5.1 1 .69. A: Central tooth and first lateral teeth. 
B: 6th— 12th lateral teeth. C: Extreme marginal 
teeth. 



multilobate gonad (not illustrated) and ending 
in talon (i.e. fertilization chamber plus seminal 
receptacle complex); talon lying on surface of 
inner side of large albumen gland. Talon (Fig. 
5C-D) with wide lateral fertilization chamber 
embracing basal portion of seminal recepta- 
cle complex. Receptacle complex very slen- 
der, only slightly enlarged apically and con- 
taining 2-3 very small distinct sperm tubes. 
Ovispermiduct wide, plurilobate, consisting of 
prostatic and uterine portions. Prostatic por- 
tion continuing anteriorly into vas deferens 
and proximal portion of penis complex. Penis 
complex (Figs. 3A-B, 4A) consisting of penial 



flagellum, epiphallus (i.e. from end of vas def- 
erens to point of attachment of penial retrac- 
tor muscle) and penis (i.e. from point of at- 
tachment of penial retractor muscle to genital 
atrium). Penial flagellum long (4.9-5.5 mm; N 
= 3) and slender along entire length. Epiphal- 
lus longer (3.0-4.0 mm; N = 3) than penis 
(2.0-2.3 mm; N = 3) and narrower. Penis 
lacking distinct penial sheath but with thin 
bands of tissue arising where penial retractor 
muscle ends and terminating near penis end. 
Penis distally enlarged where it contains pe- 
nial papilla (= glans). Wide area yellow in 
colour and covered with glandular tissue (also 
evident internally) on external side of terminal 
portion of penis walls. Penis walls level with 
base of penial papilla sometimes appear to 
contain glandular tissue. Penial papilla (Figs. 
3C, 4B-C) cylindrical, consisting of a wide 
tube. Penial papilla lumen continuous with 
proximal penis and epiphallus. T-shaped (in 
transverse section) pilaster running alongside 
penial papilla for entire length, joined to it by a 
peduncle. Ejaculatory pore at apex of penial 
papilla, slit-like, variable in width. T-shaped 
pilaster basally connected to penis wall for 
short distance at penial papilla base (Figs. 
3C, 4C). Penial retractor muscle usually 
short. Uterine portion of ovispermiduct con- 
tinuing anteriorly into short (approximately 1 
mm; N = 3), wide uterine canal (i.e. free ovi- 
duct) ending in vagina level with point of entry 
of duct (4-6 mm in length; N = 3) of bursa 
copulatrix. Bursa copulatrix (i.e. gametolytic 
gland) (Figs. 3A-B, 4A) sac-like and variable 
in shape. Duct of bursa copulatrix 4-6 mm 
long (N = 3), its initial portion slightly flared, 
internally figured with series of fine pleats. 
Dart-sac complex consisting of two pairs of 
stylophores, each pair consisting of a smaller 
inner dartless stylophore and a larger outer 
dart-bearing stylophore. Opening of each pair 
of stylophores clearly visible half way along 
vagina on opposite sides (Figs. 3A-B, 4A, 
5A-B). Two pairs of digitiform glands also 
opening on opposite sides of proximal vagina 
close to dart-sac complex. Digitiform glands 
irregular in shape and sometimes apically bi- 
furcated. Each digitiform gland and dart-sac 
pair opens into the vagina within a furrow lat- 
erally bordered by two large, distally fused 
folds (Figs. 3D, 4D). Other smaller folds run 
longitudinally on the internal vagina walls be- 
tween two systems of larger folds. All folds 
taper distally to end on internal genital atrium 
walls. Dart of each outer stylophore very 
small (1 mm in length; N = 2), basally circular 



A NEW HYGROMIIDAE FROM ITALY 



119 



in transverse section, apically flattened-oval 
or rhombic, tip arrowhead-shaped (Fig. 5G). 

Other Anatomical Characters: Penial nerve 
apparently originating in right cerebral gan- 
glion (according to Franc, 1968: 473, even if it 
comes from cerebral ganglion the penial 
nerve originates in the pedal ganglion). 

Right ommatophore retractor independent 
of penis and vagina (i.e. not passing between 
the penis and vagina). 

Radula (Fig. 6A-C): Radula consisting of 
many rows of teeth each according to formula 
20-22 + С + 20-22. Central tooth with wide 
basal plate and raised pointed upper vertices. 
Body of tooth with very large mesocone and 
two small ectocones, 1/3 of mesocone length. 
First lateral teeth having wide basal plate but 
with inner vertex missing; body with large 
pointed mesocone and small pointed ectone 
1/4 of mesocone length. At about 6th or 7th 
lateral tooth, inner side of mesocone showing 
slight protuberance developing into pointed 
cusp in following lateral and marginal teeth. 
Moving laterally, teeth maintaining same 
shape but progressively smaller, with more 
pointed cusps and reduced basal plates. Last 
marginal teeth having mesocone apex with 
2-3 cusps and ectotone split into 2-4 smaller 
points. 

Jaw: Jaw odontognathous, strongly ribbed 
and devoid of central denticle. 



Type Locality 

Aeolian Islands, Alicudi Island: Perciato 
[UTM references: 33SVC4465]. 

Typical Series 

Holotype (shell) (Fig. 1A) and 7 paratypes 
(2 shells + 3 dissected specimens + 2 spirit 
specimens) from the type locality, F. G. leg. 
24.10.69. Holotype and all the paratypes in 
Giusti Collection, Dipartimento di Biología Ev- 
olutiva, Université degli Studi di Siena, Via 
Mattioli 4; 1-53100 Siena, Italy. Other material 
examined (all from Aeolian Islands [UTM réf.: 
33SVC, WC]) 

Alicudi Island [33SVC46]: Spano [[4467], F. 
G. leg. 26.10.69 (2 spirit specimens + 2 dis- 
sected specimens). 

Basiluzzo Islet [33SWC07]: S. Bruno leg. 



25.2.67 (7 spirit specimens), F. G. leg. 5.1 1 .69 
(1 spirit specimen + 1 dissected specimen); F. 
G. leg. 31 .3.71 (3 spirit specimens). 

Filicudi Island [33SVC66, 67]: Siccagni 
[6070], F. G. leg. 29.10.69 (1 spirit specimen); 
Stimpagnato [6168], F. G. leg. 28.10.69 (4 
spirit specimens + 2 dissected specimens); 
Between Canale and Monte Guardia [6268, 
6368], F. G. leg. 28.10.69 (3 spirit speci- 
mens); Zueco Grande [6270, 6370], F. G. leg. 
30.10.69 (2 spirit specimens + 1 shell), F. G. 
leg. 23.3.72 (2 shells). 

Lipari Island [33SVC95, 96]: G. Marcuzzi 
leg. 13.4.68 (1 shell + 1 spirit specimen); 
Capistello [9556, 9656], F. G. leg. 27.4.70 (1 
shell); Monte Sant'Angelo [9360, 9460], F. G. 
leg. 23.10.69 (5 shells). 

Lisca Bianca Islet [33SWC07]: F. G. leg. 

5.10.69 (7 spirit specimens) 

Panarea Island [33SWC07]: D. Caruso & I. 
Marcellino leg. 27.5.67 (5 shells), Punta del 
Corvo [0576], F. G. leg. 30.3.71 (1 shell + 1 
spirit specimen); Punta Milazzese [0575], F. 
G. leg. 5.11.69 (1 spirit specimen + 1 dis- 
sected specimen). 

Salina Island [33SVC86, 87]: Capo Faro 
[8870], F. G. leg. 25.4.70 (2 spirit specimens); 
Lingua [8865], F. G. leg. 25.4.70 (2 spirit 
specimens + 1 dissected specimen); Malta 
[8570], R. Arcidiacono leg. 17.9.66 (1 spirit 
specimen); Monte dei Porri [8368, 8468], F. 
G. leg. 26.4.70 (3 spirit specimens); Pollara 
[8369], R. Arcidiacono leg. 21.9.66 (1 shell), 
F. G. leg. 25.4.70 (1 spirit specimen); Rinella 
[8566], F. G. leg. 26.4.70 (1 shell); Valle del 
Santuario [8568], F. G. leg. 25.4.70 (1 spirit 
specimen). 

Stromboli Island [33SWC19, 20]: G. Mar- 
cuzzi leg. 16.4.68 (28 spirit specimens). 

Vulcano Island [33SVC95]: G. Marcuzzi 
leg. 12.4.68 (2 shells); Porto [9652], F. G. leg. 

27.4.70 (2 spirit specimens + 1 shell). 

All the material in Giusti Collection, Dipar- 
timento di Biología Evolutiva, Université degli 
Studi di Siena, Via Mattioli 4; 1-53100 Siena, 
Italy. 

Locality names and UTM references based 
on the Official Map of Italy 1 :25.000 Series M 
891. 



Origin of the Name 

The new species is dedicated to Prof. Do- 
menico Caruso, Director of the Department of 
Animal Biology, University of Catania, Italy, in 
token of highest esteem and companionship. 



120 



GIUSTI, MANGANELLI & CRISCI 



DISCUSSION 
Generic Status 

As outlined in the introduction, the su- 
praspecific status of the new species was dif- 
ficult to establish. Some anatomical features 
(2 + 2 stylophores and right ommatophore 
retractor independent of penis and vagina) 
suggested relationships with more than one 
genera of Hygromiidae found in the western 
Mediterranean and Europe: Xerotricha Mon- 
terosato, 1 892 (type species: Helix conspur- 
cata Draparnaud, 1801), Helicella Fèrussac, 
1821 (type species: Helix itala Linnaeus, 
1758) and Helicopsis Fitzinger, 1833 (type 
species: Helix striata Müller, 1774) (see Git- 
tenberger, 1969; Gittenberger & Manga, 
1977; Schileyko, 1978a; Gittenberger & 
Raven, 1982; Giusti & Manganelli, 1989; Git- 
tenberger et al., 1989; Hausdorf, 1988). 

Following the scheme of classical evolu- 
tionary systematics a first comparison was 
made with Xerotricha, the type species of 
which, X. conspurcata (Draparnaud, 1801), 
has a small hairy shell that differs from Heli- 
cotricha carusoi n. sp. only by virtue of its 
longer periostracal hairs and its narrower um- 
bilicus. 

Xerotricha conspurcata, like both its conge- 
neric species, X. apicina (Lamarck, 1822) and 
X. nubivaga (Mahille, 1882) (Hausdorf, 1988; 
Gittenberger et al., 1989, Giusti & Manganelli, 
1989), shows a differently structured vaginal 
complex characterized by a large dart-sac 
complex consisting of two pairs of stylo- 
phores, each constituted by a larger outer 
dart-bearing stylophore and a very small inner 
dartless stylophore (i.e. 2 + 2) (see Giusti & 
Manganelli, 1989: 51, for a discussion of ho- 
mology and terminology of these structures). 
The latter open into the vagina inside the slit 
delimited by two large tongue-like structures 
that are apically independent of each other 
(Hausdorf, 1988: fig. 9; Giusti & Manganelli, 
1989: figs. 3, 5, 9A-B). The penial papilla in 
Xerotricha is also different, being simple and 
without lateral pilaster (Giusti & Manganelli, 
1989: figs. 2D, 4E). 

Anatomically the new species is also simi- 
lar to some small Helicella with shells that 
have persistent postembryonic hairs (e.g. H. 
cordero! Gittenberger & Manga, 1977 — see 
Gittenberger & Manga, 1977; Manga Gonza- 
les, 1983; H. mangae Gittenberger & Raven, 
1982— see Gittenberger & Raven, 1982; ac- 
cording to Prieto, 1986, this nominal species 



is a junior synonym of H. gonzalei Azpeitia, 
1924). Nevertheless, Helicella has a dart-sac 
complex which, although similar (consisting of 
two opposite pairs of stylophores, each 
formed by a very large apically pointed outer 
stylophore and a very reduced, almost vesti- 
gial, inner dartless stylophore, i.e. 2 + 2), 
opens internally into a continuous pleated 
tube-like structure contained in the vagina 
(Hausdorf, 1988: fig. 8; Giusti & Manganelli, 
1989: figs. 6A,E, 7, 9C). In Helicella, the pe- 
nial papilla is also different, being simple and 
without lateral pilaster (Hausdorf, 1988: fig. 8; 
Giusti & Manganelli, 1989: fig. 6F). 

Detailed comparison was then made with 
Helicopsis 4 , the species of which have a very 
similar genital scheme (Hesse, 1934; Gitten- 
berger, 1969; Damjanov & Likharev, 1975; 
Schileyko, 1978a; Grossu, 1983; Giusti & 
Manganelli, 1989; present paper: Figs. 5F, 
8A-C). Helicopsis (s. str.) has so far been 
identified with certainty only in central and ori- 
ental Europe, and subgenus Xeroleuca Ko- 
belt, 1877 (type species: Helix turcica Holten, 
1802) has been reported in northwestern Af- 
rica (Hesse, 1934; Zilch, 1960). There are two 
records of Helicopsis (s. str.), one in Tunisia 
(Ktari & Rezig, 1976) and one at Huelva, 
Spain, (Gasull, 1972, 1985) that must be 
checked. The species briefly studied by Ktari 
& Rezig (1976) may correspond to a species 
found by one of us in Morocco (Figs. 7A, 8A- 
C). Helicopsis (s. str.) has a small, thick, ro- 
bust, ribbed and hairless shell and a vagina 
with a dart-sac complex constituted by two 
opposite pairs of stylophores, each formed by 
an outer and an inner stylophore (i.e. 2 + 2). 
The outer stylophores are smaller and more 
slender than those in Xerotricha and Helicella 
and are more clearly distinguished from the 
inner dartless stylophores which, in their turn, 
are larger than those in Xerotricha and Heli- 
cella. Moreover, the vagina in Helicopsis is 
clearly different from that in Xerotricha and 
Helicella, being internally devoid of tongue- 
like or tube-like structures into which the sty- 
lophores open (Giusti & Manganelli, 1989: 
figs. 8C, 9D). In view of the above and be- 
cause the new species and those of Helicop- 



4 While the paper was in press, Hausdorf (1990b) de- 
scribed the genitalia of three species-group taxa supposed 
to belong to genus Helicopsis: H. gittenbergeri n. sp., H. s. 
subcalcarata (Naegele, 1903) and H. subcalcarata neu- 
berti n. subsp. Comparison with the latter taxa has been 
omitted because no information about the penial papilla 
structure, the only sure diagnostic character for Helicopsis. 
was furnished. 



A NEW HYGROMIIDAE FROM ITALY 



121 




FIG. 7. A: Shell of Helicopsis sp. collected at the foot of Mount Zerboum, Moulay Idris (Morocco). B: Neotype 
of Helix aetnea Benoit, 1857, Nicolosi sull'Etna, С Caroti leg. 1877 (Museum of Zoology, University of 
Florence, Italy, MZUF no. 5049/1). 



122 



GIUSTI, MANGANELLI & CRISCI 




FIG. 8. Genitalia of Helicopsis sp. in specimens collected at the foot of Mount Zerboum, Moulay Idris 
(Morocco). A: Genital apparatus (gonad excluded). B: Vagina opened to show its inner structure. C: Part of 
the penial complex with the penis opened to show the penial papilla. 



sis (Schileyko, 1978a: fig. 237; Giusti & Man- 
ganelli, 1989: fig. 8C) have stylophores 
opening in a slit bordered by two large but 
simple pleats running along the vagina walls, 
it seems plausible to include the new species 
in the genus Helicopsis. Nevertheless, the 
structure of the penial complex in the new 
species differs from that in Helicopsis. The 



species of the latter genus, about which there 
is detailed knowledge of genital structure, 
have a penial papilla consisting of a central 
tube wrapped in an external sheath that is 
more or less laterally and basally fenestrated 
and which basally expands to reach and to 
fuse with the penial walls (Schileyko, 1978a: 
figs. 235, 238-242, 244; Giusti & Manganelli, 



A NEW HYGROMIIDAE FROM ITALY 



123 



1989: figs. 8D, F-H; present paper: Fig. 8C). 
In so doing, the sheath distinguishes the cav- 
ity of the distal penis (containing the penial 
papilla) from that of the proximal penis, al- 
though it does not impede communication 
through its fenestrations. The cavity of the 
proximal penis is traversed by a tube that has 
sometimes a lateral pilaster connected by 
frenula to the penis walls and which is basally 
continuous with the epiphallus and apically 
with the central tube of the penial papilla. This 
peculiar penis structure, present not only in 
the European species but also in the Magh- 
rebian species studied herein, can be consid- 
ered diagnostic for Helicopsis. Consequently, 
the present new species cannot be included 
in the genus Helicopsis. 

One can argue that the penis structure in 
the present new species may be derived from 
that in Helicopsis through the reduction of the 
penial papilla sheath (the lateral T-section pi- 
laster may be a residue of the sheath) and the 
loss the peculiar inner structure of the proxi- 
mal penis. However this hypothesis seems 
less probable than the following one. 

Helicotricha carusoi n. sp. has a shell clearly 
resembling in overall structure, microsculpture 
and colour that of a well-known Mediterranean 
species: Microxeromagna armillata (Lowe, 
1852) (Giusti & Manganelli, 1989: pi. 7, figs. 
A-E). The latter species, moreover, has a gen- 
ital structure corresponding to that of H. caru- 
soi n. sp. (similar talon, similar length and pro- 
portions of the parts of the penial complex, a 
yellow glandular area on distal penis walls; see 
Ortiz de Zarate Y Lopez, 1 950: fig. 22; Forcart, 
1976: fig. 3; Clerx & Gittenberger, 1977: figs. 
1 02-1 03; Falkner, 1 981 : fig. 2; Aparicio, 1 982: 
fig. 3; Manga Gonzales, 1983: fig. 12; Haus- 
dorf, 1988: fig. 13; Manganelli & Giusti, 1988: 
figs. 11A-F, 14H; present paper: Fig. 5E). It 
only differs in the dart-sac complex, which has 
+ 2 stylophores (instead of 2 + 2), and in 
some details of the penial papilla. Other mono- 
phyletic groups have been hypothesized to be 
formed by genera with 2 + 2 and + 2 sty- 
lophores (i.e. Helicella-Candidula and Xero- 
lenta-Xeromunda; Hausdorf, 1988, 1990a, 
Giusti & Manganelli, 1989; Manganelli & 
Giusti, 1989). Similarly, we hypothesize that 
Helicotricha n. gen. forms a monophyletic 
group with Microxeromagna. The former ge- 
nus can have originated the latter by reduction 
of the dart-sac complex; the origin of Helicot- 
richa from Microxeromagna by duplication of 
the dart-sac complex seems less probable ac- 
cording to Schileyko's (1978a, 1984) recon- 



struction of the phylogenetic relationships in 
the Hygromiidae. 

Many other genera of the European and 
Russian Hygromiidae resemble Helicotricha 
n. gen. in having the vaginal complex with 
digitiform glands and dart-sac complex con- 
stituted by two opposite pairs of stylophores, 
each formed by a large, dart-bearing outer 
stylophore and a smaller dartless inner stylo- 
phore (i.e. 2 + 2): Hygrohelicopsis Schileyko, 
1977 (type species: H. darevskii Schileyko, 
1977), Leucozonella Lindholm, 1927 (type 
species: Helix rubens von Martens, 1874), 
Kokotschashvilia Hudec & Lezhawa, 1969 
(type species: Helix holotricha Boettger, 
1874), Caucasigena Lindholm, 1927 (type 
species: Helix eichwaldi Pfeiffer, 1846), Pli- 
cuteria Schileyko, 1977 (type species: Helix 
lubomirskii Slosarski, 1881), Trichia Hart- 
mann, 1840 (type species: Helix hispida Lin- 
naeus, 1758), and Edentiella, Polinski, 1929 
(type species: Helix edentula Draparnaud, 
1801) (see Schileyko, 1978a, 1978b). Of 
these, the genus closest to Helicotricha n. gen. 
is Hygrohelicopsis because it shows the right 
ommatophore retractor independent of penis 
and vagina. Nevertheless, Hygrohelicopsis 
can be easily distinguished by the structure of 
distal genitalia showing inner stylophores ex- 
tremely reduced, not visible from outside, dis- 
tal vagina absent, and penial papilla bulbous 
but simple without external sheath or lateral 
pilaster. All the other genera are more distin- 
guished from Helicotricha n. gen. because 
they have the right ommatophore retractor 
passing between penis and vagina, different 
and larger shells, different penial papillae and 
many other minor differences in the structure 
of the genitalia (Schileyko, 1978a, 1978b). 

Cladistic Analysis 

The entire set of character states (Table 2) 
utilized in the traditional approach was used 
for cladistic analysis. The latter was at- 
tempted even if the characters were few and 
limited to genitalia structure. To avoid compli- 
cating the analysis, only a limited number of 
eastern European genera was considered: 
those for which better description exist and 
which are at least apparently more closely re- 
lated to the new genus. 

A total of 105 most parsimonious hypothe- 
ses were generated by our data matrix. All 
have 37 steps with a consistency index of 
0.67 and rescaled retention index of 0.72 after 
non-informative characters were excluded. 



124 



GIUSTI, MANGANELLI & CRISCI 





OUT 


CAUC 
EDEN 
HELP 
KOKO 
LEUC 
PLIC 
TRIC 


HVfin 












































£1 X V3fl 




HYGR 


























K* JCilvTl 


CAND 

HELL 
PXER 






































XERM 
















XERL 
NANA 






















XERT 
HELT 



































MICR 
XERS 



FIG. 9. The Nelsen consensus tree of 105 cladograms. 



The successive weighting procedure did not 
discriminate among them, the main difference 
being the position of the set of taxa HELP- 
LEUC. The Nelsen consensus tree of 105 cla- 
dograms (Fig. 9) showed that 1 1 monophyl- 
etic groups appear in all of them, listed with 
their synapomorphies as follows: 



(1) 



(2) 



(3) 



(4) 



(5) 

(6) 
(7) 



All the taxa, except HELP, TRIC, 

KOKO, PLIC, CAUC, EDEN, LEUC: 2 

(1) (parallel with HELP and a reversion 

in HYGM). 

All the taxa, except HELP, TRIC, 

KOKO, PLIC, CAUC, EDEN, LEUC, 

HYGH: 15(6). 

HYGM, CERN: 3 (1), 9 (1), 10 (4) (the 

first parallel with CAND, XERM, MICR, 

XERS). 

CAND, HELL, PXER, XERL, XERM, 

XERT, NANA, HELT, MICR, XERS: 8 

(1) (parallel with HELP, TRIC). 

CAND, HELL: 1 (1), 5 (4), 10 (3) (the 

first parallel with CERN). 

PXER, XERL, XERM: 7(1). 

XERL, XERM: 5 (2). 



(8) XERT, NANA, HELT, MICR, XERS: 13 
(1) (with a reversion in XERS). 

(9) XERT, NANA: 5(1), 10(2). 

(10) HELT, MICR, XERS: 4 (0), 15 (2) (the 
former is a reversion). 

(11) MICR, XERS: 3 (1) (parallel with 
HYGM, CERN, CAND, XERM). 



Character 6 represented a synapomorphy 
for the entire set of terminal taxa with a rever- 
sion in MICR. Figure 10 is one of the 105 
minimum-length trees. It is not a preferred 
tree and is simply given to illustrate the evo- 
lution of the characters. 

As is evident, the main result of cladistic 
study has been that of apparently supporting 
the conclusion reached in the preceeding 
paragraph, i.e. that Helicotricha is a sister 
group of Microxeromagna-Xerosecta. It also 
appeared to confirm that Helicotricha is not 
closely related to Helicopsis as hypothesized 
in the same paragraph. 

Relationships with Xerosecta suggested by 
cladogram seem logical: Xerosecta is very 



A NEW HYGROMIIDAE FROM ITALY 



125 





OUT 




КЗ 




II 


з 


II 






00 




4 






— 




I 










о 










I 





HELP 

TRIC 
KOKO 

PLIC 



CAUC 

EDEN 
LEUC 



HYGH 



««5 



HYGM 



CERN 



~ Ü о 



CAND 

HELL 
PXER 






XERM 

XERL 
NANA 

XERT 
HELT 



MICR 



XERS 



FIG. 10. One of the most parsimonious trees generated by the data matrix given by way of illustration of the 
evolution of the characters (lines: synapomorphies; double lines: homoplasies; X reversions). 



126 



GIUSTI, MANGANELLI & CRISCI 



close to Microxeromagna (Hausdorf, 1988, 
1990c; Manganelli & Giusti, 1988). 

The Suprageneric Systematics of 
the Hygromiidae 

The phylogenetic hypothesis concerning 
Helicotricha n. gen. formulated above, in ad- 
dition to those concerning Xerolenta- 
Xeromunda and Helicella-Candidula, dis- 
agrees with Schileyko's proposal to 
distinguish the Trichiinae (hygromiids with 2 
+ 2 stylophores) from the Hygromiinae (hy- 
gromiids with + 2 stylophores). 

The present data supports the idea that the 
supposed members of one of these "subfam- 
ilies" independently evolved from supposed 
members of the other so that their derived 
status is due to parallelism (Giusti & Manga- 
nelli, 1987). 

The consideration of a larger number of 
characters, when eventually available (ge- 
netic, cytological, etc.) could promote better 
understanding of the phylogeny of Hygromi- 
dae (and Helicoidea in general) and verify the 
contention of Giusti et al. (1991) who, on the 
basis of spermatozoa fine morphology, sug- 
gested there are too many family group cat- 
egories in the Helicoidea and that it is inop- 
portune to produce new schemes and create 
new taxa on the basis of old and insufficient 
anatomical (or even shell!) characters. 

Comparison with Old and Uncertain Taxa of 
the Species Group 

A search of the taxa described for Sicilian 
fauna did not produce positive results. Apart 
from Xerotricha conspurcata, X. apicina and 
the two species recently revised by us and 
recognized to belong to a distinct genus — 
Schileykiella: S. reinae Pfeiffer, 1 856, and S. 
parlatoris, Bivona, 1839 (Manganelli et al., 
1989) — only one small hygromiid with a hairy 
shell, Helix aetnea Benoit, 1857, is known in 
Sicily. 

Some shells were found in the Paulucci 
Collection which, according to the label, were 
collected in the type locality of H. aetnea, 
studied by Benoit himself, and confirmed by 
him to fully correspond to his H. aetnea (Mu- 
seum of Zoology, University of Florence, Italy, 
MZUF no. 5049). Our study of the minute 
shell characters confirms Paulucci's (1878: 6, 
32) and Westerlund's (1889: 302) identifica- 
tion of H. aetnea as a juvenile of X. conspur- 
cata. Because Benoit's typical series is un- 



traceable and there is no other possible 
typical material in the principal malacological 
collections, and because Benoit clearly con- 
firmed that Paulucci's material totally corre- 
sponded to his species, we have selected a 
neotype for this species (MZUF no. 5049/1) 
illustrated in Figure 7B. Helix aetnea Benoit, 
1857, consequently becomes a junior syn- 
onym of X. conspurcata. 

Geographic Distribution 

All the Aeolian Islands. A few shells from 
the island of Ustica (NW of Sicily) [UTM réf.: 
33SUC38, 48] may belong to the new spe- 
cies, but since they are sexually immature 
and anatomical study is impossible, no con- 
clusions can be drawn. Although its presence 
in Sicily is highly probable, a search of the 
literature and of our materials from Sicily, and 
some Sicilian and Maltese Islands did not 
bring to light any useful information. 

Ecology 

Helicotricha carusoi n. sp. has been found 
to live under stones, dry leaves and pieces of 
wood in many different places on single is- 
lands, frequently together with Xerotricha 
conspurcata. Like the latter species, the 
present species is thus a xeroresistant ele- 
ment, well adapted to Mediterranean habitats. 



ACKNOWLEDGMENTS 

We thank Mr. L. Gamberucci and Mrs. A. 
Daviddi for technical assistance and Mrs. H. 
Ampt for linguistic revision. This research was 
supported by CNR, MPI 40% and MPI 60% 
grants, and was carried out as part of a 
project of the Escuela de Specialization en 
Ambiente y Patología Ambientale of the Uni- 
versidad Nacional de La Plata, Argentina, and 
Université di Siena, Italy. 

Thanks also to two anonymous revisers for 
the detailed analysis of manuscript which im- 
proved the quality of the paper. 



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A NEW HYGROMIIDAE FROM ITALY 



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MALACOLOGIA, 1992, 34(1-2): 129-141 

STRUCTURE AND COMPOSITION OF THE SHELL OF THE 
ARCHAEOGASTROPOD LIMPET LEPETODRILUS ELEVATUS ELEVATUS 

(MCLEAN, 1988) 

Stephen Hunt 

Department of Biological Sciences, University of Lancaster, Bailrigg, Lancaster, 

LA1 4YQ England 

ABSTRACT 

Shells from the hydrothermal vent-dwelling archaeogastropod limpet Lepetodrilus elevatus 
elevatus are described in terms of their mineralogy, fine structure and composition. The shell is 
mainly crossed-lamellar aragonite, with narrow inner and outer prismatic layers and no nacre. 
The thick proteinaceous periostracum has an unusually high glycine content. Sulphur, detected 
in the periostracum by X-ray microprobe analysis, cannot be attributed to either sulphur-con- 
taining amino acids or free elemental sulphur. Periostracum and shell both contain phosphorus, 
but metals other than calcium are present in traces only. Most shells examined show erosion in 
the form of pits rich in sulphur and phosphorus. The significance of the unusual periostracal 
composition in relation to the shell mineralogy, the propensity for shell erosion, and the chem- 
istry of the vent water environment, is discussed. 

Key words: archaeogastropod, Lepetodrilus, shell, calcification, aragonite, cross-lamellar, pe- 
riostracum, polyglycine. 



INTRODUCTION 

Exploration of deep ocean hydrothermal 
sites discovered in the 1980s revealed an un- 
expected rich fauna with molluscs well repre- 
sented. While large bivalves attracted much 
attention, gastropods are most numerous, 
mainly as small limpet forms, new and unique 
to this special environment. Hickman (1983) 
noted the presence in the Galapagos Rift and 
East Pacific Rise (13°N and 21 °N) sites of 
three new archaeogastropod groups that 
could not be assigned to then-recognised su- 
perfamilies. These have been described by 
McLean (1985, 1988) and by Fretter (1988). 
Here I describe the architecture and compo- 
sition of the shell of Lepetodrilus elevatus el- 
evatus, a member of the archaeogastropod 
superfamily Lepetodrilacea (McLean, 1988). 



MATERIALS AND METHODS 

Specimens of L. elevatus elevatus (see 
McLean, 1988, for the systematic description 
and list of the principal repositories of the type 
material) were collected by Dr. F. Gaill of the 
Centre du Biologie Cellulaire, CNRS, Ivry sur 
Seine, France, at 2600 m depth during the 
1984 Biocyarise cruise using the submersible 
Cyana at 12°48'N, 103°56'W. 



Scanning electron microscopy was carried 
out on shells that had been rinsed with dis- 
tilled water, air dried, mounted with graphite 
paste cement onto aluminium support stubs, 
and sputter coated with gold. Microscopy was 
performed with a Jeol JSM50A scanning elec- 
tron microscope at 15kv. For microprobe 
analysis, specimens were mounted on graph- 
ite support stubs and were rotary coated with 
carbon. Analyses were carried out at 1 5kv ex- 
citation voltage by a Kevex energy dispersive 
analysis system at 20ev per channel gain with 
a preset live time of 100 sec. 

For thin sectioning, periostracum was ob- 
tained by shell dissolution with 0.1 M hydro- 
chloric acid, washed in distilled water, fixed 
overnight in pH 7.0, 0.1 M cacodylate buffer 
containing 5% glutaraldehyde, and post-fixed 
in cacodylate-2% osmium tetroxide for 30 
min. Following dehydration through alcohols, 
fixed material was embedded in Spurr's resin 
and sectioned at 50 nm or less using a dia- 
mond knife. Sections were stained with uranyl 
acetate and Reynold's lead citrate for 30-60 
min each. Sections were examined at 100kv 
in a Jeol 100CX STEM microscope. 

X-ray diffraction patterns were determined 
using finely ground specimens of whole shell 
held in glass capillaries mounted in a helium- 
filled flat camera of working distance 3.5 cm 
with Ni-filtered Cu/Ka radiation and pinhole 



129 



130 



HUNT 





FIG. 1. Lepetodrilus elevatus elevatus. A. Lateral 
view. B. Dorsal view. С Ventral view. 



FIG 2. Inner surface of the shell (i) showing the 
folding over of the periostracum (p) at the shell 
edge and the granular structure of the shell with 
large and small scale irregularities (li and si) ar- 
rowed. A. Low magnification. B. Same region at a 
higher power. 



about 4 mm by 3 mm (Fig. 1 A-C). The exter- 
nal surface has low concentric, closely 
spaced growth ridges and a smoother apex. 
An olive-brown periostracum folds over the lip 
of the shell with a 0.2-0.3 mm inner overlap 
(Figs. 1С, 2A). The inner surface of the shell 
is smooth though higher magnifications re- 
veal regularly distributed granules and shal- 
low pitted irregularities (Fig. 2A, B). 



collimation. Fibre diffraction patterns were 
made using small strips of wet material 
clamped in holders and slightly tensioned be- 
fore drying. Standard patterns were made 
with powdered calcite (Iceland spar) and ara- 
gonite (scleractinian exoskeleton). 

Amino acid analyses were carried out on 
specimens hydrolysed in evacuated tubes 
with 3M methane sulphonic acid containing 
0.2% w/v tryptamine at 100°C for 48 hours. 
Neutralized aliquots were analysed using an 
LKB model 4101 amino acid analyser. 



RESULTS 

General Description of the Shells 

Shells of L elevatus elevatus are typically 
about 2.25 mm high, with an oval aperture 



Shell Mineralogy 

Figure 3 shows the diffraction pattern of 
powdered shell. The principal reflections are 
aragonitic, with little or no calcitic contribution. 

Ultrastructure of the Mineral Phase 

Fractured edges of the shell show three 
major shell layers — outer, middle and inner, 
of which the middle is the major and com- 
posed quite uniformly of the crossed-lamellar 
structure type (Figs. 4-6). Between this layer 
and the periostracum is a columnar layer of 
irregular fibrous prismatic organization (Fig. 
7). The innermost shell layer is columnar in 
irregular simple prismatic form (Fig. 8). Here a 
thin zone, no more than 2 |лт deep and 
formed of either short prisms or granules, de- 
marcates this layer from the crossed lamellar. 



LEPETODRILUS SHELL STRUCTURE AND COMPOSITION 



131 





FIG 3. Powder diffraction patterns of A. Aragonite. 
B, Lepetodrilus shell. C. Calcite. 



There is no evidence of an innermost nacre- 
ous layer being present. 

Elemental Analysis 

X-ray microprobe analyses of the main min- 
eralized layers show calcium to be the only 
detectable element present in significant 
amounts (Fig. 9, Table 1), presumably as the 
carbonate. The traces of sulphur and phos- 
phorous would be unlikely to make an impor- 
tant contribution to shell mineralogy even if 
they were present as calcium phosphate and 
sulphate. Although vent waters are transition 
and heavy metal rich, none are detectable in 
the shell mineral. Traces of magnesium, sili- 
con and aluminium could indicate chance in- 
corporation clay-forming suspensates. Roux 
et al. (1985) have noted that concentrations of 
trace elements in the aragonitic shell of the 
hydrothermal vent bivalve Calyptogena did 
not reflect vent-water trace element concen- 
trations but were similar to those found in ara- 
gonitic shell of other marine bivalves. 



Periostracum 

Figure 10 is a low-power transmission elec- 
tron micrograph of sectioned periostracum. 
No laminated, orthogonal or helicoidal fibrous 
architecture of the type found in some marine 
and terrestrial prosobranch gastropod perios- 
traca (Bevelander & Nakahara, 1970; Hunt & 
Oates, 1970, 1978, 1985) was found. The 
faint bands in the plane of the periostracum 
are probably depositional irregularities with- 
out compositional or orientational signifi- 
cance. The darker and more homogeneous 
zone adjacent to the shell may represent a 
phase of rapid deposition and tanning. The 
inner surface has an irregular profile and 
forms a thin, more electron-luscent layer than 
the main body of periostracal protein into 
which it merges via a narrow, more densely 
staining zone. The outer surface is also irreg- 
ular. What may be knobs or ridges adhere to 
a thick layer of vacuolated granular material 
that may be a deposit of microorganisms and 
detritus. The remnants of bacteria are present 
in the vacuoles. 

Table 2 gives the amino acid analysis of the 
periostracum, and Table 3 lists values for key 
features of the composition enabling compar- 
isons with other structural proteins. The peri- 
ostracum has a high glycine content, over 70 
residue percent. Glycine-rich proteins are 
common among structural proteins, and 
whereas many periostracal proteins are of 
this type, it is unusual for those of marine gas- 
tropods to be glycine-rich. Figures 11 and 12 
show the composition of this periostracum 
placed in relation to other gastropod periost- 
raca. Its glycine content is higher than in any 
other gastropod (62 species) recorded by us. 
This, coupled with its hydrophobic index 
( + 47), ranks it with those of terrestrial proso- 
branchs. The only periostraca with higher 
glycine levels (over 80 residue percent) are 
from bivalve molluscs at hydrothermal vent 
sites. But, unlike the latter, the sulphur-con- 
taining amino acid levels here are low or ab- 
sent (Hunt, 1987). The high beta turn poten- 
tial value, coupled with low values for helix 
and beta sheet potential, point to a protein 
secondary structure high in random folding. 

Figure 13A shows the typical X-ray emis- 
sion profile of the periostracum probed to 
avoid surface deposits. Noteworthy is the 
large sulphur peak. There are traces of iron 
and copper and a little phosphorus (Table 4). 
In the absence of calcium, the latter cannot be 
accounted for as hydroxyapatite, though this 



132 



HUNT 




FIG. 4. Fractured edge of shell with the dorsal surface at the top and showing the periostracum (p) and the 

three main calcified layers, outer (o), middle (m) and inner (i). Cross-lamellation (cl) is evident in the deeper 

middle layer. 

FIGS. 5, 6. Two views of the crossed-lamellar organization of the middle layer of the shell. 

FIG. 7. The outer layer of the shell with the periostracum (p) overlying an irregular layer of aragonite 

columns. 

FIG. 8. The innermost layer of the shell composed of irregular columns sharply separated from the overlying 

cross-lamellar material by a narrow zone of short prisms or granules. 



is present in some bivalve periostraca 
(Waller, 1983). The absence of sulphur-con- 
taining amino acids demands another expla- 
nation of the high sulphur content. Many mi- 
croorganisms present at the hydrothermal 
vents have an energy metabolism based on 
oxidation of sulphur compounds. Deposition 



of elemental sulphur as part of this process is 
common and occurs in the gills of Calypto- 
gena as part of a symbiotic relationship (Fel- 
beck et al., 1985). Extraction, however, of the 
periostracum with carbon disulphide only re- 
moves traces of elemental sulphur (Fig. 
13B.C, Table 4). 



LEPETODRILUS SHELL STRUCTURE AND COMPOSITION 



133 



Ca 



м ч1аИ 



Ca 




FIG. 9. Analysis of the shell by X-ray microprobe. 
For this analysis, the beam was centred in the 
crossed-lamellar layer, but closely similar patterns 
were obtained for the outer and inner calcified lay- 
ers. The analysis is energy dispersive, and the ab- 
scissal axis is in terms of X-ray energy and is a 
measure of increasing atomic number, whereas the 
ordinate axis is a measure of received X-ray photon 
counts at the detector. 



X-ray diffraction patterns prepared from 
flattened strips of periostracum show no ori- 
ented fibre reflections. There is considerable 
diffuse scatter and a weak ring corresponding 
to a 0.387 nm spacing, which tends to confirm 
the secondary structure as being a compact 
and highly folded random coil. The helical 
coiled polyglycines I and II are absent. 

Shell Erosion 

All specimens examined had erosion pits, 
most commonly near the shell apex. Many of 
these pits penetrate the shell and are usually 
partially filled or closed by brown or black ma- 
terial. Figure 14 shows several such pits with 
characteristic rounded profiles around single 
or multiple coalescing sites of penetration. 
The pitting is often associated with areas of 
periostracal cracking that follow the pit profile 
(Fig. 15). 

Pits penetrate the shell in a series of ter- 



raced steps. In several, the first layer re- 
vealed beneath the periostracum is more 
finely granular than the deeper terraces or the 
coarser pit-bottom material (Figs. 16, 17). 

Figure 18 and Table 5 show X-ray micro- 
probe analyses of eroded and pitted areas. A 
characteristic feature are the high levels of 
sulphur and phosphorus in the pits and, as 
might be expected, increasingly high calcium 
counts from lower within them. 



DISCUSSION 

Leptodrilus elevatus elevatus is found in 
huge numbers in close association with the 
giant vestimentiferan tube worm Riftia at 
depths of around 2600 m, close to the hydro- 
thermal vents, where the sea water has high 
concentrations of such reduced gases as hy- 
drogen sulphide, together with dissolved sul- 
phides of iron, manganese, zinc and copper 
and low oxygen tension. Many of the animals 
living there take advantage of chemo- 
lithotrophic bacteria that base their metabo- 
lism on oxidation of hydrogen sulphide and 
other reduced forms of sulphur. Endosymbi- 
otic relationships exist between vent bacteria 
and bivalve molluscs (Felbeck et al., 1985), in 
which hydrogen sulphide, concentrated from 
the sea water by the mollusc, is transferred to 
bacteria in the gills and used as the energy 
source for synthesis of organic compounds 
used by the animal. De Burgh & Singla (1984) 
describe how a vent archaeogastropod limpet 
also obtains nutrients by endocytosis of bac- 
teria growing upon its gills. 

The need to absorb oxygen at low concen- 
trations from waters containing high metal 
concentrations, as well as nutritional strate- 
gies involving consumption of bacteria, which 
may have absorbed metallic elements in their 
cell coats (Juniper et al., 1986), might lead to 
expectation that mineralization would yield 
shells reflecting the sea water chemistry, but 
this seems not to be so. Although traces of 
magnesium, aluminium and silicon are nearly 
always present in the shells, none of the 



TABLE 1 . Elemental composition of the main mineralized layer of the Lepetodrilus shell determined by 
X-ray microprobe analysis. Values are as emitted X-ray counts for each integrated elemental peak as a 
percentage of total counts integrated across the whole energy spectrum. For conditions, see text. These 
values relate to Figure 9, on which the traces of magnesium and phosphorus barely show. 



Element 
counts %total 



Na 
1.11 



Mg 
0.54 



AI 
3.67 



Si 
1.19 



P 
0.48 



S 
4.75 



CI 
1.28 



Ca 
86.99 



134 



HUNT 




FIG. 10. Transmission electron micrograph of a transverse section through the periostracum (p). The outer 
surface with bacterial debris contamination (be) is at the left and the inner shell contact surface (if) at the 
right. The specimen has been decalcified. 



TABLE 2. Amino acid composition of the periostracum of L. elevatus elevatus reported as residues of 
each amino acid per thousand total amino acid residues. 



Aspartic acid 


17 


Alanine 


10 


Tyrosine 30 


Threonine 


6 


Cysteine 





Phenylalanine 12 


Serine 


8 


Valine 


34 


Histidine 14 


Glutamic acid 


11 


Methionine 


2 


Lysine 41 


Proline 


8 


Isoleucine 


23 


Arginine 22 


Glycine 


713 


Leucine 


49 


Tryptophan — Not determined 



heavier metallic elements appear to be in- 
cluded during calcification. The only elements 
achieving any prominence against the domi- 
nant calcium component are phosphorus and 
sulphur. 

Traces of phosphorous in the shells might 
be accounted for by phosphoprotein in the 
matrix. Petit et al. (1980) have found calcium 
"spherites" containing phosphorus in the vas- 
cular channels and connective tissue imme- 
diately adjacent to mantle edge cells of the 
freshwater bivalve Amblema, implicating 
these in shell formation. Marsh & Sass (1983) 
have demonstrated calcium-binding phos- 
phoprotein particles in the extrapalial fluid 
and innermost shell lamella of estuarine and 
marine bivalves. 

Shell sulphur could be due either to traces 



of calcium sulphate or perhaps sulphated 
polysaccharides. Nautilus shell, for example, 
contains polysaccharide sulphates in the 
membrane overlying the nacreous crystals 
(Crenshaw & Ristedt, 1976). Crassostrea has 
sulphated proteins in the matrix (Wilbur & 
Salleuddin, 1983). Organic ester sulphate 
could also account for the high sulphur con- 
tent of the periostracum. 

Aragonite appears to be the predominant 
form of crystaline calcium carbonate in the 
shell. There are no precedents that would in- 
dicate if very high hydrostatic pressures can 
affect shell mineralization in molluscs, though 
in vitro work shows that calcium carbonate 
crystalization equilibria are pressure sensi- 
tive — aragonite becomes the most stable form 
at higher pressure and temperature (Mellor, 



LEPETODRILUS SHELL STRUCTURE AND COMPOSITION 



135 



TABLE 3. Side-chain characteristics and conformational predisposing properties of the amino acid 
composition of the Lepetodrilus and other molluscan periostracal proteins calculated from the data of 
Table 2. 







Av. other 


Av. 


Side Chain 


Lepetodrilus 


marine gastropods 9 


marine bivalves 10 


Acidic 1 


28 


237 


60 


Basic 2 


77 


103 


71 


Polar 3 


149 


501 


264 


Apolar 4 


136 


300 


189 


Polar/Apolar 


1.1 


1.6 


1.4 


Hydrophobic index 5 


+ 47 


-226 


+ 235 


Helix potential 6 


215 


513 


270 


Sheet potential 7 


229 


428 


352 


Turn potential 8 


746 


420 


620 



a Asp + Glu. 2 His + Lys + Arg. 3 Acidic + Basic + Thr + Ser + Туг. 4 Pro + Ala + Val + He + Leu + Phe. Calculated 
according to the method of Andersen (1979) by ranking each residue according to the characteristic energy change involved 
in transfer of it between water and ethanol. Hydrophilic residues have negative values and hydrophobic positive. Summation 
of the product of the i m amino acid content and AG for the i, h transfer, for all amino acids, gives the hydrophobic index. 
Calculated from data in Chou & Fasman (1977) using the conformational frequency parameters P as weighting factors to 
load the overall numbers of helix-promoting amino acids in the periostracal proteins. Thus, "helix potential" is given by the 
number of residues (per 1000 total) of an amino acid multiplied by its Pa value and summed for the residues Glu, Met, Ala, 
Leu, Lys, Phe, He and Val (in decreasing order of helix-promoting ability). Typical values for a "random coil" protein (resilin), 
a nearly 100% a-helical protein (tropomyosin), and a globular protein with low helix (2%) and high beta sheet (57%) content 
(concanavalin A) are 350, 956 and 561, respectively (calculated from amino acid compositions in residues per 1000 total 
residues) (Andersen & Weis-Fogh, 1964; Huddart & Hunt, 1975; Wang et al., 1971 ; Reeke et al., 1975). 7 As for 6 , but using 
the beta sheet-promoting conformational parameters P ß to weight and sum the contents of Val, He, Tyr, Phe, Leu, Cys, Thr 
and Met (Chou & Fasman, 1977). Values for resilin, tropomyosin and concanavalin A are 226, 344, and 532, respectively. 
8 Asp + Ser + Pro + Gly. These residues show high frequency in the four amino acids at a beta turn (Chou & Fasman, 
1977). Values for resilin, tropomyosin and concanavalin A are 636, 174 and 388, respectively. 9 38 non-hydrothermal 
species (Hunt, 1987). 10 29 non-hydrothermal species (Hunt, 1987). 



1960). Probably the chemistry of the organic 
matrix is more important. Commenting upon 
the entirely aragonitic form of the shell in the 
hydrothermal vesicomyid С magnifica and 
the mixed aragonitic and calcitic form of a hy- 
drothermal mytilid, Lutz (1982) observes that 
in the eastern Pacific regions, the depths of 
2000-3000 m, where sampling has taken 
place, are well above reported calcite com- 
pensation depths but substantially below 
those for aragonite compensation. 

Because the limpet is in a chemically hos- 
tile niche, the aragonitic mineralogy might 
seem to be a disadvantage being less stable 
and slightly more soluble than the calcitic. 
The unreliability of aragonite is seen where 
the periostracum has been damaged and 
rapid dissolution begins to penetrate the shell. 

Clearly the periostracum offers much pro- 
tection for the shell and probably because its 
composition is atypical of marine gastropods. 
Hunt & Oates (1985) note that there are prob- 
ably two main families of periostracal proteins 
in gastropods — those with a low glycine con- 
tent and negative hydrophobic index, and 



those with a medium to high glycine content 
and positive hydrophobic index. The latter is 
typical also of most marine and freshwater 
lamellibranchs. The former have easily wet- 
ted periostraca with high permeabilities to wa- 
ter and ions; the latter have the opposite char- 
acteristics. I have commented elsewhere 
(Hunt, 1987) on the resistance of shells of the 
latter type to attack by 2M hydrochloric acid 
through an intact periostracum; Lepetodrillus 
behaves in this way, showing only periostra- 
cal blistering, carbon dioxide evolution, and 
shell erosion where the periostracum is al- 
ready damaged. 

Marine prosobranch periostraca, for which 
we have analyses available, do fall into both 
groups, but the hydrophilic class are more 
common. Assuming that the primary role of 
periostraca is in shell deposition, then given 
the type of massive calcitic shell formed 
by many marine gastropods, an ion-imper- 
meable barrier would not normally be an es- 
sential protective feature where acid pH is a 
less frequently encountered environmental 
pressure. But here the environment is more 



136 



HUNT 



Π
< 

О 

CL 

< 



500 Г 



400 - 



300 - 



200 - 



1 00 - 




100 



200 



300 



400 



500 



600 



POLAR 

FIG. 11. The relationship between apolar and polar amino acids, as defined in Table 3, for gastropod 
periostraca. The different groups of gastropod are designated as follows:- TP, terrestrial prosobranchs; TPu, 
terrestrial pulmonates; FWP, freshwater prosobranchs; FWPu, freshwater pulmonates; MP, marine proso- 
branchs. Amino acid analyses are drawn partly from Meenakshi et al. (1969) and Degens et al. (1967) and 
from analyses performed in this laboratory but not yet published. Numerals after the designations represent 
numbers of species sampled, and the enclosed areas are the limits of apolar/polar value clusters — some 
gastropod groups have periostraca of more than one type. The graphical line demarcates apolar periostraca 
on the left from polar on the right. The star represents the position of the Lepetodrilus analysis. 



extreme and shell mineralology unstable, de- 
manding an impermeable and strong perios- 
tracum. 

Account may also have to be taken of the 
lower oxygen tension at which the limpets 
function. Operating metabolism partly anaer- 
obically may mean that consistently elevated 
levels of succinic acid could form in the extra- 
pallial fluid within the shell (Crenshaw & Neff, 
1969; Lutz & Rhoads, 1980), demanding a 
constant internal regeneration of the shell to 
compensate for acid dissolution. Better exte- 
rior protection would then be advantageous. 

Although the composition of the perios- 
tracum is unusual, its hydrophobicity is still 



inside the range for marine gastropods (Fig. 
12). It is the glycine content that pulls it away 
from the major groupings, and this factor too 
contributes to the anomalous position on the 
polarity distribution diagram. The almost 
equal balance of polar and apolar character is 
shared with some terrestrial and freshwater 
prosobranchs, though not with most marine 
species (Fig. 11). A high glycine content may 
promote a densely packed, highly folded ran- 
dom structure that density of which could be 
as helpful in reducing ion-permeability as hy- 
drophobicity. Lamellibranch periostraca from 
hydrothermal species also have unusual 
amino acid compositions with high glycine 



LEPETODRILUS SHELL STRUCTURE AND COMPOSITION 



137 

















/трД 
/б \ 












F> 


lip/ 






4 00 
















£ 200 












TP 3 




5 








'MP / 
7 / 








a. 
о 




TP n 6 










о 


















-200 








V_" TPu 


5 l/ 








-400 


■ 


MP 


1 8 


Г WPu / 

L 4 y' 










-600 


- 
















- 700 




' 


' 


1 


1 


' 





О 100 200 300 400 500 600 700 
Glycine 

FIG. 12. The relationship between hydrophobic in- 
dex and glycine content for gastropod periostraca. 
Hydrophobic index is defined in the footnote to Ta- 
ble 3. Because glycine has only a hydrogen in the 
side chain, it is assigned zero hydrophobicity (arbi- 
trarily, as thermodynamically it will have a finite AG 
value for transfer between water and a less polar 
solvent) and is assumed to contribute neither net 
polar or apolar value. Designations, numerals and 
symbols have the same meanings as in Figure 1 1 . 



contents. That from Bathymodiolus thermo- 
philus contains 85 residue percent though the 
few remaining amino acids give it more hy- 
drophobic character (Hydrophobic index: 
+ 156) (Hunt, 1987). 





FIG. 13. Analysis of the periostracum by X-ray mi- 
croprobe. A. Periostracum untreated other than by 
dilute acid decalcification to remove the shell. B. 
Periostracum extracted with carbon disulphide. C. 
The carbon disulphide extract dried onto the graph- 
ite stub. 



TABLE 4. Elemental composition of the periostracum of Lepetodrilus determined by X-ray microprobe 
analysis. A. Untreated material removed from the shell with dilute acid. B. Another specimen after 
removal from the shell and extraction with carbon disulphide. C. The carbon disulphide extract 
concentrated by evaporation at room temperature and dried onto the graphite-coated stub. These 
compositions relate to Figure 13A-C. Traces of aluminium and silicon may originate from clay particles 
on the surface of the material. Low levels of material recovered by CS 2 extraction do not make count 
integration meaningful. 



Element 


AI 


Si 


P 


S 


Fe 


Cu 


Counts %total 














A 


— 


1.01 


3.19 


93.41 


2.04 


0.34 


В 


1.22 


1.26 


1.97 


93.96 


0.94 


0.11 


С 


trace 


trace 


— 


trace 


— 


trace 



138 



HUNT 




FIG. 14. Scanning electron micrograph of the shell surface in the apical region showing erosion pits pen- 
etrating the periostracum and the underlying calcified material. 

FIG. 15. Detail of an erosion pit from the lower center of Figure 14. Note the cracking of the periostracum 
at the right which seems to follow the profile of the pit. 

FIG. 16. Detail of the pit shown in Figure 15 which illustrates the stepped progression of erosion. 
FIG. 17. Detail from Figure 16 illustrating what appears to be residual strands and sheets of organic matrix 
in and around the deepest pit. 



LEPETODRILUS SHELL STRUCTURE AND COMPOSITION 



139 




FIG. 18. Analysis of erosion pits by X-ray micro- 
probe. A. Interior of a shell outside surface pit of the 
type shown in Figure 17. B. Interior of a pit that has 
penetrated the shell to the inside and probed from 
within the shell. 



oldest and curved more sharply to conform to 
the shell's contour. Aging, chemical deterio- 
ration, non-specific cross-linking could all 
contribute to progressive stiffening and brittle- 
ness causing cracking and porosity. Another 
possibility is biodégradation. Baross & Dem- 
ing (1985) note extensive colonization of the 
limpets' shell surfaces by microorganisms. 
McLean (1988) remarks on heavy infestations 
by an unidentified sedentary organism form- 
ing globular irregularities on the shells of this 
species and the related L pustulosus. These 
organisms may etch and damage the under- 
lying periostracum. Herrera-Duvault & Roux 
(1986) have studied corrosion in mussel 
shells from the 13°N thermal vent site. After 
predator damage or microboring, bacterial ac- 
tivity destroys the organic matrix of the shell, 
making the dissolution of the aragonite layers 
easier. 

High phosphorous and sulphur levels, 
found here in the erosion pits, are difficult to 
account for. The dark material seems to be 
organic, as the microprobe detects no metals 
other than calcium. Aragonite dissolution will 
leave behind shell matrix proteins. Possibly 
these are being secondarily tanned to form 
products interacting with sulphur and phos- 
phorous-containing inorganic species in the 
water. Fibrous and sheet-like residues are 
visible in the pit shown in Figure 17. A more 
likely explanation is that pits provide a growth 
site for microorganisms. Accumulated bacte- 
rial material could be rich in sulphur and phos- 
phorus as sulpho- and phospholipids and as 
the phosphate groups of nucleic acids and 
cell wall polymers. 



Most of the limpet shells are eroded by 
small pits showing that if the periostracum is 
penetrated the vulnerable aragonitic shell will 
be damaged. At 2000 m depth, wave action 
can hardly be the cause of periostracal dam- 
age. Most pits and periostracal cracks occur 
near the shell apex, where the protein sheet is 



ACKNOWLEDGEMENTS 

I am indebted to Dr. Françoise Gaill for pro- 
vision of specimens of Lepetodrilus, to Dr. K. 
Oates for assistance with X-ray microprobe 
analysis, and to Mr. Haydn Morris for carrying 
out amino acid analyses. 



TABLE 5. Elemental composition of eroded pits in the shell of Lepetodrilus determined by X-ray 
microprobe analysis. A. Within a pit on the outside surface of the shell. B. Within a pitted area on the 
inside surface where erosion has penetrated the shell from the outside. These analyses relate to Figures 
18Aand 18B. 



Element 


Na 


Mg 


AI 


Si 


P 


S 


Cl 


Ca 


Counts %total 

A 

В 


0.36 
0.35 


3.58 
2.11 


2.57 


1.27 


39.08 
13.94 


28.33 
27.06 


1.07 
0.84 


25.57 
51.87 



140 



HUNT 



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WANG, J. C, B. A. CUNNINGHAM & G. M. EDEL- 



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WILBUR, K. M. & A. S. M. SALEUDDIN, 1983, Revised Ms accepted 3 February 1992 



MALACOLOGIA, 1992, 34(1-2): 143-342 



THE POMATIOPSIDAE OF HUNAN, CHINA 
(GASTROPODA: RISSOACEA) 

George M. Davis 1 , Cui-E Chen 2 , Chun Wu 3 ,Tie-Fu Kuang 4 , Xin-Guo Xing 5 , Li Li 6 , 

Wen-Jian Liu 7 , Yu-Lun Yan 8 



ABSTRACT 

This is a monograph involving the systematics of the 1 7 species of pomatiopsid snails thus far 
found in Hunan Province, the People's Republic of China. The Triculinae and Pseudobythinellini 
(new tribe) of the Pomatiopsinae dominate the pomatiopsid fauna. Four new species are de- 
scribed, three of Neotricula and one of Tricula. Detailed anatomical data are the basis for 
describing each species. The anatomies of "Akiyoshia" and Lithoglyphopsis are presented for 
the first time. Guoia is a new genus for some species previously considered to be Lithoglyphus 
(a strictly European hydrobiid genus) or Lithoglyphopsis (restricted to China). The anatomical 
data are the basis for phenetic and cladistic analyses to provide insight into those characters and 
character-states that serve to distinguish taxa, and to assess relationships among genera of the 
tribes Pachydrobiini (Halewisia, Neotricula, Pachydrobia, Jinhongia, Robertsiella, Guoia, Wu- 
conchona, Gammatricula) and Triculini (Tricula, Delavaya, Fenouilia, Lithoglyphopsis, Lacunop- 
sis). Hubendickia served as the outgroup genus of the remaining tribe of the Triculinae, the 
Jullieniini. Biogeographic deployment of the genera of all three tribes of the Triculinae is mapped 
on an area cladogram of relevant river systems. Genera of the Jullieniini dominate the lower 
Mekong River. Tricula extends from Northern India down the Yangtze and upper Mekong rivers. 
Tricula and Neotricula primarily flourish along the Yangtze River drainage. Four genera have 
shells that are so similar that shell characters cannot be used to distinguish among the genera: 
Tricula, Neotricula, Jinhongia and Gammatricula. Most of the species treated here are of medical 
importance because they either transmit or are suspected of transmitting lung flukes of the 
genus Paragonimus, or blood flukes of the genus Schistosoma. 

Key words: Pomatiopsidae, biogeogrpahy, phenetics, cladistics, China, Hunan, Schistosoma, 
Tricula, Neotricula, Paragoniumus, Yangtze River 



INTRODUCTION 

This work is one of a series of papers ded- 
icated to establishing the detailed anatomy 
and systematic relationships of genera of 
southeast Asian and Chinese freshwater 
prosobranch snails suspected of being mem- 
bers of the Pomatiopsidae as defined in Davis 
(1979, 1980). It is one more step in docu- 
menting the extensive speciation that has oc- 
curred within the tribes Triculini and Pachy- 
drobiini of the Triculinae, which occur 
primarily throughout southern China, in con- 
trast to the extensive Jullieniini adaptive radi- 
ation documented in the lower Mekong River 



in Thailand, Laos, and Cambodia (Davis, 
1979). 

In this work, Guoia is established as a new 
genus. The anatomies of "Akiyoshia" and 
Lithoglyphopsis are presented for the first 
time. Anatomical data are provided for clarifi- 
cation of the genus Pseudobythinella. Several 
nomenclatural problems are clarified or pre- 
sented relative to necessary future work. Sev- 
enteen nominal species are treated. 

Phenetic and cladistic analyses are done to 
provide insight into those characters and 
character-states that serve to distinguish 
taxa, and to assess relationships among gen- 
era of the tribes Pachydrobiini and Triculini, 



1 The Academy of Natural Sciences of Philadelphia, 1900 Benjamin Franklin Parkway, Philadelphia, Pennsylvania 19103- 

1195 U.S.A. 

institute of Parasitic Diseases, Zhejiang Academy of Medical Sciences, Hangzhou, Zhejiang Province, People's Republic 

of China 

3 Hunan Medical University, Changsha, Hunan Province 

"Hengshan County Anti-Epidemic Station, Hengshan, Hunan, People's Republic of China 

5 Anhua County Anti-Epidemic Station, Zhuzhou, Hunan, People's Republic of China 

6 Zhuzhou City Anti-Epidemic Station, Zhuzhou, Hunan, People's Republic of China 

7 Cili County Anti-Epidemic Station, Cili, Hunan, People's Republic of China 

8 Shimen County Anti-Epidemic Station, Shimen, Hunan, People's Republic of China 



143 



144 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



with one genus of the Jullieniini serving as the 
outgroup. Biogeographic deployment of the 
Triculinae is considered in conjunction with an 
assessment of the evolution of river systems 
in continental south Asia. 

Most of the taxa treated here are of medical 
importance because they either transmit or 
are suspected of transmitting lung flukes of 
the genus Paragonimus or blood flukes of the 
genus Schistosoma. 



MATERIAL AND METHODS 

Localities are provided with the synonymy 
section for each species. Sites 1 to 12 are 
found on Figure 1 . With reference to Figure 1 , 
be aware that maps published at different 
times may provide different spellings for stan- 
dard Chinese characters. Relevant here are: 
Hsiang = Xiang; Tzu = Zi; Lin = Li. The 
Yangtze River = the Chang Jiang (jiang = 
river; shui = river). Dong Ting Lake (Lake = 
Hu) = Tung fing Hu. 

Abbreviations for institutions housing 
voucher species are: ANSP = Academy of 
Natural Sciences of Philadelphia (the A- num- 
ber sequence indicates alcohol-preserved 
specimens); IZAS = Institute of Zoology, Ac- 
adémica Sínica; SMF = Senckenberg Mu- 
seum, Frankfurt A.M. Germany; ZAMIP = 
Zhejiang Academy of Medical Sciences, Insti- 
tute of Parasitic Diseases, Hangzhou, China. 
Voucher catalog numbers are provided in the 
synonymy section for each species. 

Methods are those of Davis & Carney 
(1973), Davis et al. (1976), Davis (1979), and 
Davis & Greer (1980). All dissections were 
done with living material in Hunan, China us- 
ing a Wild dissecting microscope and a Nikon 
high intensity lamp. Scanning electron micro- 
scopical analyses, photography of shells, rad- 
ular preparations and computer analyses 
were done in Philadelphia. 

Descriptions of taxa employ only those 
character and character-states useful to dis- 
criminate among species of Pomatiopsidae. 
Character-states common to all are not re- 
peated here but are described elsewhere 
(Davis, 1979, 1980). 

Relative sizes of organs or shells are de- 
fined by range in size (as in shells) or ratios to 
facilitate comparison among taxa using terms 
such as "small, " "elongate, " etc. This prac- 
tice follows Davis et al. (1986) and all later 
papers in this series of studies. The sizes and 
ratio used are: 



1. Shell length: large (> 5.0 mm), medium 
(4.0-4.9 mm), small (2.1-3.9 mm), minute (< 
2.0 mm) (based on mean size of the mature 
size class with the greatest number of individ- 
uals, realizing that mature snails may occur 
with different numbers of whorls, e.g. 5.0, 5.5, 
6.0 whorls). 

2. Operculum attachment pad: width of at- 
tachment pad -s- width of operculum: narrow 
(< 0.35), wide (0.36-0.55), very wide 
(> 0.56). 

3. Osphradium length: length of osphra- 
dium *■ length of gill (Davis et al. 1976, 1982, 
1983): long (> 0.40), short (< 0.35). 

4. Length of gill filament section Gf 2 : length 
of Gf 2 * length of G^ and Gf 2 : long (> 0.51), 
medium ( = normal) (0.31-0.50), short 
(< 0.30). 

5. Gill filament number: few (< 15), numer- 
ous moderate (16-25), numerous (> 26). 

6. Bursa copulatrix length: length of bursa 
h- length of the palliai oviduct: long (> 0.40), 
short (< 0.38). 

7. Albumen gland length: length of the al- 
bumen gland (Ppo) ■*■ length of the entire pal- 
liai oviduct: standard (> 0.45), short (< 0.42). 

8. Male gonad length: gonad length 
-e- length of digestive gland: short (< 0.50), 
long (> 0.50). 

9. Seminal receptacle duct length: very 
short or absent (< 0.02 mm), long (> 0.02 
mm). 

10. Radula length: short (< 0.40 mm), me- 
dium (0.41-0.59 mm), long (0.60-0.79), very 
long (> 0.80 mm). 

1 1 . Radula; mean rows of teeth: few (< 59), 
moderate (60-69), many (70-84), very many 
(> 85). 

12. RPG ratio: length of the supraesoph- 
ageal connective ■*■ sum of the length of the 
right pleural ganglion, the supraesophageal 
connective, and the supraesophageal gan- 
glion; concentrated (< 0.29), moderately con- 
centrated (0.30-0.49), elongate (0.50-0.67), 
extremely elongate (> 0.68) (Davis et al. 
1984, 1985). 

13. Subesophageal connective length: 
usual or standard triculine (0.02-0.09 mm); 
none; long (> 0.10 mm) [based on average of 
several measurements]. 

Multivariate Analysis 

Shell data were analyzed for populations of 
Guoia. The first data matrix involved 33 OTUs 
(operational taxonomic units) (individual 
shells) and 11 characters. The OTUs were: 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 145 





HUNAN 

p 25 50 75 km 




114' 




















HUBEi 




29 
28 
27 
26 




..Д!-У_ Cj 

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v '«-/ /К Í^S^w / \s— ^ ^V( cs î' .Г ^ V 

S _.\ V ?*«еКУ — 7к ' ' ч \ Гх '-^ Г / > 

Í ~~~4„ c-^Shîmen ^ V ¿< УУ Ь-4.. f Г ^ 

\ ? (it) улан уЗУУ í';^/ v >r 

} »Shenglot» — ^ (Sil) ) Ч А '«Л / JpP 
\ «¿^»Xikou f \ / Г{\* V » ? 

\ Listel 8 ( ' 5ni <зш ,X Л Щ\\1 wL \ 

Xiao^uanping У^ Щу^Л^^/ 

У/ (5 ю ^~-^ \ 

j Annua о, — ¿lIV \ 
^_У _^ю Jiangnan д 

~? ¿ (;1¡li) Ichangsha 

у ? л (i ¡>) 

V, /^~1 HUNAN V 


с - 

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v < fife ( 
; ш s 


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w Í \) fr 

z ""•' ^ ^ c 

1 .Г / ( 


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) 

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s 

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3aisha(è,W Uv " 
ngshan((^ llj/ ) 

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( \ S'— A i* Л ' — Sanhe 


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111' 112" 113' 


114 



FIG. 1 . Map of Hunan, China, showing localities from which specimens described in this monograph were 
collected. 



146 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Guoia viridulus, large class snails, males (1- 
7), females (8-12); ANSP cataloged speci- 
mens of Guoia viridulus collected over 90 
years ago (18-21); small class snails, males 
(22-26), females (27-32). Guoia fuchsianus 
males (13, 14), females (15-17). ANSP cat- 
aloged historic G. fuchsianus (33). 

The characters were: 

1. Number of whorls 

2. Length 

3. Width 

4. Length of body whorl 

5. Length of penultimate whorl 

6. Width of penultimate whorl 

7. Width of 3rd whorl 

8. Length of last three whorls 

9. Length of aperture 

10. Width of aperture 

1 1 . Width of columellar shelf 

The second matrix included individuals of 
two populations from historic collections, 
Guoia fuchsianus cataloged in the ANSP col- 
lections: ANSP 98205 (34-38), ANSP 45961 
(39-41). These OTUs were added to the 
OTUs of matrix one to give a total OTU count 
of 41 . The character matrix was reduced be- 
cause all OTUs 34-41 had eroded apices, 
thus characters 1,2,7, and 8 could not be 
measured for all individuals. 

Computations were made using the 1974 
version of NT-SYS (Rohlf et al., 1972). Char- 
acters were standardized (standard deviation 
and mean values). Similarity and distance co- 
efficients were calculated and phenograms 
produced using UPGMA. Cophenetic correla- 
tions were calculated. Principal Component 
Analysis (PCA) was done with components 
extracted until eigenvalues became less than 
1 .0. A transposed matrix of the first three prin- 
cipal components with their character loading 
was postmultiplied by the standardized matrix 
to yield a matrix of OTU projections in the 
principal component space (Rohlf et al., 
1972). The OTU locations in the three-dimen- 
sional space were used as the initial configu- 
ration for a nometric multidimensional scaling 
(MDS) placement of Q-mode taxonomic dis- 
tances between OTUs (Kruskal, 1964). Ordi- 
nation was done followed by a Prim Network 
(minimum Spanning Tree = MST). Subse- 
quently a phenogram was produced based on 
MDS ai.d the cophenetic correlation calcu- 
lated. 

A similar multivariate analysis was used to 
assess phenetic relationships among 30 



OTUs on the basis of shell data. The OTUs 
involved species of Tricula, Neotricula or spe- 
cies the shells of which resembled those of 
species of Tricula. The analysis was done as 
an aid for distinguishing among species on 
the basis of shell characters. 

Another similar multivariate analysis was 
done to assess phenetic relationships among 
the same species for which the shell analysis 
was done. However, this analysis involved 48 
anatomical character-states. 

Cladistic Analysis 

Scorings were not done in binary alone but 
involved six multistate characters (Table 85). 
Analysis was first done using the computer- 
mediated program Hennig-86 (Farris, 1988). 
Hubendickia served as the outgroup. Options 
run were cc-. (character-states all unordered); 
ie; tplot; tsave; nelsen. A final non-computer- 
mediated cladogram was made after weight- 
ing one character following establishing the 
direction of evolution of its states. 

Shell Characters 

Shell character-states is illustrated in Figs. 
2, 3. Aside from shell measurements there 
are a number of diagnostic qualitative char- 
acter-states of use to discriminate among 
shells (Tables 1 , 2). 

Triculine shells have three predominant 
shapes (Fig. 2A): globose, ovate-conic, 
ovate-turreted. Many character-states involve 
the aperture. The adapical end may be regu- 
larly rounded or slightly constricted to form an 
adapical apertural notch (Adn, Fig. 3D). The 
constriction may be less pronounced (Fig. 
3E). The adapical aperture and apertural 
notch may be extended to form the adapical 
apertural beak (Adb, Fig. ЗА). The apertural 
notch may have a noticeable internal groove 
(Ngr, Fig. 3E) or lack a groove (Fig. ЗА). The 
adapical apertural notch may be bounded by 
an adapical tooth or node on the inner lip (Adt, 
Fig. ЗА, D); or the adapical inner lip may lack 
such a thickening (Fig. 3E, H). 

The inner lip may (Ah, Fig. ЗА) or may not 
be fused (Gap, Figs. 3D, E) to the body whorl. 
When separated from the body whorl the gap 
(Gap) may be narrow (Fig. 3E) or wide (Fig. 
3D). The inner lip may be straight, arched 
(Fig. ЗА), sigmoid or undulating (Fig. 3E), or 
angled (Fig. 3D). The adapical part of the in- 
ner lip may be thickened (Adil, Fig. 3D) or 
thin; the abapical part of the inner iip the 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 147 



TABLE 1 . Twenty-six shell character and character- 
states useful for describing shells of Triculinae as 
illustrated in Figures 2 and 3. 

Abil Abapical inner lip 

Abs Abapical spout 

Adb Adapical apertural beak 

Adil Adapical part of inner lip 

Adt Adapical tooth or node 

Adn Adapical apertural notch 

Agp Adapical apetural gap 

Aol Adapical outer lip angle 

Aols Adapical outer lip sinus 

Ari Arched inner lip 

Cr Crenulated suture 

Ct Columellar tooth 

Gap Gap between the body whorl and the 

inner lip 

L Length 

Mln Mid-lip notch 

Nab Normal adapical aperture 

Ngr Adapical notch groove 

Ol Outer lip 

Sfo Scooped forward 

Sin Sinuate outer lip 

Stl Straight lip 

Umc Umbilical chink 

Var Varix 

W Width 

x distance from base of body whorl to 

abapical end of aperture 

y distance from edge of outer lip to edge of 

body whorl 



same. The inner lip may have a mid-lip notch 
(Mln, Fig. 3D). 

The abapical end of the aperture may 
project noticeably beyond the base of the 
body whorl when the shell is viewed in aper- 
tural view (Fig. 3E); it may not (Fig. 3F). 
The abapical end of the aperture may be 
spout-like (Figs. 3D, E); it may not (Fig. ЗА, 
F). The columella may have a tooth (Ct, Fig. 
3F). It is necessary to break open the body 
whorl of some shells from a population to de- 
termine whether or not there is an internal 
columellar node, tooth, or keel. 

The adapical end of the aperture may be 
pulled away from the body whorl leaving an 
adapical gap (Agp, Fig. 3D, F) or the adapical 
aperture may be fused to the body whorl (Fig. 
ЗА). 

In side view the outer lip may be straight 
(Stl, Fig. 3H) or sinuate (Sin, 3B). The outer 
lip may be aligned with the axis of coiling (Fig. 
3C, H) or scooped forward (Sfo, Fig. 3B). In 
side view there may be an adapical depres- 
sion or sinus in the outer lip (Aols, Fig. 3H) 
caused by an adapical apertural notch. 



Rotating the shell from apertural view to the 
right (apex up) so that the inner lip is perpen- 
dicular to the horizontal, the abapical part of 
the inner lip may be deflected away from the 
columella causing a lip deflection angle (Fig. 
3G, 2B). Such an angle may not occur. 

Gill Filament Shape 

Considering the longest (thus mid-gill) gill 
filaments, three shapes are found when the 
filaments are examined so as to see the entire 
leaflet (Fig. 4). The Gf 2 section may be flat, 
have a modest dome, or be high domed. 

Vital Staining 

Davis (1967) discussed using aqueous 
methylene blue and neutral red as vital stains. 
The exact methods are as follows: small 
quantities of powdered stain are dumped into 
a quantity of tap water (approximately 500 ml) 
and stirred until dissolved. Sufficient powder 
is added so that when the solution is placed 
before a light the solution is opaque. Wide- 
mouth jars are used, one each for the two 
stains. Wide mouth containers are specified 
because the stain is used over and over 
again. With the living animal pinned out on the 
black parafin-wax layer in a 9-cm Petri dish, 
the water covering the animal is poured off 
and the stain poured on to cover the animal. 
The stain is left on for 30 seconds to a minute, 
then poured back into the storage bottle. The 
animal is rinsed five to ten times to remove 
excess stain and then covered with water to 
continue the dissection. 

Neutral red is used first to deliniate glandu- 
lar tissue and the ducts of the female repro- 
ductive system. The two regions of the palliai 
oviduct are differentiated; the oviduct is 
stained as well as the gonad and oocytes. 

Methylene blue is used next; it seems to 
harden and delineate the ducts of the female 
reproductive system. The slender duct of the 
seminal receptacle and the various other 
components of the bursa copulatrix complex 
of organs are made to stand out. 

Bouins Solution 

When all work with the living animal is fin- 
ished, Bouins solution is most useful to elim- 
inate problems caused by mucus, to delineate 
mantle cavity structures, such as the gill fila- 
ments, and to dissect the nervous system. 
The nerves and ganglia stain bright yellow en- 
abling one to see them better and to difieren- 



148 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 2. Shell character-states scored for 29 characters used for a multivariate analysis of phenetic 
relationships among 21 species of Triculinae, the shells of which either are classified as Tricula or 
Neotricula or resemble the shells of these two genera. 



1. Size: 

2. Shape: 

3. Aperture shape: 

4. Umbilicus: 

5. Whorl at suture: 

6. Teleoconch sculpture: 

7. Spiral sculpture at mid body- 
whorl to shell base 

8. Protoconch whorls: 

9. Adapical aperture: 

10. Adapical aperture: 

1 1 . Adapical aperture: 

12. Abapical aperture: 

13. Inner lip: 

14. Outer lip-side view: 

15. Outer lip-side view: 

16. Inner lip: 

17. Inner lip: 

18. Base of inner lip, side view: 

19. Inner lip: 

20. Inner lip: 

21 . Adapical aperture: 

22. Columella within body whorl: 

23. Varix: 

24. Adapical aperture: 

25. Adapical outer lip angle: 

26. Base of body whorl: 

27. Base of shell at umbilicus: 

28. Base of shell at abapical lip: 

29. Shell attains 7.0 whorls: 



small (0), medium (1), minute (2), long (3) 

ovate-conic (0), ovate-turreted (1), turreted (2), cylindrical-conic 

(3), globose-conic (4) 
ovate (0), pyriform (1), distorted (2) 
none (0), chink (1), clearly open (2) 
smooth (0), crenulated (1) 
none (0), spiral microsculpture at suture (1) 

none (0), with (1) 

smooth (0), wrinkled (1), malleated and/or pitted (2) 

normal (0), with notch (1), with beak (2) 

normal (0), with internal groove (1) 

normal (0), with sinus (1) 

normal (0), with spout (1) 

straight (0), arched (1), sinuate (2), angled (3) 

straight (0), sinuate (1) 

parallel with axis of coiling (0), scooped forward (1), slanted back 

(2) 
no tooth (0), with tooth (1) 
no notch (0), with notch (1) 
straight (0), angled to form lip deflection angle (1) 
thin (0), thick [> 0.10mm] (1), differentially thickened (2) 
fused to body whorl (0), partly separated (1), totally separated by 
narrow gap (2), widely separated (3) 
fused to body whorl (0), slightly separated (1), widely separated 

(2) 
smooth (0), tooth or node (1), spiral keel (2), lamellae on aperture 

side of columella (3) 
none (0), slight (1), pronounced (2) 
normal (0), with beak tubercle (1) 
none (0), slight (1), extended (2) 

normal (0), with keel spiraling down from umbilical area (1) 
normal (0), with wide columellar shelf (1) 
normal (0), with basal post (1) 
no (0), yes (1) 



tiate nerves from muscle fibers. Bouins solu- 
tion strengthens the nerves such that they 
can be handled better. Dissections of the 
head involving the buccal mass, salivary 
glands and associated nerves are facilitated. 

Body Measurements 

How the lengths of the gonad, digestive 
gland, and body are measured is illustrated in 
Davis & Carney (1973). Lengths of mantle 
cavity organs are measured with the mantle 
cut along the right side of the neck from the 
mantle edge to the rear of the mantle cavity. 
The mantle is then reflected to the left and 
pinned out as shown in Figure 5. How lengths 
of mantle cavity structures are measured is 
illustrated in Figure 5. The ocular micrometer 
is positioned and rotated along the dotted tra- 
jectory. As shown in Figure 5, not all gill fila- 
ments may be illustrated. 



Abbreviations 

a line perpendicular to mid-line "x" at 

posterior edge of eye lobes 

a' Line perpendicular to mid-line "x" at 

the anterior edge of penial base 

Adb Adapical apertural beak 

Ae Abapical embayment 

Af Anterior foot; 

Algo Anterior lobes of gonad 

Als Adapical outer lip sinus 

Apg Anterior pedal glands. 

Apo Anterior palliai oviduct = capsule 
gland 

Apo-1 Anterior palliai oviduct = capsule 
gland 

Apo-2 Different tissue type anterior to cap- 
sule gland. 

Ast Anterior chamber of stomach 

Au Auricle 

Be Basal crescent 

Bg Beak groove 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 149 



Bm 


Buccal mass 




pressed down on a horizontal sur- 


Bp 


Base of penis 




face 


Bu 


Bursa copulatrix 


LOs 


Length of osphradium 


Cc 


Cerebral commissure 


Lpl 


Left pleural ganglion 


CI 


Columellar muscle 


Lpw 


Length of penultimate whorl 


Coi 


Oviduct coil 


Ma 


Mantle collar 


Cr 


Crescent ridge 


Ne 


Neck 


Cs 


Columellar shelf 


Ngr 


Adapical notch groove 


Csd 


Common sperm duct 


Odi 


Opening from stomach to digestive 


Dbu 


Duct of bursa 




gland 


Di 


Digestive gland 


Oki 


Opening of kidney into mantle cavity 


Dsr 


Duct of seminal receptacle 


Ol 


Outer lip 


Ebr 


Eyebrow 


Orne 


Opening of spermathecal duct into 


Ebv 


Efferent branchial vein 




mantle cavity 


Edi 


Anterior edge of digestive gland 


Omn 


Osphradiomantle nerve 




(Figs. 141, 142) 


Ooc 


Oocyte(s) 


Edi 


Dashed line indicates anterior limit 


Oop 


Opening into albumen gland (Ppo) 




of digestive gland 


Oov 


Opening of oviduct to albumen gland 


Щ 


Ejaculatory duct 




(enlarged in Fig. 10) 


Emc 


Posterior end of mantle cavity 


Ope 


Opening for sperm entry to pericar- 


Epr 


Dashed line indicates posterior limit 




dium 




of prostate gland 


Opo 


Opening of oviduct into albumen 


Es 


Esophagus 




gland 


Ey 


Eye 


Opr 


Opening to vas deferens 


Eyb 


Eyebrow 


Os 


Osphradium 


Fp 


Fecal pellet 


Osd 


Opening of spermathecal duct to 


Gf 1 


Thickened basal bar of gill filament 




mantle cavity (Figs. 11, 21 ) 


Gf 2 


Slender terminal part of gill filament 


Osd 


Point where oviduct fuses to, and 


Glo 


Glandular lobe 




opens into pericardium 


Go 


Gonad 


Osm 


Osphradiomantle nerve 


Go-a 


Section of gonad, the extent of which 


Osr 


Oviducal seminal receptacle 




is indicated by the dashed line, re- 


Ov 


Oviduct 




moved to show seminal vesicle 


Pa 


Papilla 


Go-b 


Gonad 


Pbu 


Pericardial bursa 


Gr 


Patch of white granules 


Pc 


Pellet compressor ( = ln 2 ) 


Gra 


Granules or glands, white to lemon 


Per 


Pericardium 




yellow 


Pe 


Penis 


Grv 


Groove 


Pf 


Penial filament 


Gs 


Grey streak on Ast 


Ppo 


Posterior palliai oviduct = albumen 


II 


Inner lip 




gland 


In 


Intestine 


Pr 


Prostate 


In, 


Intestine looping around anterior 


Psc 


Pleurosupraesophogeal connective 




end of style sac 


Pst 


Posterior chamber of stomach 


ln 2 


Fecal pellet compressor section of 


Reg 


Right cerebral ganglion 




intestine 


Ri 


Yellow ridge 


ln 3 


Anterior intestine 


Rpl 


Right pleural ganglion 


Ki 


Kidney 


Rs 


Radular sac 


L 


Length 


Sd 


Spermathecal duct 


Lap 


Length of aperture 


Sdo 


Opening of oviduct into the pericar- 


Lbw 


Length of body whorl 




dium 


LGf, 


Length of gill filament section Gf, 


Sdu 


Sperm duct 


LGf 2 


Length of gill filament section Gf 2 


Sec 


Pleurosupraesophageal connective 


LGi 


Length of gill 


Seg 


Supraesophageal ganglion 


LMa 


Length (= width) of mantle collar 


sg 


Salivary gland 




anterior to gill 


Sn 


Snout 


Lm 


Dashed line is trajectory used to 


Sr 


Seminal receptacle 




measure length of gill and length of 


Sts 


Style sac 




mantle cavity 


Sty 


Stylet 


LofA 


Length of shell with aperture 


Sue 


Pleurosubesophageal connective 



150 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Sug 

Sv 

Tn 

Twv 

Uc 

Vd 

Vd, 

Vd 2 

Ve 

Vei 

Ven 

W 

Wap 

Wgr 

Wg 

WofA 

W 
x 

x 

Yri 



Subesophageal ganglion 
Seminal vesicle 
Tentacle 

Thin walled vestibule 
Umbilical chink 
Vas deferens 
Posterior vas deferens 
Anterior vas deferens 
Vas efferens 
Vein 

Ventricle 
Width (Fig. 110) 
Width of aperature 
White granular inclusions 
White granules 

Width of shell with aperature pressed 
down on a horizontal surface 
Wall of neck 

Mid-line of snout-neck with snout an- 
terior (up). 

Marks the same point in Figs. 20, 21 
A and В; 117A, 118 
Yellow ridge along anterior and pos- 
terior ends of the Ast 



SYSTEMATICS 



Taxa Treated 



POMATIOPSIDAE 

Pomatiopsinae 
Pomatiopsini 
Oncomelania 
Oncomelania hupensis Gredler, 1881 
Pseudobythinellini, Davis & Chen new 
tribe 

Akioyshia 
"Akiyoshia" chinensis Liu, Zhang & 
Chen, 1982 
Pseudobythinella 
Pseudobythinella chinensis (Liu & 

Zhang, 1979) 
P. shimenensis Liu, Zhang & Chen, 
1982 
Triculinae 
Pachydrobiini 
Guoia Davis & Chen gen. nov. 
G. fuchsianus (Moellendorff, 1885) 
G. viridulus (Moellendorff, 1888) 
Neotricula 
N. cristella (Gredler, 1887) 
N. dianmenensis Davis & Chen, sp. 

nov. 
N. duplicata Davis & Chen, sp. nov. 



N. //7/7 Chen & Davis, sp. nov. 

N. minutoides (Gredler, 1885) 

Triculini 
Lithoglyphopsis 
L. modesta (Gredler, 1886) 

Tricula 

T. gredleri Kang, 1986 
7. maxidens Chen & Davis, 
sp. nov. 

T. odonta Liu, Zhang & Wang 1983a 

Nomina nuda: 

Tricula gredleri 
Akiyoshia odonta 



Nomina Nuda 

Two names were introduced into the litera- 
ture for presumed Pomatiopsidae from Hunan 
Province but without photographs, illustra- 
tions, designation of types, or descriptions 
that would permit one to identify the species 
in question. Accordingly these are nomina 
nuda. The nominal taxa involved are: 

Tricula gredleri Feng et al., 1985 [Kang sp. nov.] 
Tricula gredleri Feng et al., 1986 [Kang sp. nov.] 
Akiyoshia {Saganoa) odonta Feng et al., 1985 
[Kang sp. nov.] 

Akiyoshia (Saganoa) odonta Feng et al., 1986 
[Kang sp. nov.] 

Higher Taxa Defined 

The family Pomatiopsidae was differenti- 
ated from the Hydrobiidae in Davis (1979); 
two subfamilies were recognized, the Poma- 
tiopsinae and Triculinae. Both subfamilies 
have a spermathecal duct, a pomatiopsid 
type central tooth (square to rectangular) with 
pronounced basal cusps arising from the face 
of the tooth (an exception includes the 
Pseudobythinellini discussed below), cover 
eggs (laid singly) with sand grains, and do not 
brood young. The penis is without complex 
glands or lobes. The differentiation of the po- 
matiopsid subfamilies from other higher taxa 
with spermathecal ducts was given in Davis et 
al. (1985: 74-75). These higher taxa are the 
Hydrobiidae: Littoridininae and Amnicolinae 
(also see Hershler & Thompson, 1988). 

The Pomatiopsinae are Gondwanian in dis- 
tribution. Pomatiopsinae snails have an elon- 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 151 






GLOBOSE 



OVATE-CONIC OVATE-TURRETED 



В 




FIG. 2. Shell characters used in describing species of Hunan Triculinae. A. Three major shell shapes. B. 
Inner lip in side view showing a pronounced lip deflection angle. 



152 

A 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 

в с 




Nab 



Abs 



Var 



FIG. 3. Shell characters used in describing species of Hunan Triculinae. Labels are defined in Table 1 . 



gated spermathecal duct extending to the an- 
terior end of the mantle cavity. In the tribe 
Pomatiopsini, the eyes are in pronounced 
bulges. These snails have a pedal crease, su- 
prapedal fold and omniphoric groove. They 
move by a step-wise mode. They have 
evolved from freshwater to an amphibious 
mode of existence, and in Japan Blanfordia 
has become terrestrial (Davis, 1979, 1981). 
The tribe Pseudobythinellini is placed in the 
Pomatiopsinae because of the elongated 



spermathecal duct. The central tooth is of the 
Hydrobia type with a pair of cusps arising 
from the lateral angles. The eyes are not in 
pronounced eye lobes. There is no su- 
prapedal fold, omniphoric groove or pedal 
crease. Animals move by ciliary glide. The pe- 
nis of species classified here as Pseudo- 
bythinellini lacks a penial lobe with functioning 
duct (in addition to the vas deferens) that wid- 
ens to form a wide coiled mass or penial 
gland in the cephalic haemocoel. The accès- 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 153 




MODEST DOME 



HIGH DOMED 



FIG. 4. Shapes of the Gf 2 segment of the largest gill 
filaments. 




FIG. 5. Mantle reflected to reveal mantle cavity or- 
gans and how their lengths were measured. The 
dashed line is the trajectory for measuring the 
length of the gill. The length of the mantle cavity is 
measured from Emc along the LGi plus LMa. 



sory penial lobe and penial gland are charac- 
teristic of the European genus Bythinella and 
North American genus Amnícola of the Hy- 
drobiidae: Amnicolinae (Davis et al., 1985; 
Hershler & Thompson, (1988). 

The Triculinae are Asian with a distribution 
from northern India throughout South China 
and southeast Asia. No Triculinae are found 
in Japan (Shikoku to Okinawa), loganzen & 
Starobogatov (1982) refer their new genus 
Sibirobythinella to the Triculidae. However, 
given the extensive convergences in structure 
such as the spermathecal duct and central 
tooth of the radula (reviewed in Davis et. al., 
1985) it is not at all certain that taxa from 
Siberia or northeastern Russia relegated to 



the Triculidae by Russian workers are Tricu- 
linae sensu Davis (1979, 1980). Sibiro- 
bythinella is not, based on data provided, a 
triculine snail. The shell and radula might in- 
dicate placement in the Pseudobythinellini. 
loganzen & Starobogatov (1982) place their 
taxon in the family Triculidae apart from taxa 
they erected to family status such as Littori- 
dinidae, Pomatiopsidae and Stenothyridae, 
all characterized by having a spermathecal 
duct running independently of the palliai ovi- 
duct from the mantle cavity to the bursa cop- 
ulatrix. Unfortunately, they do not provide suf- 
ficient data to differentiate their taxon from 
European Hydrobiidae: Littoridininae, Amni- 
colinae, or Pomatiopsinae: Pseudobythinel- 
lini. 

Triculinae snails have a short spermathecal 
duct opening to the posterior end of the man- 
tle cavity. The snails are aquatic, do not have 
a suprapedal fold, pedal crease, or om- 
niphoric groove. Only in a few derived genera 
are the eyes in pronounced lobes. Snails 
progress by ciliary glide. 

Tribal, Generic and Species Descriptions 



POMATIOPSINAE 

Pomatiopsini 

Type genus. Pomatiopsis Tryon 1 862 

Diagnosis. Ovate-conic to turreted shells. 
Shell length greater than 2.5 mm; most 
greater than 3.0 mm. Apical whorls are not 
flattened. Spermathecal duct runs from the 
bursa copulatrix to the anterior end of the 
mantle cavity. Spermathecal duct not tightly 
pressed to palliai oviduct. Sperm duct running 
posteriorly from oviduct to the bursa or the 
spermathecal duct at the bursa. Eyes are in 
pronounced eye lobes. There is a suprapedal 
fold, an omniphoric groove, and pedal crease. 
Basal cusps of the central tooth of the radula 
arise from the face of the tooth. 

Oncomelania Gredler 1881 

Type Species. Oncomelania hupensis Gred- 
ler 1881: 120-121; pi. 6, fig. 5 

Type locality. U — tschang — fu, March 1879. 
[= Hubei Province; Wu — Tshan — fu; Yen, 
1939]. 



154 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Designation. By monotypy 

Types. Bozen; lectotype and two paralecto- 
types 

Bozen No. 89. No. types at SMF (Zilch, 
1974: 197) 

Oncomelania hupensis Gredler, 1881 

No detailed study of this well-known spe- 
cies is presented here. This species is in- 
cluded for completeness sake in monograph- 
ing the Pomatiopsidae of Hunan, China. This 
species is the vector for Schistosoma japoni- 
cum and abounds in the marshes around 
Dong Ting Lake and along the Yangtzee 
River (Fig. 1) (Lou et al., 1982; Liu et al., 
1981 ). Detailed anatomical data for Oncomel- 
ania have been published elsewhere (Davis 
1967, 1968a, 1969a; Davis & Carney 1973). 

Pseudobythinellini Davis & Chen 
New Tribe 

Type genus. Pseudobythinella Liu & Zhang 
1979. 

Diagnosis. Ovate shells less than 2.5 mm 
long, with flattened apical whorls. The sperm 
duct runs anteriorly from the oviduct to the 
spermathecal duct; it is so tightly pressed to 
the palliai oviduct that it is difficult to differen- 
tiate it. Unlike the Pomatiopsini, the sperm 
duct enters the spermathecal duct far anterior 
to the bursa copulatrix; the duct of the bursa is 
thus elongated (the continuation of the sper- 
mathecal duct to the bursa from the point of 
entry of the sperm duct). Eyes are not in pro- 
nounced eye lobes. There is no suprapedal 
fold, no omniphoric groove or pedal crease. 
Animals are aquatic and move by ciliary glide. 
Basal cusps of the central tooth of the radula 
arise from the lateral angles of the tooth. 

Synonymy. Erhaiini Davis et al. 1985: 69. 

Discussion. The genera included in this tribe 
are Akiyoshia and Pseudobythinella from 
China. Liu & Zhang, 1979, described certain 
species from China and classified them as 
Bythinella and Pseudobythinella. They pro- 
vided no anatomical data aside from penis 
and radular illustrations. Pseudobythinella Liu 
& Zhang, 1979, was described as different 
from so-called Bythinella of China by (1) hav- 
ing a tooth or node on the inner lip, and (2) the 
central tooth having two pairs of basal cusps, 
those on each side on the same level (not 
Hydrob/a-like). 



The so-called Bythinella of China lack the 
complex male reproductive system charac- 
ters that serve to define the Hydrobiidae: Am- 
nicolinae of Europe and North America in- 
cluding genuine Bythinella (Europe) and 
Amnícola (North America). Because Bythi- 
nella does not occur in China, Davis & Kuo, 1 
(in Davis et al., 1985) described the genus 
Erhaia to accommodate new species that Liu 
and Zhang would have considered to be 
Bythinella. Davis & Kuo (1985) raised the 
tribe Erhaiini to accommodate Erhaia. 

As a result of this study, Erhaia is placed in 
the synonymy of Pseudobythinella. It is clear 
that at least one species described as 
Bythinella from China {B. chinensis Liu & 
Zhang, 1979) has a tooth on the columella; it 
is observed when the aperture is tilted. Upon 
breaking open the shell it is seen that this 
"tooth" is the terminus of a thick, glassy, spi- 
ral columellar shelf or ledge. In summary, 
there appears to be no basis for placing those 
taxa with an overall similar anatomical ground 
plan in separate genera on the basis of pres- 
ence or absence of a tooth on the columella, 
or where the tooth is prominently seen in the 
aperture contrasted with the presence of a 
columellar thickening seen only upon break- 
ing the shell. The type species of Erhaia, E. 
daliensis Davis & Kuo (in Davis et al., 1985) 
has no columellar "tooth" in evidence in the 
aperture even when the aperture is rotated to 
examine as deeply as possible into the shell. 
However, upon breaking open the body 
whorl, a pronounced thickened columellar 
ridge is seen. This glassy ridge is also found 
in Erhaia kunmingensis Davis & Kuo 1985, 
but it is not very pronounced and forms no 
node. 

The same situation involves the basal 
cusps of the central tooth. The two basal 
cusps on a side being on the same level or 
one above the other is irrelevant to generic 
definition. Most species of Pseudobythinella 
(including the Chinese taxa described as 
Bythinella) have only one pair of basal cusps 
on the lateral angles. 

The Pseudobythinellini are not classified as 
Hydrobiidae but are in the Pomatiopsidae for 
the following reasons: (1) There is evidence 
that the position of the basal cusps of the cen- 



1 Due to a change in China affecting the manner of spelling 
Chinese words in English, Dr. Y. H. Kuo in the literature 
until 1985 changed the spelling of his name to Y. H. Guo 
(see Davis et al., 1985, 1986). 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 155 



tral tooth derived from the pomatiopsine con- 
dition, i.e. from the face of the tooth. In 
Pseudobythinella kunmingensis from Yun- 
nan, China most central teeth of most snails 
had a single pair of basal cusps. However, 
some central teeth had two pairs (Davis et al., 
1985: fig. 16F), the innermost arising from the 
face of the tooth and lower than the outermost 
pair arising from the laternal angle. This has 
not, to our knowledge, been seen in any taxon 
of Hydrobiidae that has a spermathecal duct. 
(2) The position of the connections of the 
spermathecal duct, duct of the bursa, and ovi- 
duct are unlike any seen in the Hydrobiidae. 
These Chinese species lack the complex pe- 
nial lobes, three to four differentiated regions 
of the palliai oviduct, and tendency to brood- 
ing seen in the Hydrobiidae: Littoridininae. (3) 
There is the biogeographic overlap of these 
snails with at least one genus of the Pomati- 
opsini, more if Akiyoshia, originally described 
from Japan, is also a member of the Pseudo- 
bythinella clade. 

Akiyoshia Kuroda & Habe 1954 

Type Species. Akiyoshia uenoi Kuroda & 
Habe, 1954: 71-73, figs. 1-4 

Type locality. Akiyoshi limestone cave, Ya- 
maguchi Prefecture, Japan 

Designation. By monotypy 

Saganoa Kuroda, Habe & Tamu, 1957 

Type species. Akiyoshia (Saganoa) kishiiana 
Kuroda, Habe & Tamu in Kuroda & Habe, 
1957: 186-187, figs. 5-8. 

Type locality. Saga (= Arashiyama), western 
foothills of Kyoto City, Japan 

Designation. By original designation 

Discussion. Species from China were de- 
scribed as Akiyoshia (Saganoa) by Liu et al. 
(1982). They were thus described because 
the shells of those species are minute, tur- 
reted, with roundish apertures, i.e. generally 
corresponded to the conchological descrip- 
tions given by Kuroda & Habe (1957). We 
doubt that the Chinese species are Akiyoshia, 
s.S., or Akiyoshia (Saganoa) due to the dis- 
tance and differences in geological events 
creating environments now inhabited by the 
Chinese and Japanese snails. We are re- 
minded that species once described as Tric- 
ula from India, China, and the Ryukyu Archi- 
pelago are now classified in four genera on 



the basis of detailed anatomical data. We 
note that Saganoa species are blind cave- or 
well-dwelling snails; those collected from 
China are stream-dwelling and not blind. 
However, we will not describe a new genus 
until the relevant Japanese snails are studied 
and the data dictate such a course. 

Akiyoshia (Saganoa) chinensis 
Liuetal., 1982 

Holotype. IZAS, HN 798002; Liu, Zhang & 
Chen, 1982: 367, figs. 2-5 (in Liu et at. 1982). 

Type Locality. Guzhang, Hunan Province, 
Sept. 1979 

Assigned Species. China only: Akiyoshia (Sa- 
ganoa) chinensis Liu, Zhang & Chen, 1982; 
A. (S.) yunnanensis Liu, Wang & Zhang, 
1982. N = 2 (in Liu et at. 1982). 

Habitat 

Refer to Tricula gredleri, D87-3. This spe- 
cies was sympatric with T. gredleri and T. 
maxidens. Snails were collected from a vil- 
lage close to the type locality, approximately 5 
km; the type locality is across the valley and 
on the opposite hillside from D87-3. 

Depository 

Specimens are catalogued into the collec- 
tions of ZAMIP, M0010; ANSP 373142, 
A12658. 

Description 

Shell. Shells are minute, smooth, turreted 
with deep sutures and slightly convex whorls 
(Figs. 6A-E, 7A-E, 8). Shells of mature ani- 
mals have 4.5-5.5 whorls; the majority are 
5.0-5.5 whorls. Shell lengths of mature snails 
of 5.0 or more whorls range from 1 .60-1 .86 
mm (Table 3). There is a slight umbilical 
chink. The aperture is sub-circular; there is a 
trace of a varix on some shells. In side view 
the outer lip is slightly sinuate (Fig. 7B). With 
the outer lip down (90° to the horizontal), the 
outer lip is either straight or angled. There are 
no apertural teeth or adapical notches. 

Under SEM the following are observed: (1) 
Spiral microsculpture can be seen at the base 
of the shell (Fig. 7C), and on some whorls 
(Fig. 7D). (2) The protoconch is smooth (Fig. 
7D, E). (3) Breaking open the body whorl re- 
veals no internal columellar teeth or spiral 
ledges (Fig. 8A-E). The columella of the pen- 



156 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 3. Shell measurements (mm) of topotypical Akioyshia chinensis. Mean 
(range). N = number measured. 



standard deviation 





Large class 




Small class 






N = 3 


N = 5 


N = 1 


N = 5 


N = 1 


No. Whorls 


5.0-5.25 


5.5 


4.5 


5.0 


5.5 


Length (L) 


1.77±0.02 


1.84±0.01 


1.64 


1.66±0.04 


1.68 




(1.76-1.80) 


(1.82-1.86) 




(1.60-1.68) 




Width (W) 


0.73±0.01 


0.70±0.02 


0.70 


0.66±0.02 


0.66 




(0.72-0.74) 


(0.68-0.74) 




(0.64-0.68) 




L last three 


1.53±0.02 


1.51 ±0.03 


1.44 


1.44±0.02 


1.40 


whorls 


(1.52-1.56) 


(1.48-1.56) 




(1.40-1.46) 




L body whorl 


0.91 ±0.03 


0.88±0.03 


0.86 


0.85±0.02 


0.80 




(0.88-0.94) 


(0.84-0.92) 




(0.84-0.88) 




L penultimate 


0.36 


0.37±0.01 


0.34 


0.34±0.02 


0.32 


whorl 


No. Var. 


(0.36-0.38) 




(0.32-0.36) 




W penultimate 


0.59±0.01 


0.58±0.01 


0.58 


0.56±0.01 


0.54 


whorl 


(0.58-0.60) 


(0.58-0.60) 




(0.54-0.56) 




W 3rd whorl 


0.48±0.02 


0.49±0.04 


0.48 


0.46 


0.46 




(0.46-0.50) 


(0.44-0.54) 




No Var. 




L aperture 


0.61 ±0.02 


0.59±0.02 


0.60 


0.55±0.02 


0.54 




(0.60-0.64) 


(0.56-0.60) 




(0.52-0.58) 




W aperture 


0.49±0.01 


0.46±0.04 


0.46 


0.31 ±0.01 


0.44 




(0.48-0.50) 


(0.40-0.48) 




(0.44-0.48) 




X 


0.20 


0.15±0.04 


0.16 


0.17±0.08 


0.18 




No Var. 


(0.10-0.22) 




(0.16-0.24) 




У 


0.09±0.03 


0.10±0.04 


0.12 


0.07±0.04 


0.12 




(0.06-0.12) 


(0.04-0.14) 




(0.06-0.08) 





ultimate whorl has minute calcareous nodes 
(Fig. 8F). The inner edge of the inner lip has 
raised calcareous pitted nodes (Figs. 8G, H); 
the inner surface of the aperture is regularly 
pitted (Fig. 8G, I). 

External features. The head is white; there 
are no white granules about the eye. There is 
no Oncomelania-Xype "eyebrow." The oper- 
culum is corneous and paucispiral (Fig. 7F.). 
The inner-lip edge is straight, i.e. the opercu- 
lum shape is sub rectangular, not ovate. The 
inner muscle attachment callus is compara- 
tively small and weakly developed, about 
27% the diameter of the operculum. 

Mantle cavity. Measurements and counts of 
mantle cavity organs are given in Table 4; see 
Figure 9. The gill is central in the mantle cav- 
ity and symmetrical about the osphradium. 
The osphradium is short in females, long in 
males relative to the length of gill. Being long 
in this case is an artifact of the much reduced 
gill. If the gill was the usual length, i.e. filling 
most the length of the mantle cavity, the os- 
phradium would (and should) be classified as 
short. 

There are only 11-14 gill filaments. Gf 2 is 
long in females, medium (or normal) in males. 
The total lengths of the longest gill filaments 



are 0.22 ± 0.03 mm. There is no patch of 
white granules anterior to the osphradium 
near to or partially on the mantle collar. 

Female reproductive system. The body of an 
uncoiled female without head or kidney tissue 
is shown in Figure 10A. Measurements of rel- 
evant organs are given in Table 4. Important 
features are: (1) The gonad (Go) is a simple 
sac located posterior to the stomach. (2) The 
bursa copulatrix (Bu) is clearly seen posterior 
to the palliai oviduct (Ppo). It is small. (3) 
There are three histologically different sec- 
tions to the palliai oviduct; the albumen gland 
(Ppo), capsule gland (Apo-1), and a dense 
white anterior region (Apo-2). These areas 
are clearly discernable in gross dissection of 
living animals. (4) The bursa complex of or- 
gans is shown in Figure 1 1 in the same rela- 
tive position as in Figure 1 0. The usual poma- 
tiopsid seminal receptacle is lost; the function 
of the seminal receptacle is relocated within a 
U-shaped bend or twist of the oviduct (Osr, 
Figs. 10, 1 1). (5) There is a pronounced, elon- 
gated sperm duct (Sdu) connecting the ovi- 
duct (Ov) to the spermathecal duct. The ovi- 
duct enters the albumen gland (Ppo) just 
posterior to the posterior end of the mantle 
cavity (Emc). (6) The spermathecal duct (Sd) 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 157 




FIG. 6. Shells of Akiyoshia chinensis (A-E); Pseudobythinella shimenensis (F-l), and P. chinensis (J-N) 
The length of shell A is 1.77 mm; others are printed to same scale. 



158 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 7. SEM pictures of shell (A-E) and operculum (F) of Akiyoshia chinensis. С Enlargement of base of 
shell showing rough growth lines crossed by fine spiral microsculpture. D, E. The apical whorls are smooth; 
teleoconch roughened sculpture starts at 1 .5 whorls. Spiral microsculpture is seen at 1 .5 to 1 .75 whorls. F. 
Inner surface of operculum to left showing modestly developed attachment pad for muscles. Outer surface 
is to right showing paucispiral coil. 



runs to the anterior end of the mantle cavity to 
open independently of the palliai oviduct. 



Male reproductive system. An uncoiled male 
is shown in Figure 12 without head and with 
kidney tissue removed. Measurements of rel- 



evant organs are given in Table 4. Important 
features are: (1) The anterior lobes of the go- 
nad (Go) are ventral to the posterior chamber 
of the stomach. (2) There are numerous lobes 
of the gonad that drain into a vas efferens 
(Ve). (3) The vas deferens leaves the vas ef- 
ferens at mid-gonad to slightly posterior to 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 159 




FIG. 8. SEM analysis of columella inside body whorl of Akiyoshia chinensis and sculptural attributes of 
columella and inner lip. See text for details. 



mid-gonad to begin coiling as the seminal 
vesicle (Sv) dorsal to the gonad. (4) The pros- 
tate (Pr) overlies the posterior end of the man- 
tle cavity. (5) The prostate consists of dis- 
cernable lobes. (6) The anterior vas deferens 
(Vd 2 ) leaves the prostate close to mid-pros- 



tate. (7) The penis (Fig. 13 C) is simple with a 
long penial filament (Pf) and small papilla 
(Pa). A penial filament is defined as a terminal 
length of penis that is distinctly narrowed 
compared with the rest of the penis. This nar- 
rowing is not simply due to the gradual taper- 



160 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 9. Mantle cavity structures of Akiyoshia chin- 
ensis 



ing of the penis. No ejaculatory duct was 
seen. An ejaculatory duct is a distinctly swol- 
len muscle wrapped section of the vas defe- 



rens found in the base of the penis or neck. 
(8) The base of the penis (Bp) is to the right at 
the snout-neck mid-line (x, Fig. 13A, B) and 
oriented 65°-90° to the mid-line. 

Digestive System. The digestive gland (Di) 
covers the posterior chamber of the stomach 
(Fig. 12). The stomach is shown in Figure 14 
in the same relative position as in Figure 12. 
There is no caecal appendix; the style sac 
(including the intestinal loop) is 48% the 
length of the stomach. 

Radular statistics are given in Tables 5 and 
6. Refer to Figures 15, 16. The cusp formula 
most frequently encountered is 

4(5)-1-(5)4; 4-1-4; 19-24; 16-21. 
1(2)-1(2) 

There are numerous cusps on the marginals 
but there are significantly more on the inner 




Apo-1 



Apo-2 



FIG. 10. Uncoiled female of Akiyoshia chinensis with head and kidney tissue removed (A) and gonad 
showing oocytes (B). 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 161 



TABLE 4. Lengths (mm) or counts of non-neural organs and structures of Akiyoshia chinensis. N = 
number of snails used. Mean ± standard deviation (range). 



Females (N = 4) 



Males (N = 2) 



Body 
Gonad 
Digestive gland 

Posterior palliai oviduct 

(= albumen gland) 
Anterior palliai oviduct 

(= capsule gland) 
Total palliai oviduct 

= OV 
Bursa copulatrix 

BU 
Duct of BU 

BU -e- OV 

Seminal receptacle 

Duct of seminal receptacle 

Mantle cavity 

Gill (G) 

Osphradium (OS) 

OS 4- G 

No. of filaments 

Gf 2 

Gfi 

Total Gf = TGF 

Gf 2 -s- TGF 

Prostate 

Seminal vesicle 
Penis 



3.27: 

(3.04- 

0.40: 

(0.28- 

1.59: 

(1.40- 

0.57: 

(0.50- 

0.52: 

(0.46- 
1.09: 
(0.96- 

0.24: 

(0.20- 

0.43: 

(0.32- 

0.22: 

(0.20- 



0.20 (N = 3) 

3.40) 

0.11 (N = 3) 

0.50) 

0.19 (N = 3) 

1.78) 

0.06 (N = 3) 

0.60) 

0.07 (N = 3) 

0.60) 

0.10 

1.20) 

0.03 

0.28) 

0.08 

0.50) 

0.02 

0.25) 



0.73±0.05 
(0.70-0.80) 

0.52±0.04 
(0.50-0.58) 

0.17±0.04 
(0.12-0.20) 

0.32±0.07 

(0.25-0.39) 

12±1.4 

(11-14) 

0.11 ±0.03 (N = 5)M + F 
(0.08-0.12) 

0.10±0.02(N = 5)M + F 
(0.08-0.14) 

0.22±0.03 
(0.18-0.26) 

0.54±0.08 
(0.46-0.67) 



3.06 
(3.02, 3.10) 

1.36 

(1.30, 1.42) 

0.80 (N = 1) 



0.70 
(0.60, 0.80) 

0.44 
(0.40, 0.48) 

0.21 
0.20, 0.22 

0.48 
(0.42, 0.55) 

11.5 
(11,12) 

0.12 
(0.10,0.14) 

0.11 
(0.10,0.12) 

0.23 
(0.20, 0.26) 

0.48 
(0.46, 0.50) 

0.73 
(0.70, 0.76) 

0.76 (N = 1) 



marginals (X of 21.3 vs. 18.1 for the outer 
marginals). The central tooth is featured in 
Figure 15C, E, H. The central raised "tongue" 
running anterior — posterior on the face of the 
tooth is flanked by deep eye-socket-like de- 
pressions (contrast lack of such deep holes 
on each side of the "tongue" in Pseudo- 
bythinella kunmingensis and P. daliensis 
(Davis et al., 1985: figs. 9, 16). Whether one 



or two basal cusps, they arise from the lateral 
angle of the central tooth. The bases of the 
lateral angles are flared as in the above men- 
tioned species of Pseudobythinella. 

The shape of the lateral teeth is empha- 
sized in Figure 15G, H. Marginals are fea- 
tured in Figure 16. The outermost cusp of the 
outer marginal is extremely elongated, a 
unique feature (Fig. 16F-H). 



162 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Osr 




FIG. 11. Bursa copulatrix complex of organs of 
Akiyoshia chinensis positioned as in Figure 10. 

Nervous system. No data. 

Remarks 

This species differs from those classified as 
Pseudobythinella by possessing a slender 



high-turreted shell without a trace of a tooth or 
node on the columella as examined in the ap- 
erture or within the body whorl. The standard 
seminal receptacle is lost; its function is 
moved into the oviduct. The bursa is a com- 
paritively small sphere clearly visible posterior 
to the palliai oviduct in contrast to the elon- 
gated tubular bursa mostly, if not entirely, bur- 
ied dorsal to the albumen gland in Pseudo- 
bythinella. The central tooth of the radula has 
deep-set socket-like depressions on either 
side of the "tongue" on the face of the tooth; 
these deep holes are lacking in Pseudo- 
bythinella. 

Pseudobythinella Liu & Zhang, 1979 

Synonymy. Erhaia Davis & Kuo, 1 985. 

Type species. Pseudobythinella jianouensis 
Liu & Zhang, 1979: 135-136, figs. 1-3. 



Ast 




0.5 mm 



FIG. 12. Uncoiled male of Akiyoshia chinensis without head and kidney tissue removed. Some lobes of 
gonad are removed to reveal seminal vesicle (Sv). Dashed line shows contour of gonad. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 163 

A * Ast Re 




FIG. 13. Position of base of penis of Akiyoshia chi- 
nensis on neck (A, B) relative to snout-neck mid- 
line (x); penis (C). 



Type locality. Jian'ou, Fujian Province, Peo- 
ple's Republic of China. 




FIG. 14. Stomach of Akiyoshia chinensis positioned 
as in Figures 10 and 12. 



Types. 
types. 



IZAS, FJ767701 , holotype, plus para- 



Designation. 
type species 



By monotypy with designated 



Assigned Species. China only: P. jianouensis 
Liu & Zhang, 1979; Bythinella chinensis Liu & 
Zhang, 1979; B. hubeiensis Liu, Zhang & 
Wang, 1983b; B. gongjianguoi Kang, 1983b; 
B. wufengensis Kang, 1983a; B. watanensis 
Kang, 1983a; P. liui Kang, 1983b; P. shime- 
nensis Liu, Zhang & Chen, 1982; ß. //'/' Kang, 
1985; Erhaia daliensis Davis & Kuo, 1985; E. 
kunmingensis Davis & Kuo, 1 985. N = 11. 

Diagnosis. Bythinella — like shell, minute, with 
smooth, flattened apical whorls. Apical whorls 
may have faint spiral microsculpture. Col- 
umella with spiral glassy thickening that may 
form a considerable keel or spiral ledge; the 
ledge may terminate in the aperture appear- 
ing as a "tooth" on the inner lip. Anatomical 
criteria are based on four species for which 



TABLE 5. Radular statistics for Akiyoshia chinensis. Mean ± standard deviation (range). N = number 
used. In mm except for width of central tooth in цтп. 



Females (N = 7) 



Males (N = 4) 



Shell length 

Radular length 

Radular width 

Total rows of teeth 

No. rows of teeth 

forming 
Central tooth width 



1.71 ±0.09 
(1.60-1.78) 
0.39±0.02 
(0.36-0.42) 
0.04±0.003 
(0.038-0.048) 
90.4±4.3 

(84-96) 

33±1.6 

(31-35) 
10.5±0.5(N = 21; 

(9.5-10.7) 



0.39±0.02 
(0.37-0.40) 
0.04±0.005 
(0.036-0.046) 
84.5±5.1 
(81 -92) 
31.3±1.3 
(30-33) 
9.8±0.5 (N 
(8.9-10.2) 



19) 



164 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 15. Radula of Akiyoshia chinensis. A-E, males; F-H, females. С, E, H. features central teeth; G, H. 
Lateral teeth. D. features inner marginals. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 165 




FIG_ 16. Radula olAkiyoshia chinensis. A, B, F = males; C-E, G, H = females. A. features inner marginals 
b, D. Basal morphology of attachment zone of marginals. F-H. Outer marginals. See text for details. 



166 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 6. Cusp formulae for the radular teeth of Akiyoshia chinensis with the percent of radulae in which 
a given formula was found at least once. 



Central Teeth 



Lateral Teeth 



Inner Marginal 
teeth 



Outer Marginal 
teeth 



4-1-4 
1-1 

5-1-5 
1-1 

5-1-4 
1-1 

5-1-5 
2-2 

4-1-4 
2-2 

5-1-5 
1-2 



57% 



57% 



29% 



29% 



14% 



14% 



4-1-4 



100% 



14 


— 




14% 


15 


— 




14% 


16 


— 




29% 


17 


— 




71% 


18 


14% 




43% 


19 


43% 




71% 


20 


71% 




43% 


21 


86% 




29% 


22 


71% 




— 


23 


86% 




— 


24 


57% 




— 


25 


29% 






X* = 


21.3±1.8 


18.1±1.7 


N = 70 


N 


= 70 



*Mean ± standard deviation of cusp number for all teeth counted. 



TABLE 7. Shell measurements (mm) of Pseudo- 
bythinella chinensis. Mean ± standard deviation 
(range). Number measured = 5. All shells of 
mature animals had 4.5 whorls. 

Length (L) 1.77±0.06 (1.68-1.84) 

Width (W) 0.85±0.03 (0.83-0.90) 

L last three whorls 1 .64 ±0.06 (1 .56- 1 .68) 

L body whorl 1 .07±0.06 (1 .00- 1 .1 6) 

L penultimate whorl 0.36±0.01 (0.34-0.36) 

W penultimate whorl 0.69±0.07 (0.56-0.74) 

L 3rd whorl 0.53±0.01 (0.52-0.54) 

L aperture 0.70±0.03 (0.66-0.72) 

W aperture 0.54±0.04 (0.52-0.60) 

x 0.16±0.03 (0.12-0.20) 

y 0.04±0.01 (0.02-0.06) 



we have data; the two species of this study 
and P. daliensis and P. kunmingensis de- 
scribed in Davis et al. (1985). The female go- 
nad is a simple tube or tube with low lobes. 
The oviduct has a wide 360° loop dorsal to 
the bursa copulatrix. There is the standard 
generalized seminal receptacle. Hydrobiid — 
type central tooth. The lateral tooth lacks a 
pronounced intermediate cusp. A lateral tooth 



has a pronounced curved process projecting 
posterior from the face of the tooth. The pro- 
cess is slight, or if large it is straight in Akiy- 
oshia chinensis. Stomach without caecal ap- 
pendix. There is a tendency for hypertrophy of 
the radular sac. 

Pseudobythinella chinensis (Liu and Zhang) 

Holotype. IZAS, HN766602, Liu & Zhang, 
1979: fig. 4. 

Type locality. Lengshuijiang, Xinhua, 
Hunan Province; July 1976. (approxi- 
mately 27°4'N, 111°25'E) 

Synonymy. Bythinella chinensis Liu & Zhang, 

1979: 134, 136. 

Erhaia chinensis, Davis et al., 1985 
Pseudobythinella chinensis, this paper 

Habitat 

Snails were collected from Shen Keng Vil- 
lage, Sanhe Town, Lingxian County, Zhu 
Zhou Prefecture; 26°17'18" N, 113°31'50" E; 
Figure 1, site 6. The field collection number 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 167 




FIG. 17. SEM picture of shells (A-E) and opercula (F, G) of Pseudobythinella chinensis. Note that in A, no 
"tooth" is seen on columella. Tilting shell to look inside body whorl reveals a swelling or "tooth" on columella. 
C, D reveal no spiral microsculpture. See text for details. The apical whorls are smooth (E). Inner surface of 
operculum (F); outer surface (G). 



168 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 18. SEM of shells of Pseudobythinella chinensis broken to show raised spiral ridge and "tooth" on 
columella inside body whorl. See text for discussion of sculpture on columella and inner lip. 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 169 



TABLE 8. Lengths (mm) or counts of non-neural organs and structures of Pseudobythinella chinensis. N 
= number of snails used. Mean ± standard deviation (range). 



Females (N = 2) 



Males (N = 1) 



Body 
Gonad 
Digestive gland 

Posterior palliai oviduct 

(= albumen gland) 
Anterior palliai oviduct 

( = capsule gland) 
Total palliai oviduct 

= OV 
Bursa copulatrix 

BU 
Duct of BU 
BU + OV 

Seminal receptacle 

Duct of seminal receptacle 

Mantle cavity 

Gill (G) 

Osphradium (OS) 

OS - G 

No. of filaments 

Gf 2 

Gfi 

Total Gf = TGF 
Gf 2 ■*■ TGF 
Prostate 
Seminal vesicle 
Penis 



3.68 

(3.42,3.94) 
0.32 

(0.30,0.34) 
1.72 
(1.70,1.74) 



1.20 
(No. Var.) 

0.38 
(0.36,0.40) 

0.32 

(0.30,0.33) 
0.04(N = 1) 

0.12(N = 1) 



3.28 
0.90 
1.80 



0.52 
0.24 



12 
0.10 
0.10 
0.20 
0.50 
0.64 
0.40 
0.52 



assigned was D85-81. Snails came from a 
small stream that flowed between the peaks 
of big mountains; the elevation was 500 m. 
The stream was heavily shaded by vegetation 
lining the banks. The stream was 20-30 cm 
wide, 10-20 cm deep with a bottom paved 
with small rocks, leaves and mud. 

Depository 

Specimens are housed at ZAMIP, M0011; 
ANSP, 373139, A1 2655. 

Description 

Shells. Shells are minute, smooth, and 
conic-turreted (Figs. 6J-N; 17A-E; 18A-H). 



They are 4.5 whorls ranging in length from 
1 .68 to 1 .84 mm (Table 7); they are umbilicate 
with a sub-ovate to sub-circular aperture. The 
sutures are moderately deep; whorls with 
slightly convex whorls. No tooth is visible in 
the aperture in 60% of the shells while a thick- 
ening on the columella is seen in the aperture 
of 40% of the shells. Tipping the aperture up, 
one can see a considerable tooth-like swell- 
ing on the columella, the terminus of the pro- 
nounced internal columellar keel. In side view 
the outer lip is straight to slightly sinuate; in 
the contrasting view the inner lip is straight 
(outer lip down, 90° to the horizontal). 

SEM analyses show the apical whorls to be 
smooth (Fig. 17E). With the aperture tilted, 



170 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




surements and counts of relevant organs are 
given in Table 8. The gill is centrally located in 
the mantle cavity and the osphradium (Os) is 
centered against the gill. The osphradium by 
definition is long but this is an artifact of the 
much reduced size of the gill. The usual po- 
matiopsid gill nearly fills the length of the 
mantle cavity. On this criterion the osphra- 
dium is here considered short (circa 0.32 
ratio). The Gf 2 is medium (= normal) length. 
The length of the longest gill filament is ap- 
proximately 0.20 mm. 



Emc 




FIG. 19. The buccal mass of Pseudobythinella chi- 
nensis with pronounced radular sac (A) and re- 
flected mantle showing mantle cavity organs (B). 



the columeller "tooth" is seen (Fig. 17C, D); 
note the numerous calcareous pitted pustules 
on the columella and "tooth". With the shell 
broken open, it is seen that the "tooth" seen 
in the aperture is the terminus of a pro- 
nounced columellar keel starting at mid-body 
whorl and spiralling down to the aperture (Fig. 
1 8A-E). An enlargement of the area of highest 
concentration of pitted calcareous pustules 
on the "tooth" (Fig. 18G) is given in Fig. 18H. 
The entire surface of the keel is roughened 
with calcareous pustules or micro-ridges. 

External features. The head-neck is white; 
there are no white granules about the eyes. 
The operculum is corneous and paucispiral 
(Fig. 17F, G). The shape is sub-rectangular. 
The columeller edge is somewhat concave, 
especially where the operculum passes over 
the columeller "tooth" (Fig. 17C). The callus 
on the inner surface is weakly developed, but 
wide, 69% the width of the operculum (Fig. 
17F). 

Mantle cavity. The opened and reflected 
mantle cavity is shown in Figure 19B. Mea- 



Female reproductive system. The body of an 
uncoiled female snail is shown in Figure 20 
without head and with kidney tissue removed. 
Measurements of relevant organs are given in 
Table 8. Important features are: (1) The go- 
nad (Go) is posterior to the stomach (Pst). (2) 
The gonad consists of a simple sac with four 
to five small undivided lobes protruding from 
the posterior end. (3) The bursa (Bu) is either 
completely covered by the albumen gland 
(Ppo) or only protrudes slightly posterior to 
the albumen gland (as in Fig. 20). (4) The 
oviduct (Ov) makes a pronounced bend 
pressing against the posterior edge of the 
bursa (Bu). (5) The spermathecal duct (Sd) 
runs to the anterior end of the mantle cavity to 
open independently of the palliai oviduct. (6) 
The sperm duct (Sdu) is unique in that it 
branches off the spermathecal duct near the 
anterior end of the spermathecal duct, far for- 
ward of the posterior end of the mantle cavity 
(Fig. 21). (7) The seminal receptacle (Sr) is 
unique in that it branches from the sperm duct 
(Sdu), not the oviduct, and in that it is entirely 
anterior to the posterior end of the mantle 
cavity (Fig. 21). 

Male reproductive system. An uncoiled male 
is shown in Figure 22 without head or kidney 
tissue. Part of the gonad (Go) has been cut 
away to reveal the seminal vesicle (Sv) coiled 
dorsal to it. Measurements of relevant organs 
are given in Table 8. Important features are: 
(1 ) The gonad is posterior to the stomach. (2) 
There is no vas efferens in the usual sense. 
Thick lobes arise from a wide collecting gutter 
that functions as a vas efferens. (3) The sem- 
inal vesicle (Sv) arises from mid-gonad and 
makes a small knot dorsal to the gonad. (4) 
The posterior vas deferens (Vd,) runs as a 
wide tube, swollen with sperm, from the sem- 
inal vesicle to become a slender duct only at 
the style sac. (5) The prostate overlies the 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 171 

Ast 



Sts 




FIG. 20. Uncoiled female Pseudobythinella chinensis with head and kidney tissue removed. 




Ov Osd 



Osd 



FIG. 21. Details and variation of bursa copulatrix 
complex of organs of Pseudobythinella chinensis. 
Figure 21 A is in same orientation as in Figure 20. In 
B, oviduct cut and reflected in direction of arrow to 
show opening of oviduct (Oov) that attaches to al- 
bumen gland 



172 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




Eme 



0.5 mm 

FIG. 22. Uncoiled male of Pseudobythinella chinensis without head or kidney tissue. Some of lobes of 
anterior gonad are cut away ( — ) to show knot of seminal vesicle (Sv) dorsal to gonad. 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 173 

TABLE 9. Radular statistics for Pseudobythinella 
chinensis. Mean ± standard deviation (range). N 
= number used. In mm except for width of 
central tooth in p.m. 




Sex Unknown (N = 4) 



Shell length 

Radular length 

Radular width 

Total rows of teeth 

No. rows of teeth forming 

Central tooth width 



1.79±0.001 
0.48±0.01 
0.06±0.004 
97.3±6.7 (N = 3) 
10.3±4.0 
12.0±0.5(N = 9) 




(Fig. 24). The most commonly encountered 
cusp formula is (4)5-1-5(4) ; 4(5)-1-(5, 6)4; 
1-1 

24-29; 17-22. 

The inner marginal teeth have significantly 
more cusps(mean of 26) than do the outer 
marginals (mean of 20). The two outer cusps 
of the outer marginals are specialized to form 
a pincer-like process (Fig. 24G, H). The 
"tongue" on the face of the central tooth is 
broad and is not flanked by deep-set holes 
(Fig. 24A, B). 

Nervous system. No data. 



0.2r 



FIG. 23. A. The orientation of base of penis (Bp) of 
Pseudobythinella chinensis to snout-neck mid-line 
(x) and lobes of eyes (a). B. Penis. 



posterior end of the mantle cavity. (6) The 
anterior vas deferens (Vd 2 ) separates from 
the prostate at mid-prostate. (7) The penis 
(Fig. 23B) has an enormous muscular ejacu- 
latory duct (Ej) in the base that continues to 
coil for a short distance in the neck. (8) The 
penis has an enormously elongated papilla, a 
unique feature. (9) The base of the penis (Bp, 
Fig. 23A) arises to the right of the snout-neck 
mid-line x and at 90° to it. 

Digestive system. The digestive gland (Di) 
covers the posterior chamber of the stomach 
(Pst, Figs. 20, 22). The radular sac (Rs) loops 
up over the buccal mass (Bm, Fig. 19 A). 
Radular statistics are given in Tables 9, 10. 
The radula is minute, with an extraordinary 
number of rows of teeth (97 per 0.48 mm) 



Remarks 

This species differs from the other three for 
which we have anatomical data by the follow- 
ing: (1) The bursa may somewhat protrude 
posterior to the albumen gland. (2) the oviduct 
makes a pronounced bend near the bursa; 
the ascending and descending arms of the 
bend are pressed together (contrast the open 
360° loops or complex oviduct loops of the 
other species). (3) The sperm duct is elon- 
gated and extends anterior to the posterior 
end of the mantle cavity to enter the sper- 
mathecal duct, a unique character-state. (4) 
The seminal receptacle branches off the 
sperm duct, not the oviduct, a unique charac- 
ter-state. (5) There is no vas efferens. (6) The 
penis has an extremely elongated papilla, a 
unique character-state. 

Pseudobythinella shimenensis Liu, 
Zhang & Chen 

Holotype. IZAS, HN797904; Liu et al., 1982: 
254, 256, fig. 1 . 



174 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 24. Radula of Pseudobythinella chinensis. B. Central and inner marginal teeth emphasized. С, D. 
Details of morphology of lateral teeth. E, F. Inner marginals emphasized. G, H. Outer marginals. See text for 
details. 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 175 

TABLE 1 0. Cusp formulae for the radular teeth of Pseudobythinella chinensis with the percent of the four 
radulae in which a given formula was found at least once. 



Central Teeth 



Lateral Teeth 



Inner Marginal 
Teeth 



Outer Marginal 
Teeth 



5-1-5 75% 
1-1 

5-1-4 



1-1 

4-1-4 
1-1 



50% 



25% 



4-1-4 


100% 


17 


— 


75% 


5-1-5 


75% 


18 


— 


75% 


6-1-5 


50% 


19 


— 


75% 


4-1-5 


50% 


20 


— 


75% 


5-1-4 


50% 


21 


— 


50% 


6-1-4 


25% 


22 


— 


50% 






23 


— 


25% 






24 


25% 


25% 






25 


100% 


— 






26 


100% 


— 






27 


75% 


— 






28 


25% 


— 






29 


25% 


— 






X* = 


= 26±1.0 


20.0±2.1 






N = 


40 


N = 33 



'Mean ± standard deviation of cusp number for all teeth counted. 



Type locality. Shimen, Hunan Province, Oc- 
tober 1979. 

Synonymy. Pseudobythinella shimenensis 
Liu, Zhang & Chen, 1982: 254-256. 
Pseudobythinella shimenensis Davis et al., 
1985:68. 

Habitat 

Snails were collected from Qingguandu Vil- 
lage, Nanzhen Town, Shimen County, 
Changde Prefecture; 29°56'36" N, 1 10°41 '42" 
E; Figure 1 , site 3; topotypes. Snails were col- 
lected from under stones and leaves at the 
edge of a small pool of a stream. The water 
was clean, clear, cool. 

Depository 

Specimens are housed at ZAMIP, M0012, 
ANSP 373137, A12653. 



TABLE 1 1 . Shell measurements (mm) of Pseudo- 
bythinella shimensis. Mean ± standard deviation 
(range). Five shells measured, all 4.0 whorls. 

Length (L) 1 .98±0.04 (1 .94-2.04) 

Width (W) 1.17±0.02 (1.14-1.20) 

L last three whorls 1 .95±0.04 (1 .92-2.02) 

L body whorl 1 .44±0.03 (1 .41 - 1 .48) 

L penultimate whorl 0.38±0.01 (0.38-0.40) 

W penultimate whorl 0.82±0.02 (0.80-0.84) 

L 3rd whorl 0.48±0.02 (0.45-0.50) 

L aperture 0.99±0.02 (0.96-1.00) 

W aperture 0.76±0.03 (0.73-0.80) 

x 0.24±0.12 (0.10-0.38) 

y 0.07±0.04 (0.04-0.12) 



Description 

Shells. Shells are minute, smooth, ovate with 
flattened apex (Figs. 6F-I, 25, 26). Shell mea- 
surements are given in Table 1 1 ; shell lengths 
range from 1 .94 to 2.04 mm for shells of 4.0 



176 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 25. SEM photographs of shells of Pseudobythinella shimenensis. A. Note slight swelling or tooth on 
columella. С Apical whorls showing spiral microsculpture starting at 0.5 whorls. D. Aperture rotated to 
expose fully tooth on columella. E, F. Columella highly magnified to show sculpture pattern on inner lip, 
columella, and tooth. 



whorls. The aperture is broadly ovate to sub- 
circular. The sutures are deep and the whorls 
shouldered, convex. The inner lip is straight to 
saddle-shaped with a narrow umbilicus above 
the center of the saddle of 33%; no umbilicus, 
66%. The columella has a tooth plainly in view 



in the aperture. The face of the body whorl is 
flattened in most specimens. The outer lip is 
straight to slightly arched (Fig. 25B). In side 
view, the adapical outer lip is fused to the 
body whorl in 40%; separated by 0.05 mm ± 
0.03 mm (N = 5) from the body whorl in 60%. 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 177 




FIG. 26. SEM photographs of shells of Pseudobythinella shimenensis broken open to reveal raised spiral 
ridge on columella of body whorl that terminates as tooth. C, D. Large scale-like sculptural plates on 
columella at aperture contrast with thin line-like raised ridges on spiral ridge within body whorl (G) 



178 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




В 




FIG. 27. Head (A) and penis (В) of Pseudo- 
bythinella shimenensis. A. Relationship of base of 
penis to mid-line of snout-neck (x) and to posterior 
end of eye lobes (a). 



The inner lip is straight (outer lip down 90° to 
horizontal). 

SEM examination shows that the tip of the 
apical whorl is smooth with fine spiral sculp- 
ture seen starting in less than half a whorl 
(Fig. 25C). Tilting the aperture somewhat, the 
columellar "tooth" is clearly seen (Fig. 25D- 
F). It is covered with raised calcareous lap- 
pets (Fig. 25E), contrasted with the pustules 
seen in P. chinensis. Upon breaking open 
several shells, it is seen that the "tooth" is the 
terminus of a columellar keel that starts mid- 
body whorl (Fig. 26A, B, E-G). The calcare- 
ous lappets begin on the columellar keel and 
extend to the aperture (Fig. 26D, G); above 
the keel there are minute raised calcareous 
ridges (Fig. 26G). 



External features. The head is white with a 
small cluster of white granules about the me- 
dial edge of the eye, i.e. an "eyebrow" (Ebr, 
Fig. 27). There is a scattering of white gran- 
ules posterior to the eyebrow at the side of the 
neck. The operculum is corneous, paucispiral 
and kidney-bean shaped with the columel- 
lar — side concavity corresponding to the tooth 
on the columella (Fig. 28). On the inner sur- 
face of the operculum the attachment pad for 
the muscle has a pronounced ridge (Fig. 28A, 
C). The pad is relatively narrow, some 40% 
the width of the operculum. 

Mantle cavity. The reflected mantle showing 
mantle cavity organs is given in Figure 29; 
measurements and counts are given in Table 
12. There are relatively few gill filaments (12- 
13) yet these take up 78% the length of the 
mantle cavity. The osphradium is mid-gill and 
is long. Gf 2 is long; the longest gill filaments 
are 0.36 mm long. There is no cluster of white 
granules just anterior to the osphradium close 
to the neck (Ne) — mantle collar (Ma) junction. 

Female reproductive system. The body of an 
uncoiled female with head and kidney tissue 
removed is shown in Figure 30. Measure- 
ments and counts of organs are given in Ta- 
ble 12. Important features, to note are: (1) 
The gonad is located posterior to the stom- 
ach. It is a single small sac that has three or 
four small protruding lobes created by pres- 
sure of individual oocytes at the posterior end 
of the gonad. (2) The albumen gland (Ppo) 
covers most of the bursa copulatrix (Bu) and 
extends beyond the bursa to cover most of 
the style sac. The albumen gland curves 
around the bursa leaving the posterior end of 
the bursa exposed. Only in a few cases is the 
bursa completely covered by the albumen 
gland. (3) The spermathecal duct (Sd) is 
tightly pressed to the palliai oviduct and 
opens into the mantle cavity close to the an- 
terior end of the capsule gland (Apo). (4) The 
bursa copulatrix complex of organs is shown 
in Figure 31 oriented in the same relative po- 
sitions as in Figure 30. The bursa is short. (5) 
The oviduct enters (Oov) the albumen gland 
posterior to the posterior end of the mantle 
cavity (Emc). The sperm duct is short and 
branches off the spermathecal duct posterior 
to the posterior end of the mantle cavity. (6) 
The duct of the seminal receptacle (Dsr) 
branches off the oviduct posterior to the 
sperm duct (Fig. 31). The seminal receptacle 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 179 



FIG. 28. Opercula of Pseudobythinella shimenensis. A, С Inner surface. B, D. Outer surface. Note prom- 
inent ridge on inner surface running along inner edge of attachment pad. 



180 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 12. Lengths (mm) or counts of non-neural organs and structures of Pseudobythinella 
shimenensis. N = number of snails used. Mean ± standard deviation (range). 





Females (N = 5) 


Males (N = 1) 


Body 


3.62±0.34 
(3.26-4.04) 


3.94 


Gonad 


0.47±0.09 
(0.34-0.60) 


2.00 


Digestive gland 


1.78±0.15 
(1.68-2.04) 


2.10 


Posterior palliai oviduct ( = albumen gland) 


— 


— 


Anterior palliai oviduct ( = capsule gland) 


— 


— 


Total palliai oviduct 


1.52±0.09 


— 


= OV 


(1.40-1.60) N = 4 




Bursa copulatrix 


0.49±0.11 


— 


BU 


(0.36-0.60) N = 4 




Duct of BU 


0.26 (N = 2) 
No. var. 




BU -e- OV 


0.33±0.08 
(0.23-0.40) N = 4 


— 


Seminal receptacle 


0.11 ±0.02 
(0.08-0.13) N = 4 


— 


Duct of seminal receptacle 


0.16±0.05 
(0.10-0.22) N = 4 


— 


Buccal Mass 


0.46 (N = 1) 


— 


Mantle cavity 


0.98±0.11 
(0.84-1.10) N = 4 


.90 


Gill (G) 


0.76±0.07 
(0.70-0.84) N = 4 


.70 


Osphradium (OS) 


0.31 ±0.08 
(0.20-0.40) 


.28 


OS - G 


0.40±0.08 
(0.29-0.48) N = 4 


.40 


No. of filaments 


12.3±0.5 
(12-13) 


.28 


Gf 2 


0.20 (N = 2) 


— 


Gf 1 


0.16 (N = 2) 


— 


Total Gf = TGF 


0.36 (N = 2) 


— 


Gf 2 -s- TGF 


0.55 (N = 2) 


— 


Prostate 


— 


1.00 


Seminal vesicle 


— 


.80 


Penis 


— 


.90 



TABLE 13. Radular statistics for Pseudobythinella shimenensis. Mean 
= number used. In mm except for width of central tooth in |xm. 



standard deviation (range). N 



Sex Unknown 



Shell length 

Radular length 

Radular width 

Total rows of teeth 

No. rows of teeth forming 

Central tooth width 



2.1 ±0.12 (1.9-2.2) N = 9 
0.79±0.46 (0.71 -0.84) N = 5 
0.071 ±0.005 (0.064-0.076) N = 6 
113±1.7 (111-115) 
9±3(6-14) 
14.1 ±0.82 (13.2-15.3) N = 13 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 181 




FIG. 29. Mantle cavity of Pseudobythinella shime- 
nensis. Mantle cut and reflected to show mantle- 
cavity structures. 




FIG. 31. Details and variation of bursa copulatrix 
complex of organs of Pseudobythinella shimenen- 
sis. Figure 31 A is in same orientation as in Figure 
30. B. Bursa cut away to show seminal receptacle 
(Sr) and oviduct coil (Coi). 




Qj 0.5mm 

FIG. 30. Uncoiled female Psuedobythinella shimenensis with head and kidney tissue removed. 



182 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 
Di 




FIG. 32. Uncoiled male of Pseudobythinella shimenensis without head or kidney tissue. Some lobes of 
gonad removed (iine with arrows) to reveal seminal vesicle (Sv). 



and the duct of the seminal receptacle are 
well developed. (7) Posterior to the duct of the 
seminal receptacle the oviduct makes an ir- 
regular loop or double coil that is dorsal to the 
bursa. In Figure 31 A, the coil has been pulled 
out a little from beneath (dorsal to) the bursa 
to show the pattern of coiling. The coil is 
shown in actual position in Figure 30. 

Male reproductive system. The body of an un- 
coiled male is shown in Figure 32 with head 
and kidney tissue removed. Measurements 
and counts of organs are given in Table 12. 
Important features to note are: (1) The gonad 
covers the stomach. (2) A portion of the go- 
nadal lobes has been cut away to show the 
seminal vesicle (Sv) that coils regularly dorsal 
to the gonad. (3) There is a well-defined vas 
efferens (Ve) with the posterior vas deferens 
arising from the vas efferens at mid — to 



slightly posterior to mid-gonad. (4) There are 
a number of bundles of testicular lobes arising 
from the vas efferens. (5) The prostate over- 
lies the posterior end of the mantle cavity 
(Emc). (6) The anterior vas deferens (Vd 2 ) 
leaves the prostate (Pr) close to the posterior 
end of the mantle cavity. (7) The penis is sim- 
ple and without discernable papilla (Fig. 27B). 
(8) The base of the penis (Bp) arises from the 
neck to the right of the snout-neck mid-line (x) 
and at 90° to it (Fig. 27A). There is no dis- 
cernable ejaculatory duct. 

Digestive system. The digestive gland covers 
the posterior chamber of the stomach of fe- 
males (Di, Fig. 30), and the entire stomach in 
males (Fig. 32). The radular sac (Rs, Fig. 33) 
is highly elongated, coiling dorsal to the buc- 
cal mass (Bm). 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 183 




0.33mm 

FIG. 33. Dorsal aspect of buccal mass of Pseudo- 
bythinella shimenensis with dorsal part of nerve 
ring. Note radular sac (Rs) coiling dorsally over 
buccal mass (Bm). 



Radular statistics are given in Tables 13 
and 14. The most commonly encountered 
cusp formula is 



4(5H-(5)4: 2(3H- 
1-1 



3(4); 17-20, 16-18. 



34C, D, F and 35A, В, E. Two points are 
important. (1) The basal process of the lateral 
tooth is prominent and curved towards mid- 
radula as also seen in P. chinensis but in con- 
trast to the weak or slighly developed straight 
basal process seen in Akiyoshia chinensis. 
(2) As in the above mentioned taxa, there is a 
gradation in size of the cusps on the lateral 
tooth. The "1 " of the 3-1 -3 is not considerably 
larger than the flanking cusps. Inner marginal 
teeth (Figs. 34B, C, F; 35C, D, E, F) and outer 
marginals (Figs. 34C, E; 35C, G) have numer- 
ous small cusps. No one cusp is extra long 
(i.e. with derived specialization). 

Nervous system. A segment of the nervous 
system is shown in Figure 33. The cerebral 
commissure (Cc) is elongated. The dorsal 
nerve ring is moderately concentrated (RPG 
of 0.38, Table 15). Otherwise the dorsal as- 
pect of the nerve ring is typical for the Tricu- 
linae. 

Remarks 

This species differs from the others for 
which we have anatomical data as follows: (1 ) 
The glands about the eye form an "eyebrow". 
Pseudobythinella chinensis and P. daliensis 
lack any glands about the eyes; there is a 
scatter of glands in P. kunmingensis. (2) A 
tooth is clearly visible in the aperture of the 
shell. (3) The bursa is an elongated tube; it is 
an elongated ovoid sac in the other species. 
(4) The penis lacks an ejaculatory duct. 

Pseudobythinella daliensis differs from the 
other three species by having a small, not 
minute shell. It does not have an elongated 
radular sac as do the others. 

Pseudobythinella kunmingensis differs 
from the other three species by having a scat- 
ter of glands about the eyes. The spermath- 
ecal duct shares a common opening with the 
capsule gland (Apo) (not shown in Fig. 31 
where the spermathecal duct is cut short). 



There is a very large number of rows of teeth 
for the comparatively short length of radula 
(113 per 0.79 mm). The radula is illustrated 
in Figures 34, 35. Fig. 34B, D, E feature the 
central tooth. The face of the tooth is moder- 
ately raised as a tongue with a slight concav- 
ity on either side extending beneath the basal 
cusps. The lateral angles have the flared 
ends typical of Pseudobythinella. 
The lateral teeth are featured in Figures 



TRICULINAE 

Pachydrobiini Davis & Kang, 1990 

Type genus. Pachydrobia Cross & Fischer, 
1876 

Diagnosis. Genera of Triculinae in which the 
spermathecal duct bypasses the pericardium 
and the oviduct does not make a closed 360° 
twist. With the exception of Wuconchona, 



184 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 





/4X 




FIG. 34. Radula of Pseudobythinella shimenensis featuring central and lateral teeth (B-D, F) and outer 
marginal teeth (E). 



there ¡s a sperm duct. The seminal receptacle 
arises from the bursa or the duct of the bursa 
in the plesiomorphic state; the seminal recep- 
tacle is lost in the derived state and its func- 
tion taken over by new structures. 

Genera assigned. Guoia, Halewisia, Neotric- 
ula, Pachydrobia, Robertsiella, Jinhongia, 
Gammatricula, Wuconchona (N = 8). 



The Lithoglyphopsis Problem 

Considering all Asian genera of freshwater 
rissoacean snails, it has been especially im- 
portant to locate and study the type species of 
Lithoglyphopsis for two reasons. (1) Lithogly- 
phopsis was the taxon that caused early Eu- 
ropean workers to include within the family 
Hydrobiidae those Asian taxa now known to 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 185 




FIG. 35. Radula of Pseudobythinella shimenensis featuring entire lateral tooth (А, В, E), inner marginals 
(C-F), outer marginals (G). 



be Pomatiopsidae: Triculinae. (2) The snail 
transmitting Schistosoma mekongi Voge et 
al., 1978, was originally described as Litho- 
glyphopsis apena Temcharoen, 1971. 

Considering the first, the shells of certain 
Chinese species from Hunan Province so 
much resembled shells of European Lithogly- 
phus that they were described as species of 



Lithoglyphus (Gredler, 1881, 1886; Moellen- 
dorff, 1888). Subsequently Thiele (1928) 
noted that the morphology of the central tooth 
of L. modestus differed slightly from that of 
European Lithoglyphus and he therefore es- 
tablished the genus Lithoglyphopsis, with L. 
modestus Gredler, 1886, as its type species 
by original designation. However, overall shell 



186 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 1 4. Cusp formulae for the radular teeth of Pseudobythinella shinenensis with the percent of the 
radulae in which a given formula was found at least once. 



Central Teeth 


Lateral Teeth 


Inner Marginal 
Teeth 


Outer Marginal 
Teeth 


4-1-4 
1-1 


80% 


2-1-3 


60% 


14 


— 




20% 


5-1-5 
1-1 


40% 


2-1-4 


60% 


15 


— 




40% 


5-1-4 
1-1 


20% 


3-1-3 


60% 


16 


— 




60% 


4-1-4 
1-1 


20% 


3-1-4 


40% 


17 

18 
19 
20 

21 


80% 

100% 

100% 

60% 

40% 




100% 

80% 
40% 
20% 










X* = 
N = 


= 18.7±1.2 
50 


17.2±1.3 
N = 49 



*Mean ± standard deviation of cusp number for all teeth counted. 



TABLE 15. Lengths of neural structures of Pseu- 
dobythinella shimenensis. The mean is given and 
with data when N = 2. N = number used. 



Cerebral ganglion 


0.20 (N = 2) 




(No. Var.) 


Cerebral commissure 


0.09 (N = 2) 




(0.08, 0.10) 


Pleural ganglion 




Right (1)* 


0.10 (N = 1) 


Left 


— 


Pleuro-supraesophageal 


0.12 (N = 1) 


connective (2)* 




Pleuro-subesophageal 


— 


connective 




Supraesophageal ganglion (3)* 


0.10 (N = 1) 


Subesophageal ganglion 


— 


Osphradio-mantle nerve 


0.04 (N = 1) 


RPG ratio* = 2 + 1+2 + 3 


0.38 (N = 1) 



and radular characters persuaded Thiele 
(1928) to include Lithoglyphopsis of the Hy- 
drobiidae along with 1 2 other Asian Triculinae 
genera within the Tribe Lithoglypheae along 
with European Lithoglyphus. As late as 1974, 
Brandt included the Asian genera in question 
in the family Hydrobiidae, subfamilies Triculi- 
nae, Cohilopinae, Rehderiellinae, and Litho- 
glyphinae. 

As for the second, Davis et al. (1976) stud- 
ied this species and stated that it was most 
closely related to Tricula, especially Tricula 



burchi Davis, 1968b; they stated that Tricula 
might be a suitable genus for "L" apena. In 
1980, Davis referred L. aperta to Tricula. 
Davis et al. (1976) pointed out that on the 
basis of both shell and radula, Tricula aperta 
could not be considered a species of Litho- 
glyphopsis. However, the questions have re- 
mained: What is Lithoglyphopsis? To which 
genera is Lithoglyphopsis most closely re- 
lated? Can one establish once and for all that 
Lithoglyphopsis is a member of the Triculinae, 
not a member of the Hydrobiidae: Lithoglyph- 
inae? What is the potential for species of 
Lithoglyphopsis to transmit a species of 
Schistosoma? 

As a result of our studies in Hunan Prov- 
ince, it was clear that species historically con- 
sidered to be Lithoglyphus in Hunan belong to 
two genera: Lithoglyphopsis and a new genus 
described here as Guoia. 



Guoia Davis & Chen, genus nov. 

Type Species. Lithoglyphus viridulus Moel- 
lendorff, 1888 

Etymology. Named for Dr. Guo Yuan Hua, 
Institute of Parasitic Diseases, China National 
Center for Preventive Medicine, Shanghai, for 
his tireless efforts to discover and understand 
species of snails involved in disease trans- 
mission. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 187 



Diagnosis. Shells small (<3.6 mm long), and 
globose-conic. The spermathecal duct opens 
into the rear of the mantle cavity. A long, slen- 
der sperm duct connects the spermathecal 
duct (at a position close to the posterior end of 
the mantle cavity) to the oviduct close to 
where the oviduct opens into the albumen 
gland. Duct of the bursa is massive, running 
directly anterior from the large bursa to form 
the short spermathecal duct opening at the 
rear of the mantle cavity. There is no seminal 
receptacle; sperm are stored in a swelling of 
the oviduct just posterior to the juncture of the 
sperm duct and oviduct, or they are stored in 
an outpocketing of the sperm duct at the junc- 
ture to the oviduct. There is a thin corneous 
stylet at the tip of the penis. The penis has a 
glandular lobe. The ejaculatory duct is mas- 
sive, extending posteriorly along the neck 
from the base of the penis. Radula Tricula- 
like. 

Relationships. A member of the Neotricula 
clade by virtue of (1) the oviduct travels from 
gonad to albumen gland straight, without twist 
or coil (contrast the Tricula clade), and (2) the 
spermathecal duct does not enter the pericar- 
dium. The closest generic similarity is with 
Robertsiella of Malaysia in that (1) the penis 
of the males has a similar stylet, and (2) the 
spermathecal duct-duct of bursa-bursa con- 
nections and relative postitions are the same. 
There are differences. In Robertsiella, the 
seminal receptacle is encapsulated in the 
muscular wrapped duct of the bursa {Guoia 
lacks a seminal receptacle). A section of the 
oviduct of Guoia serves as a seminal recep- 
tacle. The shell of Robertsiella is ovate-conic, 
not globose-conic. The sperm duct of Guoia is 
elongated, twisting over the bursa; it is very 
short in Robertsiella. The duct of the bursa is 
very short in Robertsiella, elongated in Guoia. 

Assigned Species. 

Lithoglyphus fuchsianus Moellendorff, 
1885 
Lithoglyphus viridulus Moellendorff, 1888 



cies and named "L. modesta" as the type 
species. Yen (1929) placed all Chinese taxa 
described as Lithoglyphus in the genus Litho- 
glyphopsis including L. liliputanus Gredler, 
1881 from "Kwangtung" (= Guangdong). 
The shell of this last species is a miniature 
version of L. modestus. Until the anatomy is 
known its generic placement is incertae sedis. 

Guoia viridulus (Moellendorff, 1888). 

Lithoglyphus viridulus Moellendorff, 1888: 
141, pi. 4, fig. 6, 6a-b 

Lithoglyphus viridulus (Moellendorff, 1888). 
Thiele 1928 

Lithoglyphopsis viridulus, Yen, 1939 

Types: S.I.; Lectotype, 4129, fig. in Yen, 

1939: pi. 4, fig. 9 

Paralectotypes, 4130, figured here; Figure 36 

A-C. 

Type Locality. Hunan 

Habitat 

Anhua County, Anhua Town, Zijiang River; 
28°23'46" N, 1 1 1 °1 2'41 " E., Figure 1 , Site 1 0. 
Collected by Chen and Davis, 16 March 1987; 
field collection number D87-1. Catalog num- 
bers are: large class, ANSP 373147, A12663; 
small class, ANSP 373148, A12664. D85-78, 
small class, ANSP 373149, A 12665; ZAMIP 
M0055. Snails were collected 1.6 km up- 
stream from the town boat landing, along the 
shores of a small island in the middle of the 
river. Water flows through a stone breakwater 
at the upstream end of the island. Between 
the breakwater and the cobbles of the island 
was a protected area with water some 30 cm 
deep and with emergent vegetation. On the 
undersides of rocks at the breakwater on the 
protected side were numerous Guoia viridu- 
lus and Lithoglyphopsis modesta. Associated 
snail fauna included Stenothyra hunanensis 
(Davis et al. 1988), Gyraulus sp., Radix sp., 
Semisulcospira sp., and a viviparid. 



Thiele (1928: 365) noted that of Asian spe- 
cies described as Lithoglyphus, L. fuchsianus 
and L. viridulus, had radulae that corre- 
sponded to the European Lithoglyphus, 
whereas the radulae of L. modestus and L. 
tonkinianus Bavay & Dautzenberg (a Viet- 
namese species) had an entirely different 
type of central tooth. He created the genus 
Lithoglyphopsis to include the latter two spe- 



Introduction 

There were two size classes of fully mature 
snails living intermixed at the site (Fig. 37). 
Throughout the description these will be re- 
ferred to as the large class and small-class 
snails. There are slight shape differences be- 
tween the classes, yet we have felt it prudent 
not to consider them belonging to different 



188 DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG 36 Lectotype and paralectotypes of species of Guoia. A-C. Paralectotypes of Guoia viridulus (SMF 
4130); A = 3.50 mm long. Guoia fuchsianus, D-F. D. Lectotype, L = 3.24 mm. E, F. Paralectotypes. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 189 




FIG. 37. Four aspects of each of six shells of Guoia viridulus. A-C. Large class; A = 3.6 mm long D-F 
Small class; D = 2.8 mm long. A, B, D, E = females; C, F = males. 



190 




DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 

Adb 

Ngr 

Als à 











FIG. 38. Illustrated details of the apertural region of shells of Guoia viridulus characteristic of the species. 



species as we could find no anatomical differ- 
ences between them, and as they live in mi- 
cro-sympatry. The greater amount of anatom- 
ical data was from the large-class snails 
based on living specimens. Subsequently, 
anatomical data were gathered in Philadel- 
phia from alcohol-preserved small class 
snails. 

As there is considerable similarity among 
shells of the two size classes of snails that we 
refer to as G. viridulus, as well as G. fuch- 
sianus, we did a multivariate analysis of shell 
measurements of these taxa as well as older 
museum specimens of these species in order 
to attempt to assess differences among them. 
The following anatomical description is based 
on large class snails. Small class snails are 
contrasted with large class individuals in the 
remarks section for this species. 



Description 

Shells. Figures 36-39. Large-class (Figs. 
37A-C, 39A-C). Shells small, globose-conic, 
4.0-4.5 whorls, smooth but with rough growth 
lines. Aperture sub-circular with complete 
peristome, with pronounced apertural beak 
(Fig. 38, Adb) and a pronounced beak groove 
(Ngr). There is a wide columellar shelf (Cs), a 
small basal crescent (Be) with crescent ridge 
(Cr). Some shells have an umbilical chink 
(Uc), others do not. The abapical lip has an 
embayment (Ae). The adapical outer lip is 
flared out making somewhat of an angle with 
the remaining outer lip. In side view, the lip is 
sinuate forming a slight sigmoid embayment 
(Als). 

Females are larger than males (Table 16). 
Size ranges (lengths) are: males, 2.8 to 3.6 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 191 




FIG. 39. SEM photos of shells of Guoia viridulus (A-F) and G. fuchsianus (G-l). A-C. Large form; D-F. 
Small form. B, C, E, F, H, I = enlargements of apical whorls. 



mm; females, 2.8-3.8 mm. Our samples com- 
pare well with two lots of G. viridulus from the 
collections (Table 16, column 3). They com- 
pare well with paralectotypes (Figs. 36A-C 
compared with Fig. 37A-C). 

Small-class (Figs. 37D-F). Shells as in the 
large class with the following differences. 
Length is substantially smaller (Table 16) with 
males 2.0-2.1 mm long, females 2.2-2.8 mm 
long. The small-class shells appear some- 
what more conic than large class shells (com- 
pare Fig. 37E vs. 37A). However, the width to 
length ratio is not significantly different be- 
tween classes; the length of the body whorl to 
total height is slightly greater in the large size 
class (0.84 ± 0.03, 0.84 ± 0.05 for large- 
class; 0.80 ± 0.02, 0.80 ± 0.03 for small 
class). The basal crescent and columellar 
shelf are deeply depressed in small class 
shells (Fig. 37D). The outer lip in side view 
has a deep sigmoid embayment (Fig. 38F). 



SEM pictures of the apical whorls are 
shown in Figure 39B, C, E, F. The large class 
individuals inevitably had eroded apices; the 
small class shell apices were smooth. Some 
spiral microsculpture was noted at the shoul- 
der of the body whorl of small class shells. 

Multivariate analysis. The phenogram based 
on distance coefficients (cophenetic correla- 
tion 0.77) is given in Figure 40. Note the fol- 
lowing: (1) Historic G. viridulus (nos. 18-21) 
form a discrete unit within the wider variance 
of 9 large-class Anhua G. viridulus collected by 
us. (2) One Anhua G. viridulus (no. 5) clusters 
with Baisha G. fuchsianus collected by us. (3) 
The one historic G. fuchsianus that had com- 
plete whorls so that it could be included in this 
analysis clusters in the middle of the G. fuch- 
sianus variance. (4) Two small-class Anhua 
G. viridulus (nos. 27, 28) and two snails clas- 
sified by us as large-class Anhua G. viridulus 



192 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



01 Д 

ОбЛ 

■ огЛ 
-11 A 

ю A 

18 V 
-19 V 
-21 V 

-20 V 

-03 Л 

-oeA 
-04Д 

-09 A 
-05 Д 
-16 « 
-17 # 
-33® 
-13 О 
-15 • 
-14 О 
-07 Л 
-12 A 
-27 В 
-28 1 
-22 D 
-23 D 
-25 D 
-24 □ 
-26 D 



-29 I 
-30 I 
32 I 



Guoia viridulus -Anhua 

V -historic material 
/\ - large class males 
Д -large class females 
И -small class males 
■ -small class females 

Guoia fuchsianus 
(^) - historic material 
Q -males 
Л -females 



—311 



FIG. 40. UPGMA phenogram of distances for shell measurements of two species of Guoia. See text for 
details. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 193 




Guoia viridulus - Anhua 
\/ - historic material 
/\ - large class males 
A - large class females 
I I -small class males 
H "small class females 

Guoia fuchsianus 
Çy -historic material 
О - males 
Щ - females 



-1.522 -1.154 -0.786 -0.418 -0.050 0.318 0.686 1054 1.422 



FIG. 41 . Ordination diagram following three dimensional scaling. The first two principal components are 
given. Taxa are connected by a MST See text for details. 



(nos. 7, 12) form a sub cluster with G. fuch- 
sianus. (5) Most (82%) of the small-class G. 
viridulus form a discrete cluster at the bottom. 
In the PCA analysis, there were three sig- 
nificant components (Table 17); the first with 
76.6% of the variance, the second with 13.49, 
the third with 5.69 (total of 95.8%). Character 
loadings showed the first component to be 
one of size (Table 18); characters loading 
heavily on the second component-axis are 
number of whorls and length of the penulti- 
mate whorl. Ordination on the first two PCA 
axes following MDS is given in Figure 41 . The 
MST connects OTUs. Size increases from left 
to right. Along the second axis, relative to 
length, shells with more whorls, longer penul- 
timate whorls, and wider 3rd whorls are to the 



bottom. Proportionally shorter penultimate 
whorls and narrower 3rd whorls are to the top. 
The second axis is one of changes in trans- 
lation, whorl expansion, and whorl number. 
We deduce the following from the ordination 
diagram and table of character loadings. (1) 
We consider the small-class and large-class 
snails to belong to the same species, G. viri- 
dulus. With only two exceptions (nos. 5, 20) 
the variance along the second axis ranges 
from -0.554 to +0.182 for both size classes 
showing the same considerable variance in 
whorl translation and expansion. The only dif- 
ferences between the size classes are clear- 
cut differences in size, and the qualitative 
differences of deeper lip sinuation and 
depressed lip sinuation and depressed eres- 



194 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 16. Shell measurements for populations of Guoia viridulis. 





Large Class (D87-1) 


ANSP 45501 
& 98206 (N = 4) 


Small Class (D87-1) 




Males (N = 7) 


Females (N = 5) 


Males (N = 5) 


Females (N = 6) 


No. Whorls 


4.0-4.5 


4.0-4.5 


4.5 


4.0-4.25 


4.5 


Length (L) 


3.21 ±0.28 


3.36±0.38 


3.44±0.16 


2.12±0.06 


2.48±0.24 




(2.76-3.56) 


(2.84-3.84) 


(3.28-3.60) 


(2.02-2.16) 


(2.24-2.80) 


Width (W) 


2.57±0.16 


2.61 ±0.29 


2.89±0.19 


1.66±0.08 


1.86±0.14 




(2.32-2.76) 


(2.12-2.88) 


(2.72-3.12) 


(1.54-1.76) 


(1.68-2.08) 


L body whorl 


2.69±0.28 


2.82±0.31 


2.99±0.11 


1.70±0.06 


1.99±0.22 




(2.12-2.92) 


(2.32-3.12) 


(2.88-3.12) 


(1.62-1.76) 


(1.80-2.32) 


L penultimate 


0.33±0.07 


0.40±0.05 


0.29±0.02 


0.24±0.02 


0.28±0.03 


whorl 


(0.20-0.38) 


(0.36-0.48) 


(0.28-0.32) 


(0.22-0.28) 


(0.24-0.32) 


W penultimate 


1.16±0.13 


1.22±0.13 


1.11 ±0.04 


0.80±0.04 


0.90±0.10 


whorl 


(0.96-1.28) 


(1.04-1.36) 


(1.08-1.16) 


(0.76-0.84) 


(0.78-1.04) 


W 3rd whorl 


0.56±0.08 


0.56±0.08 


0.53±0.04 


0.44±0.04 


0.51 ±0.06 




(0.48-0.68) 


(0.48-0.68) 


(0.48-0.56) 


(0.40-0.50) 


(0.40-0.56) 


L last 3 


3.15±0.26 


3.28±0.34 


3.38±0.17 


2.05±0.06 


2.41 ±0.23 


whorls 


(2.72-3.48) 


(2.80-3.68) 


(3.20-3.52) 


(1.96-2.12) 


(2.20-2.72) 


L aperture 


1.95±0.19 


2.00±0.19 


2.29±0.12 


1.26±0.05 


1.44±0.11 




(1.56-2.12) 


(1.68-2.20) 


(2.16-2.44) 


(1.20-1.32) 


(1.32-1.60) 


W aperture 


1.56±0.17 


1.62±0.22 


1.83±0.12 


0.94±0.05 


1.10±0.09 




(1.24-1.76) 


(1.24-1.76) 


(1.68-1.96) 


(0.88-1.00) 


(0.96-1.20) 


W columellar 


0.33±0.04 


0.31 ±0.10 


0.33±0.05 


0.13±0.02 


0.11 ±0.03 


shelf 


(0.28-0.40) 


(0.14-0.40) 


(0.28-0.40) 


(0.12-0.16) 


(0.09-0.16) 


W + L 


0.80±0.03 


0.78±0.03 


0.84±0.02 


0.78±0.03 


0.75±0.03 




(0.77-0.84) 


(0.75-0.81) 


(0.82-0.87) 


(0.76-0.82) 


(0.71-0.79) 



TABLE 17. Percent of variance accounted for by 
each principal component (PC). 



PC 



% 



76.6 

13.5 

5.7 



Accumulated % 



76.6 
90.1 
95.8 



cent-columellar callus in the small-class 
snails. Note that two shells (nos. 7, 12) where 
originally selected as large-class snails based 
on visual impression of shape and apertural 
characters, yet are clearly small-class snails 
when measured. The length variance for 
small-class shells was 2.02-2.84 mm, for 



TABLE 1 8. Character loading on each PC axis. 







Components 






1. 


2. 


3. 


1 . No. Whorls 


0.293 


-0.724 


-0.624 


2. Length 


0.996 


0.012 


-0.015 


3. Width 


0.965 


0.210 


-0.092 


4. Length of body whorl 


0.970 


0.192 


-0.059 


5. Length of penultimate whorl 


0.700 


-0.579 


0.307 


6. Width of penultimate whorl 


0.944 


-0.209 


0.210 


7. Width of 3rd whorl 


0.745 


-0.525 


0.249 


8. Length of last three whorls 


0.994 


0.054 


-0.026 


9. Length of aperture 


0.994 


0.271 


-0.113 


10. Width of aperture 


0.949 


0.270 


-0.098 


1 1 . Width of columellar shelf 


0.874 


0.274 


-0.034 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 195 



TABLE 19. Lengths (mm) or counts of non-neural organs and structures of large-class Guola viridulus. 
Mean ± standard deviation (range). N = number of snails used. 



Females 



Males (N = 1) 



Body 

Digestive gland 
Gonad 

Total palliai oviduct 

PO 
Bursa copulatrix 

= Bu 
Bu * PO 

Duct of bursa 

Buccal mass 

Mantle cavity 

Osphradium 

= Os 
Gill 

= G 
Os - G 

No. gill filaments 

Gf 2 

of, 

Total Gf 

= TGF 
Gf 2 + TGF 

Prostate 
Seminal vesicle 
Penis 



5.69: 

(4.36- 

2.23: 

(1.80- 

0.89: 

(0.46- 

2.09: 

(1.80- 

0.61: 

(0.50- 

0.29: 

(0.27- 

0.12: 

(0.06- 

0.67: 

(0.64- 
1.51: 
(1.30- 

0.53: 

(0.46- 

1.35: 
(1.14- 
0.40: 

(0.32- 

24.4: 

(22- 

0.25: 

(0.12- 

0.37: 

(0.30- 

0.57: 

(0.42- 

0.39: 

(0.29- 



1.29 

7.24) N = 4 

0.49 

2.9) N = 4 

0.46 

1.44) 

0.21 

2.30) N = 4 

0.11 

0.76) N = 4 

0.03 

0.33) N = 4 

0.06 

0.18) N = 3 

0.03 

0.70) N = 3 

0.18 

1.66) N = 5 

0.09 

0.68) N = 5 

0.17 

1.50) N = 5 

0.05 

0.45) N = 5 

2.1 

27) N = 5 

0.09 

0.34) N = 8 

0.07 

0.42) N = 8 

0.17 

0.74) N = 8 

0.08 

0.49) 



6.0 
2.8 
1.4 



2.30 

1.00 

2.00 

0.50 
(0.26-0.36) 
25 

male + female 

male + female 

male + female 

male + female 

1.80 

1.20 

3.33 (N = 2) 
(3.06-3.60) 



large-class, 2.92-3.84 mm; thus for the spe- 
cies there is a 90% increase in size from 
shortest to longest mature specimen. 

2) We consider the shells at the top of the 
ordination clustered around mid-axis 1 as be- 
longing to G. fuchsianus. They differ consid- 
erably from G. viridulus in that all have 4.0 
whorls, relatively narrower 3rd whorls and 
shorter penultimate whorls. Only 1 individual 
(no. 5) of G. viridulus (7%) grouped with 
shells of G. fuchsianus. 

External features. The head is shown in Figure 
42. The snout is transparent but flecked with 
spots of melanin pigment on the dorsal snout 
and bars of pigment along the tentacles. 



There is a small dense eyebrow (Eyb) about 
each eye comprised of yellow glands. The 
operculum is shown in Figures 43, 44C-F. It 
is ovate, paucispiral, with a modest internal 
attachment pad. 

Mantle cavity. The reflected mantle is shown 
in Figure 45A. The mantle cavity structures are 
typical for taxa of the Neotricula clade. The 
opening of the spermathecal duct (Osd) is next 
to the pericardium (Pe). The osphradium (Os) 
is long and situated mid-gill. The terminal gill 
filaments (Gf 2 ) are normal length (Tables 19, 
20). The length of the longest filaments is 0.57 
± 0.1 7 mm long. The filaments are not pleated 
and lack a pronounced crest (Fig. 45B). 



196 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 20. Lengths (mm) or counts of non-neural organs and structures of alcohol-preserved small-class 
Guoia viridulus. N = number of snails used. 



Females 



Males (N = 1) 



Body 
Digestive gland 

Gonad 

Posterior palliai 

oviduct ( = albumen gland) 
Anterior palliai 

oviduct ( = capsule gland) 
Total palliai oviduct 

Bursa copulatrix 

Mantle cavity 

Osphradium 

= Os 
Gill 

= G 
Os- G 

No. gill filaments 

Gf 2 

Gf n 

Total Gf 

= TGF 
Gf 2 - TGF 

Prostate 
Penis 



6.3 

(5.60, 6.98) N = 2 
2.75 

(2.4, 3.1) N = 2 
1.10(N = 1) 

1.2 
(1.1, 1.3) N = 2 

1.3 
(1.2, 1.34) N = 2 

2.28±0.33 
(1 .9-2.5) N = 3 

0.43±0.16 
(0.25-0.56) N = 3 

1.81 
(1.62, 2.00) N = 2 

0.67 
(0.58, 0.76) N = 2 

1.61 
(1 .42, 1 .80) N = 2 

0.41 
(0.41, 0.42) N = 2 

22 
(21,23)N = 2 

0.23±0.07 
(0.16-0.30) N = 3 

0.30±0.08 
(0.24-0.36) N = 3 

0.53±0.13 
(0.40-0.66) N = 3 

0.43±0.03 
(0.40-0.46) N = 3 



1.50 

0.62 

1.24 

0.41 

19 

male & female 

male & female 

male & female 

male & female 

1.04 
2.24 




FIG. 42. Head and penis of Guoia viridulus. 



Female reproductive system. The body of an 
uncoiled female without head and with kidney 
tissues removed is shown in Figure 46A. Mea- 
surements of relevant organs are given in Ta- 
bles 1 9, 20. Important features to note are: (1 ) 
The body is not squat, but regularly tubular 
(contrast Lithoglyphopsis modesta). (2) The 
posterior palliai oviduct (Ppo) does not bend 
over the style sac (contrast L modesta). (3) 
The gonad (Go) is posterior to the stomach. 
(4) Sperm enter the system at the rear of the 
mantle cavity (Emc) through the opening of 
the spermathecal duct. The spermathecal 
duct does not enter the pericardium (contrast 
taxa of the Tricula clade). (5) The pericardium 
does not swell out into the mantle cavity. The 
bursa copulatrix complex of organs is shown 
in Figure 47. (6) The terminal end of the mus- 
cular spermathecal duct (Sd) is a thin walled 
vestibule (Twv). The spermathecal duct is 
short or long; it is that section of duct between 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 197 




FIG. 43. Opercula of Guoia viridulus (A-D) and Guoia fuchsianus (E, F). А, В from large-class specimens; 
C, D from small-class specimens. A, C, E. Outer surfaces; B, D, F. Inner surfaces. 



198 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 21. Radular statistics for Guoia viridulus snails. Mean 
except for width of central tooth in (im. 



standard deviation (range). In mm 



Large Class 



Females (N = 4) 


Males (N = 3) 


0.96±0.11 


0.94±0.02 


(0.84-1.08) 


(0.92-0.96) 


0.12±0.01 


0.12±0.01 


(0.104-0.132) 


(0.112-0.128) 


70±4 


71.7±2.5 


(66-75) 


(69-74) 


10±1.8 


9.7±0.06 


(8-12) 


(9-10) 


27±3.5 


26±2 


(24-32) 


(24-28) 


Small Class 


0.67±0.05 


0.61 ±0.05 


(0.64-0.72) 


(0.60-0.62) 


0.09±0.01 


0.08±0.01 


(0.08-0.10) 


(0.07-0.09) 


69±3.6 


69.7±6.7 


(66-73) 


(64-77) 


7.5±0.6 


8.7±0.6 


(7-8) 


(8-9) 


18±1.6 


17.3±1.2 


(16-20) 


(16-18) 



Radular length 
Radular width 
Total rows of teeth 
Rows of teeth forming 
Central tooth width 

Radular length 
Radular width 
Total rows of teeth 
Rows of teeth forming 
Central tooth width 



the end of the mantle cavity (Emc) and the 
point where the sperm duct (Sdu) arises from 
the spermathecal duct. (7) The spermathecal 
duct runs directly posterior from the mantle 
cavity to become the duct of the bursa (Dbu) 
directly posterior to which the thin-walled 
bursa copulatrix (Bu) swells as a large oval 
sac. (8) The sperm duct (Sdu) is a long con- 
voluted duct transporting sperm from the 
spermathecal duct to the oviduct close to 
where the latter (Opo) enters the albumen 
gland (Ppo). (9) The usual seminal receptacle 
has been lost. When dissecting living speci- 
mens one often observes a bright pink sheen 
or glittering within a section of the oviduct 
(Ov) just posterior to the juncture of the sperm 
duct (Sdu) and oviduct (Osr, Fig. 47). This sec- 
tion of oviduct assumes the function of the 
seminal receptacle and is called the oviducal 
seminal receptacle (Osr). The duct may be 
considerably swollen with sperm (Fig. 47D, 
E). (10) Posterior to the sperm storage area 
(Osr) the oviduct is convoluted (Fig. 47B). (1 1 ) 
The bursa is posterior to the posterior palliai 
oviduct (Bu, Fig. 46). (12) The bursa is short. 



Male reproductive system. The body of an un- 
coiled male is shown in Figure 48 without 



head but with kidney tissue (Ki) left in place. 
Measurements of relevant organs are given in 
Tables 19, 20. Important features are: (1 ) The 
gonad consists of relatively large lobes drain- 
ing into a vas efferens (Ve, Fig. 49). (2) The 
seminal vesicle arises from the vas efferens 
(Ve) approximately one third of the way pos- 
terior from the anterior end of the gonad (Fig. 
49). The juncture of the vas efferens and the 
beginning seminal vesicle may be swollen 
(Fig. 48). The coils of the seminal vesicle may 
be ventral to or dorsal to the lobes of the go- 
nad. (3) The gonad is posterior to the stom- 
ach. (4) The prostate (Pr) overlaps the poste- 
rior end of the mantle cavity (Emc). (5) The 
penis has a glandular lobe (Fig. 42 Glo) on 
the concave edge near the base. (6) The pe- 
nis has a stylet (Sty, Fig. 42). 

The stylet requires some comment. It is 
very much like the stylet observed in Robert- 
siella (Davis & Greer, 1980). It is corneous 
and very fragile. In the living-moving male, it 
is very obvious at 50X, projecting from the tip 
of the penis (Fig. 42). However, in relaxed 
and fixed specimens it is not in evidence. 
Rarely at 50X one observed a pin-prick of 
light reflecting from the tip of the stylet mostly 
retracted into the penis. To observe the stylet 
in preserved specimens, the penis is cut from 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 199 




FIG. 44. Opercula of Guoia viridulus (C-F) and G. fuchsianus (A, B). C, D from large-class snails; E, F from 
small-class snails. A, C, E. Outer surfaces; B, D, F. Inner surfaces. 



the animal, mounted in water on a slide with 
cover slip, and examined at 400 X (Fig. 50). 
The stylet is quickly dissolved when the penis 
is placed in Clorox (0.5% Na hypochlorite). 
Thus the stylets of Robertsiella and Guoia are 
not robust as those found in Stenothyra 
(Davis et al. 1986, 1988). 

(7) The ejaculatory duct is massive and 
highly musculahzed (Ej, Figs. 42, 51). It ex- 
tends out of the base of the penis and from 
the posterior penis along the entire length of 
the dorsal neck. The circular ejaculatory duct 
is slightly left (Fig. 52), central, or to the right 
of the snout-neck mid-line (Fig. 51). 



Digestive system. The digestive gland covers 
the posterior chamber of the stomach. The 
paired salivary glands are the standard tricu- 
line type. The radular sac does not coil up 
over the buccal mass. 

The radulae of large-class snails are shown 
in Figures 53E-H and 54A-H. Teeth counts 
and statistics are given in Tables 21, 22. The 
most frequently encountered formula is: 

3-1-3; 3(4)-1[2]-3(4); 10-13; 8-10. 
3-3 

The inner pair of basal cusps of the central 
tooth are comparatively enormous. The cen- 



200 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 22. Cusp formulae for the radular teeth of Guoia viridulus with the percent of the radulae in which 
a given formula was found at least once. 



Central Teeth 



Lateral Teeth 



Inner Marginal Teeth 



Outer Marginal Teeth 



Large Class (N = 5 radulae) 



3-1-3 
3-3 

3-1-3 
4-3 

3-1-4 
4-4 

4-1-3 
3-3 

4-1-4 
3-3 

5-1-5 
3-3 

5-1-4 
3-3 

5-1-5 
3-3 



60% 
20% 
20% 
20% 
20% 
20% 
20% 
20% 



3-1 [2]-4 60% 

4-1[2]-3 60% 

3-1[2]-3 40% 

3-1[2]-5 20% 

4-1[2]-4 10% 



8 

9 20% 

10 60% 

1 1 80% 

12 60% 

13 40% 

14 20% 

X = * 1.5±1.6 
N = 50 

Small Class (N = 5 radulae) 



8 40% 

9 60% 

10 40% 

1 1 20% 

12 

13 20% 

14 20% 
10.4±2.3 



4-1-4 40% 



3-3 




4-1-3 


40% 


4-3 




4-1-5 


40% 


3-3 




3-1-5 


40% 


4-4 




3-1-3 


20% 


4-3 




3-1-4 


20% 


4-4 




3-1-3 


20% 


4-4 




4-1-4 


20% 


4-4 




4-1-3 


20% 


3-4 




4-1-4 


20% 



4-3 



4-2-3 


100% 


3-2-4 


80% 


4-2-4 


60% 


5-2-3 


60% 


5-2-4 


60% 


3-2-3 


40% 


4-2-4 


40% 


4-2-5 


20% 



1 1 20% 

12 60% 

13 80% 

14 100% 

15 80% 

16 60% 



X = *7.40±0.9 

N = 30 



11 





12 


40% 


13 


80% 


14 


100% 


15 


60% 


16 





6.3±0.6 


N = 


= 30 



'Mean ± standard deviation of cusp number for all teeth counted. 



tral cusp at the anterior edge of the central 
tooth is frequently grooved or split. 

The radulae of small-class snails are 
shown in Figures 54E-H and 55. Teeth counts 
and statistics are given in Tables 21 , 22. The 
most frequently encountered formula is: 



3(4)-1-(4)3; 3(4)-1[2]-3(4); 12-16; 12-15. 
3-3 
Comparing both size classes, as one would 
expect, the small-class has a smaller radula 
except for total rows of teeth, which are not 
significantly different between classes. The 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 201 

_Gf, 




FIG. 45. Cut and reflected mantle to show mantle cavity organs of Guoia viridulus (A). Not all gill filaments 
are shown. B. Single gill filament. 



morphologies of the teeth of both size classes 
are the same. 

Nervous system. Measurements are given in 
Table 23. The RPG ratio is 0.229 ±0.11, that 
is, the pleuro-supraesophageal connective is 
concentrated. The pleuro-subesophageal 
connective is lost in the fusion of ganglia. The 
osphradio-mantle nerve is elongated (>0.12 
mm) thus compensating for the low RPG ra- 
tio. 

Remarks 

In comparing small- and large-class snails, 
no qualitative anatomical differences were 
found. The sizes of the bodies and organs 
were not significantly different. However, the 
small class animals were relaxed with sodium 
nembutol prior to fixing in 8% formalin and 
grading up to 70% ethyl alcohol. Presumably, 
the extended fixed bodies would measure 
longer than animals removed living from the 
shells and pinned out in a contracted state. 
Because the animals of the two size classes 
do not differ in details of anatomy, and be- 



cause they live in microsympatry, they are 
considered the same species. 

While the shells are significantly different in 
size, the range of variance along the second 
principal component is the same for both size 
classes indicating that the substantive differ- 
ence is only one of size. What could account 
for this? Has this phenomenon been seen be- 
fore? A similar situation was reported for Tric- 
ula xianfengensis Davis & Guo, 1 986, in which 
large and small class mature snails with the 
same anatomy and shell shape were found in 
a narrow ditch alongside a kitchen garden in 
Xiaguan City, Yunnan Province. In both of 
these cases, we suspect that the size classes 
resulted from two different cohorts when egg 
laying occurred at different seasons. It is pos- 
sible that eggs laid late in the season with 
growth extending into the cold weather months 
resulted in a small size class due to stunting 
caused by decreased rate of growth in the fall 
and winter. Presumably egg laying and growth 
in the late spring and summer would result in 
optimal growth. A comparative molecular ge- 
netic analysis of these size classes would be 
most instructive and desirable. 



202 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 
Ast 4 i n 




FIG. 46. Uncoiled female of Guoia viridulus with head and kidney tissue removed (A). B. Variation in gonad 
morphology. 



TABLE 23. Lengths of neural structures from four individuals of large class Guoia viridulus. Mean 
standard deviation (range). 



Cerebral ganglion 
Cerebral commissure 
Pleural ganglion 
Right (1) 

Left 
Pleuro-supraesophageal 

connective (2) 
Pleuro-subesophageal 

connective 
Supraesophageal 

ganglion (3) 
Subesophageal 

ganglion 
Osphradio-mantle nerve 



0.31 ±0.03 
0.07±0.02 

0.15±0.04 

0.11 ±0.01 
0.08±0.04 



0.12±0.02 

0.12±0.02 

0.15±0.03 



(0.26-0.32) 
(0.06-0.10) 

(0.10-0.12) 

(0.10-0.12) 
(0.04-0.14) 



(0.10-0.14) 

(0.10-0.14) 

(0.12-0.18) 






THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 
Bu 



203 



Osr 



Twv 




FIG. 47. Bursa copulatrix complex of organs of Guoia vihdulus. Organs in A are in same orientation as in 
Figure 46. Figures В, С shows bursa flipped over to show dorsal aspect to reveal origin of elongated and 
twisting sperm duct (Sdu). Figure D most closely approximates organ position in Figure 46 but with tissue 
cleared away to show relationships of ducts relative to end of mantle cavity (Emc) and albumen gland (Ppo). 
E. Blown up section of oviduct within which sperm are stored (Osr). 



204 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 
Di Ki 




FIG. 48. Uncoiled male of Guoia viridulus with head removed. 



Guoia fuchsianus (Moellendorff, 1885) 

Lithoglyphus fuchsianus Moellendorff, 1885: 

169, 170 

Lithoglyphus, fuchsianus, Moellendorff, 1888: 

140. pl. 4, fig. 5, 5a-b. 

Lithoglyphus fuchsianus, Thiele, 1 928 

Lithoglyphopsis fuchsianus, Yen, 1939 

Type locality. Moellendorff, 1885: Hsiang-tan 

provinciae sinensis Hunan. 

Moellendorff, 1888: Hsiangtan and 

Hêngshan-hsien; Hunan. 

Yen, 1929: Heng-dshou-fu, Hunan 

Types. Lectoype: SI, 4127. Figured by Yen 
(1939: pl. 4, fig. 10); paralectotypes: SMF, 
4128. Figured here, Figure 36E, F. 

Habitat 

Xiangjiang River at Baisha, Hengshan 
County, 25°58'22"N, 112°45'55"E. Figure 1, 




FIG. 49. Male gonad with anterior lobes cut away to 
show origin of vas deferens from vas efferens (Ve), 
and seminal vesicle (Sv) that lies dorsal to gonad. 
Dashed line indicates extent of gonad cut away. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 205 

x 




FIG. 50. Tip of penis of preserved Guoia viridulus 
as observed under compound microscope at 400X. 
Sty, Stylet; Vd, Vas deferens. 





FIG. 51 . Relationship of ejaculatory duct to mid-line 
(x) of snout-neck of Guoia viridulus (from small- 
class specimen). 



FIG. 52. Ejaculatory duct of Guoia viridulus slightly 
to left of snout-neck mid-line. 



locality 9. Collected by Davis, Chen, & Wu, 
October 1985, field number D85-75; by Chen 
& Wu, 1986. Snails sympatric with Lithogly- 
phopsis modesta on stones at the bottom of 
the river in 2.0-2.5 m depth. Catalog numbers 
are: D86-B, ANSP 373145; A12661. D85-75, 
ANSP 373146; A1 2662. 



Description 

Shell. Shells (Figs. 36D-F, 39G-I, 56C-F, 
57, 58) are small (Table 24), and generally as 
described for large class L viridulus with the 
following differences. The outer lip of G. fuch- 
sianus is only slightly sinuate. There is a more 
pronounced umbilical depression. The inner 
lip is separated slightly from the body whorl by 
a narrow depressed groove, the basal cres- 
cent (Fig. 57, Be) paved with shell growth in- 
crements from body whorl to the inner lip 
(Figs. 57, 58). The penultimate whorl of the 
species is proportionally shorter than that of 
G. viridulus and the third whorl less wide. 

Overall, however, the shells have such in- 
trapopulation variance that the species are 
difficult to differentiate on the basis of shells 
alone. Some 1 0% of the shells from one pop- 
ulation are virtually indistinguishable from 
shells of the other. An example of variance in 
shape for G. fuchsianus is shown in Figure 
58A-D, in which camera lucida drawings of 
two shells from collection D86-B are illus- 
trated with two shells from historic ANSP 
98205. Because most of the historic G. fuch- 



206 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 53. Radulae of large size-class Guoia viridulus (E-H) and G. fuchsianus (A-D). A, E. = portions of 
radulae. B-C, F-G. Central, lateral and inner marginal teeth; D, H. Outer marginal teeth. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 207 




m 



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E 

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CD 

CO 



с 

CO 

■8 

1 

CD 

T3 

с 
со 

Й • 

-Э £ 

-Э ® 
лэ ф 

с ** 

•S £ 
о о) 

Ti 



СО - 

ÜI 

N О 



ES 
ся _ 

= 1 

СО О) 

.— . СО 

ш Е 
< ф 

со g 

СО .- 

«тэ 

О С 

ф СО 
N -= 



ф 



со 

o>J5 

со _- 

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s"-" 



208 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 







FIG. 55. Radula of small size-class Guoia viridulus. A. Segment of radula; В, С Central, lateral and inner 
marginal teeth; D. Outer marginal teeth. 



sianus had eroded shells so that characters 1 , 
7, and 8 could not be measured, the multivari- 
ate analyses involving all populations of G. 
viridulus and G. fuchsianus without these 
three characters did not provide differentia- 
tion of species or populations. Individuals 
were simply spread out along a size gradient. 
The historic G. fuchsianus specimens are 
larger than field-collected specimens. Still, 
basing size on the length of body whorl and 
width, this species is somewhat smaller than 
G. viridulus (compare Tables 24 and 16). 

Anatomy. In 1985, most of the snails were 
sexually immature so that complete anatom- 
ical data could not be gathered. Organ mea- 
surements and counts that could be made are 
presented in Table 25. Sufficient observations 
were made to demonstrate that Guoia fuch- 
sianus is a distinct species, not to be con- 
fused with G. viridulus. Only anatomical fea- 
tures unique to this species will be presented 
here. 

(1) The duct of the bursa (Dbu, Fig. 59B) is 
a considerably swollen duct that narrows be- 
fore opening into the capacious bursa (Bu). 



(2) The duct of the bursa is comparatively 
long: 0.35 mm for G. fuchsianus, 0.12 for G. 
viridulus. (3) The sperm storage area (Sr) is a 
swelling that bulges out of the sperm duct 
(Sdu) where the sperm duct opens into the 
oviduct (Ov, Fig. 59). (4) The buccal mass is 
longer (0.64-0.70 mm for G. viridulus, 0.70- 
0.72 for G. fuchsianus). (5) The central ante- 
rior cusp of the central tooth of the radula and 
flanking cusps are considerably more poste- 
riorly projecting than those of G. viridulus 
(compare Fig. 54C, G, K). The anterior cusp 
support and cusps of G. fuchsianus form a 
prominent triangular-shaped projection poste- 
riorly. This, coupled with the elongated central 
cusp, causes the tip of the central cusp to 
overlap the tooth immediately posterior. A ra- 
tio will demonstrate the difference between 
taxa. The length from anterior central edge to 
posterior tip of the central cusp divided by 
width of the anterior central cusp yields 0.75- 
0.88 for G. fuchsianus and 0.57-0.71 for G. 
viridulus. (See Tables 26, 27 for radular sta- 
tistics.) (6) Measurements of neural struc- 
tures are given in Table 28. The RPG ratio is 
0.42, that is, the pleuro-supraesophageal 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 209 



connective is moderately concentrated (con- 
trast the concentrated condition in G. viridu- 
lus, RPG ratio = 0.23). 

The penis of two individuals had a well-de- 
veloped stylet but no glandular lobe or pro- 
nounced ejaculatory duct. The prostate was 
fully developed and penis length was 1 .5-1 .6 
mm long. Because the penis of G. viridulus 
with a pronounced glandular lobe exceeded 
3.0 mm in length, it is possible that the ones 
we observed for G. fuchsianus were imma- 
ture, underdeveloped, or, conversely, atro- 
phied. We are not convinced that the penis of 
G. fuchsianus lacks a glandular lobe or ejac- 
ulatory duct. 

Remarks 

Conchological characters that separate this 
species from G. fuchsianus are few and dis- 
cussed above. Refer also to the multivariate 
analysis presented under G. viridulus. 

Wheareas it is clearly desirable to have 
more anatomical data, the available data sub- 
stantiate the placement of this species in the 
genus Guoia. The six anatomical differences 
provided above convince us that we are deal- 
ing with a distinct species. The penial data are 
not of sufficient quality to permit us to make a 



definitive comment except that a stylet is def- 
initely present. 

Neotricula Davis, 1986 

Type Species. Lithoglyphopsis apena Tem- 
charoen, 1971: 103-104, pi. 7, fig. 14. 

Tricula apena (Temcharoen), Davis 1979, 
1980. 

Type locality. Mekong River at Ban Na on 
Khong Island, Laos 

Assigned Species. N. aperta (type for genus 
assigned in Davis et al., 1986a); N. burchi 
(Davis, 1968) [Thailand]; N. cristella (Gredler, 
1887); N. dianmenensis Davis & Chen, sp. 
nov.; N. duplicata Davis & Chen, sp. nov.; N. 
¡tin Chen & Davis, sp. nov.; N. minutoides 
(Gredler, 1885); N = 7. 

Diagnosis. Shells small to medium sized, 
ovate conic, smooth. Central tooth of radula 
with several anterior cusps (contrast single tri- 
angular blade as in Delavaya). The oviduct 
runs from the gonad to the palliai oviduct with- 
out making a loop or twist. The spermathecal 
duct does not enter the pericardium but opens 
into the posterior mantle cavity; it is a narrow 
duct throughout. The duct of the bursa enters 
a U-shaped bend to run into a discrete sem- 



TABLE 24. Shell measurements (mm) for populations of Guoia fuschianus. Mean 
(range). 



standard deviation 







Baisha 


ANSP 


98205, 45961 




Males (N = 2) 


Females (N = 3) 


Whole Shell (N = 


1) Eroded Shell (N = 8) 


1. 


No. Whorls 


4.0 


4.0 


4.0 


— 


2. 


Length (L) 


2.26 
(2.48, 2.76) 


2.79±0.12 
(2.72-2.79) 


2.96 


— 


3. 


Width (W) 


2.20 


2.36±0.07 


2.32 


2.44±0.12 






(2.08, 2.32) 


(2.32-2.44) 




(2.28-2.60) 


4. 


L body whorl 


2.26 


2.45±0.02 


2.60 


2.70±0.12 






(2.16,2.36) 


(2.44-2.48) 




(2.56-2.88) 


5. 


L penultimate 


0.24 


0.23±0.05 


0.24 


0.30±0.04 




whorl 


(0.20, 0.28) 


(0.20-0.28) 




(0.24-0.36) 


6. 


W penultimate 


0.90 


0.92±0.07 


0.86 


1.01 ±0.08 




whorl 


(0.80, 1.00) 


(0.88-1.00) 




(0.92-1.12) 


7. 


W 3rd whorl 


0.46 
(0.40, 0.52) 


0.47±0.02 
(0.44-0.48) 


0.40 


— 


8. 


L last three whorls 


2.60 
(2.48, 2.72) 


2.77±0.09 
(2.72-2.88) 


2.92 


— 


9. 


L aperture 


1.68 


1.85±0.08 


2.00 


1.93±0.08 






(1.56, 1.80) 


(1.76-1.92) 




(1.80-2.04) 


10. 


W aperture 


1.38 


1.45±0.02 


1.52 


1.54±0.08 






(1.28, 1.48) 


(1.44-1.48) 




(1.40-1.60) 


11. 


W columellar shelf 


0.18 


0.25±0.02 


0.28 


0.28±0.04 






(0.16,0.20) 


(0.24-0.28) 




(0.20-0.32) 


W 


*■ L 


0.84 
No var. 


0.85±0.01 
(0.84-0.85) 


0.78 





210 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 56. Shells of small-class Guoia viridulus (A, B) for comparison with shells of G. fuchsianus (C-F). Shell 
A is 2.36 mm long; others printed at same scale. All are from females except E, which is from a male. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 21 1 



TABLE 25. Lengths (mm) or counts of non-neural organs of female Guoia fuschianus. 



1. 



2. 



X. 



Body 

Digestive gland 
Bursa copulatrix 
Duct of bursa 
Buccal mass 
Mantle cavity 
Gill 

Osphradium 
Osphradium -н gill 
No. gill filaments 
Gf 2 
Gf, 
Total Gf 



4.9 
2.1 
0.68 
0.34 
0.70 
2.0 
1.8 
0.9 
0.50 
25 
0.16 
0.48 
0.64 



0.54 
0.36 
0.72 
1.70 
1.50 
0.7 
0.47 
22 
0.22 
0.46 
0.68 



2.20 
2.00 
0.84 

25 



0.61 
0.35 
0.71 

1.97±0.25 
1.77±0.25 
0.81 ±0.10 
0.48 

24.0±1.7 
0.19 
0.47 
0.66 




FIG. 57. SEM photograph of a shell of Guoia fuch- 
sianus. 



inal receptacle in most species. The duct of 
the seminal receptacle may shorten or be- 
come lost. The spermathecal duct joins the 
duct of the bursa (contrast joining the bursa 
as in Halewisia). A slender duct, the sperm 
duct, connects the U-shaped duct to the ovi- 
duct. 

Neotricula cristella (Gredler, 1887) 

Paralectotypes. SMF 4242; plate 4, fig. 3, in 
Yen, 1939 

Type locality. Kiangshi Province, "in Quell- 
wasser." 

Synonymy. Hydrobia cristella Gredler, 1887 
Tricula cristella, Yen, 1939 
Neotricula cristella, this paper 



Habitat 

Material for this paper was collected from 
Mojingtai, Hengshan Mountain, Nanyue 
Town, Hengshan County, Hengyang Prefec- 
ture; 27°15'N, 112°39'13"E; Figure 1 site 1. 
Snails came from a small stream 0.4 km down 
the mountain road from the Mojingtai Hotel 
towards the Banshan Ting Temple. Collec- 
tions number = D85-73; the collection was 
made by Dr. Chen Cui— E, 28 Sept. 1985. 

Depository 

Specimens are deposited in ZAMIP, 
M0001 ; in ANSP 368774 and A12146. 

Description 

Shell. Shells are small, narrowly ovate-conic, 
of 5.0 to 5.5 whorls (Figs. 60H-L, 61A-D). 
Lengths range from 2.40 to 2.84 mm (Table 
29). The aperture is ovate; there is no umbi- 
licus. The whorls at the suture are smooth 
(not crenulated). SEM analyses reveals a 
faint trace of spiral microsculpture on the 
adapical surface of the whorls at the suture. 
The inner lip is arched and widely separated 
from the body whorl, the distance increasing 
from abapical to adapical. Thus, the adaptical 
end of the aperture is widely separated from 
the body whorl. The adapical end of the ap- 
erture has a wide notch; there is no internal 
notch groove; there is no sinus. There is no 
abapical spout. In side view, the outer lip is 
slightly sinuate; it is not scooped forward. The 
inner lip lacks nodes, teeth or notches. The 
inner lip is thin throughout. In side view, the 
inner lip is straight on some shells, angled to 
form a deflection angle in others. Within the 



212 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 






2.0mm 



FIG. 58. Illustration of four shells of Guoia fuchsianus aided by camera lucida. Shells A, D from ANSP 98205 
(historic collections); shells В, С from D86-B. 



body whorl, the columella is smooth. There is 
a slight varix on some shells, no varix on oth- 
ers. In apertural view, the lip projects beyond 
the base of the shell 0.34 ± 0.06 mm. SEM 
analysis shows the protoconch to be slightly 
wrinkled (Fig. 61 C, D). 



External features. The head is not pigmented. 
There are no granules about the eyes (Fig. 
62). There is a patch of white granules where 
the mantle margin meets the neck. The oper- 
culum is corneous, paucispiral and appears 
to have two layers, a larger outer layer and a 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 213 



TABLE 26. Radular measurements (mm) and counts for Guoia fuchsianus. Mean 
(range). In mm except for width of central tooth in u,m. 

Female (N = 5) 



standard deviation 



Male (N = 2) 



Radular length 
Radular width 
Total rows of teeth 
No. rows of teeth forming 
Central tooth width 



0.90±0.05 
(0.84-0.98) 
0.11 ±0.003 
(0.11-0.12) 
72.6±4.3 

(66-78) 
23.8±1.1 

(23-25) 
23.2±2.3 

(22-26) 



0.80 
(no variation) 

0.11 

(0.10-0.11) 

68.5 

(68-69) 
22.5 

(22-23) 
22 
(no variation) 



TABLE 27. Cusp formulae for the radular teeth of Guoia fuschianus with the percent of radulae in which 
a given formula was found at least once. N = 3 radulae. 



Central Teeth 



Lateral Teeth 



Inner Marginal Teeth 



Outer Marginal Teeth 



4-1-4 



66% 



2-2 

3-1-3 33% 
3-3 



3-1[2]-4 


100% 


4-1[2]-3 


66% 


3-1[2]-3 


66% 


4-1[2]-4 


33% 


2-1[2]-4 


33% 



10 33% 

1 1 66% 

12 100% 



X = * 11.4±0.7 
N = 26 



10 


33% 


11 


66% 


12 


100% 


13 


33% 


11.3 


±0.8 


N = 


30 



*Mean ± standard deviation of cusp number for all teeth counted. 



TABLE 28. Lengths (mm) of neural structures of 
Guoia fuchsianus. Mean (data). N = 2 



Cerebral ganglion 
Cerebral commissure 

Pleural ganglia 

Right (1) 

Left 
Supraesophageal 

connective (2) 
Subesophageal connective 

Supraesophageal 
ganglion (3) 

Subesophageal ganglion 

Osphradio-mantle nerve 

RPG ratio (2-1+2 + 3) 



0.29 (0.28, 0.30) 
0.12 (no variation) 

0.11 (0.10, 0.12) 
0.10 (no variation) 
0.16(0.12, 0.20) 



0.11 (0.10,0.12) 

0.10 (no variation) 
0.12(0.10,0.14) 
0.42 (0.38, 0.46) 



narrower inner layer (Fig. 61 E, F). The inter- 
nal attachment pad is prominent but only 40 
to 50% the width of the operculum. 

Mantle cavity. Mantle cavity organs are 
shown in Figure 63A. Organ measurements 
and counts are given in Table 30. The osphra- 
dium is slightly anterior to mid-gill; it is oval 
and small. There are 1 3 to 1 5 gill filaments, of 



which only seven or eight are fully developed, 
that is, with both Gf , , and Gf 2 elements prom- 
inent. The anterior five filaments are widely 
separated and without the Gf 2 part. Gf 2 is 
long. The longest gill filaments are 0.38 ± 
0.09 mm long. The largest gill filaments in lat- 
eral view are modestly domed (Fig. 63B). The 
pericardium (Pe), opening of the kidney into 
the mantle cavity (Oki) and the opening of the 
spermathecal duct (Sd) into the mantle cavity 
are shown in relationship to each other (Fig. 
63 A). 

Female reproductive system. An uncoiled fe- 
male without head and with kidney tissue re- 
moved is shown in Figure 64. Measurements 
of organs are given in Table 30. Important 
features are: (1) The gonad (Go) is posterior 
to the stomach, is small and consists of few 
lobes. (2) The bursa copulatrix (Bu) is round 
and situated directly posterior to the albumen 
gland (Ppo). (3) The albumen gland is of nor- 
mal size. (4) The bursa copulatrix complex of 
organs is shown in Figure 65. Organs in Fig- 
ure 65A have the same orientation as in Fig- 
ure 64. The bursa is short. (5) The duct of the 
seminal receptacle (Dsr) is a U-shaped con- 



214 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 

Ppo 




0.5mm 




Sdu 
Dbu 



FIG. 59. Aspects of female reproductive system of Guoia fuchsianus. A. Female reproductive system 
oriented in same position as in Figure 46A. Anterior palliai oviduct not shown. B. Bursa of A turned 180° over, 
as indicated by the arrow, to show the relationships of ducts attaching to the bursa (Bu) and oviduct (Ov). 



tinuation of the duct of the bursa (Fig. 65B, C). 
It turns to the right side and tucks dorsal to the 
bursa. (6) The duct of the seminal receptacle 
(Dsr) is long. The seminal receptacle (Sr) is a 
comparatively minute bulb. (7) The spermath- 
ecal duct (Sd) is short and opens into the pos- 
terior end of the mantle cavity (Emc). (8) The 
spermathecal duct opens into the bottom of 



the U-shaped bend formed by the duct of the 
bursa at a point opposite the opening of the 
duct of the seminal receptacle (Fig. 65A, B). 

Male reproductive system. An uncoiled male 
snail without head and with kidney tissue re- 
moved is shown in Figure 66. Measurements 
of organs are given in Table 30. Important 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 215 




FIG. 60. Shells: Neotricula lilii, A, B. Paratypes. N. minutoides, C-G. N. cristella, H-L. Length of shell A 
3.12 mm; others printed to same scale. 



216 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 61 . SEM photographs of shells (A-D) and opercula (E, F) of Neotricula cristella. С, D. Details of apical 
whorls. In E, F, inside surface of opercula shown on right side. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 217 

TABLE 29. Shell measurements (mm) for male and female Neotricula cristella. Mean ± standard 
deviation (range). N = number measured. 





Females 




Males 


Whorls 


5.0-5.25 (N = 4) 


5.5 (N = 2) 


5.0 (N = 3) 


Length (L) 


2.64±0.10 


2.78 


2.37±0.02 




(2.52-2.76) 


(2.72-2.84) 


(2.36-2.40) 


Width (W) 


1.33±0.09 


1.38 


1.21 ±0.08 




(1.24-1.44) 


(1.36-1.40) 


(1.12-1.28) 


L body whorl 


1.85±0.11 


1.86 


1.61 ±0.02 




(1.76-2.00) 


(1.84-1.88) 


(1.60-1.64) 


L penultimate whorl 


0.40±0.01 


0.45 


0.36±0.03 




(0.38-0.40) 


(0.40-0.50) 


(0.34-0.40) 


W penultimate whorl 


0.90±0.02 


0.94 


0.83±0.03 




(0.88-0.92) 


(0.88-2.00) 


(0.80-0.86) 


L last three whorls 


2.46±0.11 


2.56 


2.20±0 




(2.36-2.60) 


(2.52-2.60) 


— 


L aperture 


1.21 ±0.08 


1.14 


1.07±0.06 




(1.16-1.32) 


(1.12-1.16) 


(1.00-1.12) 


W aperture 


0.81 ±0.05 


0.80 


0.69±0.02 




(0.76-0.88) 


— 


(0.68-0.72) 


Tip apical whorl (W) 


0.14±0.02 


— 







(0.12-0.16) N = 6* 


— 





Diameter 1 st whorl 


0.28±0.02 


— 


(0.26-0.30) N = 6* 


X" 


0.37±0.02 


0.34±0.06 






(0.36-0.40) N = 3* 


(0.28-0.40) 





all whorl classes 
'distance from base of body whorl to abapical tip of aperture 



features are: (1) The gonad (Go) is posterior 
to the stomach. (2) The prostate (Pr) overlaps 
the posterior end of the mantle cavity (Emc). 
(3) The seminal vesicle (Sv) arises from the 
vas efferens (Ve) about mid-gonad. (4) The 
seminal vesicle is coiled lateral to the gonad, 
like a spring or coils in a knot dorsal to the 
gonad posterior to the stomach. It does not 
continue onto the stomach. (5) The anterior 
vas deferens (Vd 2 ) leaves the prostate at the 
posterior end of the mantle cavity. (6) The 
penis is simple but with a very elongated pe- 
nial filament. (Pf, Fig. 67). (7) The vas defer- 
ens is highly coiled passing through the cen- 
ter of the penis. There is no ejaculatory duct in 
the base of the penis or in the neck. (8) The 
orientation of the base of the penis (Bp) to the 
snout-neck midline (x) and the posterior edge 
of the eye bulges is shown in Figure 62. The 
long axis of the penial base varies from 70° to 
90° from "x" as shown. 

Digestive system. The digestive gland covers 
the posterior chamber of the stomach of fe- 
males but is posterior to the stomach of 
males. Radular statistics are given in Tables 



31 and 32. There are 99 ± 3.4 rows of teeth 
along a radula 0.54 mm long. The most fre- 
quently encountered formula is 

3(2)-1-(2)3 ; 3-1[2]-3(4); 14-17; 12-15. 
3(2)-(2)3 

SEM photographs of radulae and teeth are 
given in Figures 68, 69. Central teeth are fea- 
tured in Figure 68C-F; they are typical of the 
generalized triculine type. The morphology of 
the entire lateral tooth is shown in Figure 68A, 
B. The one unusual feature seen in the radula 
of this species involves the dominant cusp of 
the lateral tooth, the "1" of the 3-1-3; it is 
deeply divided, almost forming two cusps. 
(Fig. 68A-F, 69A-C). 

Nervous system. Measurements are given in 
Table 33. The RPG ratio of 0.36 shows that 
the dorsal nerve ring is moderately concen- 
trated. 

Remarks 

Conchologically, this species differs from 
all other Neotricula by having a shell with (1) 



218 



DAVIS, CHEN, WU. KUANG, XING, LI, LIU & YAN 




( 



efficients (Fig. 153) shows linkage with Gam- 
matricula chinensis. The differences are: (1) 
The sutures of N. cristella are smooth; those 
of G. chinensis are crenulated. (2) The adapi- 
cal aperture of the former has a notch; of the 
latter, no notch. (3) The inner lip of the former 
is thin, of the latter, thick. (4) The inner lip of 
the former is widely separated from the body 
whorl, of the latter, narrowly separated. (5) 
The adapical end of the aperture of the former 
is widely separated from the body whorl, of 
the latter, slightly separated. (6) There is no 
abapical outer lip deflection angle in the 
former, a slight one in the latter. 

The minimum spanning tree (Fig. 154) 
based on distance coefficients yields a differ- 
ent closest relationship, one with Tricula hud- 
iequanensis. However, (1)7. hudiequanensis 
has a medium length shell (N. cristella has a 
small shell). (2) The adapical aperture of the 
former does not have a notch. (3) The outer 
lip of the former, in side view, is scooped for- 
ward; it is straight in the latter. (4) The inner lip 
of the former is thick; it is thin in the latter. (5) 
The inner lip and adapical aperture are not as 
greatly separated from the body whorl in the 
former as in the latter. 

The most distinguishing anatomical charac- 
ter (considering only species of Neotricula) is 
the long penial filament. This species shares 
with N. //'//'/ the character-state of having very 
many rows of teeth on the radula; with N. min- 
ima, the state that the digestive gland covers 
the posterior chamber of the stomach. The 
U-shaped continuation of the duct of the 
bursa into the duct of the seminal receptacle 
is shared with N. burchi of northwest Thailand 
and N. apena of the lower Mekong River. 



1.0mm 

FIG. 62. Head of a male snail of Neotricula cristella 
showing the orientation of long axis of the penial 
base (Bp) to snout-neck mid-line x. 



the inner lip widely separated from the body 
whorl, and (2) the adapical end of the aper- 
ture widely separated from the body whorl. 

The multivariate analysis involving all spe- 
cies of Triculinae with shells closely resem- 
bling those of Tricula or Neotricula indicates 
different affinities depending on the analysis 
used. The phenogram based on distance co- 



Neotricula dianmenensis Davis & Chen, 
sp. nov. 

Holotype: ZAMIP-M0034, Figure 70A. 
Paratypes: ANSP 373143, A12659; ZAMIP 
MOO04. Figure 70B-D; Figure 71 A, B. 

Type Locality. Jiepai Village, Dianmen Town, 
Hengshan County, Hengyang Prefecture. 
27°15'16"N, 112°33'31"E. Figure 1, Site 5. 
Field collection D85-80. 

Collection Date: October 1 985. 

Etymology: Named for the town of Dianmen. 

Habitat 

300 m above sea level. Snails were col- 
lected from a small stream 20-25 cm wide and 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 219 




FIG. 63. A. Mantle cavity structures of Neotricula cristella showing the relationship of the gill to pericardium 
(Pe) and openings of the kidney (Oki) and spermathical duct (Sd). B. Single gill filament 



Ast 



Eme 




Apo 



FIG. 64. Uncoiled female Neotricula cristella with head and kidney tissue removed. 



220 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 30. Lengths (mm) or counts of non-neural organs and structures of Neotricula cristella. Mean ± 
standard deviation, (range). 



Females (N = 5) 



Males (N = 3) 



Body 
Gonad 
Digestive gland 

Posterior palliai oviduct 

(= albumen gland) 
Anterior palliai oviduct 

(= capsule gland) 
Total palliai oviduct 

= OV 
Bursa copulatrix 

= BU 
Duct of BU 

BU * OV 

Seminal receptacle 

Duct of seminal 

receptacle 
Mantle cavity 

Gill (G) 

Osphradium (OS) 

OS - G 

No. of filaments 

Gf 2 

Gf, 

Total Gf = TGF 

Gf 2 4- TGF 

Prostate 

Seminal vesicle 

Penis 

Buccal mass 



4.60: 

(4.34- 

0.62: 

(0.54- 

2.06: 

(1.90- 

0.86: 

(0.76- 

0.96: 

(0.90- 

1.82: 

(1.66- 

0.38: 

(0.34- 
0.19: 
(0.14- 

0.21: 

(0.17- 

0.10: 

(0.08- 

0.19: 

(0.16- 
1.11: 
(0.76- 

0.92: 

(0.60- 

0.23: 

(0.16- 

0.26: 

(0.18- 

13.6: 

(13- 

0.23: 

(0.16- 

0.16: 
(0.12- 
0.38: 

(0.26- 

0.61: 

(0.56- 



0.17 

4.76) 

0.08 

0.70) 

0.17 

2.30) 

0.06 

0.90) 

0.09 

1.10) 

0.13 

2.00) 

0.05 

0.46) 

0.05 

0.26) 

0.04 

0.20) 

0.02 

0.12) 

0.04 

0.24) 

0.28 

1.44) 

0.28 

1.28) 

0.08 

0.36) 

0.08 

0.37) 

0.9 

15) 

0.06 

0.30) 

0.03 

0.20) 

0.09 

0.50) 

0.04 

0.65) 



4.17±0.32 
(3.80-4.36) 

1.03±0.21 
(0.80-1.20) 

1.93±0.31 
(1.60-2.20) 



1.00: 

(0.96- 

0.85: 

(0.80- 

0.24: 

(0.22- 

0.29: 

(0.26- 

13.3: 

(12- 



0.07 
■1.08) 
0.05 
0.90) 
0.02 
0.26) 
0.03 
0.33) 
1.2 
■14) 



0.50±0.06 
(0.44-0.56) 



0.81 ±0.02 
(0.80-0.90) 
0.38±0.07 
(0.40-0.44) 
1.53±0.18 
(1.40-1.74) 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 221 
Bu 




Emc 



FIG. 65. Details and variation of bursa copulatrix complex of organs of Neotricula cristella. Figure A is in 
same orientation as in Figure 64. B. Bursa removed to show entire seminal receptacle (Sr) and duct (Dsr). 
С Bursa rotated to show interconnection of Dbu, Dsr and the spermathical duct (Sd). 




Emc 



FIG. 66. Uncoiled male of Neotricula cristella without head or kidney tissue. 



222 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 31. Radular statistics for Neotricula 
cristel la. Mean ± standard deviation (range). N = 
number used. In mm except for width of central 
tooth in u.m. 

Males and females (N = 4) 



Shell length 
Radular length 
Radular width 
Total rows of teeth 
No. rows of teeth 

forming 
Central tooth width 



2.58±0.30 (2.16-2.88) 
0.54±0.05 (0.48-0.59) 
0.06±0.002 (0.056-0.060) 

99 ±3.4 (95-103) 

23±6.9 (16-32) 

12.9±0.8 (12.3-14) 



TABLE 33. Lengths (mm) of neural structures of 
Neotricula cristella. Mean ± standard deviation 
(range). N = 3. 



Cerebral ganglion 
Cerbral commissure 

Pleural ganglion 

Right (1)* 

Left 
Pleuro-supraesophageal 

connective (2)* 
Pleuro-subesophageal 

connective 
Supraesophageal 

ganglion (3)* 
Subesophageal ganglion 

Osphradio-mantle nerve 

RPG ratio* = 
2 - 1+2 + 3 



0.23±0.06 (0.16-0.28) 
0.05±0.01 (0.04-0.06) 

0.08±0.02 (0.06-0.10) 
0.10±0.04 (0.06-0.14) 

0.11 ±0.01 (0.10-0.12) 

0.09±0.06 (0.02-0.14) 
0.11±0.01 (0.10-0.12) 

0.11 ±0.01 (0.10-0.12) 
0.05±0.03 (0.02-0.08) 
0.36±0.01 (0.36-0.38) 




FIG. 67. Penis of Neotricula cristella. 



1 0-1 5 cm deep. The flow was slow; the water 
was clean and cool. The stream flows down 
from Nanyue Mountain. The bottom of the 
stream was paved with small rocks, sand, and 
leaves. At stream side was short, scrubby 
vegetation. There were some 50 snails per 
stone. 



Description 

Shell. The shells are small, ovate-conic, of 5.0 
to 5.5 whorls (Figs. 70A-D, 71 A, B). Because 
the apices of most mature snails are eroded, 
it is not possible to be precise about lengths; 



TABLE 32. Cusp formulae for the radular teeth of Neotricula cristella with the percent of radulae in which 
a given formula was found at least once. 



Central Teeth 



Lateral Teeth 



Inner Marginal Teeth 



Outer Marginal Teeth 



3-1-3 93% 
3-3 

2-1-2 21% 
2-2 



4-1-4 
3-3 



9% 



3-1[2]-3 


79% 


11 


— 


3-1[2]-4 
4-1[2]-3 


21% 
14% 


12 
13 
14 


14% 
43% 


4-1[2]-4 


7% 


15 


71% 






16 


71% 






17 


50% 






18 


21% 






X* = 15.1 ±2.7 
N = 138 



7% 
21% 
71% 
79% 
50% 



13.6±1.2 
N = 127 



'Mean ± standard deviation of cusp number for all teeth counted. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 223 




FIG. 68. Radula of Neotricula cristella. See text for details. 



224 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 69. Radula of Neotricula cristella. See text for details. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 225 




c IG u ^ K Shel l S ,° f: Ne ° tricula dianmenensis A-D. A. Holotype. Neotricula duplicata E-l from D85 collections 
F Holotype. Neotricula ///// J-L. J. Holotype. A = 3.32 mm; other shells printed at same scale. Shells not 
designated as holotypes are paratypes. 



226 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 71 . SEM photographs of shells (А, В) and opercula (С, D) of Neotricula dianmenensis. D. Inner surface. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 227 

TABLE 34. Shell measurements (mm) for Neotricula dianmenensis. Mean ± standard deviation (range). 
All except two shells were eroded, therefore lengths are less than for entire specimens, e = eroded. 
*, probably a male. 





Shells used for dissections 




Types 










*Small 




Females (3) 


Males (1) 


Holotype 


Paratypes (5) 


Paratype (1) 


No. Whorls 


3e 


5.5 


4e 


2.0-4.0e 


5.0 


Length (L) 


2.90±0.20 
(2.76-3.12) 


2.72 


3.32 


2.89±0.25 
(2.64-3.20) 


2.76 


Width (W) 


1.50±0.08 
(1.44-1.60) 


1.36 


1.64 


1.50±0.06 
(1.44-1.56) 


1.28 


L body whorl 


2.02±0.12 
(1.92-2.16) 


1.76 


2.20 


1.96±0.08 
(1.84-2.04) 


1.68 


L penultimate 


0.54±0.10 


0.40 


0.52 


0.52±0.05 


0.44 


whorl 


(0.48-0.62) 






(0.46-0.56) 




W penultimate 


1.10+0.10 


0.96 


1.16 


1.08±0.06 


0.96 


whorl 


(1.04-1.20) 






(1.00-1.16) 




W 3rd whorl 


— 


— 


0.80 


0.77±0.05 


0.68 




— 


— 




(0.72-0.80) N = 3 




L last three whorls 


2.90±0.20 
(2.76-3.12) 


2.72 


3.12 


2.85±0.19 
(2.64-2.96) N = 3 


2.40 


L aperture 


1.47±0.08 
(1.40-1.56) 


1.24 


1.56 


1.43±0.08 
(1.32-1.52) 


1.26 


W aperture 


0.91 ±0.02 
(0.88-0.92) 


0.80 


0.96 


0.91 ±0.03 
(0.88-0.94) 


0.80 


X 


0.42±0.05 
(0.40-0.48) 


0.40 


0.50 


0.43±0.08 
(0.32-0.52) 


0.36 


У 






0.16 


0.09±0.05 
(0.04-0.16) 


0.08 




FIG. 72. Uncoiled female Neotricula dianmenensis with head and kidney tissue removed. 



228 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 35. Dimensions (mm) or counts of non-neural organs and structures of Neotricula dianmenensis. 
N = number of snails used. L = length. Mean ± standard deviation (range). 



Females (N = 3) 



Males (N = 1) 



Body L 

Digestive gland L 
Gonad L 

Palliai oviduct L. 

= PO 
Bursa copulatrix L. 

= BU 
Duct of bursa L. 

BU -s- PO 

Seminal receptacle L. 

Duct of seminal 

receptacle L. (see text) 
Buccal mass L. 

Mantle cavity L. 

Osphradium L. 

= Os 
Gill L 

= G 
Os- G 

No. filaments 

Gf 2 L 

Gf 1 L. 

Total Gf L. 
= TGF 

Gf 2 + TGF 

Prostate L. 
Seminal vesicle L. 
Penis L. 



4.9±0.53 
(4.4-5.4) 
1.99±0.36 
(1.76-2.4) 
0.75±0.08 
(0.70-0.84) 
1.58±0.16 
(0.44-0.56) 
0.50±0.06 
(0.44-0.56) 
0.18 

(0.16, 0.20) N = 2 
0.32±0.07 
(0.26-0.40) 
0.15±0.01 
(0.14-0.16) 
Oto 0.12 

0.55±0.04 
(0.52-0.60) 

1.41 ±0.20 
(1.20-1.60) 

0.35±0.05 
(0.30-0.40) 

1.23±0.21 
(1.00-1.40) 

0.29±0.03 
(0.26-0.32) N = 4 

18.3±1.5 
(17-20) 

0.25 
(0.20, 0.30) N = 2 

0.21 N = 2 

0.46 N = 2 
no variation 
0.54 
(0.44, 0.65) N = 2 



3.9 
1.9 
1.1 



1.26 
0.34 
1.06 

17 
(males & females) 
(males & females) 

(males & females) 

1.0 
0.70 

1.54 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 229 

Omc 

Emc 




FIG. 73. Details and variation of bursa copulatrix complex of organs of Neotricula dianmenensis. A, same 
orientation as in Figure 72. B-D. Bursa complex flipped over to show where seminal receptacle enters the 
base of bursa, and variation. 



we can give no data on apical whorl charac- 
ters. Lengths probably range from 2.76 to 
3.44 mm (Table 34). There is possibly sexual 
dimorphism, with the males smaller. The ap- 
erture shape is distorted due to the pro- 
nounced aperture beak. There is a narrow but 
pronounced umbilicus. The whorls at the su- 
ture are smooth (not crenulated). SEM anal- 
ysis reveals a trace of spiral microsculpture 
below the sutures (also seen at 50X). There is 
an angulation of the inner lip just abapical to 
the pronounced beak tubercle. The inner lip is 
fused to the body whorl (most shells). The 
inner lip is slightly separated from the body 
whorl opposite the beak tubercle (some 
shells). The adapical end of the aperture is 
fused to the body whorl; there is a pro- 
nounced apertural beak some 0.26 mm long 



with a wide opening into the interior (0.12- 
0.16 mm wide gap); there is no internal notch 
groove. There is an apertural sinus. In side 
view, the outer lip is straight; it is slightly 
scooped forward. Facing the inner lip in side 
view, there is a strong lip deflection angle. 
There is no varix. In apertural view, the abapi- 
cal lip extends beyond the base of the shell 
0.43 ± 0.08 mm. 

The whorls are slightly convex and the su- 
tures sharply defined but shallow. 

External features. Details of the head are not 
available. The operculum (Fig. 71C.D) is dis- 
tinctive for its long, thin shape. The width to 
length ratio is 0.44 ± 0.03. As seen in Figure 
71 D, there may be an easily detached exter- 
nal layer. The internal attachment pad is pro- 



230 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 

Ast 




0.5 mm 



FIG. 74. Uncoiled male of Neotricula dianmenensis without head or kidney tissue. Some lobes of gonad cut 
away to show seminal vesicle that coils dorsal to gonad. Dashed line shows anterior limit of gonadal lobes. 



nounced (Fig. 71 D). The shape of the oper- 
culum reflects the inner lip angulation. 

Mantle cavity. Measurements and statistics 
are given in Table 35. Mantle cavity structures 
and relationships are normal for those of 
Neotricula. The osphradium is mid-gill; it is 
short. Gill filaments are normally developed; 
Gf 2 is normal length. Gf 2 does not have a 
pronounced crest. 

Female reproductive system. The body of an 
uncoiled female without head and with kidney 
tissue removed is shown in Figure 72. Mea- 
surements of organs are given in Table 35. 
Important features are: (1) The gonad is pos- 
terior to the stomach; it consists of few lobes. 
(2) The bursa (Bu) is clearly seen posterior to 
the posterior palliai oviduct (Ppo). (3) Sperm 
enter the system at the posterior end of the 
mantle cavity (Emc) by passing into the sper- 
mathecal duct (Sd) that bypasses the pericar- 
dium. The spermathecal duct runs a short dis- 
tance to open into the bursa (Bu, Fig. 73A) 
dorsal to the duct of the bursa (Dbu). (4) The 



sperm duct (Sdu) runs from the bursa (Bu) to 
the oviduct (Ov); there is no duct of the bursa. 
(5) The seminal receptacle (Sr) arises from 
the spermathecal duct where the latter joins 
the bursa (Fig. 73B-D). (6) The duct of the 
seminal receptacle varies in length from (Sr 
fused to the spermathecal duct, Fig. 73C) to 
moderately long (Fig. 73D). (7) The oviduct 
runs from gonad to palliai oviduct without 
making a loop or twist. (8) The oviduct opens 
into the palliai oviduct close to the posterior 
end of the palliai oviduct. (9) The bursa cop- 
ulatrix is round and short. (10) The length of 
the albumen gland is standard (based on one 
individual for which this could be discerned, 
ratio of 0.47). 

Male reproductive system. The body of an un- 
coiled male is shown in Figure 74 without 
head and with kidney tissue removed. The 
anterior lobes of the gonad were removed to 
reveal the seminal vesicle. Measurements 
are given in Table 35. Important features are: 
(1) The gonad consists of numbers of finely 
divided lobes that drain into a vas efferens 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 231 




FIG. 75. The penis (B). (A), orientation of the base 
of the penis (Bp) of Neotricula dianmenensis to the 
snout-neck mid-line (x). 



TABLE 36. Radula statistics for Neotricula dian- 
menensis. Mean ± standard deviation (range). In 
mm except the central tooth width in jim. N = 4. 
Shell lengths not available. 



Radular length 
Radular width 
Total rows of 

teeth 
No. rows of teeth 

forming 
Central tooth 

width 



0.45±0.06 

0.08±0.002 

58.8±4.2 

1.90±3.6 

14.1 ±0.6 



(0.36-0.48) 

(0.76-0.080) 

(56-65) 

(14-22) 

(13.4-14.9) 
N = 11 



(Ve, Fig. 74). (2) The posterior vas deferens 
arises from the vas efferens posterior to mid- 
gonad and immediately coils as the seminal 
vesicle (Sv) that is dorsal to the lobes of the 
gonad. (3) The seminal vesicle coils posterior 
to the stomach. (4) The gonad is entirely pos- 
terior to the stomach. (5) The prostate is mas- 
sive; it overlaps the posterior end of the man- 
tle cavity. The posterior prostate overlaps the 
entire style sac (Sts). (6) The penis is simple, 
without lobes; it has a papilla (Fig. 75B). (7) 
No ejaculatory duct was found. (8) The shaft 
of the penis arises from the neck to the right of 
the snout-neck mid-line (x, Fig. 75A) at an 
angle of 22°-23°. 

Digestive system. The digestive gland is pos- 
terior to the stomach (only slight overlap on 
the posterior chamber (Figs. 72, 74). The rad- 



TABLE 37. Cusp formulae for the radular teeth of Neotricula dianmenensis with the percent of radulae (N 
= 4) in which a given formula was found at least once. 



Central Teeth 



Lateral Teeth 



Inner Marginal Teeth 



Outer Marginal Teeth 
(N = 3) 



2-1-2 50% 



2-2 




3-1-3 


25% 


2-3 




3-1-3 


25% 


2-2 




2-1-3 


25% 



2-2 



3-1-4 


75% 


4-1-3 


50% 


4-1-2 


25% 


2-1-3 


25% 


2-1[2]-2 


25% 


3-1-2 


25% 


3-1-3 


25% 


3-1-4 


25% 



7 


25% 


8 


25% 


9 


25% 


10 


25% 


11 


100% 


12 


100% 


13 


75% 


14 


25% 


X* = 

N 


11.5±1.7 
= 38 



7 





8 





9 


33% 


10 


33% 


11 


66% 


12 


100% 


13 


66% 


14 





11.6±1.2 




30 



*Mean ± standard deviation of cusp number for all teeth counted. 



232 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



í^S 4 







ик**^ 









|\W 



UrflHE. 






^P" Wfi*t IGIf«* -ita 



FIG. 76. Radula of Neotricula dianmenensis. A, E. Segments of the radula. B-D, F-H. Central, lateral and 
inner marginal (C, D, H) teeth. See text for details. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 233 








FIG. 77. Radula of Neotricula dianmenensis. Outer marginals are featured in C, E, F. Dominant cusp of 
lateral tooth (i.e. the "1" of the 3-1-2) is either massive (D) or bifurcated (B). 

TABLE 38. Lengths (mm) of neural structures of Neotricula dianmenensis. Mean ± standard deviation 
(range). N = 4. 



Cerebral ganglion 


0.23±0.01 




Cerebral commissure 


0.07±0.01 




Pleural ganglia 

Right (1) 

Left 
Pleuro-supraesophageal 

connective (2) 
Pleuro-subesophageal 

connective 


0.13±0.01 

0.12- 

0.13±0.04 

0.08 N = 2 


(no variation) 
(0.10-0.18) 

(0.06-0.10) 


Supraesophageal ganglion (3) 


0.11 ±0.01 


(0.10-0.12) 


Subesophageal ganglion 


0.12- 




Osphradio-mantle nerve 


0.04+0.01 


(0.02-0.04) 


RPG ratio = 2-1 +2 + 3 


0.35±0.01 


(0.34-0.36) 



234 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 78. Paratypes of Neotricula duplicata from D87-2. Shell in Figure A ¡s 3.52 mm long; other shells printed 
at same scale. 



ula sac does not coil dorsal to the nerve ring. 
The radula is shown in Figures 76, 77; radular 
statistics are given in Tables 36 and 37. The 
radula is typical Triculini. The enlarged cusp 
of the lateral tooth (the "1" of 3-1-2, etc.) is 
frequently bifid or massive (Figs. 77A, B, D). 
The typical radular formula is: 

2(3)-1-(3)2 , 3(4)-1[2]-2 to 4, 10-13, 10-13. 
2-2 

Nervous system. Standard Triculinae. Mea- 
surements are given in Table 38. The RPG 
ratio shows the supraesophageal connective 
to be moderately concentrated. 

Remarks 

Conchologically, N. dianmenensis is most 
similar to N. duplicata and N. //7/7 (Figs. 153, 
1 54). However, it differs from the other two by 
having a distorted aperture shape (Table 2, 
char. 3), an angled inner lip (char. 13), and a 
pronounced beak tubercle (char. 24). Addi- 



tionally, it differs from N. duplicata by having a 
clearly open umbilicus (char. 4), an adapical 
apertural sinus (char. 11), a sinuate outer lip 
(char. 14), and a thin inner lip (char. 19). 

Anatomically the greatest similarity is to N. 
duplicata (Fig. 157), but it differs from that 
species in seven characters (15% of the 46 
comparable characters, Table 80). Neotric- 
ula. dianmenensis has few rows of teeth 
whereas N. duplicata has many (char. 42). 
The gonad of the former is posterior to the 
stomach; it overlaps the stomach in the latter. 
The spermathecal duct is comparatively long 
in the former, short in the latter (char. 19). 
There is no duct of the bursa in the former; it 
is long in the latter (char. 16). The osphradium 
is long in the former, short in the latter (char. 
9). 

Neotricula duplicata Davis & Chen 
sp. nov. 

Holotype. ZAMIP-M0035, Figure 70A. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 235 




FIG. 79. SEM photographs of shells of Neotricula duplicata from D85-79. B, D show the abapical lip 
deflection. С, E. Enlargements of apical whorls. 



Paratypes. ANSP 373152, A12668; 373151, 
A12667; ZAMIP M0003; Figure 70E, G-l; Fig- 
ure 78B-E. 

Type Locality. Huang-sha-ping Village, Jiang- 
nan Town, Anhua County, Yiyang Prefecture. 
28°21'24"E, 110°15'11"N. Figure 1, site 4. 
Collection numbers D85-79, 5 October 1985, 
Davis, Hoagland & Chen; D87-2, 18 March 
1987, Davis & Chen. 

Etymology. Named for the duplication of an- 
atomical research effort on specimens col- 



lected from this population in both 1985 and 
1987, not realizing in 1987 that the population 
had been dissected and analyzed in 1 985. 

Habitat 

Some 400 m from Zijiang River, a small 
stream flowed from a hill through rice fields. 
The stream was about 10 cm deep, 14 cm 
wide with a mud substratum. Snails were col- 
lected in the upper narrow shaded part of the 
stream from under stones. Associated mol- 
luscan fauna: Gyraulus sp. 



236 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 80. SEM photographs of shells of Neotricula duplicata from D87-2. C-E. Enlargements of apical 
whorls. 



Description 

Shell. Shells are illustrated in Figures 70E- 
I, 78A-E, 79, 80. They are small, ovate-conic 
(Tables 39-41). Because most snails are 
eroded, it is difficult to analyze size classes 
based on whorl numbers. It is clear that there 
are two size classes of mature snails (Table 
39); females are larger than males (Table 41 ). 
Additionally, there appear to be two size 
classes within each sex (Table 41). The size 
range based on uneroded shells is 2.72-3.88 



mm. The aperture is pyriform. Adapically 
there is a strong apertural beak; there is no 
beak tubercle. There may or may not be an 
internal notch groove. 

The whorls at the suture are smooth (not 
crenulated). There is an umbilical chink. SEM 
analysis reveals faint spiral microsculpture at 
the shoulders of the whorls after the second 
whorl (Fig. 80D, E). 

The inner lip is straight to arched, narrowly 
separated from the body whorl, and uniformly 
thick. The adapical end of the aperture is only 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 237 



TABLE 39. Measurements in mm of D85-79 shells of Neotricula duplicata. Mean 
(range), e = eroded apical whorls; ( ) = number measured. Sex unknown. 



standard deviation 



Holotype 



Paratypes 



large class 


large class 


small class 


4e 


3e-4e (3) 


3e-5e(5) 


3.44 


3.03±0.06 


2.87±0.10e 




(3.00-3.10) 


(2.72-3.00) 


1.72 


1.57±0.10 


1.46±0.04 




(1.50-1.68) 


(1.44-1.48) 


3.12 


2.91 ±0.16 


2.67±0.06 




(2.80-3.10) 


(2.56-2.72) 


2.24 


2.12±0.14 


1.94±0.02 




(2.04-2.28) 


(1.92-2.96) 


0.52 


0.45±0.02 


0.47±0.02 




(0.44-0.52) 


(0.44-0.48) 


1.10 


0.99±0.07 


0.97±0.02 




(0.92-1.06) 


(0.96-1.00) 


0.76 


0.66±0.05 


0.65±0.02 




(0.60-0.70) 


(0.64-0.68) 


1.60 


1.51±0.12 


1.36±0.04 




(1.44-1.64) 


(1.32-1.40) 


1.08 


0.95±0.08 


0.86±0.04 




(0.88-1.04) 


(0.80-0.88) 


0.48 


0.48±0.07 


0.44±0.06 




(0.40-0.52) 


(0.36-0.52) 


0.20 


0.15±0.06 


0.15±0.04 




(0.08-0.16) 


(0.10-0.20) 



No. Whorls 
Length (L) 

Width (W) 

L last three whorls 

L body whorl 

L penultimate whorl 

W penultimate whorl 

W 3rd whorl 

L aperture 

W aperture 

x 

У 



TABLE 40 Measurements in mm of paratype shells of Neotricula duplicata from D87-2. Mean 
deviation (range), e = eroded apical whorls; ( ) = number measured. Sex unknown. 



standard 



No. Whorls 


5e 


5.5 (3) 


6.0 (2) 


Length (L) of eroded shell 


3.44 






Length of uneroded shells 




2.89±0.20 


3.53 






(2.72-3.12) 


(3.46,3.60) 


Width (W) 


1.60 


1.37±0.09 


1.61 






(1.32-1.48) 


(1.60,1.62) 


L last three whorls 


2.92 


2.59±0.15 


3.10 






(2.48-2.76) 


(3.00,3.20) 


L body whorl 


2.08 


1.87±0.08 


2.18 






(1.80-1.96) 


(2.08,2.28) 


L penultimate whorl 


0.56 


0.45±0.03 


0.54 






(0.42-0.48) 


(0.52,0.56) 


W penultimate whorl 


1.06 


0.91 ±0.05 


1.10 






(0.88-0.96) 


(1.08,1.12) 


W 3rd whorl 


0.72 


0.62±0.05 


0.72 






(0.58-0.68) 


No variation 


L aperture 


1.52 


1.33±0.09 


1.52 






(1.28-1.44) 


(1.44,1.60) 


W aperture 


1.00 


0.85±0.06 


0.96 






(0.84-0.92) 


No variation 


X 


0.52 


0.37±0.05 


0.42 






(0.32-0.40) 


(0.40,0.44) 


У 


0.16 


0.10±0.02 


0.14 






(0.08-0.12) 


(0.12,0.16) 



238 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 41 . Measurements (mm) of shells of Neotricula duplicata from both collections where animals were 
used for dissections. Mean ± standard deviation (range). All except two shells were eroded, therefore lengths 
are less than for entire specimens, e = eroded. *, probably a male. ( ) = number measured. 





Females 






Males 




No. Whorls 


2e-5e(3) 


5.5(1) 


6.0(1) 


4e-5e (2) 


5.0(1)* 


5.5 (2) 


Length (L) of eroded 


3.31 ±0.45e 






3.22 






shells 


(2.80-3.68) 






(3.12,3.32) 






Length of 




3.60 


3.88 




2.88 


3.12 


uneroded shells 












No var. 


Width (W) 


1.77±0.08 
(1.68-1.84) 


1.76 


1.76 


1.57 
(1.56, 1.58) 


1.48 


1.56 
No var. 


L last three whorls 


3.24 (N = 2) 
No var. 


3.20 


3.36 


2.88 
(2.76, 3.00) 


2.62 


2.76 
(2.72, 2.80) 


L body whorl 


2.36±0.08 
(2.28-2.44) 


2.36 


2.36 


2.10 
(2.08,2.12) 


1.96 


1.96 
(1.92,2.00) 


L penultimate 


0.69±0.34 


0.52 


0.62 


0.49 


0.40 


0.46 


whorl 


(0.48-1.08) 






(0.42, 0.56) 




(0.44, 0.48) 


W penultimate 


1.12±0.04 


1.12 


1.16 


1.05 


0.96 


1.01 


whorl 


(1.08-1.16) 






(1.02, 1.08) 




(1.00, 1.02) 


W 3rd whorl 


0.64 (N = 2) 
(0.56, 0.72) 


0.78 


0.82 


2.88 
(2.76, 3.00) 


2.62 


2.76 
(2.72, 2.80) 


L aperture 


1.60±0.06 
(1.56-1.68) 


1.60 


1.64 


1.46 
(1.42, 1.48) 


1.32 


1.38 
(1.36, 1.40) 


W aperture 


1.07±0.08 
(0.88-0.92) 


1.08 


1.06 


0.92 
No var. 


0.88 


0.94 
(0.92, 0.96) 




0.5 mm 

FIG. 81. Head of a male Neotricula duplicata from 
D85-72 showing the relationship of the base of the 
penis (Bp) to the mid-line of the snout-neck (x). 



slightly separated from the body whorl. There 
is no apertural sinus. In side view, the outer lip 
is straight; the outer lip is slightly scooped for- 
ward. In side view, the inner lip has a strong 
deflection angle, some 140°. There is no 
varix. The abapical lip just beyond the base of 
the shell 0.48 ± 0.07 mm; the abapical lip has 
a spout. 

SEM examinations of the apical whorls 
(Figs. 79C-E; 80C-E) show that the tip of the 
apical whorl is smooth, but that just beyond 
the tip the shell is minutely wrinkled. Coarse 
growth lines begin just before 1 .75 whorls. 

External features. The head is dark grey to 
black. There may or may not be white gran- 
ules close to the eyes extending posteriorly 
back along the neck (Wg, Fig. 81). When 
granules occur, they do not form a dense lu- 
nate mass or "eyebrow." 

Opercula from both collections are shown 
in Figures 82, 83. They are elongate-ovoid 
and are composed of discernable flaky lay- 
ers. The muscular attachment pad (Fig. 82B, 
D) is pronounced and wide (66% width of the 
operculum). 

Mantle cavity. Mantle cavity organs are 
shown in Figure 84; not all gill filaments are 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 239 




FIG. 82. Opercula of Neotricula duplicata from D85-79. A, С Outer surface; B, D. Inner surface. Note the 
external loose, flaky layer (A, C, D). 



240 DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 83. Opercula of Neotricula duplicata from D87-2. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 241 

TABLE 42. Lengths (mm) or counts of non-neural organs of Neotricula duplicata. Mean ± standard 
deviation (range). For D87-2 snails, N = 5 for females, N = 4 for males; for D85-79 snails, N = 5 for 
females, N = 3 for males unless stated otherwise. 







Females 




Males 




D87-2 




D85-79 




D87-2 


D5-79 


Body 


4.99±0.45 




4.90±0.51 




4.43±0.14 


4.31 ±0.02 




(4.46-5.70) 




(4.20-5.46) 




(4.30-4.56) 


(4.30-4.34) 


Digestive gland 


1.95±0.46 




2.03±0.20 




2.02±0.14 


1.92±0.14 




(1.30-2.50) 




(1.74-2.30) 




(1.86-2.20) 


(1.76-2.00) 


Gonad 


0.61 ±0.13 




0.75±0.05 




1.50±0.38 


1 .23 N = 2 




(0.48-0.74) N 


= 4 


(0.70-0.80) 




(1.20-2.10) 


(1.16, 1.30) 


Total palliai oviduct 


1.70±0.20 




2.04 ±0.34 




— 


— 


(= TPO) 


(1.50-1.90) N 


= 3 


(1.78-2.50) N 


= 4 






Bursa copulatrix 


0.47±0.12 




0.48±0.05 




— 


— 


(= Bu) 


(0.30-0.60) N 


= 4 


(0.44-0.56) 








Duct of bursa 


0.17±0.06 




0.10 




— 







(0.10-0.20) N 


= 3 


No var. N = 


2 






Bu 4- TPO 


0.32±0.12 
(0.20-0.43) N 


= 3 


0.23±0.04 
(0.18-0.30) 




— 


— 


Seminal 


0.21 ±0.07 




0.25±0.05 




— 





receptacle 


(0.16-0.30) 




(0.18-0.26) 








Duct of seminal 


0.06 




0.06 




— 





receptacle 


No var. N = 


2 


N = 1 








Mantle cavity 


1.51±0.18 




1.48±0.07 




1.33±0.04 


1.35±0.19 




(1.26-1.70) 




(1.40-1.56) N 


= 4 


(1.30-1.38) 


(1.14-1.52) 


Ctenidium 


1.27±0.19 




1.28±0.09 




1.07±0.09 


1.15±0.19 


(= CT) 


(1.00-1.50) 




(1.20-1.36) N 


= 4 


(1.00-1.20) 


(0.94-1.30) 


Osphradium 


0.37±0.13 




0.37±0.03 




0.35±0.06 


0.36±0.13 


(=OS) 


(0.20-0.48) 




(0.34-0.40) N 


= 3 


(0.28-0.36) 


(0.24-0.50) 


No. gill 


19.3±1.7 




18.0±1.4 




17.0±1.2 


23.6±7.1 


filaments 


(17-21) 




(16-19) N = 


4 


(16-18) 


(16-30) 


OS - CT 


0.30±0.11 




0.29 ±0.04 




0.33±0.07 


0.32±0.10 




(0.17-0.44) 




(0.23-0.28) N 


= 3 


(0.23-0.40) 


(0.20-0.38) 


Gf 1 


0.20±0.09 




0.28±0.05 




— 


— 




(0.12-0.34) N 


= 8 


(0.24-0.36) N 


= 4 






Gf 2 


0.24±0.07 




0.23 ±0.03 




— 


— 




(0.20-0.30) N 


= 8 


(0.20-0.26) N 


= 4 






Total Gf 


0.46±0.08 




0.50±0.05 




— 







(0.32-0.56) N 


= 8 


(0.44-0.50) N 


= 4 






Gf 2 h- G^ + Gf 2 


0.43±0.15 
(0.25-0.63) 




0.54±0.07 
(0.48-0.64) N 


= 4 


— 


— 


Prostate 










0.85±0.13 
(0.70-1.00) 


0.87±0.06 
(0.80-0.90) 


Seminal vesicle 


— 




— 




0.53±0.22 
(0.20-0.70) 


0.45±0.09 
(0.40-0.56) 


Penis 






— 




1.16±0.27 
(0.90-1.54) 


1.51 ±0.22 
(1.36-1.76) 



illustrated. Measurements and counts are 
given in Table 42. The osphradium is approx- 
imately mid-gill; it is short (ratio of 0.29). The 
shape of the osphradium is lunate, not oval. 
The number of gills ranges from 17 to 30. Gill 



filament section Gf 2 is normal; the length of 
the longer gill filaments is 0.46 to 0.50 mm 
long. The shape of the gill leaflet in side view 
is moderate to high domed. There is no cir- 
cular patch of white or yellowish granules just 



242 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Eme 




FIG. 84. Mantle cavity organs of Neotricula duplicata from D89-79. Dashed line is the trajectory for mea- 
suring length of gill and mantle cavity. Only three central gill filaments are illustrated. 



Apo 




DÍ 0.5mm 

FIG. 85. Uncoiled female Neotricula duplicata with head and kidney tissue removed; from D85-79. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 
Pst 
Di 



243 



Ast 




FIG. 86. Uncoiled female Neotricula duplicata with head and kidney tissue removed from D87-2. 



anterior to the osphradium next to the neck- 
mantle collar (Ma) juncture. 

Female reproductive system. The bodies of 
uncoiled females without head or kidney tis- 
sue are shown in Figures 85, 86. Organ mea- 
surements are given in Table 42. Important 
features are: (1 ) The gonad is posterior to the 
stomach; it is relatively short and of few lobes. 
(2) The albumen gland (Ppo) is of normal 
length. (3) The bursa copulatrix (Bu) is clearly 
visible posterior to the albumen gland; the 
bursa is small and round to oval. (4) The ovi- 
duct runs from the gonad to the albumen 
gland without coiling. (5) The bursa copulatrix 
complex of organs is shown in Figures 87, 88. 
Figure 87A is positioned exactly as in Figures 
85, 86. The spermathecal duct (Sd) opens 
into the posterior mantle cavity; it is short and 
swollen in most specimens. (6) The seminal 
receptacle (Sr) arises from the dorsal surface 
at the juncture of the duct of the bursa and the 
spermathecal duct. (7) The duct of the semi- 
nal receptacle (Dsr) slowly increases in diam- 
eter to form a club-shaped (not spherical) 
storage sac that lies dorsal to the bursa cop- 



ulatrix. In some specimens, it is clear that the 
seminal receptacle is a U-shaped continua- 
tion of the duct of the bursa and that the sper- 
mathecal duct attaches at the bottom of the 
"U" (Fig. 88C). (8) The sperm duct (Sdu) 
arises from the duct of the bursa on the ven- 
tral side just posterior to the duct of the sem- 
inal receptacle. 

Male reproductive system. The posterior sec- 
tion of an uncoiled male with kidney tissue 
removed is shown in Figure 89. It is the same 
in specimens from both D85 and D87 collec- 
tions. The outline of the gonad (Go) is shown 
(dashed line) but most of the lobes of the go- 
nad are removed to show the seminal vesicle 
(Sv) coiled dorsal to the gonad. Measure- 
ments of organs are given in Table 42. Impor- 
tant features are: (1) The gonad overlaps the 
posterior part of the posterior chamber of the 
stomach (Pst). (2) The prostate (Pr) overlaps 
the posterior end of the mantle cavity (Emc) 
and covers the style sac. (3) The seminal ves- 
icle, while coiling dorsal to the gonad, does 
not extend over the stomach. (4) The anterior 
vas deferens leaves the prostate (Pr) close to 



244 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 87. Details and variation of bursa copulatrix complex of organs of Neotricula duplicata from D85-79. 
Figure A is in same orientation as in Figures 85 and 86. B. Swelling of spermathecal duct (Sd) seen in most 
individuals. С Bursa complex seen in В flipped over 180° so that Emc is to left; this view shows juncture of 
duct of the bursa (Dbu), seminal receptacle (Sr) and spermathecal duct (Sd). D. As in A, but with oviduct (Ov) 
pulled to left to show the juncture of Dsr and Sd with duct of bursa. 



TABLE 43. Radular statistics for Neotricula duplicata. Mean á 
for the width of the central tooth in (im. N = number counted. 



standard deviation (range). In mm except 



D85-77 



D87-2 



Shell length (not eroded) 
Radula length 
Radula width 
Total rows of teeth 
No. rows of teeth forming 
Central tooth width 



3.20±0.26 
(2.84-3.60) N = 8 
0.55±0.04 
(0.66-0.60) N = 11 
0.08±0.004 
(0.072-0.088) N = 11 
70.4±4.1 
(61 -74) N = 8 
9.0±2.1 

(7-13) N = 8 
15.5±0.8 
(14.6-16.8) N = 9 



3.43±0.21 
(4.44-5.20) 
0.57±0.04 
(0.52-0.62) N = 10 
0.08±0.006 
(0.078-0.092) N = 10 
60.1 ±8.0 
(46-69) N = 10 
10.3±3.3 

(6-15) N = 10 
17.1 ±1.5 
(15.8-19.0) N = 7 



the posterior end of the mantle cavity (Emc). 
(5) The penis is simple, with a small evertible 
papilla (Fig. 90B). In some specimens, the pa- 



pilla is withdrawn and cannot be seen (living 
tissue, fresh microscope mount). In others, 
only the tip of the papilla can be seen being 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 245 

Emc 



Dbu /Oov 




Sdu 



Dbu 




В 




Sdu Sd 



0.5mm 

FIG. 88. Details and variation of bursa copulatrix complex of organs of Neotricula duplicata from D87-2. 
Figure A is in same orientation as in Figures 85 and 86. B-C. Variation in configuration of seminal receptacle 
(Sr). B. Bursa complex rotated in the direction of arrow to show interconnections of Dbu, Csd, Sr and Sd. С 
Dorsal side of bursa complex to show that duct of bursa (Dbu) makes a U-shaped bend into duct of seminal 
receptacle (Dsr). Swollen short spermathecal duct (Sd) enters bottom of U-shaped bend. Note: Common 
sperm duct (Csd) is that portion of duct extending from duct of bursa between Sdu and Sd. Duct of bursa 
is defined in all papers as that duct extending from bursa copulatrix (Bu) anteriorly to point where another 
sperm duct connects to it, e.g. the Sdu. 



pushed out and then withdrawn. (6) The base 
of the penis (Bp, Figs. 81 , 90A) is oriented on 
the neck in diverse ways. The angle of the 
long axis of attachment varies from parallel to 
the mid-line of the snout-neck (x) through 90°. 
The base may overlap the mid-line or be to 
the right of mid-line. (7) No ejaculatory duct is 
seen in the base of the penis or in the neck. 

Digestive system. The digestive gland is pos- 
terior to the stomach in both sexes. However, 
in some males of the D87-2 collection both 
digestive gland and gonad covered the pos- 
terior chamber of the stomach. 



Radular statistics are given in Tables 43 
and 44. The most commonly encountered for- 
mula in the D85-79 collection was: 

2-1-2 ; 3-1[2]-3(2); 12-15; 11-14; 
2(3)-(3)2 

in the D87 2 collection it was: 

3(2)-1-(2)3; 3(4)-1-3; 12-14; 11-13. 
3(2)-(2)3 

SEM photographs of radulae from snails of 
both collections are given in Figures 91-94 
No significant differences occur between the 



246 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 
Pst Ast 




FIG. 89. Uncoiled male of Neotricula duplicata from D85-79. Anterior part of mantle cavity omitted. Posterior 
lobes of the gonad (Go) removed to show coiled seminal vesicle (Sv) dorsal to gonad. Dashed line indicates 
extent of gonad. 



snails of the two collections; the tooth mor- 
phologies are the same and are typical of the 
generalized triculine type. Of note is the split 
or fork in the dominant central cusp of the 
lateral tooth (i.e. the "1" of the 3-1-3) (Fig. 
91 C, 92C, D, 93G, 94C). 

The stomach has a noteworthy character. 
The anterior chamber of the stomach (Ast, 
Fig. 95) has at the anterior and posterior 
edges prominent yellow ridges (Yri). 

Nervous system. Measurements of neural 
structures are given in Table 45. The RPG 
ratio indicates a moderately concentrated 
dorsal aspect of the nerve ring. 

Remarks 

These detailed anatomical studies of a sin- 
gle population collected two years apart pro- 
vide a unique opportunity to assess (1) how 
well methods provide an adequate assess- 
ment of qualitative and quantitative character- 
stokes. The 1987 study is a control for the 
1985 study, particularly as it was not remem- 
bered in 1987 that the 1985 assessment had 



been made. (2) Variation due to sampling (in 
part) and actual quantitative changes that 
could have occurred in two years. This is the 
first time that we have obtained an indication 
of the amount of variation that might be ex- 
pected in such a study. This is of considerable 
help in assessing how much difference may 
actually exist when comparing two popula- 
tions, each studied only once, e.g. between 
N. cristella and N. dianmenensis of this 
monograph. 

Overall, there was excellent agreement in 
qualitative anatomy when comparing the two 
year classes. There was significant variation 
in cusp counts between year classes, partic- 
ularly involving the marginal teeth. More vari- 
ation occurred in central tooth cusp numbers 
in the 1987 year class. 

Conchologically, N. duplicata is most simi- 
lar to N. dianmenensis, N. //7/7, and Tricula 
gredleri (Figs. 153, 154). Differences from N. 
dianmenensis are given on page 234. Neotric- 
ula duplicata differs from T. gredleri in three 
characters (10%). The former has spiral mi- 
crosculpture lacking in the latter (char. 6). The 
former has an abapical spout lacking in the 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 247 



TABLE 44. Cusp formulae for the radular teeth of Neotricula duplicata with the percent of radulae in 
which a given formula was found at least once. 



Central Teeth 
(N = 10) 


Lateral Teeth 


Inner Marginal 
Teeth 


Outer Marginal 
Teeth 


2-1-2 
3-3 


70% 


3-1[2]-3 


D85-79 
100% 


9 


— 


9 


10% 


2-1-2 
2-2 


30% 


3-1[2]-2 


50% 


10 


10% 


10 


20% 


2-1-2 
3-2 


30% 


4-1[2]-3 


30% 


11 


20% 


11 


60% 


2-1-2 
2-3 


20% 


2-1[2]-3 


20% 


12 


50% 


12 


70% 


3-1-3 
2-2 


10% 






13 


100% 


13 


60% 


3-1-2 
2-2 


10% 






14 


90% 


14 


40% 










15 


50% 


15 


— 



3-1-3 


60% 


3-3 




2-1-2 


50% 


3-3 




2-1-2 


40% 


2-3 




3-1-3 


30% 


2-2 




2-1-2 


20% 


3-2 




2-1-2 


10% 


2-2 




3-1-3 


10% 


2-3 




3-1-2 


10% 


3-3 




2-1-2 


10% 


3-2 




2-1-3 


10% 



D87-2 



3-1[2]-3 100% 

4-1[2]-3 40% 

3-1[2]-4 30% 

2-1[2]-3 10% 

3-1[2]-2 10% 



16 30% 

1 7 20% 

X* = 13.6±1.5 
N = 108 



11 — 

12 70% 

13 80% 

14 60% 

15 20% 

X = 12.9±0.7 
N = 100 



16 — 

17 — 

12.0±1.1 
120 



1 1 40% 

12 90% 

13 30% 

14 20% 

15 — 

12.1±1.3 
N = 100 



3-3 



'Mean ± standard deviation of cusp number for all teeth counted. 



248 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Bp- 




FIG. 90. A. The position of base of the penis of Neo- 
tricula duplicata relative to the snout-neck mid-line 
(x). B. Penis. 



oval operculum with two or more layers (chars. 
2, 3); the latter has an ovate operculum with 
one layer. The opercular attachment pad is 
very wide in the former, wide in the latter 
(char. 4). Gill filament section Gf 2 is of me- 
dium length in the former, long in the latter 
(char. 8). The bursa shape is ovoid in the 
former, round in the latter (char. 15). The 
spermathecal duct is short in the former, long 
in the latter (char. 19). The spermathecal duct 
runs directly anterior from the duct of the 
bursa to the mantle cavity in the former; it 
slants away at an angle in the latter (char. 20). 
The vas deferens bends away from the pros- 
tate at the posterior end of the mantle cavity in 
the former, at mid-prostate in the latter (char. 
31). Neotricula lilii has very many rows of 
teeth on the radula; N. duplicata has many 
(char. 42). 

Neotricula lilii Chen & Davis, sp. nov. 

Holotype. ZAMIP-M0002, Figure 70J. 

Paratypes. ANSP 373138, A12654, Figure 70 
K, L Figure 96A-D. 

Type Locality. Chang Wang Village, Chuan- 
xing Town, Lingxian County, Zhuzhou Prefec- 
ture. 26°17'18"N, 113°41'3"E. Figure 1, Site 
2. 

Etymology. Named for Dr. Li Li, Zhuzhou Pre- 
fectural Epidemic station, who collected this 
species. 

Habitat 



latter (char. 12). The latter has an adapical 
beak tubercle lacking in the former (char. 24). 
Anatomical differences are considerable, 
those used to define the genera among oth- 
ers (Fig. 157). 

Conchologically, N. duplicata differs from N. 
lilii by having a wrinkled protoconch, not a 
smooth one (char. 8); having an adapical 
beak instead of a notch (char. 9); having a 
straight to arched inner lip, not a sinuate one 
(char. 13); having a thick inner lip, not a thin 
one (char. 19); by lacking a slight varix found 
in the latter (char. 23); and by lacking an 
adapical outer lip angle seen in the latter 
(char. 25). Anatomically, these species differ 
in nine characters (19%) (Figs. 157, 158). The 
differences are: The former has an elongated 



The assigned field collection number was 
D85-74. Snails came from a mountain stream 
550 m above sea level. The stream was 10 to 
15 cm deep, with small rocks, leaves and 
mud. There was low vegetation beside the 
stream. 

Description 

Shell. The shells are small, ovate-conic, with 
5.5 to 6.0 whorls (Figs. 60A-B, 70J-L, 96A- 
B). Measurements are given in Tables 46, 47. 
Limited data suggest that females are larger 
than males (Table 47) with the lengths of the 
last three whorls of females ranging from 
2.98-3.28; those for males, 2.72-2.80. The 
aperture is pyriform. Adapically there is a wide 
apertural notch; there is no beak tubercle. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 249 




m 






зъ 




FIG. 91 . Radula of Neotricula duplicata from D85-79. Centrals, laterals and inner marginals featured in В, С 
D. Outer marginals. 



TABLE 45. Lengths (mm) of neural structures of Neotricula duplicata. Mean ± standard deviation 
(range). * = neural elements measured to calculate the RPG ratio. N = number measured. 





D85-79 (N = 4) 


D87-2 (N = 5) 


Cerebral ganglion 


0.26±0.03 


0.24+0.03 




(0.22-0.30) 


(0.24-0.26) 


Cerebral commissure 


0.05+0.02 


0.04+0.01 




(0.02-0.06) 


(0.03-0.06) 


Pleural ganglion 






Right (1)* 


0.14+0.03 


0.13+0.02 




(0.10-0.16) 


(0.10-0.16) 


Left 


0.12+0.03 


0.12+0.01 




(0.08-0.14) 


(0.10-0.12) 


Pleuro-supraesophageal 


0.14+0.04 


0.11+0.02 


connective (2)* 


(0.10-0.18) 


(0.10-0.14) 


Pleuro-subesophageal 


0.06+0.05 


0.06+0.02 


connective 


(0.00-0.10) 


(0.03-0.08) N = 4 


Supraesophageal 


0.13+0.02 


0.12+0.02 


ganglion (3)* 


(0.10-0.14) 


(0.10-0.14) 


Subesophageal 


0.11+0.02 


0.11+0.01 


ganglion 


(0.10-0.12) 


(0.10-0.12) N = 4 


Osphradio-mantle nerve 


0.08+0.01 


0.08+0.04 




(0.06-0.08) 


(0.00-0.12) 


RPG ratio (2-1+2 + 3)* 


0.34+0.08 


0.31+0.03 




(0.26-0.45) 


(0.28-0.35) 



250 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 92. Radula of Neotricula duplicata from D85-79. Centrals, laterals and inner marginals featured in B-D. 
E, F. Outer marginals. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 251 




FIG. 93. Radula of Neothcula duplicata from D87-2. Centrals, laterals and inner marginals featured in B, C, 
E, G. D, H = outer marginals. A-D = males; E-H = females. 



252 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 





£!2S^2 


/TÍ 4 


№¡ 


1 ""»IT < 


jF^* 


?3 




ш^кк-^. 


4^^^.\ \ 


/i4*l 


!.-^^ж\ 




IffW/^i 




FIG. 94. Radula of female Neotricula duplicata from 
D87-2. 



Pst Ast, 




Od¡ 



FIG. 95. Ventral aspect of the stomach of Neotricula 
duplicata in same orientation as in Figures 85, 86. 



There is no internal notch groove. There is a 
pronounced adapical outer lip angle. 

The whorls at the sutures are smooth (not 
crenulated). There is an umbilicus. SEM anal- 
ysis reveals faint spiral microsculpture at the 
shoulders of the whorls after 1 .75 whorls (Fig. 
96A, C). 

The inner lip is slightly sinuate and clearly 
separated from the body whorl in most spec- 
imens; it is uniformly thickened. In some 
(< 20%), the adapical inner lip is fused to the 
body whorl. The adapical end of the aperture 
is only slightly separated from the body wall. 
An apertural sinus is apparent examining the 
outer lip in side view. In side view, the outer 
lip, abapical to the sinus, is straight. The outer 
lip is slightly scooped forward. In side view, 
the inner lip has a strong deflection angle. 
There is a slight varix. There is an abapical 
spout. The abapical lip projects beyond the 
base of the shell 0.50 ± 0.05. SEM examina- 
tion of the shell reveals that the apical whorl is 
smooth. (Fig. 96D). 

External features. The head is grey. There 
are no granules about the eyes and no 
"eyebrow." The operculum is corneous and 
paucispiral. The inner lip edge is straight abapi- 
cally and slightly sinuate towards the aper- 
tural beak end (adapically). The muscle at- 
tachment callus is pronounced and narrow, 
only 40% of the operculum width. A porous, 
regularly pitted surface of the callus was 
found in most specimens (75%) (Fig. 96E-H). 

Mantle cavity. Measurements and counts of 
mantle cavity structures are given in Table 48. 
The ovoid osphradium is mid-gill; it is short. 
There are 20-24 gill filaments. Gf 2 is long. 
The length of the longest filament is 0.51 mm. 
The larger gill filaments are moderately 
domed. There is no spherical patch of white 
granules anterior to the osphradium where 
the mantle collar joins the neck. 

Female reproductive system. The body of an 
uncoiled female without head or kidney tissue 
is shown in Figure 97. Organ measurements 
are given in Table 48. Important features are: 
(1) The gonad (Go) is posterior to the stom- 
ach; it is short and consists of few groups or 
bundles of lobes. (2) The bursa copulatrix 
(Bu) is spherical and prominent posterior to 
the albumen gland (Ppo). (3) The bursa is 
short. (4) The albumen gland is standard ( = 
normal) length. (5) The bursa copulatrix com- 
plex of organs is shown in Figure 98A, В in 
the same orientation as in Figure 97. The nar- 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 253 




FIG. 96. SEM photographs of shells and operculum of Neotricula lilii. C, D. Enlargements of apical whorls. 
E-H. Inner surfaces of four opercula; insets provide enlargements of attachment pads to reveal micro- 
stucture. 



254 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 46. Shell measurements (mm) for the types of Neotricula lilii. Mean 
e = eroded apical whorls. N = number measured. 



standard deviation (range). 



Holotype 



Paratypes 



Larger (N = 1) 


Smaller (N = 4) 


4e 


4e to 5.5 


3.20 


2.98±0.18 




(2.76-3.20) 


1.68 


1.51±0.10 




(1.38-1.62) 


3.00 


2.70±0.11 




(2.54-2.80) 


2.20 


1.90±0.09 




(1.76-1.96) 


0.48 


0.49±0.02 




(0.48-0.52) 


1.12 


1.03±0.05 




(0.96-1.08) 


0.80 


0.72±0.04 




(0.68-0.76) 


1.60 


1.39±0.10 




(1.24-1.48) 


1.04 


0.93±0.06 




(0.86-1.00) 


0.56 


0.50±0.05 




(0.44-0.56) 


0.20 


0.15±0.04 




(0.12-0.20) 



No. Whorls 
Length (L) 

Width (W) 

L last three 

whorls 
L body whorl 

L penultimate 

whorl 
W penultimate 

whorl 
W 3rd whorl 

L aperture 

W aperture 

x 

У 



5.5 
3.20 

1.52 

2.76 

1.96 

0.48 

0.98 

0.72 

1.40 

0.92 

0.52 

0.12 



TABLE 47. Shell measurements (mm) for Neotricula lilii used for anatomical work. Mean ± standard 
deviation (range), e = eroded apical whorls. N = number measured. 



Females 



Larger Size Class (N = 3) 



Small Size Class (N = 1) 



Males 



(N = 3) 



No. Whorls 


4e-6.0 


Length (L) 


3.43±0.19 




(3.20-3.56) 


Width (W) 


1.63±0.08 




(1.56-1.72) 


L last three 


3.11±0.15 


whorls 


(2.98-3.28) 


L body whorl 


2.21 ±0.12 




(2.08-2.28) 


L penultimate 


0.55±0.04 


whorl 


(0.52-0.60) 


W penultimate 


1.13±0.06 


whorl 


(1.08-1.20) 


W 3rd whorl 


— 


L aperture 


1.59±0.10 




(1.48-1.68) 


W aperture 


0.99±0.02 




(0.96-1.00) 


X 


0.52±0.06 




(0.44-0.56) 



5.75 
3.12 

1.48 

2.72 

1.92 

0.48 

1.00 

1.32 
0.96 
0.52 



3e-4e 



2.89: 

(2.76- 
1.47: 

(1.40- 
2.76: 

(2.72- 
1.95: 

(1.88- 

0.49: 

(0.44- 

1.03: 

(1.00- 



0.12 
3.00) 
0.06 
1.52) 
0.04 
2.80) 
0.06 
2.00) 
0.04 
0.52) 
0.04 
1.08) 



1.35±0.08 
(1.28-1.44) 
0.89±0.02 
(0.88-0.92) 
0.45±0.02 
(0.44-0.48) 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 255 



Ast 




1.0 mm 

FIG. 97. Uncoiled female Neotricula lilii with head and kidney tissue removed. 



256 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




Eme 




FIG. 98. Details and variation of bursa copulatrix 
complex of organs of Neotricula lilii. Figure A is in 
same orientation as in Figure 97. B. Bursa flipped 
over to show piace where seminal receptacle (Sr) 
enters common sperm duct (Csd). С Variation in 
size and shape of seminal receptacle. 



row spermathecal duct dilates to form the 
common sperm duct (Csd) from which opens 
the sperm duct (Sdu) and the seminal recep- 
tacle (Sr) (Fig. 98 B). (6) The sperm duct is 
very short and to the left of the common 
sperm duct. (7) The seminal receptacle arises 
some 45° from the opening of the sperm duct; 
it arises from the right dorso-lateral side of the 
sperm duct. (8) The seminal receptacle varies 
in shape from elliptical to sub-spherical (Fig. 
98C); it has a very short duct (Dsr). (9) The 
duct of the bursa (from the bursa to openings 
of Dsr and Sdu) is extremely short. (10) The 
distance from the posterior end of the mantle 
cavity (Emc) to the bursa is short, only 0.16 
mm to 0.32 mm. The spermathecal duct — 
common sperm duct run straight posteriorly 
from the end of the mantle cavity to the bursa. 



Male reproductive system. A section of an un- 
coiled male without kidney tissue is shown in 
Figure 99. The anterior gonadal lobes are re- 
moved to show the seminal vesicle (Sv). Or- 
gan measurements are given in Table 48. Im- 
portant features are: (1) The gonad (Go) 
slightly overlaps the posterior chamber of the 
stomach. (2) The seminal vesicle (Sv) arises 
from the vas efferens (Ve) at mid-gonad to 
slightly posterior to mid-gonad. (3) The pros- 
tate (Pr) overlaps the posterior end of the 
mantle cavity and covers half the style sac 
(Sts). (4) The anterior vas deferens (Vd 2 ) di- 
verges from the prostate slightly anterior to 
the posterior end of the the mantle cavity. (5) 
The penis is simple with a papilla (Pa) that 
everts (Fig. 100B). There is a massive con- 
centration of white granules along the anterior 
convex edge. (6) The penis arises to the right 
of the snout-neck mid-line (Fig. 100A). The 
long axis of the penial base (Bp) is swollen at 
its base increasing the diameter from 0.14 
mm to 0.22 mm. (7) No ejaculatory duct is 
seen in the penial base or neck. 

Digestive system. The digestive gland is pos- 
terior to the stomachs of both males and fe- 
males. Radular statistics are given in Tables 
49 and 50. The radula is illustrated in Figures 
101 , 102. The central tooth is the generalized 
triculine type (Fig. 101B, E). The dominant 
cusp of the lateral tooth (the "1" of the 3-1-3) 
is almost always bifurcated (Fig. 101). The 
most commonly encountered cusp formula is 

3(2)-1-(2)3; 3-[2]-3(4); 13-15; 12-14. 
2-2 

Nervous system. It is standard triculine. Mea- 
surements are given in Table 51. The RPG 
ratio of 0.44 indicates that the dorsal nerve 
ring is moderately concentrated. 

Remarks 

Conchologically, N. lilii is most similar to N. 
duplicata and N. dianmenensis (Figs. 153- 
155). The comparison of these three species 
is given in the remarks section for the two 
previously treated species. Anatomically, N. 
lilii is most similar to N. cristella, a species that 
has a very different shell (Figs. 154, 156-158). 
There are five differences (Tables 80, 81) 
(10%): N. lilii has an operculum with a single 
layer whereas the operculum of N. cristellás 
has two or more layers (char. 3). The former 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 257 



TABLE 48. Lengths (mm) or counts of non-neural organs and structures of Neotricula lilii. N = number 
of snails used. Mean ± standard deviation (range). 



Females (N = 4) 



Males (N = 3) 



Body 
Gonad 
Digestive gland 

Posterior palliai oviduct 

(= albumen gland) 
Anterior palliai oviduct 

( = capsule gland) 
Total palliai oviduct 

= OV 
Bursa copulatrix 

= BU 
Duct of BU 
BU ■*■ OV 

Seminal receptacle 

Duct of seminal receptacle 

Mantle cavity 

Gill (G) 

Osphradium (OS) 

OS ^ G 

No. of filaments 

Gf 2 

Gf, 

Total Gf = TGF 

Gf 2 - TGF 

Prostate 

Seminal vesicle 

Penis 



4.96±0.82 
(3.92-5.86) 
0.80±0.12 
(0.70-0.96) 
2.03±0.23 
(1.80-2.30) 
1.00±0.08 
(0.96-1.10) N = 3 
1.14±0.16 
(0.96-1.26) N = 3 
2.15±0.22 
(1.92-2.36) N = 3 
0.52±0.04 
(0.50-0.56) N = 3 

0.10 (N = 1) 
0.24±0.02 
(0.23-0.26) 
0.19±0.02 
(0.16-0.20) 
0.04 ±0.02 
(0.02-0.06) 
1.54±0.20 
(1.20-1.56) 
1.19±0.15 

(1.0-1.36) 
0.32±0.05 
(0.28-0.38) 
0.27±0.07 
(0.22-0.38) 
21.8±1.7 

(20-24) 
0.32±0.03 
(0.28-0.36) N = 6 
0.19±0.03 
(0.16-0.20) N = 6 
0.51 ±0.05 
(0.46-0.58) N = 6 
0.63±0.04 
(0.58-0.69) N = 6 



4.27±0.08 
(4.64-4.80) 
1.42±0.07 
(1.36-1.50) 
2.13±0.19 
(2.00-2.34) 



1.37: 

(1.30- 

1.22: 

(1.16- 

0.38: 

(0.36- 

0.31: 

(0.28- 

21.4: 

(21- 



0.07 

1 .44) 

0.07 

1.30) 

0.02 

0.40) 

0.02 

0.33) 

0.6 

22) 



Males & Females 

Males & Females 

Males & Females 

Males & Females 
1.02±0.13 

(0.90-1.16) 
1.00±0.20 

(0.80-1.20) 
1.15±0.01 

(1.50-1.52) 



TABLE 49. Radular statistics for Neotricula lilii. 
Mean ± standard deviation (range). N = 4. In 
mm except for width of central tooth in (im. 



Shell length 
Radular length 
Radular width 
Total rows of teeth 
No. rows of teeth 

forming 
Central tooth width 



3.00±0.17 (2.80-3.16) 
0.58±0.01 (0.57-0.60) 
0.08 (0.072-0.080) 
87 (84, 90) (N = 2) 
19 (16, 22) (N = 2) 

17.2(16.2-17.9) N = 3 



has a moderate number of gill filaments whilst 
the latter has few (char. 6). The male gonad 
overlaps the stomach of the former; it is pos- 
terior to the stomach in the latter (char. 27). 
The anterior vas deferens leaves the prostate 
at mid-prostate in the former; from the pros- 
tate at the posterior end of the mantle cavity in 
the latter (char. 31). The penial tip of the- 
former has a papilla; that of the latter a long 
penial filament. 



258 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 

Ast 




FIG. 99. Uncoiled male of Neotricula lilii without head or kidney tissue. Anterior part of mantle cavity omitted 
as is posterior end of digestive gland. 



TABLE 50. Cusp formulae for the radular teeth of Neotricula lilii with the percent of the three radulae in 
which a given formula was found at least once. 



Central Teeth 


Lateral Teeth 


Inner Marginal 
Teeth 


Outer Marginal 
Teeth 


3-1-3 
2-2 


66% 


3-1[2]-3 


100% 


12 


— 


33% 


2-1-2 
2-2 


33% 


3-1-3 


66% 


13 


100% 


66% 


3-1-3 
2-3 


33% 


4-1[2]-3 
3-1-4 


33% 
33% 


14 
15 
16 

X* 

N = 


100% 

66% 

33% 

14.2±0.9 
= 30 


100% 
33% 

13.8±1.0 
N = 30 



'Mean ± standard deviation of cusp number for all teeth counted. 




THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 259 

Neotricula minutoides (Gredler, 1885) 

Paralectotypes. SMF 4155; pi. 4, fig. 1, in 
Yen, 1939. 

Type locality. "Aus Quellen bei Hensan"; 
Heng-shan-hsien, Hunan (Yen, 1939). Near 
site 1, fig. 1 (Hengshan) 

Synonymy. Bithynia minutoides Gredler, 

1885 

Hydrobia minutoides (Gredler, 1887) 

Tricula minutoides, Yen, 1939 

Neotricula minutoides, this paper 

Habitat 

Specimens collected for this paper came 
from Tong Meng Village, Xikou town, Cili 
County, Changde Prefecture; 29°13'58"N, 
1 10°44'12"E; Figure 1 site 8. The field collec- 
tion number was D85-83 on 7 October 1985. 
Specimens were collected from Tong meng 
village, Xikou Tow, Cili County, Changde Pre- 
fecture; 29°13'58"N, 110°44'12"E; Figure 1 
site 8. The assigned collection number was 
D85-83. The habitat was a small perrenial 
stream 15-25 cm wide and 5-10 cm deep at 
an altitude of 550 m above sea level. The 
stream was shaded; the flow was slow. The 
bottom of the stream was paved with small 
stones in mud. The water was clean and cool. 
The sides of the stream had weeds and short 
shrubery. 



0.5mm 




FIG. 100. A. Relationship of base of the penis of 
Neotricula lilii to mid-line of snout-neck (x) and to 
posterior end of eye lobes (a). B. Penis. 



TABLE 51. Lengths of neural structures of Neotricula lilii. Mean ± standard deviation (range). N 
* = neural elements measured to calculate the RPG ratio. 



Cerebral ganglion 

Cerebral commissure 

Pleural ganglion 
Right (1)* 

Left 

Pleuro-supraesophageal connective (2)* 

Pleuro-subesophageal connective 

Supraesophageal ganglion (3)* 

Subesophageal ganglion 

Osphradio-mantle nerve 

RPG ratio* = 2 + 1+2 + 3 



0.25+0.01 (0.24-0.26) 
0.09+0.05(0.04-0.16) 

0.11+0.01 (0.10-0.12) 
0.10 (N = 3) Novar. 
0.17+0.03(0.12-0.18) 
0.08+0.03 (0.04-0.10) N = 3 
0.11+0.03(0.10-0.14) 
0.08+0.04(0.10-0.12) N = 3 
0.02 (0, 0.04) N = 2 
0.44+0.08 (0.33-0.50) 



260 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 101. Radula of Neotricula lilii. A. Segment of radula. B, E. Central teeth. C, D, F. Lateral and inner 
marginal teeth. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 261 




FIG. 102. Radula of Neotricula lilii. A. Right lateral and inner marginal teeth. B-F. Outer marginal teeth; B, 
D, F = right; C, E = left outer marginal teeth. 



262 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 52. Shell measurements (mm) of Neo- 
tricula minutoides. Mean ± standard deviation 
(range). N = 5; all shells 5.5 whorls. 



Length (L) 


3.41 ±0.20 (3.14- 


-3.60) 


Width (W) 


1.68±0.09(1.60- 


-1.80) 


L last three whorls 


3.07±0.18 (2.84- 


-3.28) 


L body whorl 


2.28±0.15(2.12- 


-2.48) 


L penultimate whorl 


0.50+0.04 (0.44- 


0.56) 


W penultimate whorl 


1.09±0.05(1.04- 


-1.16) 


W 3rd whorl 


0.72±0.03 (0.68- 


0.76) 


L aperture 


1.53±0.13(1.40- 


-1.70) 


W aperture 


1.02±0.07(0.96- 


-1.12) 


X 


0.44±0.05 (0.40- 


-0.52) 


У 


0.18±0.05(0.12- 


-0.24) 



Depository 

Specimens are deposited in ZAMIP, 
M0006; ANSP, 373136, A12652. 

Description 

Shells. The shells are small, ovate-conic with 
5.5 whorls (Figs. 60C-G, ЮЗА -E). Lengths 
range from 3.14-3.60 mm (Table 52). The 
aperture is pyriform. Adapically there is no 
notch or beak, nor is there an internal notch 
groove. There is no adapical outer lip angle. 

The whorls at the suture are smooth (not 
crenulated). There ¡s a pronounced umbilicus. 
SEM analysis reveals faint traces of spiral mi- 
crosculpture on the adapical part of the 
whorls of the teleoconch. 

The inner lip is thick and arched. It is clearly 
separated from the body whorl by a narrow 
gap. The adapical apertural lip is pulled away 
from the body whorl but the interspace is filled 
with shell layers (as is the inner side of the 
inner lip). There is no apertural sinus. In side 
view, the outer lip is straight and slightly 
scooped forward. In side view, the inner lip is 
straight. There are no apertural teeth or 
notches, no varix, no spout. The abapical lip 
projects beyond the base of the shell 0.44 ± 
0.05 mm. 

As seen with SEM, the protoconch is mi- 
nutely wrinkled (Fig. 103C-E). Growth lines 
begin on the teleconch at 2.0 whorls. 

External Features. The head is dark grey, 
with few or no white granules about the eyes. 
The operculum is corneous and paucispiral 
(Fig. 103F). It appears to grow in layers. The 
internal attachment pad is prominent, some 
49% the width of the operculum. 

Mantle cavity. Mantle cavity structures are 
typically triculine. Organ measurements are 
given in Table 53. The osphradium is located 
mid-gill; it is small. Only one gill filament was 



measured; it was 0.58 mm long. For this fila- 
ment Gf 2 was long. 

Female reproductive system. An uncoiled fe- 
male without head and with kidney tissue re- 
moved is shown in Figure 104. Measure- 
ments of organs are given in Table 53. 
Important features are: (1) The gonad is pos- 
terior to the stomach, is small, and consists of 
one or two bundles of lobes. (2) The bursa 
copulatrix (Bu) is round, small and situated 
directly posterior to the albumen gland (Ppo). 
It is not occluded by the posterior part of the 
albumen gland. (3) The albumen gland is 
short. (4) The bursa copulatrix complex of or- 
gans is shown in Figure 1 05. The orientation 
of organs in Figure 105A is the same as that 
in Figure 104. The bursa (Bu) is short. (5) The 
seminal receptacle (Sr) is spherical; it may or 
may not have a duct. In either case, it opens 
into the base of the duct of the bursa where 
the latter runs into the spermathecal duct (Sd) 
and also receives the sperm duct (Sdu). This 
opening for the seminal receptacle is on the 
dorso-lateral side of the bursa complex (Fig. 
105C). (6) When there is a discernable duct 
of the seminal receptacle, it is extremely 
short, about 0.02 mm. (7) The spermathecal 
duct is short and swollen; it is nearly a 
straight, as well as a direct continuation of the 
duct of the bursa; it opens into the posterior 
end of the mantle cavity (Emc). 

Male reproductive system. The posterior half 
of an uncoiled male with kidney tissue re- 
moved is shown in Figure 106. Measure- 
ments of organs are given in Table 53. Impor- 
tant features are: (1) The gonad (Go) is 
posterior to the stomach. It consists of several 
bundles of lobes (removed in Figure 106 to 
reveal the seminal vesicle) arising from the 
vas efferens. (2) The prostate (Pr) overlaps 
the posterior end of the mantle cavity (Emc.) 
(3) The seminal vesicle (Sv) arises from mid- 
vas efferens. (4) The seminal vesicle forms a 
small knot of ducts dorsal to the gonad; it 
does not continue onto the stomach. (5) The 
anterior vas deferens (Vd 2 ) leaves the pros- 
tate near the posterior end of the mantle cav- 
ity (Emc). (6) The penis (Figure 107B) is sim- 
ple, slender and differs from that of any other 
Hunan species by being highly extensible. (7) 
The penis has no papilla nor was an ejacula- 
tory duct found at the base of the penis or in 
the neck. (8) The orientation of the base of the 
penis on the neck is shown in Figure 107A. It 
slightly overlaps the snout-neck mid-line; it is 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 263 



FIG. 1 03. SEM photographs of shells and opercula of Neotricula minutoides. Note in В that the outer lip is 
slanted. C-E. Details of protoconch and in C, beginning teleconch. F. Opercula with outer surface (left) and 
inner surface (right). 

orientated at an angle of 40° to the snout-neck 54 and 55. There are 56.4 ± 4.8 rows of teeth 
mid-line (x). along a radula of 0.51 mm length. The most 



frequently encountered formula is 

(2)3-1-3 
sexes. Radular statistics are given in Tables 2-2 



Digestive system. The digestive gland covers 

the posterior chamber of the stomach of both (2)3-1-3(2) ; 2 to 4-[2]-2(3); 12-14; 11-14. 



264 DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 53. Lengths (mm) or counts of non-neural organs and structures of Neotricula minutoides. N = 
number of snails used. Mean ± standard deviation (range). 

Females (N = 5) Males (N = 1) 

Body 5.02 ±0.13 4.40 

(4.90-5.16) 
Gonad 0.83±0.15 1.28 

(0.70-1.00) 
Digestive gland 2.10±0.28 1.90 

(1.80-2.36) 
Posterior palliai oviduct 0.80 — 

(= albumen gland) (0.60, 1.00) N = 2 

Anterior palliai oviduct 1 .28 — 

(= capsule gland) (1.16, 1.40) N = 2 

Total palliai oviduct 2.02±0.34 — 

= OV (1.76-2.40) 

Bursa copulatrix 0.55±0.14 — 

= BU (0.40-0.66) 

Duct of BU — — 

BU + OV 0.28±0.10 — 

(0.17-0.35) 
Seminal receptacle 0.10 — 

(no var.) N = 2 
Duct of Seminal receptacle — — 

Mantle cavity 1 .56 1 .40 

(1 .36, 1 .76) N = 2 
Gill (G) 1.40 1.24 

(1.20, 1.60) N = 2 
Osphradium (OS) 0.41 0.44 

(0.36, 0.46) N = 2 
OS - G 0.29 0.35 

(0.29, 0.30) N = 2 — 

No. of filaments 17 18 

(13, 21) N = 2 
Gf 2 0.36 N = 1 — 

Gf, 0.22 N = 1 — 

Total Gf = TGF 0.58 N = 1 — 

Gf 2 -s- TGF 0.62 N = 1 — 

Buccal mass 0.57 N = 1 — 

Prostate — 1.10 

Seminal vesicle — 0.48 

Penis — 2.60 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 265 




1.0mm 



FIG. 104. Uncoiled female Neotricula minutoides with head and kidney tissue removed. 



266 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Dbu Про 




TABLE 54. Radular statistics for Neotricula 
minutoides. Mean ± standard deviation (range). 
N = number used. In mm except for width of 
central tooth in ц.т. 

Females (N = 6) 

Shell length 3.67±0.30 (3.40-4.00) 

Radular length 0.51 ±0.06 (0.44-0.57) 

Total rows of teeth 56.4±4.8 (50-60) N = 7 

No. rows of teeth 10.9±6.3 (5-21) N = 8 

forming 

Central tooth 15.1 ±1.1 (13.4-16.8) N = 13 

width 



Nervous system. Measurements are given in 
Table 56. The RPG ratio is 0.45; the dorsal 
nerve ring is thus moderately concentrated. 

Remarks 

Conchologically, this species most closely 
resembles Tricula bambooensis (Figs. 153- 
155). Neotricula minutoides has a proportion- 
ally much wider shell (compare illustrations 
here with Davis et al., 1986: fig. 20A-E). Ad- 
ditionally, the outer lip of the former, in side 
view, is scooped forward; it is parallel with 
the axis of coiling in the latter (char. 15). The 
adapical aperture of the former is fused to 
the body whorl; it is slightly separated from 
the body whorl in the latter (char. 21). The 
aperture of the former is pyriform; it is oval in 
the latter (char. 3). 

The most prominent anatomical differences 
are those that separate the two genera. 



1.0mm 

FIG. 105. Details and variation of the bursa copu- 
latrix complex of organs of Neotricula minutoides. 
Figure A is in the same orientation as in Figure 1 04. 
B. Section of oviduct cut away at "x" to show con- 
tinuation of duct of bursa (Dbu) into spermathecal 
duct (Sd) with latter opening (Osd) into posterior 
end of mantle cavity. С Bursa complex flipped over 
to show relationships of seminal receptacle to 
sperm duct (Sdu), spermathecal duct (Sd) and duct 
of bursa. 



The radula is shown in Figure 108. A pro- 
nounced bifurcation of the blade of the pri- 
mary cusp of the lateral tooth may be found 
(Fig. 108C, D, E). Otherwise tooth morpholo- 
gies are standard triculine. 



TRICULINI Davis, 1979 

Type genus. Tricula Benson, 1 843 

Diagnosis. Those genera of Triculinae in 
which the spermathecal duct enters the peri- 
cardium. The oviduct makes a 360° closed 
twist. In the plesiomorphic state, the seminal 
receptacle arises from the oviduct; in the de- 
rived state, the seminal receptacle is lost and 
its function taken over by derived structures 
attaching to the oviduct-spermathecal duct 
juncture. There is no sperm duct. 

Genera assigned. Delavaya, Fenouilia, La- 
cunopsis, Lithoglyphopsis, Tricula. (N = 5). 



Lithoglyphopsis Thiele, 1928 

Type species. Lithoglyphus modestus Gred- 
ler, 1886(1887] 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 267 

Ast ln 




1.0mm 



FIG. 106. Uncoiled male of Neotricula minutoides without head or kidney tissue. Anterior part of mantle 
cavity omitted. Outline of gonad (Go) represented by a dashed line; lobes of gonad removed to show knot 
of tubes of seminal vesicle (Sv). 



TABLE 55. Cusp formulae for the radular teeth of Neotricula minutoides with the percent of radulae in 
which a given formula was found at least once. N = 7 radulae. [ ] indicates one cusp support with a 
split blade. 



Central Teeth 



Lateral Teeth 



Inner Marginal 
Teeth 



Outer Marginal 
Teeth 



3-1-3 
2-2 


71% 


3-[2]-2 


57% 




11 


29% 


11 


100% 


2-1-2 
2-2 


57% 


2-[2]-3 


29% 




12 


71% 


12 


71% 


3-1-3 
3-2 

3-1-3 
2-3 


29% 
14% 


4-1-3 

4-[2]-3 

3-1-2 


29% 
29% 
14% 


X 


13 
14 


86% 

71% 

12.5+1.9 


13 
14 
12.1 


71% 
43% 
±1.6 


2-1-3 
2-2 


14% 


3-1-3 


14% 


N 


= 64 


N = 


64 






4-[2]-2 


14% 













*Mean ± standard deviation of cusp number for all teeth counted. N = number of teeth counted on 7 radulae. 



268 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




TABLE 56. Lengths (mm) of neural structures of 
Neotricula minutoides. Mean ± standard deviation 
(range). N = number used = 2. * = neural 
elements measured to calculate the RPG ratio. 



Cerebral ganglion 


0.28 (0.26, 0.30) 


Cerebral commissure 


0.06 (0.5, 0.6) 


a Pleural ganglion 




Right (1)* 


0.13(0.12,0.14) 


a' Left 


0.11 (0.10,0.12) 


Pleuro-supraesophageal 




connective (2)* 


0.20 (no var.) 


Pleuro-subesophageal 




connective 


0.07 (0.04, 0.10) 


Supraesophageal 


0.11 (0.10,0.12) 


ganglion (3)* 




Subesophageal ganglion 


0.11 (0.10,0.12) 


Osphradio-mantle nerve 


0.05 (0.04, 0.06) 


RPG ratio* = 2 * 1+2 + 3 


0.45 (no var.) 



0.5 mm 




FIG. 107. A. Relationship of base of the penis of 
Neotricula minutoides to mid-line of the snout-neck 
(x) and to posterior end of the eye lobes (a). B. 
Penis. 



Type locality. Hen-Kiou-fee bis Pe-shang 
(Gredler, 1886), = Heng-dshou-fu bis Pe- 
shang, Hunan (Yen, 1939). 

Designation. By Thiele, 1928 



Lithoglyphopsis modesta (Gredler) 

Types. Paralectotypes SMF 4214a. Figure 
109A = specimen figured by Yen (1939). Fig- 
ure 109B-D = additional type specimens. 

Type locality. See above 

Synonymy. Lithoglyphus modestus Gredler, 

1886 

Lithoglyphopsis modesta, Thiele, 1928 

Lithoglyphopsis modesta, Wenz, 1939: 580, 

Figure 1581 

Lithoglyphopsis, modestus Yen, 1939: 43, pi. 

4, Figure 7. 

Habitat 

The type locality is today known as Baisha, 
Hengshan County from the Xiangjiang River; 
25°58'22"N, 112°45'55"E, Figure 1, locality 9. 
Snails were collected by diving to obtain 
stones from the river bottom at a depth of 
about 2.0-2.5 m. Collected by Chen and Wu 
in 1986, collection number 86-B, ANSP 
373144, A12660, ZAMIP M00054. 

The locality from which snails were used for 
most of the dissections was Anhua town, An- 
hua County, Zijiang River; 28°23'46"N, 
111°,12'41"E, Figure 1, locality 10. The col- 
lection number was D87-1, by Davis, Chen 
and Xing on 16 March 1987, ANSP 373150, 
A12666. Snails were collected 1.6 km up- 
stream from the town boat landing. Collec- 
tions came from the shores of a small island 
in the middle of the river. Water flows through 
a stone breakwater at the upstream end of the 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 269 




{ A v « 



4 y 1ц 



kL^ фк 



\?ч^ 



*■*■' Vi 




FIG. 108. Radula of Neotricula minutoides. A. Section of radula. B-E. Central, lateral and inner marginal 
teeth. F. Outer marginal teeth. 



270 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 1 09. Shells of Lithoglyphopsis modesta. A-D, paralectotypes, SMF; A. specimen figured by Yen (1 939). 
E, F. Specimens from Anhua, D87-1. A = 4.56 mm long; other shells printed at same scale. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 271 

TABLE 57. Shell measurements (mm) of Anhua and Baisha populations of Lithoglyphopsis modesta. 
Mean ± standard deviation (range). N = numbered measured. 







Anhua 


Baisha 






Female (N = 


5) Male (N = 6) 


Female (N = 5) 


Male (N = 5) 


No. Whorls 


3.0-3.5 


3.0-3.5 


3.0-4.0 


3.5-4.0 


Length (L) 


5.18±0.04 


5.10±0.60 


4.53±0.27 


4.46±0.31 




(4.58-5.57) 


(4.16-5.66) N = 5 


(4.16-4.91) 


(4.15-4.90) 


Width (W) 


4.87±0.33 


4.68 ±0.42 


4.49±0.23 


4.58±0.32 




(4.74-4.99) 


(3.91-5.16) 


(4.24-4.83) 


(4.23-4.98) 


L body whorl 


4.91 ±0.40 


4.76±0.54 


4.27±0.22 


4.19±0.32 




(4.37-5.41) 


(3.91 -5.24) 


(4.07-4.65) 


(3.90-4.65) 


L penultimate 


0.15±0.04 


0.21- 


0.20±0.05 


0.252- 


whorl 


(0.08-0.17) 


No. var. 


(0.17-0.25) 


No. var. 


W penultimate 


1.12±0.15 


1.06±0.10 


1.00±0.09 


1.04±0.11 


whorl 


(0.99-1.33) 


(0.99-1.25) N = 5 


(0.91-1.04) 


(0.91-1.16) 


W 3rd whorl 


0.44±0.04 


0.42±0.06 


0.44±0.04 


0.46±0.50 




(0.42-0.50) N 


= 3 (0.33-0.50) N = 4 


(0.42-0.50) 


(0.42-0.50) 


L aperture 


4.43±0.19 


4.20±0.31 


4.05±0.17 


4.03±0.20 




(4.16-4.66) 


(3.66-4.58) 


(3.90-4.07) 


(3.90-4.32) 


W aperture 


3.62±0.23 


3.52±0.31 


3.29±0.15 


3.32±0.30) 




(3.24-3.83) 


(3.00-3.83) 


(3.15-3.49) 


(3.07-3.74) 


L crescent 


2.31 ±0.19 


2.25±0.35 


2.04±0.19 


2.20±0.35 




(2.08-2.50) 


(1.83-2.58) 


(1.83-2.32) 


(1.83-2.57) 


W crescent 


0.42±0.06 


0.35±0.06 


0.38±0.09 


0.58±0.17 




(0.33-0.50) 


(0.25-0.42) 


(0.25-0.50) 


(0.49-0.83) 


W columellar 


0.73±0.06 


0.77±0.17 


0.68±0.07 


0.72±0.08 


plate 


(0.67-0.83) 


(0.50-1.00) 


(0.58-0.75) 


(0.66-0.83) 


L "A" 


5.53±0.25 


5.26±0.44 


4.95±0.22 


5.08±0.30 




(5.16-5.74) 


(4.58-5.66) 


(4.73-5.23) 


(4.81-5.48) 


W "A" 


4.20±0.27 


4.08±0.33 


3.64±0.22 


3.74±0.26 




(3.91-4.58) 


(3.58-4.49) 


(3.49-3.94) 


(3.49-4.07) 




island. Snails were collected from the bottom 
surface of stones along the protected inner 
edge of the breakwater where there was an 
expansive shallow water area some 30 cm 
deep and with emergent vegetation. Associ- 
ated fauna included: Stenothyra hunanensis, 
Semisulcospira sp, a species of Viviparidae, a 
planorbid, and Radix sp. 




FIG. 110. Illustration to show three shell orienta- 
tions for measurements. 



Description 

Shells. Shells measurements are given in Ta- 
ble 57 for the two populations. Shells are il- 
lustrated in Figures 109-113. How measure- 
ments are made for these globose shells is 
given in Figure 110. Shells are medium to 
long in length. They are nearly round in out- 
line with the apex to base alignment as in 
Figure 11 0C; they are globose and smooth. 
The shell is dominated by the body-whorl. 



272 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 111. Shells of Lithoglyphopsis modesta from Anhua (D87-1) A-C; "L" liliputinus, D; and Guoia viri- 
dulus, paralectotypes, E, F. Shell A is 4.72 mm long; other shells printed at same scale. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 273 




FIG. 112. Shells of Lithoglyphopsis modesta from Baisha (86-B), A-D. А, В = males; C, D = females. "L 
liliputinus, E, F. A = 4.72 mm; other shells printed to same scale. 



274 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 113. SEM photographs of shells of Lithoglyphopsis modesta. A-C. Enlargements of apical whorls. 
Spiral microsculpture is seen on second and third whorls. E-H. Enlargement of apertural areas showing 
basal crescent (Be), apertural beak (Ab), and beak or adapical apertural groove (Agr). 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 275 



TABLE 58. Lengths (mm) or counts of non-neural organs and structures of Lithoglyphopsis modesta. 
Mean ± standard deviation (range). N = number of snails used. *See text for discussion of palliai 
oviduct length. 



Females 



Males (N = 2) 



Body 

Digestive gland 
Gonad 

Posterior palliai 

oviduct 

(= albumen gland) 
Anterior palliai 

oviduct 

( = capsule gland) 
Total palliai oviduct 

= PO 
Bursa copulatrix 

Duct of bursa 

Bu -s- PO 

Seminal receptacle 

Duct of seminal 

receptacle 
Buccal mass 

Mantle cavity 

Osphradium 

= Os 
Gill 

= G 
Os - G 

No. filaments 

Gf 2 

Gf, 

Total Gf 

= TGF 
Gf 2 -г- TGF 

Prostate 

Seminal vesicle 

Penis 



7.52±1.52 
(4.96-9.40) N = 6 

3.01 ±0.60 
(2.16-4.0) N = 6 

1.96±0.21 
(1.10-2.20) N = 6 

1.43±0.06 
(1.40-1.50) N = 6 

2.89 
N = 1 



4.23d 
(3.70- 

0.87 

(0.80 

0.31 d 
(0.22- 

0.22 
(0.18 

0.19 

(0.18, 

0.33± 
(0.30- 

1.60± 
(1.50- 

4.57± 
(4.28- 

1.94± 
(1.76- 

4.20± 
(4.28- 

0.47± 

(0.39- 

39 

(38 

0.43 
(0.20 

1.04 
(0.70 

1.48± 
(1.06- 

0.29± 
(0.13- 



0.33 
4.60) 

0.94) N = 2 
±0.09 
-0.40) N = 3 

-0.25) N = 2 

0.20) N = 2 

0.04 

0.38) N = 3 

0.13 

1.78) N = 4 

0.26 

4.86) N = 5 

0.22 

2.30) 

0.26 

4.86) 

0.05 

0.48) 

2 

41) N = 4 

0.19 

0.80) N 

0.14 

1 .30) N 

0.21 

1 .90) N 

0.12 

0.41) N = 16 



16 



16 



15 



7.85 
(7.50, 8.20) 

3.65 
(3.10,4.20) 

3.35 
(2.70, 4.00) 



4.08 
(3.96, 4.20) 

1.15 
(1.0, 1.30) 

3.70 
(3.60, 3.80) 

0.31 
(0.26, 0.36) 

41 
no variation 
male + female 

male + female 

male + female 

male + female 

2.52 
(1.84,3.20) 

1.60 
no variation 

3.67 
(3.60, 3.74) 



276 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 59. Radula statistics for Lithoglyphopsis modesta. Mean ± standard deviation (range). In mm 
except for the width of the central tooth in p.m. N = number measured. 



Females (N = 10) 



Males (N = 7) 



Greatest shell dimension 
Radular length 
Radular width 
Total rows of teeth 
No. rows of teeth forming 
Central tooth width 

Greatest shell dimension 
Radular length 
Radular width 
Total rows of teeth 
No. rows of teeth forming 
Central tooth width 



Anhua Population (D87-1) 

4.92±0.47 

(4.00-5.36) 

2.52±0.20 

(2.26-2.90) 

0.23±0.02 

(0.20-0.27) 

69±5 

(62-77) 

31 ±4 

(25-36) 

55±4 

(48-58) 

Baisha Population (1986) 

4.79±0.25 

(4.60-5.16) 

2.60±0.36 

(2.10-2.96) 

0.21 ±0.01 

(0.20-0.22) 

66±8 

(55-77) 

34±7 

(23-40) 

51 ±5 

(48-60) 



4.91: 
(4.44- 

2.44: 

(2.20- 

0.23: 

(0.22- 
66: 

(61- 
25: 

(19- 
55: 

(54- 

4.77: 
(4.28- 

2.78: 

(2.10- 

0.21: 

(0.20- 

74: 

(71- 

37: 

(35- 
49: 
(48- 



0.26 

5.20) 

0.16 

2.66) 

0.01 

0.24) 

3 

70) 

:5 

32) 

:1 

56) 

0.32 

5.12) 

0.13 

3.00) 

0.01 

0.22) 

:3 

80) 

2 

39) 

2 

52) 



TABLE 60. Cusp formulae or the radular teeth of Lithoglyphopsis modesta with the percent of radulae in 
which a given formula was found at least once. N = number used and/or counted. 



Central Teeth 



Lateral Teeth 



Inner Marginal Teeth 



Outer Marginal Teeth 



Anhua Population (N = 10) 



1 

2-2 


60% 


0-1-3 


80% 


6 


30% 


1 
3-3 


50% 


2-1-0 


70% 


7 


90% 


1 
3-2 


40% 


3-1-0 


60% 


8 


90% 


1 
2-3 


10% 


0-1-2 
4-1-0 


40% 
10% 


9 

10 

X* = 
N = 


30% 


= 7.4±0.8 
100 



Baisha Population (N = 3) 



1 


67% 


0-1-3 


100% 


5 — 


2-2 














2-1-0 


67% 


6 33% 


1 


67% 








2-3 




0-1-2 


67% 


7 67% 


1 
3-2 


33% 


3-1-0 


33% 


8 67% 

X* = 7.40±0.9 

N = 30 



6 20% 

7 90% 

8 70% 

9 20% 
10 10% 

7.5±0.9 

5 33% 

6 100% 

7 100% 

8 33% 
6.3±0.6 



"Mean ± standard deviation of cusp number for all teeth counted. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 277 



TABLE 61. Dimensions of neural structures from five individuals of Lithoglyphopsis modesta. Mean ± 
standard deviation (range). In mm; L = length. * = neural elements measured to calculate the RPG 
ratio. 



Cerebral ganglion L 
Pleural ganglion L 

Right (1)* 

Left 
Cerebral commissure L 

Pleuro-supraesophageal 

connective L (2)* 
Pleuro-subesophageal 

connective L 
Supraesophageal 

ganglion L (3)* 
Subesophageal 

ganglion L 
Osphradio-mantle nerve L 

Pedal commissure L 

RPG ratio* (2 + 1+2 + 3) 



0.55+0.07 

0.26+0.04 
0.18+0.02 
0.30+0.07 
0.19+0.03 

0.02 

0.29+0.03 

0.20+0.03 

0.27+0.08 
0.23+0.11 
0.26+0.03 



(0.46-0.64) 

(0.20-0.30) 
(0.16-0.20) 
(0.20-0.38) 
(0.16-0.24) 

(n variation) 

(0.24-0.32) 

(0.18-0.24) 

(0.20-0.30) 
(0.10-0.30) 
(0.23-0.30) 




FIG. 114. Head of Lithoglyphopsis modesta from a 
snail collected from D87-1. 



The apex is a protruding nipple, a scant 7 to 
8% of the overall shell length. In side view, 
with axis of coiling vertical, the outer lip is 
straight and slanted back (to the right) 35° to 



the axis of coiling (Fig. 109, column 3). The 
shell length (L) is always with the axis of coil- 
ing vertical. Greatest overall length (L of A) is 
with the shell resting on the aperture (Fig. 
1 1 0C). The aperture is measured by tilting the 
axis of coiling so that the plane of the aperture 
is horizontal (Lap, Fig. 1 10B). There is a wide 
columellar shelf 0.32 to 0.36 mm wide (Cs, 
Fig. 1 1 0B); the edge at the aperture is straight 
to slightly arched. There is a pronounced 
crescent-shaped ridge to the left of the col- 
umellar shelf with a somewhat depressed 
concavity, the basal crescent (Be), between 
the ridge and the columella. The peristome is 
complete. There is no umbilicus. The adapical 
end of the aperture is produced into a beak- 
like projection (Ab, Fig. 113G). The aperture 
shape is round to broadly pyriform. The apex 
of some individuals may have a reddish color; 
the cleaned shell is horn yellow. 

The SEM pictures of varied aspects of the 
shell are shown in Figure 113. The apical 
whorls are shown in Figure 1 13A-D. They are 
invariably eroded. Spiral microsculpture is ev- 
ident (Fig. 1 1 3А, В, D). Figure 1 1 3E-H shows 
variation in the size of the basal crescent (Be). 
The apertural beak (Ab) is featured in Figure 
1 13F-G showing the groove (Agr) running in- 
side it. 

External features. The head is shown in Fig- 
ure 1 14; it is broad and squat; there are pro- 
nounced bulging eye lobes at the base of 
each tentacle. In these character-states the 
head is similar to those of Fenouilia and La- 



278 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 







FIG. 115. Opercula of Lithoglyphopsis modesta. А, В from Baisha snails; C, D from Anhua snails. A, C. 
External surfaces; B, D. Internal surfaces. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 279 




1.0 mm 



FIG. 116. Masses of anterior pedal glands (Apg) in 
anterior foot seen through dorsal surface. 



cunopsis. The granules shown on the head 
vary in color from bright lemon yellow to light 
yellow to white. There is a pronounced om- 
niphoric groove; no suprapedal fold. The 
operculum is shown in Figure 1 15; it is regu- 
larly ovate, corneous, paucispiral without dis- 
cernable internal pad; without ridges. The at- 
tachment area is barely distinguishable. 

Opening the anterior foot dorsally and re- 
moving the buccal mass and supporting mus- 
cle bands, one sees a mass of tubular ante- 
rior pedal glands (Figure 116). These come 
up as a bunch over the elongated pedal com- 
missure. 



Mantle cavity. The reflected mantle is shown 
in Figure 1 1 7A. The mantle cavity is typical for 
those of species of Triculini in which the sper- 
mathecal duct (Sd) enters the pericardium 
and the pericardium (Pe) swells out into the 
mantle cavity. Organ measurements and sta- 
tistics are given in Table 58. 

The osphradium (Os) is long; the posterior 
end may be considerably narrowed (Figs. 
117, 119C). Variation in osphradial shape is 
shown in Figure 119C. The terminal gill fila- 
ment Gf 2 is short, that is the ratio is 0.29 ± 
0.12 (Table 58). The length of the longest fil- 
aments is 1.48 ± 0.21 mm. In lateral view the 
filament is pleated and Gf 2 has a pronounced 
dome (Fig. 117B). 



The pericardium bulges out into the mantle 
cavity and the opening into the pericardium 
for sperm entry is clearly observable (Ope, 
Fig. 117A). The mantle cavity organs and ar- 
rangement is typical of the taxa of the Tricula 
clade of Triculini. 



Female reproductive system. The body of an 
uncoiled female without head and with kidney 
tissue removed is shown in Figure 118. Mea- 
surements of the relevant organ systems are 
given in Table 58. Important features to note 
are: (1 ) The body is squat and fat as would be 
expected from shell shape. (2) The dorsal 
surface is densely pigmented with melanin. 
(3) The posterior palliai oviduct (Ppo) makes 
a pronounced bend over the style sac. (4) The 
gonad covers the stomach. (5) The sperm en- 
ter the system at the rear of the mantle cavity 
through the pericardium (Ope, Fig. 117A). 
The pericardium swells out into the mantle 
cavity. The spermathecal duct (Sd) is a short 
tube running from the pericardium to the swol- 
len section of the oviduct (Fig. 1 19A). (6) The 
oviduct makes a tight twist or coil dorsal to the 
bursa copulatrix. (7) The seminal receptacle 
(Dsr) arises from the oviduct posterior to the 
duct of the bursa (Dbu, Fig. 119A). (8) The 
bursa complex is dorsal to the posterior palliai 
oviduct (= albumen gland). (9) The oviduct 
opens into the palliai oviduct close to the pos- 
terior end of the mantle cavity, i.e. not into the 
posterior end of the palliai oviduct. (10) The 
bursa is short. (11) The albumen gland length 
is short if the albumen gland is measured 
from the posterior edge on the stomach 
thereby not including the bend over the style 
sac (ratio of 0.35 ± 0.02). However, measur- 
ing along the bend the actual length (func- 
tional length) averages 1 .83 mm and this di- 
vided by that palliai oviduct length averages 
0.45 ± 0.03, i.e. the functional length is stan- 
dard. 

Character-states 5-7 are the same as those 
found in Tricula, Fenouilia, and Delavaya. 



Male reproductive system. The body of an un- 
coiled male is shown in Figure 120 without 
head and with kidney tissue removed. Mea- 
surements of relevant organs are given in Ta- 
ble 58. Important features are: (1) The gonad 
consists of moderately large lobes draining 
into a vas efferens (Ve). (2) The posterior vas 
deferens arises from the vas efferens at mid- 
gonad or slightly posterior to mid-gonad and 



280 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 281 




FIG. 118. Uncoiled female Lithoglyphopsis modesta with head and kidney tissue removed. 



282 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Dbu 




° v ^%n 




FIG. 119. Bursa copulatrix complex of organs; А, В. 
Variation in shape of the osphradium, C; refer also 
to Os, Figure 117A. In Figure A, organs are in the 
same orientation as in Figures 1 1 7 and 1 1 8. How- 
ever, here the bursa copulatrix is cut away to show 
the oviduct coil (Coi) and the seminal receptacle 
(Sr) where it attaches to oviduct. B. Variation in the 
shape of seminal receptacle. 



and run posteriorly over the nerve ring. The 
radular sac (Rs) is extremely elongate and 
coils up dorsally along the buccal mass (Bm) 
between the buccal mass and the right cere- 
bral ganglion (Reg, Fig. 122B). 

The radula is shown in Figure 123. Teeth 
counts and statistics are given in Tables 59 
and 60. Diagnostic features are (1) the large, 
singular triangular anterior cusp of the central 
tooth, (2) the massive basal cusps on the face 
of the central tooth, and (3) the massive dom- 
inant cusp on the lateral tooth. 

The stomach (Fig. 124) has a relatively 
slender posterior chamber (Pst) that is 
slanted from ventral to dorsal along its whole 
length and is buried dorsal to the digestive 
gland. The digestive gland covers part of the 
anterior chamber (Ast) as does the prostate. 
There is no caecal appendix. 



Nervous system. There are several notewor- 
thy features of the nervous system (Fig. 
122B). (1) The RPG ratio has a mean of 0.26 
(Table 61); the pleuro-supraesopheageal 
connective is short. (2) The cerebral commis- 
sure is elongate (exceeds 0.12 mm). (3) The 
cerebral ganglion have melanin splotches 
(Fig. 122B). (4) The pedal commissure is 
elongate (exceeds 0.12 mm). (5) The osphra- 
dio-mantle nerve is elongate (exceeds 0.12 
mm). 



forms a loosely coiled seminal vesicle (Sv) 
visible to the left side of the gonad. (3) The 
gonad covers the stomach (is ventro-lateral to 
it). (4) The prostate is comparatively massive, 
covering the style sac and part of the anterior 
chamber of the stomach. (5) The penis is sim- 
ple, without lobes (Fig. 121B). The anterior 
end is extremely slender, that is, a long penial 
filament (Pf). There is no papilla. (6) There is 
a slender muscular ejaculatory duct (Ej) be- 
neath the base of the penis in the neck. (7) 
The shaft of the penis arises to the right of the 
snout-neck mid-line (x, Fig. 121 A at an angle 
of 30° ± 10°). 

Digestive system. The digestive gland covers 
the posterior chamber and most of the ante- 
rior chamber of the stomach (Figs. 118, 120) 
in males and females. The buccal mass and 
salivary glands are shown in Figure 122A. 
The paired salivary glands (Sg) are massive 



Remarks 

Lithoglyphopsis modesta could only be 
confused with various species of Lacunopsis 
on the basis of shell. Anatomically they differ 
as the latter has several accessory seminal 
receptacles; the generalized seminal recepta- 
cle is lost. 

Liu et al. (1980) described Lithoglyphopsis 
ovatus, L grandis, Lacunopsis yunnanensis 
and L. auris as new species from Yunnan, 
China. On the basis of the shell illustrations 
given, and in the absence of anatomical data, 
it is not possible to ascertain generic status or 
discuss relationships. Lacunopsis yunnanen- 
sis has a shell with a pronounced keel. That 
together with the central tooth morphology 
given indicates a possibility that a species of 
Fenouilia is involved. 

The shells of Lythoglyphopsis liliputanus 
(Gredler, 1881) look like a miniature L. mod- 
esta (Figs. 1 1 1D, 1 12E, F). Nothing is known 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 283 




FIG. 120. Uncoiled male Lithoglyphopsis modesta without head and with kidney tissue removed. Two 
bundles of gonadal lobes were cut away. 



284 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



V 



Л 



Bp 




FIG. 121. A. Relationship of base of penis (Bp) of 
Lithoglyphopsis modesta to snout-neck mid-line (x). 
B. Penis 



of this species except for the shells. The type 
locality is Lien-dshou-ho in Kwangtung [ = 
Guangdong Province], immediately south of 
Hunan (Yen, 1939). The lectotype and para- 
lectotypes are in the Bozen Museum, No. 109 
(Zilch, 1974). 

It is probable that there are additional spe- 
cies of Lythoglyphopsis. The closest relation- 
ship of L. modesta to any other Triculinae thus 
far studied is with Fenouilia kreitneri (see 
Davis et al. 1983). Both species have a fat 
body, thus fitting a shell shape that is globose 
or rather trochoid. The shells are quite differ- 
ent. Lythoglyphopsis modesta has a globose- 
conic shell where as Fenouilia has a trochoid 
to trochoid-ovate shape. The shell of the 
former is smooth, that of the latter has keels. 
The characters of the head are the same. The 
female reproductive system is virtually the 
same except for two substantive differences. 
One is the very short spermathecal duct of the 
former bridging the oviduct to the pericardium 
where the oviduct turns towards the bursa 
complex. The spermathecal duct is elongated 





FIG. 122. A. Dorsal aspect of the buccal mass of 
Lithoglyphopsis modesta. В. Dorsal aspect of nerve 
ring. 



in the latter. The other is the bending around 
of the posterior palliai oviduct thus covering 
the bursa complex of organs (in the former). 
In the latter, the palliai oviduct does not bend; 
the bursa is posterior to the palliai oviduct. 
The male reproductive system is the same in 
both taxa with one substantive difference; the 
penis of the former has a long penial filament 
lacking in the latter. The radular sac is ex- 
tremely elongated and coils up dorsally along 
the buccal mass; it is short, that is, general- 
ized in length in the latter (as also in Lacunop- 
sis). The cerebral commissure of the former, 
is over double that of the latter. The gill fila- 
ments are different. In the former they are of 
moderate length with a short section of Gf 2 
clearly seen. In the later, the gill filament Gf., 
is elongated; Gf 2 is not visible with the mantle 
reflected. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 



285 




FIG. 123. Radula of Lithoglyphopsis modesta. A, E segments of radula. C, D, G, H. Central and lateral teeth. 
B, E, F. Lateral, inner and outer marginal teeth. 



286 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 

Epr 




FIG. 124. Ventral aspect of the stomach of Litho- 
glyphopsis modesta. 

However, given this series of differences 
between the two taxa, without more data for 
more species, it is not possible to define ge- 
neric limits clearly. 

Tricula Benson, 1843 

Type Species. Tricula montana Benson, 
1843:465 

Type locality. Bhimtal, N. India 

Designation. By monotypy 

Assigned species. Based on anatomical 
study; type species and Chinese species 
only. T. montana Benson, 1843; T. bam- 
booensis Davis & Zheng, 1986 (in Davis et 
al., 1986b); T. bollingi Davis, 1968; T grego- 
riana Annandale, 1924; T. hudiequanensis 
Davis & Guo, 1986 (in Davis et al., 1986b); T. 
ludongbini Davis & Guo, 1986; T xianfengen- 
sis Davis & Guo, 1986; T. xiaolongmenensis 
Davis & Guo, 1986. T. gredleri Kang, 1986; T. 
maxidens Chen & Davis, sp. nov; T. odonta 
Liu, Zhang & Wang, 1983a. N = 11. 

Diagnosis. Shells small to medium, ovate- 
conic. Central tooth as in Neotricula. The ovi- 
duct makes a tight 360° twist dorsal to the 
bursa. The spermathecal duct enters the peri- 
cardium; the spermathecal duct starts as a 
wide duct when diverging from the oviduct 
and narrows to a slender duct at the pericar- 
dium. The duct of the seminal receptacle 
arises from the oviduct, from the base of the 
duct of the bursa, or from the duct of the 
bursa. The duct of the bursa runs undimin- 
ished in diameter into the oviduct close to the 
opening of the oviduct into the albumen 
gland. Some species of Tricula have a peri- 




FIG. 125. Shells of Tricula gredleri. Figure A is 3.2 
mm long; other shells are printed at same scale. 

cardial bursa (Davis et al., 1986b: 515, fig. 
40). The type species has a pericardial bursa 
(Davis et al., 1986a: 434, fig. 6A). 

Tricula gredleri Kang, 1986 

Syntypes. Hubei Medical College, Depart- 
ment of Parasitology, Wuhan City, Hubei 
Province, People's Republic of China. Fig- 
ured in Kang (1986: pl.1, fig. 2). Holotype not 
segregated in Kang's collection. SMF 
305653-305654 

Type locality. Maluxi, "(28°56'N, 109°92'E)", 
Orientalis Commune, Guzhang County, 
Hunan Province. Collected 18 Oct. 1983. 

Etymology. Named for Vincent Gredler, the 
German malacologist who first described Chi- 
nese Tricula from Hunan Province. 

Habitat 

Specimens studied for this paper were col- 
lected from a small stream at Yantuo Village, 
Xiaoguanping Town, Guzhang County, 
Xiangxi Prefecture; 28°42'7"N, 110°0'37"E 
(Fig. 1, site 12), 20 May 1987, Li Chi-Jian. 
The habitat was 700 m at the edge of the 
stream shaded by short shrubs. The name of 
the stream is You Shui He (he = small 
stream). The field number is D87-3. 

Depository 

Specimens are catalogued into the collec- 
tions of ZAMIP, M0008; ANSP 373141, 
A12657. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 287 




FIG. 126. SEM photographs of shells of Tricula gredleri. A, B. Whole shells of mature individuals; C-D. 
Magnified view of the aperture showing small but deep umbilicus and pronounced apertural notch. E, F. 
Eroded protoconchs; teleoconch begins at 1 .75 whorls. 



288 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 127. Opercula of Tricula gredleri. Outer surface at left (A); inner surface to the right (B). 



Description 

Shell. Shells are small, ovate-conic, and 
smooth (Figs. 125, 126). Virtually all shells 
are eroded at the apices so that a true under- 
standing of length is not possible. Lengths do 
exceed 3.30 mm (Table 62). The aperture is 
pyriform. Adapically there is a pronounced 
apertural beak with beak tubercle. There is no 
internal notch groove. There is no adapical 
outer lip angle. 

The whorls at the suture are smooth (not 
crenulated). There is an umbilical chink. The 
inner lip is straight abapical to the beak tuber- 
cle; it is separated from the body whorl. 
Adapically shell material may be layered be- 
tween the lip and the body whorl to the point 
of fusion. The inner lip is uniformly thin. An 
apertural sinus is seen when the outer lip is 
seen in side view. Abapical to the sinus the lip 
is straight to slightly scooped forward. In side 
view, the inner lip is straight; there is no spout, 
no varix. 

The abapical lip projects beyond the base of 
the shell 0.38 ± 0.02 mm. 

External features. The head is translucent; 
there is no deposition of melanin pigment. 
There are no white or yellow granules about 



the eyes. The operculum (Fig. 127) is cor- 
neous and paucispiral. It is unique in being 
especially thickened abapically and sitting 
cap-like on the foot. The attachment pad is 
prominent, some 56% the width of the oper- 
culum. 

Mantle cavity. Mantle cavity organs are 
shown in Figure 128. Organ measurements 
and counts are given in Table 63. The osphra- 
dium is positioned slightly anterior to mid-gill; 
it is elliptical and short. There are 13 to 19 gill 
filaments with both Gf., and Gf 2 elements 
prominent. The longest gill filaments are 0.48 
± 0.09 mm long. Gill filament section Gf 2 is 
long. In side view, the largest gill filaments are 
moderately domed. 

The portal for sperm entry into the pericar- 
dium (Ope) is shown. The pericardium (Pe) is 
considerably swollen and there is an enor- 
mous pericardial bursa (Pbu) pushing the lin- 
ing of the mantle cavity far forward into the 
mantle cavity. 

Female reproductive system. An uncoiled fe- 
male without head and with kidney tissue re- 
moved is shown in Figure 129. Measure- 
ments of organs are given in Table 63. 
Important features are: (1) The gonad is pos- 
terior to the stomach. It is small and consists 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 289 

Pbu 



Osd 




FIG. 128. Mantle cavity of Tricula gredlen showing the enormous pericardial bursa (Pbu) bulging into mantle 
cavity. 



TABLE 62. Shell measurements (mm) of Tricula gredleri syntypes and of shells of snails used for 
dissections. All shells that had eroded apices = e. Mean ± standard deviation (range). N = number 
measured. M = male, F = female. 





Syntypes (N = 


= 2) 


For Anatomy (N = 5) 


No. Whorls 


1. 
4e 


2. 
5e 


3e-4e 


Length (L) 


3.08e 


3.1 2e 


3.16 (4e,M); 3.32 (4e,F) 


Width (W) 


1.40 


1.36 


1.52±0.11 (1.36-1.64) 


L last three whorls 


2.80 


2.76 


3.01 ±0.11 (2.84-3.16) 


L body whorl 


2.04 


2.08 


2.20±0.11 (2.12-2.36) 


L penultimate whorl 


0.44 


0.40 


0.54±0.02 (0.52-0.56) 


W penultimate whorl 


0.92 


0.96 


1.06±0.05 (1.00-1.12) 


W 3rd whorl 


0.64 


0.60 


0.71 ±0.04 (0.68-0.76) 


L aperture 


1.48 


1.48 


1.50±0.08 (1.40-1.56) 


W aperture 


0.80 


0.88 


0.99±0.05 (0.92-1.04) 


X 






0.38±0.02 (0.36-0.40) 


У 






0.18±0.06 (0.10-0.24) 



290 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Eme 




Apo 



1.0mm 

FIG. 129. Uncoiled female Tricula gredlen with head and kidney tissue removed. 




10r 



FIG. 130. Variation in gonad structure of female 
Tricula gredleri. 



of one to four bundles of lobes Figures 129, 
130. (2) The bursa copulatrix (Bu) is round to 
sub-triangular, minute, and clearly visible 
posterior to the albumen gland (Ppo). (3) The 
albumen gland is of normal size. (4) The 
bursa copulatrix complex of organs is shown 
in Figures 131, 132. In Figure 131 the orien- 
tation of the bursa complex is the same as in 
Figure 129. The 360° twist of the oviduct (Ov) 
and the duct of the bursa arising from the ovi- 
duct are seen in Figure 131 ; these two char- 
acter-states are seen in all genera of the tribe 
Triculini. (5) The species is unique in having 
such an enlarged pericardial bursa (Pbu). (6) 
In Figure 132A and B, it is shown that the duct 
of the seminal receptacle arises from the ovi- 
duct slightly posterior to the opening of the 
duct of the bursa either slightly offset dorso- 
lateral^, or directly in line with the opening of 
the duct of the bursa (Fig. 132 C). (7) The 
seminal receptacle (Sr) is a small sphere. (8) 
There is no spermathecal duct, or, at best, a 
minute one; the oviduct appears, in gross dis- 
section, to be fused to the pericardium (Sdo. 



Males (N = 2) 


4.80 


(4.60, 5.00) 


2.60 


(2.40, 2.80) 


1.97 


(1.92, 2.02) 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 291 



TABLE 63. Lengths (mm) or counts of non-neural organs and structures of Jacula gredleri. N = number 
of snails used. Mean ± standard deviation (range). 

Females (N = 5) 

Body 4.96±0.45 

(4.30-5.44) 
Gonad 0.85±0.11 

(0.70-1.00) 
Digestive gland 2.17±0.32 

(1.70-2.50) 
Posterior palliai oviduct 1.05±0.19 

(= albumen gland) (0.80-1.20) N = 4 

Anterior palliai oviduct 1.13±0.15 — 

(= capsule gland) (1.00-1.30) N = 4 

Total palliai oviduct 2.14±0.17 — 

= OV (2.00-2.40) 

Bursa copulatrix 0.31 ±0.01 — 

= BU (0.31-0.32) 

Duct of BU 0.17±0.01 — 

(0.16-0.18) N = 3 
BU + OV 0.14±0.01 — 

(0.13-0.16) 
Seminal receptacle 0.17±0.01 — 

(0.16-0.18) N = 4 
Duct of seminal receptacle 0.08 ±0.02 — 

(0.06-0.10) N = 4 
Mantle cavity 1.42 ±0.13 1.55 

(1.30-1.60) (1.50,1.60) 

Gill (G) 1.07±0.17 1.11 

(1.30-1.60) (1.10,1.12) 

Osphradium (OS) 0.35±0.08 0.31 

(0.30-0.46) N = 4 (0.24, 0.38) 

Os * G 0.35±0.09 .28 

(0.25-0.46) N = 4 (0.22, 0.34) 

No. of filaments 16.8±2.3 16.0 

(13-19) (Novar.) 

Buccal mass 0.58 ±0.09 — 

(0.50-0.68) 
Gf 2 0.26 ±0.06* — 

(0.20-0.36) N = 6 
G^ 0.22±0.08* — 

(0.18-0.26) N = 6 
Total Gf = TGF 0.48 ±0.09* — 

(0.38-0.62) N = 6 
Gf 2 - TGF 0.54±0.03* — 

(0.50-0.58) N = 6 
Prostate — 0.93 

(0.86, 1 .00) 
Seminal vesicle — 0.70 

(0.60, 0.80) 
Penis — 2.60 



(2.00, 3.20) 



'males and females 



292 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




Pbu 



0.33 г 



FIG. 131 . Details and variation of bursa copulatrix complex of organs relative to style sac (Sts), pericardium 
(Pe) and posterior end of mantle cavity (Emc). 



TABLE 64. Radular statistics for Tricula gredleri. Mean ± standard deviation (range). N = number used. 
In mm except for width of central tooth in (xm. 



Females (N = 3) 



Males (N = 4) 



Shell length 

Radular length 

Radular width 

Total rows of teeth 

No. rows of teeth forming 

Central tooth width 



3.41 ±0.17 
(3.28-3.60) 
0.49±0.10 
(0.39-0.60) 
0.07±0.002 
(0.072-0.076) 
56 ±4.4 
(53-61) 
16.0±2.0 
(14-18) 
13.7±1.0 
(12.8-15.0) N 



3.30 

(3.10, 3.40) N 

0.51 ±0.03 

(0.47-0.54) 

0.07±0.01 

(0.060-0.072) 

63.4±4.2 

(61 -70) 

16.3±1.5 

(15-18) 

15.1±1.4 

(13.1-17.1) N 



17 



Fig. 132B). (9) Sperm enter the female sys- 
tem from the mantle cavity into the pericar- 
dium hence to the oviduct. 

Male reproductive system. An uncoiled male 
without head and with kidney tissue removed 
is shown in Figure 133. Two bundles of go- 
nadal lobes are cut away to show the seminal 



vesicle (Sv). Measurements are given in Ta- 
ble 63. Important features are: (1) The gonad 
fills the digestive gland (Di) and covers the 
posterior and anterior chambers of the stom- 
ach. (2) The prostate (Pr) overlaps the poste- 
rior end of the mantle cavity (Emc). (3) The 
seminal vesicle (Sv) arises from the vas effe- 
rens (Ve) about mid-gonad. (4) The seminal 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 293 

А В 



Oov 



Oov 




Dsr 



FIG. 132. Details and variation of bursa copulatrix complex of organs of Tricula gredleri. A. Compared with 
Figure 131 , bursa copulatrix cut off and duct system rotated 90° to show position of seminal receptacle (Sr) 
and its duct (Dsr). Complex in A is further rotated in В to show where duct of seminal receptacle (Dsr) opens 
into oviduct (Ov). С Section of oviduct oriented as in Figure 131 showing a variation in position of opening 
of duct of seminal receptacle into oviduct, the usual Tricula position. 

TABLE 65. Cusp formulae for the radular teeth of Tricula gredleri with the percent of the seven radulae 
in which a given formula was found at least once. 



Central Teeth 



Lateral Teeth 



Inner Marginal Teeth 



Outer Marginal Teeth 



3-1-3 
2-2 


71% 


2-[2]-3 


86% 




8 




29% 


3-1-3 
3-3 


14% 


3-[2]-2 


86% 




9 




57% 


3-1-2 
2-2 


14% 


2-[2]-4 


43% 




10 




71% 


2-1-2 
2-2 


14% 


4-[2]-2 


14% 




11 




57% 


4-1-3 
3-2 


14% 






X 

N 


* = 9.69±2.01 
= 70 



8 14% 

9 71% 

10 57% 

11 71% 



9.0±3.1 
N = 70 



"Mean ± standard deviation of cusp number for all teeth counted. 
[ ] = blade bifurcates thus resembling two cusps. 



294 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




0.5r 



FIG. 133. Uncoiled male of Tricula gredleri without head or kidney tissue. Two sections of gonadal lobes 
removed to reveal seminal vesicle coiled dorsal to gonad. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 295 




В 




FIG. 134. A. Relationship of base of penis of Tricula 
gredleri to mid-line of snout-neck (x) and to poste- 
rior end of eye lobes (a). B. Penis. 



vesicle coils dorsal to the gonad from mid- 
gonad anterior to cover the posterior chamber 
of the stomach. (5) The anterior vas deferens 
(Vd 2 ) leaves the prostate near the posterior 
end of the mantle cavity (Emc). (6) The penis 
is unique among triculine taxa (Figure 134B). 
The anterior end is blunt with a protudable 
papilla (Pa) emerging from the edge of the 
square penial tip at the concave side of the 
penis. (7) The ejaculatory duct is massive and 
extends as a wide muscular duct from the 
base of the penis along the dorsal aspect of 




FIG. 135. Stomach of Tricula gredleri oriented ex- 
actly as in Figures 129, 131, and 133. 



TABLE 66. Lengths of neural structures of Tricula 
gredleri. Mean ± standard deviation (range). N = 
number used = 5. * - neural elements measured 
to obtain the RPG ratio. 



Cerebral ganglion 

Cerebral commissare 

Pleural ganglion 
Right (1)* 

Left 
Pleuro-supraesophageal 

connective (2)* 
Pleuro-subesophageal 

connective 
Supraesophageal 

ganglion (3)* 
Subesophageal ganglion 

Osphradio-mantle nerve 

RPG ratio* = 
2 ■* 1+2 + 3 



0.24±0.02 (0.22-0.26) 
0.04+0.02 (0.02-0.06) 

0.12+0.02(0.10-0.14) 
0.12+0.02(0.10-0.14) 

0.14+0.02(0.12-0.16) 

0.14+0.03(0.10-0.16) 
0.12+0.02 (0.10-0.14) 

0.12+0.02(0.10-0.14) 
0.07+0.03 (0.04-0.12) 
0.37+0.04(0.33-0.41) 



the neck to the end of the neck. (8) The ori- 
entation of the base of the penis to the snout- 
neck mid-line is shown in Figure 134A. It is to 
the right of the mid-line ("x"), and at an angle 
of 45°-50° to it. 

Digestive system. The digestive gland is pos- 
terior to the stomach of both sexes. The stom- 
ach is shown in Figure 135. Two features of 
note are: (1) there is a groove (Gr) or crease 
between the anterior chamber of the stomach 
(Ast) and the section of stomach receiving the 
esophagus (Es) and duct of the digestive 
gland (Odi). (2) The anterior chamber (Ast) is 
yellow with a centrally positioned grey area. 
Radular statistics are given in Tables 64 
and 65. There appears to be sexual dimor- 
phism in the number of rows of teeth on the 



296 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 136. Radula of Tricula gredleri. A. Section of radular ribbon. B-E. Central, lateral and inner marginal 
teeth. F. Outer marginal teeth. Note pronounced bifurcation of the blade of the major cusp of lateral tooth 
(especially in Figs. C-E). 



radular ribbon; males have more (61 to 70 on 
a radula 3.30 mm long). However, the sample 
size is small. The most frequently encoun- 
tered formula is 

3-1-3 ; 2(3)-[2]-2(3); 9-1 1 ; 9-1 1 . 



(3)2-2(3) 



SEM photographs of radulae are shown in 
Figure 136. The dominant cusp of the lateral 
tooth (Fig. 136B-E) has a blade that bifur- 



cates. Otherwise the tooth morphologies are 
standard triculine. 

Nervous system. Measurements are given in 
Table 66. The RPG ratio of 0.37 indicates that 
the dorsal nerve ring is moderately concen- 
trated. The pleuro-subesophagal connective 
of this species is unusual for its length, equal 
to that of the pleuro-supraesophageal con- 
nective. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 297 



Remarks 

Conchologically, Tricula gredleri is most 
similar to N. duplicata (Figs. 153-155). Con- 
chological comparison with N. duplicata is 
given in the remarks section for that species. 
Anatomically, T. gredleri has the generic-level 
character-states that serve to differentiate it 
from N. duplicata. 

Tricula gredleri has seven unique anatom- 
ical character-states (Tables 80, 81). The 
operculum is cap-like, not a flat sheet (char. 
2). The oviduct is fused to the pericardium; 
there is no discernable length of spermathe- 
cal duct (chars. 19,20). The penial opening of 
this species arises form the concave edge of 
a blunt tip (char. 34). The anterior chamber of 
the stomach is lemon yellow (char. 40). The 
pleuro-subesophageal connective is long 
(char. 48). 

Tricula gredleri belongs to the species 
group within Tricula that has a pericardial 
bursa T. montana, T. gredleri, T. gregoriana 
and T. maxidens. Its closest relationship an- 
atomically is with T. odonta. 

Tricula maxidens Chen & Davis, sp. nov. 

Holotype. ZAMIP, M0009, Figure 137A. 

Paratypes. ANSP 373140, A12656, Figure 
137B-E. 

Type locality. Yantuo Village, Xiaoguanping 
Town, Guzhang County, Xiangxi Prefecture; 
28°42'7"N, 110°0'37"E. Figure 1, site 12. 

Etymology, contraction from the Latin maxi- 
mum (the greatest) and dens (tooth). 

Habitat 

These snails were collected in sympatry 
with Tricula gredleri and Akiyoshia chinensis 
from a small mountain stream at an altitude of 
700 m. The field number assigned was D87- 
3. 

Description 

Shell. Shells are small, smooth, and cylindri- 
cal-conic (Figs. 137A-E; 138A-D, F). They 
mature at 5.5 whorls with the largest size 
class ranging in length from 2.08-2.24 mm 
(Table 67). The sutures are deep and the 
whorls are slightly convex. Cleaned shells are 
glassy; there is no umbilicus. The aperture is 
semi-circular with the inner lip a flat-straight 
edge. The inner lip is fused to the body whorl. 



The outer lip in side view, is straight, without 
notch or sinuation. With the outer lip down 
and 90° to the horizontal, the inner lip is seen 
in side view with a slight angulation. The most 
prominent feature is the relatively gigantic col- 
umellar tooth seen in the aperture (Fig. 138B) 
that continues inside the shell wrapping the 
columella adapically. SEM analyses of the 
apical whorls indicates that they are minutely 
malleated to smooth (Fig. 138D, F). 

External Features. The head is devoid of mel- 
anin pigment. There are no white or yellow 
granules or glands around the eyes. The 
operculum (Fig. 138E) is corneous, paucispi- 
ral and kidney-bean shaped. The embayment 
on the columellar side corresponds to the 
large tooth on the columella. The attachment 
pad is narrow (33% of operculum width) and 
moderately prominent. 

Mantle cavity. Mantle cavity organs are 
shown in Figure 139A. Organ measurements 
and counts are given in Table 68. The osphra- 
dium (Os) is mid-gill, and long. There are 10 
to 15 well developed gill filaments. The long- 
est gill filament are 0.27 ± 0.01 mm long. The 
Gf 2 part of the gill is long. Sperm enter the 
pericardium at the rear of the mantle cavity 
(Ope). The pericardium (Pe) is moderately 
swollen and bulges out into the mantle cavity. 
A discrete but small and delicate pericardial 
bursa (Fig. 139D) was seen in two individuals, 
it measured 0.30 x 0.14 mm. 

Female reproductive system. An uncoiled fe- 
male without head and with kidney tissue re- 
moved is shown in Figure 140. Measure- 
ments of organs are given in Table 68. 
Important features are: (1) The gonad (Go) is 
posterior to the stomach; it is small and with 
bundles of lobes. (2) The bursa copulatrix is 
round and small; half the bursa is dorsal to the 
posterior end of the albumen gland (Ppo). (3) 
The albumen gland is of standard length. (4) 
The bursa copulatrix complex of organs is 
shown in Figure 139. The orientation in Figure 
139A is the same as that in Figure 140. The 
bursa copulatrix (Bu) is short. (5) The seminal 
receptacle (Sr) is spherical; it has a short duct 
(Dsr) opening into the right ventro-lateral 
edge of the oviduct (Ov) just posterior to the 
opening of the duct of the bursa (Dbu) into the 
oviduct. (6) The spermathecal duct (Sd) is 
moderately long; it runs to open into the peri- 
cardium (Pe). (7) The pericardium is swollen 
with sperm and distends into the posterior 
mantle cavity. In some specimens, a minute 



298 DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 137. Shells of Tricula maxidens, A-E; Tricula odonta, F-K. A. Holotype; B-E. Paratypes. Length of A 
= 1.94 mm; other shells printed to same scale. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 299 




FIG. 138. SEM photographs of shells and opercula of Tricula maxidens. A. Shells of mature individuals. Note 
straight outer lip (right photo) and prominent columellar tooth. B. Enlargement of aperture showing columel- 
lar tooth. C, D, F. Protoconch. Although encrusted, protoconch is seen to be vaguely malleated. E. Opercula: 
right operculum shows outer surface with paucispiral growth line; left operculum shows inner surface with 
narrow attachment pad. 



300 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 67. Shell measurements (mm) of Tricula maxidens. Mean 
broken apical whorls. N = number measured. 



standard deviation (range), b = 





Holotype 




Paratypes 




Large class (N = 5) 


Small Class (N = 3) 


No. Whorls 


5.5 


5.5 


5.5 


Length (L) 


1.94 


2.17±0.06 


1.97±0.06 






(2.08-2.24) 


(1.92-2.04) 


Width (W) 


0.86 


0.92±0.03 


0.82±0.03 






(0.90-0.96) 


(0.80-0.86) 


L last three 


1.82 


1.95±0.03 


1.73±0.08 


whorls 




(1.92-2.00) 


(1.68-1.82) 


L body whorl 


1.24 


1.34±0.04 


1.16±0.10 






(1.28-1.36) 


(1.08-1.28) 


L penultimate 


0.40 


0.40±0.03 


0.35±0.02 


whorl 




(0.36-0.44) 


(0.32-0.36) 


W penultimate 


0.68 


0.71 ±0.01 


0.62±0.02 


whorl 




(0.70-0.72) 


(0.60-0.64) 


W 3rd whorl 


0.48 


0.50±0.01 


0.46±0.02 






(0.48-0.52) 


(0.44-0.48) 


L aperture 


0.76 


0.86±0.02 


0.73±0.06 






(0.84-0.90) 


(0.68-0.80) 


W aperture 


0.58 


0.62±0.02 


0.55±0.05 






(0.60-0.64) 


(0.52-0.60) 


X 


0.30 


0.34±0.02 


0.32 






(0.32-0.36) 


no variation 


У 


0.08 


0.08±0.01 


0.07±0.03 






(0.06-0.10) 


(0.04-0.10) 



pericardial bursa was observed (Fig. 139D, 
Pbu). 

Male reproductive system. An uncoiled 
male with head and kidney tissue removed is 
shown in Figure 141. Lengths of non-neural 
organs are given in Table 68. Important fea- 
tures are: (1 ) The gonad (Go) covers the pos- 
terior chamber of the stomach and most of 
anterior chamber (Ast). (2) The prostate (Pr) 
is relatively massive and overlaps the poste- 
rior end of the mantle cavity (Emc); it covers 
half the style sac. (3) The vas deferens arises 
from mid-gonad to slightly anterior to mid- 
gonad (Fig. 142). (4) The seminal vesicle 
forms a short coil of tubes dorsal to the gonad 
and posterior to the stomach. (5) The anterior 
vas deferens (Vd 2 ) leaves the prostate 
slightly posterior to the anterior end of the 
prostate (Fig. 141). (6) The penis is simple, 
without papilla; it is slender (Fig. 143B). (7) 
There is a pronounced ejaculatory duct (Ej) in 
the base of the penis. (8) The orientation of 
the long axis of the penial base (Bp) to the 
snout-neck mid-line (x) is shown in Fig. 143A. 
The penial base is to the right of the mid-line 
and at an angle of 40°. 

Digestive system. The digestive gland is pos- 
terior to the stomach in both sexes. Radular 



statistics are given in Tables 69 and 70. There 
are 59.8 ± 6.4 rows of teeth along a radula of 
0.29 ± 0.04 mm ( = equivalent of 207 teeth 
per mm). The most frequently encountered 
formula is 

3-1-3; 3(4)-1-4; 12-14; 13-14. 
2-2 

The radula is shown in Figure 144; radula 
morphology is of the generalized Tricula type. 
The central anterior cusp of the central tooth 
is long and dagger-like. The dominant cusp of 
the lateral tooth is slender and dagger-like. 

The stomach (Fig. 145) is the usual type 
but with a diagnostic grey streak (Gs) running 
across the ventral surface of the anterior 
chamber (Ast). 

Nervous system. Measurements not taken. 

Remarks 

Conchologically, among species of Triculi- 
nae, T. maxidens is unique and clearly sepa- 
rated from all other species (Figs. 153-155). 
Unique character-states (Tables 2, 76) are 
the cylindrical-conic shape (char. 2) and the 
massive tooth nearly filling the aperture (char. 
16). The columella within the body whorl has 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 301 




Emc 



Pbu 



FIG. 139. Details and variation of bursa copulatrix of organs of Tricula maxidens. Figs. A and В in the same 
orientation as in Figure 1 40. С Enlarged seminal receptacle. D. Pericardium with minute pericardial bursa 
(Pbu). 



a raised spiral keel. The shell seems more 
similar to Pseudobythinella than to Tricula. 

Anatomically, this species is most closely 
allied phenetically to T. xiaolongmenensis 
(Figs. 156-158). It differs from that species in 
1 1 character-states (26%); see Tables 80, 81 . 
Some of these differences are: In T. maxi- 
dens the bursa is covered by the albumen 
gland; in T. xiaolongmenensis the bursa is 
posterior to the albumen gland (char. 14). In 
the former, the duct of the seminal receptacle 
arises from the inside edge of the oviduct; it 
arises from the outside edge in the latter 
(char. 24). The former has a slight pericardial 
bursa; the latter has no pericardial bursa 
(char. 25). The radula is short in the former, of 



medium length in the latter (char. 41). The 
central anterior cusp of the central tooth is 
long and dagger-like in the former, the stan- 
dard type in the latter (char. 44). 

Tricula odonta Liu, Zhang & Wang, 1983a 

Holotype. SX 788104 (= IZAS; Beijing, Peo- 
ple's Republic of China). Figure 1. 

Type locality Shangnan, Shaanxi Province. 
24 October 1978 

Distribution. Shangnan, Shangxian, Zhen- 
ping, Tongguan, Ningqiang and Yuanqu of 
Shanxi Province; Xingshan and Wufeng of 
Hubei Province; Henan Province 



302 DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 68. Lengths (mm) or counts of non-neural organs and structures of Tricula maxidens. N 
number of snails used. Mean ± standard deviation (range). 



Females (N = 5) Males (N = 2) 



Body 3.30±0.33 3.24 

(2.90-3.68) (3.22, 3.26) 

Gonad 0.47±0.09 1.31 

(0.36-0.60) (0.90,1.72) 

Digestive gland 1.32±0.19 1.41 

(1.00-1.48) (1.40,1.42) 

Posterior palliai oviduct 0.58±0.11 — 

(= albumen gland) (0.54-0.70) N = 3 

Anterior palliai oviduct 0.67±0.15 — 

(= capsule gland) 0.50-0.80) N = 3 

Total palliai oviduct 1.23±0.23 — 

= OV (1. 00-. 150) 

Bursa copulatrix 0.24±0.05 — 

= BU (0.22-0.30) 

Duct of BU 0.09±0.01 — 

(0.08-0.10) N = 4 

BU ■*■ OV 0.20±0.06 — 

(0.16-0.30) 

Seminal receptacle 0.07±0.01 — 

(0.06-0.08) N = 3 

Duct of seminal receptacle 0.02 — 

N = 1 

Mantle cavity 0.81 ±0.08 0.82 

(0.74-0.90) N = 3 N = ' 

Gill (G) 0.66±0.13 0.70 

(0.54-0.80) N = 3 N = 1 

Osphradium (OS) 0.28 0.26 

(0.24, 0.32) N = 2 N = 1 

OS - G 0.39 0.37 

(0.38, 0.40) N = 2 N = 

No. filaments 12.7±2.5 14 

(10-15) N = 

Gf 2 0.17±0.01 - 

(0.16-0.18) 

Gi, 0.11 ±0.01 — 

(0.10-0.12) 

Total Gf = TGF 0.27±0.01 — 

Gf 2 - TGF 0.61 ±0.04 — 

(0.57-0.64) 

Prostate — °- 75 

(0.70, 0.80) 

Seminal vesicle — °- 65 

(0.40, 0.90) 

Penis — °- 64 

(0.52, 0.76) 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 303 



Ast 



Eme 




Apo 



FIG. 140. Uncoiled female Tricula maxidens with head and kidney tissue removed. 



304 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 
Ast In 




0.5mm 

FIG. 141. Uncoiled male of Tricula maxidens without head or kidney tissue. 




FIG. 142. A. Figure to show seminal vesicle (Sv) of 
Tricula maxidens in relationship to stomach. Most 
of gonad and vas efferens cut away. 



TABLE 69. Radular statistics for Tricula max- 
idens. Mean ± standard deviation (range). N = 
number used. In mm except for width of central 
tooth in ц.т. 

Females (N = 4) 



Shell length 
Radular length 
Radular width 
Total rows of teeth 

No. rows of teeth 

forming 
Central tooth 

width (N = 18) 



2.1 9±0.08 (2.08-2.24) 
0.29±0.04 (0.240-0.204) 
0.03±0.01 (0.028-0.040) 
59.8±6.4 (55-69) 
11.3±1.5(9-12) 

9.6±0.3 (9.0-10.1) 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 

Habitat 



305 





FIG. 143. Tricula maxidens: A. Relationship of base 
of penis to mid-line of snout-neck (x) and to poste- 
rior end of eye lobes (a). B. Penis. 



Material for this study was collected on 7 
October 1985 from Shenglong Village, San- 
huokou Town, Cili County, Changde Prefec- 
ture; 29°38'35"N, 110°39'12"E; Figure 1, site 
7. The field collection number assigned was 
D85-82. Specimens came from a small 
stream flowing to the Lishui River. The collec- 
tor was Dr. Liu Wen Jian of Cili County Public 
Health Station. 

Depository 

Specimens for this study were deposited in 
ZAMIP, M0005; ANSP, 368773, A12145. 

Description 

Shell. Shell measurements are given in Table 
71; See Figures 137F-K, 146. Mature males 
and females are primarily 7.0 whorls; rela- 
tively few are 6.0 or 6.5 whorls (Table 71). 
Only one specimen of 7.5 whorls was ever 
seen (within the size range of 7.0 whorls 
snails of Table 71). Shells are medium sized 
with a length range of 3.76 to 4.36 mm for 
shells of 7.0 whorls (the dominant size class). 
Shells are ovate-turretted. The whorls are 
slightly convex; the sutures are deep. The 
peristome is complete; the inner lip is widely 
separated from the body whorl along its entire 
length (0.08 ± 0.02 mm, range 0.06-0.10; 



TABLE 70. Cusp formulae for the radular teeth of Tricula maxidens with the percent of the seven radulae 
in which a given formula was found at least once. 



Central Teeth 


Lateral Teeth 


Inner Marginal 
Teeth 


Outer Marginal 
Teeth 


3-1-3 100% 


3-1-4 


75% 


12 


75% 


12 


25% 


2-2 


4-1-4 


75% 


13 


100% 


13 


75% 


4-1-4 25% 
2-2 


2-1-3 


25% 


14 


100% 


14 


100% 


4-1-3 25% 


5-1-4 


25% 


15 


25% 


15 


25% 


2-2 
3-1-4 25% 






16 


25% 


16 


25% 


2-2 














2-1-3 25% 
2-2 






X* = 
N = 


= 13.4±0.9 
40 


' 


3.9±0.9 
N = 40 


3-1-3 25% 














2-3 















* Mean ± standard deviation of cusp number for all teeth counted. 



306 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



4W*^7Vi 



¿~а у, *2£<у:. 



jÉàl 



p7 : 






^»v 



Л »V l ч'У __2SJ-\ 1 .+JP5?s.\\ \ /^ jfc. • О 

iÉ% j 5ur1 z 



f*yw,~y~ tt 



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tu, 






FIG. 144. Radula of Tricula maxidens. A. Part of radular ribbon. В, С. Central, lateral and inner marginal 
teeth. D, E. Outer marginal teeth. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 307 




0.5mm ES 

FIG. 145. Stomach of Tricula maxidens. 

N = 5). The aperture is pyriform. The adapical 
end of the inner lip has a tooth-like swelling; 
this, plus the thickening of the outer lip oppo- 
site the swelling, enclose an adapical notch. 
Mid-columella there is a marked indentation. 
Although the inner lip is separated from the 
body whorl, there is no umbilicus in some in- 
dividuals; there is one in some. Facing the 
aperture of the shell the adapical outer lip 
may be sinuate (16%); in side view, the outer 
lip is scooped forward (83%) or straight 
(17%). When the outer lip is scooped forward 
there is an adapical sinuation or notch. There 



is no basal post (see Davis et al., 1986). The 
cleaned shell is glistening, glassy. 

The nuclear whorls (Fig. 146) have a rough, 
wrinkled surface. There are 1.5 nuclear 
whorls. The roughened surface may not be 
discerned at magnifications less than 200 or 
300X. The aperture has six diagnostic fea- 
tures: (1) There is an adapical notch or beak 
bounded by (2) a thickening or node on the 
inner lip; (3) The inner lip has an angulation 
located some 60% of the length of the inner 
lip abapically from the adapical end of the ap- 
erture. The inner lip is thickened noticeably 
from the fulcrum of the angle to the apertural 
notch. (4) There is an indentation at the ful- 
crum of the angle. Abapical to the fulcrum of 
the angle the inner lip edge is thin. (5) In side 
view, the adapical end of the outer lip is 
slightly sinuate, the remaining lip is straight. 
(6) With the outer lip down, 90° to the hori- 
zontal, the inner lip has a slight sinuation that 
corresponds to a slight trough seen on the 
abapical end of the inner lip in apertural view. 

External features. The head of this species is 
dark gray. No cluster of white granules or 
glands about the eyes were observed. 

The operculum is corneous. The typical 
paucispiral growth line is not seen; the nu- 
cleus is prominent (Fig. 147E-G). The oper- 
culum has an odd irregular shape reflecting 



TABLE 71 . Shell measurements (mm) of mixed males and females of Tricula odonta. Mean ± standard 
deviation (range). N = number measured. 



No. specimens (N) 


N = 7 




N = 2 


N = 1 


No. Whorls 


7.0 




6.5 


6.0 


Length (L) 


4.09±0.22 




3.40 


3.00 




(3.76-4.36) 




(3.32, 3.48) 


— 


Width (W) 


1.52 ±0.10 




1.40 


1.28 




(1.48-1.60) N = 


= 5 


— 


— 


L body whorl 


2.09±0.09 




1.86 


— 




(1.96-2.24) 




(1.84, 1.88) 


— 


L penultimate 


0.69±0.06 




0.58 


0.56 


whorl 


(1.16-1.26) N = 


= 5 


(1.02, 1.08) 


— 


W penultimate 


1.20±0.05 




1.06 


0.98 


whorl 


(1.16-1.26) N = 


= 5 


(1.02, 1.08) 


— 


L last three whorls 


3.25±0.17 




2.78 


2.48 




(3.04-3.48) 




(1.71,2.84) 


— 


L apertue 


1.34±0.05 




1.20 


1.04 




(1.28-1.40) N = 


= 5 


— 


— 


W aperture 


0.89±0.02 




0.85 


0.76 




(0.88-0.92) N = 


= 5 


(0.84, 0.86) 


— 


X 


0.37±0.08 
(0.28-0.48) N = 


= 4 






У 


0.19±0.07 
(0.12-0.28) N = 


= 4 







308 



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FIG. 146. SEM photos of shell apical whorls of 7"r7cu/a odonta. Teleoconch begins at about 1 .50 to 2.0 whorls 
(A, C, G). The protoconch is heavily wrinkled (B, D, F, H). 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 309 




FIGl 147. Radula (A-D) and opercula (E-G) of Tricula odonta. A. Left lateral, inner and outer marginal teeth 
B-D. Marginal teeth. E, G. Inner surfaces. Note irregular shape of opercula and thickened ridge along 
columellar edge of operculum in G. F. Outer surface. All three opercula show the double layers (see text) 



310 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 
Coi 




FIG. 148. Details and variation of bursa copulatrix complex of organs and mantle cavity structures of Tricula 
odonta. A in same orientation as in Figure 149. Not all gill filaments shown. B. Single gill filament. 



E mc 




Ma 



Di 0.75 mm 

FIG. 149. Uncoiled female Tricula odonta with head and kidney tissue removed. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 31 1 



TABLE 72. Lengths (mm) or counts of non-neural organs and structures of Tricula odonta. Mean ± 
standard deviation (range). N = number measured. 

Females (N = 3) Males (N = 2) 

Body 5.83±0.36 4.74 

(5.60-6.24) (4.7, 4.78) 

Gonad 0.74±0.15 1.55 

(0.60-0.90) (1.50,1.60) 

Digestive gland 2.45±0.19 2.04 

(2.24-2.60) (2.00, 2.08) 

Mantle cavity 1.83±0.15 1.53 

(1.80-2.00) (1.50,1.60) 

Osphradium 0.34±0.05 0.33 

(0.30-0.40) (0.30, 0.36) 

Ctenidium = Gill 1.63±0.15 1.38 

(1.50-1.80) (1.36,1.40) 

OS + G 0.21 ±0.04 0.24 

(0.17-0.25) (0.22,0.26) 

Gill filament No. 29.7±1.5 25.5 

(28-31) (24,27) 

Buccal mass 0.57±0.06 — 

(males + females; N = 3) (0.50-0.62) 

Posterior palliai oviduct 0.92 — 

(= albumen gland) (0.64, 1.20) N = 2 

Anterior palliai oviduct 1.12 — 

(= capsule gland) (1.0, 1.24) (N = 2) 
Total palliai oviduct 2.04 

= OV (1 .64, 2.44) N = 2 — 
Bursa copulatrix 0.28±0.03 

= BU (0.24-0.30) — 
Duct of BU 0.21 ±0.04 

(0.16-0.24) — 
BU *■ OV 0.14 

Seminal receptacle 0.11 ±0.03 — 

(0.08-0.14) 

Gf 2 0.26±0.02* — 

(0.24-0.28) 

Gi, 0.23±0.02* — 

(0.20-0.24) 

Total Gf = TGF 0.49±0.01* — 

(0.48-0.50) 
Gf 2 -h TGF 0.53±0.04* 

(0.50-0.58) 

Prostate — 0.75 

(0.70, 0.80) 

Seminal vesicle — 0.66 

N = 1 

Penis — 1 .62 

(1.40, 1.84) 

•males and females 



312 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



Go 




FIG. 150. Uncoiled male of Tricula odonta without head or kidney tissue. 




FIG. 151. A, B. Relationship of base of penis to 
mid-line of snout-neck (x) and to posterior end of 
eye lobes (a) of Tricula odonta; C. Penis. 



TABLE 73. Radular statistics for males and 
females of Tricula odonta. Mean ± standard 
deviation (range). N = 5. In mm except for width 
of central tooth in jim. 



Shell length: 



Radular length 

Radular width 

Total rows of teeth 

No. rows of teeth 

forming 
Central tooth width 



No shell measurements; 
radulae came from 
heads of snails used 
in dissections. 
0.54±0.04 (0.52-0.62) 

0.08±0.004 (0.08-0.09) 

61 .3±4.4 (56-69) 

7±1.7(5-10) 

19.8±2.9 (18.1 -23.6) 



the pyriform shape of the aperture and the 
mid-inner lip indentation. It is particularly nar- 
row at the abapical end, not broadly or regu- 
larly rounded as in those species with an oval 
operculum. The operculum consists of two 
layers that are readily separated if the oper- 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 313 




FIG. 152. Radula of Tricula odonta. A. Part of a radular ribbon. B. Central tooth. С Left lateral tooth. D, F. 
Left lateral and marginal teeth. Note pronounced bifurcation of the major cusp of lateral tooth. E, G, H. 
Marginal teeth. 



314 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 

a given 



74. Cusp formulae for the radular teeth of Tricula odonta with the percent of the radulae in which 
formula was found at least once. N = number of radulae used. 



Central Teeth 


Lateral Teeth 


Inner 


Marginal 


Outer 


Marginal 


N = 6 




N = 


6 


Teeth (N = 3) 


Teeth (N = 4) 


3-1-3 


50% 


2-1[2]-3 


83% 


7 


— 


7 


25% 


3-3 




















3-1-3 


33% 


8 


— 


8 


50% 


3-1-3 


33% 














2-2 




2-1-2 


17% 


9 


33% 


9 


25% 


2-1-2 


33% 














3-3 




1-1[2]-2 


17% 


10 


66% 


10 


50% 


2-1-2 


17% 






11 


33% 


11 


25% 


4-4 
























12 


66% 


12 


50% 


3-1-3 


17% 














4-4 








13 


66% 


13 


25% 










14 


33% 


14 

15 


25% 
25% 










X* = 


11.5±1.6 


9.2 


»±1.7 










N = 30 


N 


= 40 



*Mean ± standard deviation of cusp number for all teeth counted. 



TABLE 75. Lengths of neural structures of four 
individuals of Tricula odonta. Mean ± standard 
deviation (range). * = neural elements measured 
to attain the RPG ratio. 



Cerebral ganglion 


0.24±0.03 




(0.20-0.26) 


Cerebral commissure 


0.03±0.02 




(0.02-0.06) 


Pleural ganglion 




Right (1)* 


0.12+0.02 




(0.10-0.14) 


Left 


0.12+0.02 




(0.10-0.14) 


Pleuro-supraesophageal 




connective (2)* 


0.20+0.07 




(0.12-0.24) 


Pleuro-subesophageal 




connective 


0.05+0.04 




(0.02-0.10) 


Supraesophageal ganglion (3)* 


0.12+0.01 




0.10-0.12) 


Subesophageal ganglion 


0.09+0.03 




(0.06-0.12) 


Osphradio-mantle nerve 


0.05+0.02 




(0.02-0.06) 


RPG ratio* = 2 ■*■ 1+2 + 3 


0.45+0.10 




(0.32-0.55) 



culum is left too long in dilute Clorox used for 
digesting and clearing away tissue fused to 
the operculum There is an inner thickened 
ridge on the columellar side. The attachment 
callus is 54.5 ± 3.5% of the width of the oper- 



culum; it is prominent and much thickened in 
some specimens. 

Mantle cavity. Measurements and counts of 
mantle cavity structures are given in Table 72. 
See Figure 148A. The osphradium is short 
(mean ratio 0.22 ± 0.04; N = 5). The position 
of the osphradium is approximately mid-gill 
(Fig. 148A). The slender aspect of the gill fil- 
ament (Gf 2 ) is long (mean ratio 0.53 ± 0.04; 
N = 3). The length of the longest gill filament 
is 0.49 ± 0.01 mm long (N = 3). The shape of 
the filaments is shown in Figure 148B. It is 
high domed. 

Female reproductive system. The body of an 
uncoiled female with head and kidney tissue 
removed is illustrated (Fig. 149). Measure- 
ments of the relevant organs are given in Ta- 
ble 72. Diagnostic features are: (1 ) The gonad 
is posterior to the stomach. (2) The bursa 
copulatrix (Bu) is posterior to the palliai ovid- 
uct (Ppo) at mid-ventrolateral surface as 
shown. (3) The bursa copulatrix complex of 
organs is the same as seen in T. bollingi 
(Davis, 1968) in that the oviduct makes a tight 
loop (twist of 360°) dorsal to the bursa (Fig. 
148A). (4) The seminal receptacle is spheri- 
cal, posterior to the duct of the bursa (Dbu) 
and fused to the surface of the oviduct on the 
mid-ventral surface, not from the inner curva- 
ture as seen in other species. (5) The sper- 
mathecal duct enters the pericardium; the 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 315 



pericardium is swollen with sperm. (6) Sperm 
enter the pericardium at the posterior end of 
the mantle cavity (Ope, Fig. 148A) (7) The 
bursa copulatrix is minute (mean ratio, 0.15 ± 
0.04; N = 3). (8) The length of the posterior 
palliai oviduct is standard. 

Male reproductive system. The body of an un- 
coiled male with head and kidney tissue re- 
moved is shown (Fig. 1 50). Lengths of relevant 
organs are given in Table 72. Important fea- 
tures are: (1) The gonad either overlaps the 
posterior chamber of the stomach or is pos- 
terior to the stomach. (2) The prostate over- 
laps the posterior end of the mantle cavity and 
covers the anterior half of the style sac. (3) The 
vas deferens arises from mid-vas efferens to 
slightly anterior to mid-gonad. (4) The seminal 
vesicle coils on the stomach; in some individ- 
uals, the posterior vas deferens was not swol- 
len to form the seminal vesicle (presumable 
immature, just pre-sperm production individ- 
ual). (5) The anterior vas deferens (Vd 2 ) 
leaves the prostate slightly anterior to mid- 
prostate. (6) The penis is simple, without pa- 
pilla (Fig. 151C). The opening of the vas def- 
erens is comparatively large and pronounced. 
(7) The base of the penis is 0.18 ± 0.03 mm 
posterior to the eye-lobes (Fig. 151 A, B). Vari- 
ation in the position of the base of the penis to 
the snout-neck mid-line (x) is shown in Figure 
1 51 A, B. A part of the penial base is to the left 
of the mid-line. The long axis of the penial base 
varies from 65° to 90° to the snout-neck mid- 
line. (8) There is a large ejaculatory duct (Ej) 
in the base of the penis (Fig. 151C). 

Digestive System. The digestive gland is pos- 
terior to the stomach. Radular statistics are 
given in Tables 73 and 74. Radular morphol- 
ogy is the generalized Tricula type (Figs. 
147A-D, 1 52). There are 61 rows of teeth per 
radula length of 0.54 mm. The major cusp of 
the lateral teeth is massive and split thus giv- 
ing the appearance of being two cusps. The 
most commonly encountered formula is 

3(2)-1-(2)3; 2(3)-[2]-3; 9-14; 7-14. 
3(2)-(2)3 

Nervous System. Lengths of neural structures 
are given in Table 75. The RPG ratio of 0.45 
shows the dorsal nerve ring to be moderately 
concentrated. 

Remarks 

Conchologically, aside from Neotricula ap- 
erta, T. odonta has the most distinctive shell 



(Fig. 153) in that it attains 7.0 whorls (Tables 
2, 76) (char. 29). It has a beak tubercle (char. 
24) as do two other species: Tricula gredleri 
and N. dianmenensis. The adapical aperture 
is widely separated from the body whorl as is 
seen in N. cristella (char. 21). The inner lip is 
widely separated from the body whorl as it is 
in N. cristella (char. 20). A unique state, the 
inner lip is differentially thickened (char. 19). 
Another unique state, the inner lip has a notch 
(char. 17). Among species of Tricula, only this 
species of those thus far studied, has an 
abapical apertural spout (char. 12), an angled 
inner lip (char. 13), and an abapical lip deflec- 
tion angle, (char. 18). 

Anatomically, Tricula odonta stands apart 
from other species of Tricula (Tables 80, 81), 
but is most similar to T. xianfengensis (Figs. 
1 56-1 58). Tricula odonta differs from the latter 
in 17 characters (37%). Most notable are the 
difference in operculum shape that reflects 
differences in shell apertural shape (char. 2), 
the differences in Gf 2 length (char. 8), female 
gonad length (char. 12) and seminal recepta- 
cle duct length (char. 23). Tricula odonta has 
a large ejaculatory duct; the latter does not 
have an ejaculatory duct (chars. 36, 37). The 
radula of the former is of medium length, of 
the latter, long (char. 41). The dorsal nerve 
ring is moderately concentrated in the former, 
elongated in the latter (char. 42). 



MULTIVARIATE RELATIONSHIPS 
Shells 

The 29 shell characters scored are given in 
Table 2, the actual scores in Table 76. All 
species of Tricula and Neotricula, as well as 
other species having shells resembling those 
of Tricula or Neotricula and for which anatom- 
ical data are available, were included. The 
sources for data for species not treated in this 
paper are listed in Appendix I. The pheno- 
gram based on UPGMA treatment of distance 
coefficients is given in Figure 153. Distance 
coefficients were used because the cophe- 
netic correlation (phenogram to origninal ma- 
trix) was r = 0.928 for distances; r = 0.725 
using similarities. 

It is clear that one cannot sort species to 
genus on the basis of shell characters. Addi- 
tionally, of the 21 species, four (19%) could 
not be clearly distinguished on the basis of 
the shell characters used; Tricula bollingi phe- 
notypes 1 and 3 clustered with T. ludongbini, 



316 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 76. Scoring of 44 OTUs for 29 shell characters listed in Table 2. Shells from the same population 
of some species varied for some characters. Such a species was divided into as many as three OTUs, 
e.g. xin-1, xin-2, xin-3. All species for which there were adequate anatomical data were used. 
Abbreviations are : ape, N. apena; bur, N. burchi; cri, N. cristella; dia, N. dianmenenensis; dup, N. 
duplicata; NI, N. lilii; min, N. minutoides; bam, T. bambooensis; boll, T. bollingi; grd, T. gredleri; grg, 7". 
gregoriana; hud, T. hudiequanensis; lud, T. ludongbini; max, T. maxidens; mon, T. montana; odn, T. 
odonta; xin, T. xianfengensis; xia, T. xiaolongmensis; chi, G. chinensis; jin, J. jinghongensis; niz, W. 
niuzhuangensis. 















1 


Neotricula 


















Thcula (N = 


11) 










1 


2 


3 


4 


5 


6 


7 


8 


9 


10 


11 


12 


13 


14 


15 


16 


17 


18 


19 


20 


21 


22 




ape 


bur-1 


bur-2 


cri-1 


cri-2 


dia-1 


dia-2 


dup-1 


dup-2 


lil-1 


lil-2 


min-1 


min-2 


bam-1 


bam-2 


bol-1 


bol-2 


bol-3 


grd-1 


grd-2 


grg-i 


grg-2 


1 


1 



























































1 


1 


2 


4 


1 


1 



































1 























3 


1 














2 


2 


1 


1 


1 


1 


1 


1 











1 





1 


1 


1 


1 


4 








1 








2 


2 


1 


1 


2 


2 


2 


2 


2 


2 





1 


2 


1 


1 


1 


2 


5 





1 


1 



























































6 





1 


1 


1 


1 


1 


1 


1 


1 


1 


1 





1 





























7 




































































8 


2 








1 


1 








1 


1 








1 


1 


1 


2 


3 


3 


3 


1 


1 








9 


2 








1 


1 


2 


2 


2 


2 


1 


1 























2 


2 





1 


10 


1 




















1 












































11 

















1 


1 








1 


1 























1 


1 








12 





1 











1 


1 


1 


1 


1 


1 



































13 


1 


1 


1 


1 


1 


3 


3 





1 


2 


2 


1 


1 


1 


1 


1 


1 


1 








1 


1 


14 





1 


1 


1 


1 


1 


1 





























1 











1 


1 


15 


2 


1 


1 











1 


1 


1 


1 


1 


1 


1 


2 


2 





1 








1 








16 




































































17 




































































18 











1 





1 


1 


1 


1 


1 


1 



































19 























1 


1 








1 


1 


1 


1 











1 


1 








20 











3 


3 


1 





2 


2 


2 


1 


2 


2 


2 


2 





1 





2 


2 








21 











2 


2 





1 


1 


1 





1 








1 


1 





1 





1 


1 








22 




































































23 











1 


1 














1 


1 











1 























24 

















1 


1 



































1 


1 








25 





























1 


1 



































26 


1 

































































27 


1 

































































28 




































































29 





































































whereas phenotype 2 clustered with T. xia- 
olongmenensis phenotype 1. Tricula xia- 
olongmenensis phenotype 2 did not have 
close affinity to any other species in the upper 
half of the phenogram. Tricula montana phe- 
notype 3 clustered with T. gregoriana, not 
with phenotypes 1 and 2 of Tricula montana. 
Tricula xianfengensis phenotype 2 did not 
have close affinity with any other species in 
the upper half of the phenogram. While most 
shells can be classified readily on the basis of 
the characters used (exact measurements 
and whorl numbers would allow further dis- 
crimination), four of the 44 OTUs (only 9%) do 
not cleanly segregate with their other conspe- 
cific OTUs. 
It is useful to compare the phenogram with 



a plot of the Prim Network (Minimum Span- 
ning Tree [MST]) (Fig. 154) based on the dis- 
tance coefficients (drawn to scale of actual 
pathway distances). The two are useful in as- 
sessing those species most closely resem- 
bling a species of immediate concern. For ex- 
ample, what species might be conchologically 
confused with Tricula gredleri? An examina- 
tion of Figures 153 and 154 indicates that N. 
duplicata is the most closely allied pheneti- 
cally. The several differences are then readily 
found in Tables 2 and 76. With MST, only two 
OTUs of different species do not link with 
other conspecific OTUs (i.e. T. montana phe- 
notype 3; T. bollingi phenotype 2). Thus, only 
5% of the OTUs do not segregate cleanly. 
Table 77 provides a ranking of species by a 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 317 
TABLE 76. (Continued) 





































Gamma- 






Wucon- 




Tricula (N 


= 11) 












Tricula 














tricula 


Jinhongia 


chona 




23 


24 


25 


26 


27 


28 


28 


30 


31 


32 


33 


34 


35 


36 


37 


38 


39 


40 


41 


42 


43 


44 




hud-1 


hud-2 lud-1 


lud-2 


max-1 


max-2 


mon-1 


mon-2 


mon- 


3 odn-1 


odn-2 


xin-1 


xin-2 


xin-3 


xio-1 


xio-2 chi-1 


chi-2 


jin-1 


jin-2 


niz-1 


niz-2 


1 


1 


1 























1 


1 


1 


2 


1 


























2 





1 








3 


3 


1 


2 


1 


1 


1 





1 


2 


1 


1 











1 








3 























1 


1 


1 


1 











1 


1 














1 


1 


4 


1 


2 


2 


2 











1 


2 


2 


2 


1 


1 


1 





1 


1 


2 





1 





1 


5 


















































1 


1 


1 


1 








6 





















































1 


1 


1 


1 


1 


7 

















2 






































1 


1 








8 


1 


2 


1 


2 

















1 


1 


2 


2 


2 


2 


2 











1 





1 


9 





























1 


1 





1 








1 














1 


1 


10 




































































11 





























1 


1 



































12 





























1 


1 























1 


1 








13 


1 


1 


1 


1 











1 





3 


3 

















1 


1 


1 


1 


2 


2 


14 


1 


1 




















1 














1 


1 





1 


1 


1 











15 


1 


1 





1 











1 








1 




















1 


1 


1 





1 


16 














1 


1 


















































17 





























1 


1 



































18 





























1 


1 


























1 


1 


1 


19 


1 


1 























2 


2 

















1 


1 














20 


2 


2 























3 


3 

















2 


2 














21 


1 


1 


1 


1 


1 


1 


1 


1 


1 


2 


2 











1 


1 


1 


1 














22 














2 


2 


























1 


1 














2 


2 


23 















































1 


1 


1 





1 








24 





























1 


1 



































25 






































1 











1 


1 














26 




































































27 



























































1 








28 
























































1 


1 








29 





























1 


1 




































TABLE 77. Ranking species on a consistency index for shell characters. The index = 1.0-[number 
of character state differences between phenotypes in a population ■*■ 29 characters]. Numbers in ( ) 
= number of phenotypes. 



N. aperta (1) 
N. cristella (2) 
N. minima (2) 
T. gredleri (2) 
T. maxidens (2) 
T. odonta (2) 
N. burchi (2) 
N. duplicata (2) 
N. lilii (2) 
T. gregoriana (2) 



1.0 

0.97 

0.97 

0.97 

0.97 

0.97 

0.93 

0.93 

0.93 

0.93 



T. ludongbini (2) 

N. dianmenensis (2) 

7". bambooensis (2) 

7". hudiequanensis (2) 

G. chinensis (2) 

W. niuzhuangensis (2) 

T. xiaolongmenensis (2) 

T. xianfengensis (3) 

7". bollingi (3) 

T. montana (3) 

J. jinhongensis (2) 



0.93 
0.90 
0.90 
0.90 
0.90 
0.90 
0.86 
0.83 
0.79 
0.79 
0.76 



consistency index based on the character- 
state differences between OTUs of the same 
species (1.0 = no differences). This table 
helps clarify the problems of why T. montana 
and T. bollingi do not cleanly link with conspe- 
cific OTUs (i.e. three classes of phenotype 
varying over six character-states). 



Principal Component Analysis (PCA) was 
done. There are ten components that account 
for 84.6% of the vaiance (Table 78). The first 
three components account for only 45% of the 
variance. Factor loading of characters for the 
first ten PCs are given in Appendix II. Ordina- 
tion diagrams are not provided because: (1) 



318 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 78. Principal component analysis of shell data: First ten principal components are listed with the 
percentage of variance loading on each component. 



Components 



Eigenvalue 



Percent 



Cumulative 



1 
2 
3 
4 
5 
6 
7 
8 
9 
10 



6.16870 
3.56713 
3.27486 
2.59528 
2.45005 
1 .76301 
1.38014 
1.30153 
1.10735 
0.93280 



21.27 
12.30 
11.29 
8.45 
8.45 
6.08 
4.76 
4.49 
3.82 
3.22 



21.27 
33.57 
44.86 
53.81 
62.26 
68.34 
73.10 
77.59 
81.41 
84.62 



most taxa cluster too close together at the 
center of the plots; (2) the MST results (Fig. 
155) are much less satisfactory than those 
given in Figure 154; there are 14 mismatches 
of phenotypes (i.e. 32%). Three dimensional 
scaling produced even less acceptable re- 
sults. 

PCA analysis is useful in demonstrating the 
utility of shell characters used for discriminat- 
ing among species. The first component 
(Table 79) is not one of size but involves ten 
apertural characters that are most useful in 
distinguishing among species. As examples: 
(1 ) only four species (N. dianmenensis, N. hin, 
T. gredleri, T. odonta) have a pronounced 
adapical apertural sinus; (2) only three spe- 
cies have an adapical apertural beak tubercle 
(T. dianmenensis, T. gredleri, T. odonta); (3) 
only T. odonta has a mid-lip inner lip notch 
and a shell of 7.0 whorls. 

The second component is defined on the 
basis of seven components that include 
sculpture, size, vahx formation, and shell 
base characters. The third component in- 
volves three (shape, columellar and apertural 
groove) characters, while the fourth is defined 
by four characters, with teeth and internal 
keels as well as sculpture and outer lip orien- 
tation. 

The Neotricula duplicata cluster of five taxa 
(Л/, duplicata, N. lili i, N. odonta, N. dianmen- 
ensis, N. gredleri), best seen in Figures 153 
and 1 54, have four to seven of the following 
seven character states: an outer lip sinus, 
adapertural beak or notch, abapical spout, a 
beak tubercle, a thickened inner lip, the inner 
lip completely separated from the body whorl, 
and an angled or sinuate inner lip. 

All species of Neotricula thus far examined, 
except the alpha race of N. aperta, have spiral 
microsculpture on the shoulders of the teleo- 
conch whorls; not so for species of Tricula. 



Neotricula aperta is unique for its globose- 
conic shape, wide columellar shelf, and keel 
on the base of the body whorl bordering the 
wide columellar shelf. Only T. maxidens has a 
cylindrical-conic shell and a large columellar 
tooth clearly visible in the aperture. Only J. 
jinghongensis has spiral microsculpture at the 
base of the body whorl and a basal post. Only 
T. xiaolongmenensis has a domed tooth on 
the columella inside the body whorl. Only W. 
niuzhuangensis has a spiral keel on the col- 
umella inside the body whorl that does not 
extend basally so as to be seen as a tooth in 
the aperture (in contrast to T. maxidens). Only 
three species have crenulated whorls at the 
suture: N. burchi, G. chinensis, J. jinghongen- 
sis. 

Anatomy 

Thirty OTUs were scored for 48 character 
states (Tables 80, 81). A UPGMA phenogram 
based on distance coefficient is given (Fig. 
156). Distances were used as the cophenetic 
correlation because r = 0.960 is superior to 
that using similarity coefficients (r = 0.865). 
In this phenetic treatment, species do not 
group neatly into well-defined generic clus- 
ters. However, by eliminating characters with 
missing data and using the Prim Network 
(MST) (Fig. 157), well-defined generic group- 
ings resulted. 

In the PCA analysis, ten components were 
extracted before eigenvalues dropped below 
1.0; these components accounted for 89.8% 
of the variance (Table 82). Factor loadings of 
characters on each of the ten components are 
given in Appendix III. Ordination on the first 
two PC axes is given (Fig. 158) with OTUs 
connected by the MST based on the original 
distance matrix (with characters involving 
missing data removed) (Fig. 157). The spe- 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 319 

TABLE 79. Shell characters highly correlated on each of nine principal components. * = also loads > 
0.440 on another axis. 



Characters 



Loading 



COMPONENTS 
I. APERTURAL/WHORLS 



11 


-0.849 


24 


-0.733 


13 


-0.715 


18 


-0.696 


17 


-0.665 


29 


-0.665 


12 


-0.653 


20 


-0.636 


19 


-0.632 


9 


-0.606 


3 


-0.582 


6 


+ 0.812 


5 


+ 0.653 


28 


+ 0.596 


12* 


+ 0.497 


1 


-0.461 


8 


-0.461 


23 


+ 0.458 


26 


+ 0.810 


27 


+ 0.751 


10 


+ 0.728 


2 


+ 0.645 


16 


-0.715 


7 


-0.611 


22 


-0.585 


15 


+ 0.443 


3* 


+ 0.574 


5* 


-0.499 


19* 


-0.459 


9* 


+ 0.450 


23* 


-0.553 


21 


-0.496 


20* 


-0.444 


25 


-0.696 


1* 


-0.570 


14 


+ 0.734 


8* 


-0.494 



sinus — adaptical aperture 

beak tubercle — adaptical aperture 

inner lip shape 

lip deflection angle 

notch, inner lip 

shell has 7.0 whorls 

abapical spout 

inner lip separated from body whorl 

inner lip thickness 

adapical notch, beak 

apertural shape 

II. SIZE, SCULTPURE, VARIX, SHELL BASE 
spiral microsculpture 

whorl at suture crenulated 

basal post 

abapical spout 

size 

protoconch sculpture 

varix 

III. SHAPE, KEEL, COLUMELLAR SHELF: N. APERTA AXIS 
keel at external base of shell 

columellar shelf 

inner adapical apertural groove 

shape 

IV. STRUCTURE ON COLUMBELLA INSIDE OR OUTSIDE SHELL; 
OUTER LIP 

inner lip tooth 

spiral sculpture at shell base 
internal columellar keel 
outer lip scooped forward 

V. APERTURE, WHORL CRENULATION 
apertural shape 

crenulated suture 

inner lip thickened 

adaptical apertural notch, beak 

VI. VARIX, LIP SEPARATION 
varix 

adaptical aperture separated from body whorl 
inner lip separated from body whorl 

VII. LIP ANGLE, SIZE 
lip angle 

aize 

VIII. LIP SINUATION 
lip sinuation 

IX. PROTOCONCH SCULPTURE 
protoconch sculpture 



cies fall cleanly into generic clusters on the 
ordination diagram. Through use of Figures 
157 and 158, it is clear which species pairs 
are most similar anatomically; e.g., Tricula 
montana is most similar to T. gregoriana. 

The ordination diagram and an assessment 
of anatomical characters with high loadings 
on each of the ten principal components are 



useful for assessing OTU relationships. The 
first and second components clearly separate 
genera (Table 83). Characters 17, 18, 24, 21 
clearly serve to separate generic clusters. 
Species with the 360° open oviduct circle 
complex of organs with the spermathecal duct 
entering the pericardium are to the left upper 
and lower quadrants. Species that have lost 



320 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



APE 


N. 


2.25 


BUR-l 


N 


0.48 


BUR-2 


1.07 


MIN-1 


N 


0.38 


MIN-2 


0.69 


BAM-1 
BAM-2 


T. 


0.53 
0.76 


HUD-1 
HUD-2 


T. 


0.36 
0.92 


BOL-1 
BOL-3 


T. 


0.47 
0.57 


LUD-1 
LUD-2 


T. 


0.36 
0.75 


XIN-1 
XIN-3 


T. 


0.54 
0.77 


BOL- 2 


T. 


0.62 


XIO-1 


T. 


0.81 


GRG-l 
GRG-2 


T. 


034 

61 


MON-3 


T. 


0.77 


MON-1 
MON-2 


T. 


0.57 
0.84 


XIO-2 


T. 


1.13 


NIZ-1 
NIZ-2 


w 


0.43 
1.20 


XIN-2 


T. 


1.21 


CRI-1 
CRI-2 


N. 


0.41 

1.13 


CHI-1 
CHI-2 


G. 


0.54 
1.36 


DIA-1 
DIA- 2 


N. 


0.45 
1.14 


LIL- 1 


N. 


0.34 


LIL- 2 


1.25 


DUP-l 
DUP-2 


N. 


0.91 
1.10 


GRD-1 


T. 


0.30 


GRD-2 


1.68 


MAX-l 


T. 


1.03 


MAX-2 


1.70 


JIN-1 


J. 


1.19 


JIN-2 


1.99 


ODN-1 
ODN-2 


T 


0.30 



FIG. 153. Phenogram based on UPGMA treatment of distance coefficients involving shell characters from 
phenotypes (1-3) of 21 species (Tables 76, 77). G, Gammatricula; J, Jinhongia; N, Neotricula; T, Tricula; W, 
Wuconchona. APE, N. apena; BUR, N. burchi; BAM, 7. bambooensis; BOL, 7. bollingi; CRI, N. cristella; CHI, 
G. chinensis; DIA, N. dianmenensis; DUP, N. duplicata; GRD, T. gredleri; GRG, 7. gregoriana; HUD, 7. 
hudiequanensis; JIN, J.jinhongensis; LIL, N. lilii; LUD, 7. ludongbini; MAX, 7. maxidens; MIN, N. minutoides; 
MON, 7. montana; NIZ, W. niuzhuangensis; ODN, 7. odonta; XIO, 7. xiaolongmenensis; XIN, 7. xianfen- 
gensis. 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 321 




FIG. 154. Minimum spanning tree (MST) based on distance coefficients involving shell characters treated in 
Tables 2 and 76. The tree is drawn to scale using inter-taxon distances. See caption for Figure 153 for 
defining abbreviations. 



the seminal recepatacle and have the sper- 
mathecal duct opening into the mantle cavity 
are in the upper right quadrant. 

While major groupings along the 1st com- 
ponent are established by the four highly cor- 
related characters given above, species 
placements are dictated by the 14 other char- 
acters with loadings < 0.678. Tricula gredleri 
is far to the left because of character 37; it 
alone, of all the OTUs, has a massive ejacu- 
latory duct. Wuconchona niuzhuangensis is 
far to the right because it shares with Gam- 
matricula chinensis character states for char- 
acters 21 and 35; the function of the seminal 
receptacle is moved to the inside of the ovi- 



duct, and the concave edge of the penis has a 
white muscular zone. Wuconchona is dis- 
placed to the right of Gammatricula due to 
characters 17, 28, and 29. Wuconchona is 
unique in having a discrete U-shaped bend of 
the oviduct, a very short male gonad, and no 
vas efferens. 

Distribution along the second principal 
component is due to the interaction of 12 
characters. Tricula gredleri, Neotricula di- 
anmenensis, and N. minutoides are at the 
bottom of the ordination diagram because of 
characters 2, 19, 20, 34, and 40. Unique to T. 
gredleri are operculum shape, fusion of the 
oviduct to the pericardium, the lemon color of 



322 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 




FIG. 155. MST based on shell distance coefficients following PCA analysis. Abbreviations are defined in 
caption for Figure 1 53. See text for details. 



the anterior chamber of the stomach, and the 
position of the penial opening. Neotricula di- 
anmenensis and N. minutoides have few 
rows of teeth per radula. Tricula minutoids 
has a highly extensible penis. Jinhongia, Wu- 
conchona and Gammatricula are at the top 
because of the loss of the seminal receptacle 
(and duct). 

Comparing Figures 156-158, it is seen 
which species are most similar anatomically 
and the affect of intrapopulation variation on 
distance measures. Linkages at a value < 
0.40 (distance coefficients) clearly involve 
population variation. Linkages > 0.67 involve 
discrete species. 

There are species clusters of phenetically 
closely related species. The biggest cluster, 
one that has the least distances among spe- 
cies, involves the Tricula bollingi complex: T. 



bollingi, T. bambooensis, T. hudiequanensis 
and T. ludongbini. These species occur in 
Yunnan, China. Within this complex, T. hud- 
iequanensis is unique in that it has a scatter of 
a few white glands about the eyes (char. 1). 
The attachment pad of the operculum is very 
wide (char. 4). The shell is most similar to that 
of T. bambooensis (Figs. 153-155). However, 
there is a 12.5% difference between T. hud- 
iequanensis and T. bambooensis (chars. 1 , 4, 
6, 8, 14, 37). Within the T. bollingi complex, T. 
bollingi is unique in that the female gonad is 
posterior to the stomach (char. 1 1 ), the radula 
is of medium length, not short (char. 41), and 
the radula has a moderate number of rows of 
teeth, not few (char. 42). The shell of T. bollingi 
is most similar to that of T. ludongbini. 

A closely allied species pair involves T. 
montana from India and T. gregoriana from 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 323 



TABLE 80. Anatomical characters and character-states (n = 48) showing one or more differences 
among 20 species belonging to the genera Tricula or Neothcula, or where the shells indicate affinity to 
these two genera. 



1 . Glands about the eyes 

2. Operculum shape: 

3. Operculum: 

4. Operculum attachment pad: 

5. Operculum; thick internal ridge: 

6. Gill filament number: 

7. Gill filament shape: 

8. Gf 2 length: 

9. Osphradium length: 

10. Circular grandular mass anterior to 
osphradium: 

11. Gonad: 

12. Gonad: 

13. Bursa: 

14. Bursa: 



15. Bursa: 

16. Bursa; duct: 

1 7. Oviduct at bursa: 

18. Spermathecal duct: 

19. Spermathecal duct: 

20. Spermathecal duct: 



21 . Seminal receptacle: 



22. Seminal receptacle duct: 

23. Seminal receptacle duct: 

24. Seminal receptacle arises from: 

25. Pericardial bursa: 

26. Albumen bland: 

27. Gonad: 

28. Gonad: 

29. Vas efferens: 

30. Seminal vesicle: 

31 . Vas deferens — 2: 

32. Penial tip: 

33. Penis: 

34. Penial opening: 



External Features (N = 5, 10%) 
no glands (0), scatter of a few glands (1), dense 
concentration of glands (2) 

ovate (0), mid-columellar indentation (1), elongate-oval (2), 
irregular shape (3), cap-like (4) 
single layer (0), two or more layers (1) 
wide (0), narrow (1), very wide (2) 
none (0), has (1) 

Mantle Cavity (N = 5, 10%) 
few (0), moderate (1), many (2) 
flat (0), moderate dome (1), high domed (2) 
long (0), medium (1), short (2) 
long (0), short (1) 
none (0), has (1) 

Female Reproductive System (N = 16, 33%) 
behind stomach (0), overlaps stomach (1) 
long (0), short (1), very short (2) 
short (0), long (1), minute (2) 
posterior to palliai oviduct [Ppo] (0), covered wholly or 
considerably by Ppo (1), ventral to Ppo (2), postero-lateral 
to Ppo (3) 
round (0), triangular to subtriangular (1), ovoid (2) 

> 0.04 mm long (0), none (1) 

makes 360° tight twist (0), runs to Ppo without twist or bend 

(1), no twist but has discrete bend (2) 

enters pricardium (0), enters mantle cavity (1) 

long (0), short (1), none, fused to pericardium (2) 

slants at an angle from duct of bursa or oviduct to mantle 

cavity, or at an angle from anterior-posterior axis (0), runs 

directly anterior from duct of bursa to mantle cavity (1), 

none, oviduct fused to pericardium (2) 

arises from oviduct or duct of bursa (0), lost, function 

removed to inside swelling between duct of bursa and 

spermathecal duct (1), lost, function removed to inside 

oviduct (2) 

U-shaped continuation of duct of bursa (0), arises from 

oviduct (1), from juncture of duct of bursa and oviduct (2), 

from duct or bursa (3), lost (4) 

> 0.02 mm long (0), very short to ductless (1), seminal 
receptacle and duct lost (2) 

inside edg of oviduct in coil (0), outside edge of oviduct coil 
(1), from duct of bursa (2), seminal receptacle lost (3) 
none (0), slight (1), large (2), extremely enlarged (3) 
normal (0), short (1) 

Male Reproductive System (N = 11, 23%) 
posterior to stomach (0), overlaps stomach (1) 
long (0), short (1) 
has (0), does not have (1) 

coils posterior to stomach (0), coils also on stomach (1), 
coils only on stomach (2) 

leaves prostate at Emc (0), leaves prostate at mid-prostate 
(1) 

no papilla (0), has papilla (1), long penial filament (2) 
normal (0), highly extensible (1) 

center of penial tip (0), from concave edge of blunt penial 
tip (1) 



(continued) 



324 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 80. (Continued). 



35. Penis: 

36. Ejaculatory duct: 

37. Ejaculatory duct: 

38. Digestive bland: 

39. Anterior chamber of stomach 

[Ast]: 

40. Ast lemon yellow color: 

41 . Radula: 

42. Radula: 

43. Radula central tooth: 

44. Radula central tooth: 

45. Lateral tooth major cusp: 

46. Lateral tooth major cusp: 

47. RPG ratio: 

48. Length pleuro-subesophageal 
connective: 



External Features (N = 5, 10%) 
no white muscular zone on concave edge (0), has such 
zone (1) 

none (0), in base of penis (1), extends from penial base into 
the neck (2), only in neck (3) 
none (0), slight (1), large/prominent (2), massive (3) 

Digestive System (N = 9, 19%) 
posterior to stomach (0), covers Pst (1) 
no streak (0), with grey-melanin streak (1) 

no (0), yes (1) 

short (0), medium length (1), long (2), very long (3) 

no. rows teeth: few (0), moderate no. (1), many (2) 

has paired swellings at posterior face of tooth between 

innermost basal cusps (1), lacks (0) 

central cusp: generalized type (0), long and dagger-like (1) 

1 (0), 1 [2] = (1) 

normal (0), displaced towards outer edge (1) 

moderately concentrated (0), elongated (1), concentrated 

(2) 

usual (0), none (1), long (2) 



Yunnan, China. They are similar anatomically 
(Fig. 157) and conchologically (Figs. 153, 
154). They differ in seven anatomical charac- 
ters (15%). 



PHYLOGENETIC RELATIONSHIPS 

Phylogenies involving 19 genera of the 
three tribes of Triculinae were recently pub- 
lished (Davis et al. 1990; Davis & Kang, 1990; 
Davis, 1 992). To these are now added the two 
additional genera (Guoia, Lithoglyphopsis), 
additions made possible because of this 
study. In this analysis, only one genus of the 
Jullieniini is used (outgroup), and the 13 gen- 
era grouped in the Triculini and Pachydrobiini. 
The characters and character-states used are 
listed in Table 84. There are 17 characters 
listed under synapomorphies, an additional 
14 as autapomorphies. Scores are tabulated 
in Table 85. The result of the Hennig-86 anal- 
ysis was 20 equally parsimonious trees with a 
length of 37, a consistency index of 86 and an 
ri (retention index; Farris, 1989) index of 91. 
Four of the trees and the Nelsen consensus 
tree are given (Figs. 159-163). All show the 
separation of the three tribes with the same 
genera assigned to each of the tribes. The 
only difference among the 20 trees for genera 



of the Triculini was the tricotomy of Fenouilia, 
Lacunopsis, Lithoglyphopsis or, alternatively, 
the divergence of Fenouilia from Lacunopsis 
and Lithoglyphopsis. The separation {(Fen- 
ouilia) (Lacunopsis, Lithoglyphopsis)) is war- 
ranted as Fenouilia has a trochoid shell, and 
the other two genera have globose shells. 

The greatest shifting about involves genera 
of the Pachydrobiini. The consensus tree 
reduces the shifting to produce a polycotomy 
for Neotricula, Halewisia, Pachydrobia, and 
Jinhongia. In all trees, Gammatricula and Wu- 
conchona diverge from the same node sepa- 
rated from the node that supports the sub- 
cluster of ((Robertsiella) (Guoia-a, Guoia-b)). 

A single tree resolves with the ordering of 
character-states involving (1) the position of 
the seminal receptacle and its duct, (2) the 
loss of the seminal receptacle and the direc- 
tion of evolution of the location of the struc- 
tures functionally replacing the seminal re- 
ceptacle, and (3) an analysis of homoplasies 
(Fig. 164). The generalized position of the 
seminal receptacle is a branch off the oviduct 
posterior to the duct of the bursa and anterior 
to a coiling of the oviduct between the gono- 
pericardial duct and the bursa copulatrix (out- 
groups Hydrobia, Hydrobiidae; Pomatiopsis, 
Pomatiopsinae: Pomatiopsini). This position 
is seen in the genera of the tribe Triculini. In 
the Triculini and Pachydrobiini, the position of 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 325 



Q. 



CRI 


N 


0.69 


LIL 


N 


0.91 


DUP-1 


N 


0.31 


DUP-2 


1.20 


BUR 


N 


0.76 


XIO 


T 


0.97 


GRG 


T 


1. 03 


BAM-1 


T 


0.33 


BAM-2 


0.66 


BOL 


T 


0.82 


HUD-1 


T 


0.33 


HUD-2 


0.78 


LUD-1 




0.38 


LUD-2 


T 


0.19 


LUD-3 




0.94 


XIN 


T 


1.13 


ODN-1 


T 


0.31 


ODN-2 


1.28 


JIN-1 


J 


0.29 


JIN-2 




0.99 


CHI-1 


G 


0.24 


CHI-2 




1.42 


MON 


T 


1.44 


MAX 


T 


1.53 


DIA 


N 


1.47 


MIN-1 


N 


0.20 


MIN-2 




1.69 


GRD-1 


T 


0.26 


GRD-2 




2.33 



NIZ W 



2.400 



FIG. 156. Phenogram based an UPGMA treatment of distances based on all 48 anatomical characters from 
1 to 2 phenotypes of 20 species. See Tables 80, 81 . Abbreviations are defined in the caption for Figure 153. 



326 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 81 . Scoring of 30 OTUs for 48 character-states listed in Table 80: bur-1 , bur-2, etc. indicates that 
shells of Neotricula burchi exhibit more than one character-state for one or more characters, bur, burchi; 

cri, cristella; dia, dianmenesis; dup, duplicata; lit, //'///'; min, minutoides; bam, bambooensis; bol, bollingi; 
grd, gredleri; grg, gregoriana; hud, hudiequanensis; lud, ludongbini; max, maxidens; mon, montana; odo, 
odonta; xian, xianfengensis; xio, xiaolongmenensis; chi, chinensis; jin, jinhongensis; niz, niuzhuangensis. 

NC = missing data. 

12 3 4 5 6 7 8 9 10 11 12 13 14 15 

N.cri N.dia N.dupl N.dup2 N.lil N.minl N.min2 N.bur T.mon T.baml Tbam2 Tboll Tgredl Tgred2 Tgreg 

1 NC 

2 2 2 

3 11 1 

4 2 

5 

6 1 1 

7 1 NC 1 

8 1 1 

9 10 1 

10 

11 

12 1 1 1 

13 

14 О 

15 2 

16 1 О 

17 1 1 1 

18 1 1 1 

19 1 

20 1 1 

21 О О О 

22 О О О 

23 О О О 

24 2 2 2 

25 О О О 

26 О О О 

27 1 

28 О О О 

29 О О О 

30 О О О 

31 О О О 

32 2 1 1 

33 О О О 

34 О О О 

35 О О О 

36 О О О 

37 О О О 

38 1 О О 

39 О О О 

40 О О О 

41 1 1 1 

42 3 2 

43 О О О 

44 О О О 

45 1 1 1 

46 О О О 

47 О О О 

48 О О О 



1 








2 








1 





1 


2 

















1 


1 


1 


2 


1 


NC 


1 








1 


1 


1 




















1 


1 


1 




















2 

















1 


1 


1 


1 


1 


1 


1 





1 


1 





1 


























1 


2 


2 


2 

















1 


1 


1 



































1 





1 


1 











1 












































1 








1 











1 


1 


1 


2 


3 























1 


1 


1 






























12 1 2 

000 004 40 

100 000 00 

О NC О 000 00 

000 200 00 

111 2 2 11 12 

NC NC 1 2 12 1 1 NC 

О NC О 1 110 

111 1 11 11 

1 О 10 

0000 100 00 

111 1 2 1 11 
000 2 222 20 
030 2 222 20 
0222 020 12 
000 000 00 
110 
110 
100 002 20 
100 002 20 
0000 100 00 
3 1 11 13 
100 000 00 
222 000 02 
002 003 32 
100 000 00 
1 1 111 11 
000 000 00 
000 000 00 
1 1 111 11 
111 110 1 
10 1 11 11 
000 000 00 
1 10 
000 000 00 
12 1 112 2 3 

112 1 113 3 2 
10 10 
000 000 00 
10 1 10 
12 2 2 11 11 

12 2 2 11 2 2 

001 000 00 
100 000 00 
1 1 10 
000 000 00 
О NC О 000 00 
О NC NC 002 2 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 327 



TABLE 81 . (Continued) 





16 


17 


18 


19 


20 


21 


22 


23 


24 


25 


26 


27 


28 


29 


30 




Thudl 


Thud2 


Tludl 


Tlud2 


Tlud3 


Tmax 


Todol 


Todo20 


Txian 


Txiao 


Jjini 


Jj¡n2 


Wniz 


Gch¡1 


Gchi2 


1 


1 


1 

















3 


1 


2 


1 


1 


2 


1 


1 


2 

















1 


3 


1 














2 








3 








1 


1 


1 





1 





1 





1 


1 





1 


1 


4 


2 


2 











1 





1 


2 


1 








2 








5 




















1 


2 














1 








6 


1 


1 


2 


2 


2 





2 


2 


2 


1 


1 


1 





1 


1 


7 


2 


2 


2 


2 


2 


NC 


2 





NC 


NC 


NC 


NC 


NC 





1 


8 























1 


1 





1 


1 











9 


1 


1 


1 


1 


1 





1 





1 


1 


1 


1 


1 


1 


1 


10 






































1 








11 


1 


1 


1 


1 


1 








1 


1 


NC 

















12 


1 


1 











1 


1 


2 





1 





1 


2 








13 


2 


2 


2 


2 


2 





2 





2 











1 


1 


1 


14 

















1 





1 








2 


2 











15 


2 


2 


1 


1 


1 





1 





2 


2 


2 


2 


2 


2 


2 


16 















































17 
































1 


1 


2 


1 


1 


18 
































1 


1 


1 


1 


1 


19 






































1 








20 















































21 























1 








1 


1 


2 


2 


2 


22 


1 


2 


1 


3 


2 


1 


1 


1 


1 


1 


4 


4 


4 


4 


4 


23 




















1 











2 


2 


2 


2 


2 


24 











2 


2 














1 


3 


3 


3 


3 


3 


25 

















1 





























26 


























1 











1 








27 


1 


1 


1 


1 


1 


1 





1 


1 


NC 


1 


1 











28 






































1 








29 






































1 








30 


1 


1 


1 


1 


1 





2 


2 


1 











1 








31 


1 


1 


1 


1 


1 


1 








1 


1 


1 


1 


1 


1 


1 


32 


1 


1 


























1 


1 


2 


1 


1 


33 















































34 















































35 






































1 


1 


1 


36 


1 


3 


1 


2 


3 


1 


1 


1 





1 


1 


1 





1 


1 


37 


2 


2 


2 


2 


2 


2 


2 


2 





1 


2 


2 





1 


1 


38 

















1 














1 


1 


1 








39 

















1 





























40 















































41 


3 


3 


2 


2 


2 





1 


1 


2 


1 


1 


1 


1 


1 


1 


42 


2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


43 















































44 

















1 














1 


1 











45 




















1 


1 























46 






































1 








47 

















NC 








1 


1 








2 








48 

















NC 











2 








1 









the opening into the duct of the seminal re- 
ceptacle shifts as follows: along the oviduct to 
the base of the duct of the bursa (one species 
of Tricula); -> to stem off the duct of the bursa 
(one species of Tricula, nearly all Neotricula, 
Halewisia); -* to stem off the sperm duct 
(Pachydrobia). The presumption is that no 



large scale rearangements of ducts were 
made in a single step. To obtain the uncoiled 
oviduct seen in Neotricula (contrast Triculini), 
first the duct of the seminal receptacle, then 
the spermathecal duct would have had to mi- 
grate along the oviduct to move onto the duct 
of the bursa. With this accomplished, the 



328 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



NEOTRICULA 




FIG. 157. MST based on distance coefficients in- 
volving all anatomical characters treated in Tables 
80 and 81 . The tree is drawn to scale using inter- 
taxon distances. See caption for Figure 1 53 for de- 
fining abbreviations. Generic groupings have been 
circled. 



TRICULA I 






"NIZ 




2 




/ LUD.3 \ ' 
/HUD 2 04.1UD.2 \| 


Jyjj^ 






HUD jj>) mon\ s 








/> R ^^W 








BAM.1 ^~M* \Ч 








ВАМ 2 1воД %. 
LUD1 I V IN 


.BUR \ 






ODN_2\ \ I 

TÖdni \ 


\k' l \ 












/ «k*V 


\ \l VCRI 
TouPl.2 \ 

\V DIA \ 


1,2/ 




ÍGRD2 / , 




NEOTRICULA 




\ 4GRD.1 / 









FIG. 158. Ordination diagram following PCA and 
MST seen in Figure 157. All characters scored in 
Table 81 as NC were removed in this analysis to 
permit the PCA treatment. The ordination is on the 
first two PCA axes. Generic groupings have been 
circled. See text for details. See caption for Figure 
1 53 for defining abbreviations. 



TABLE 82. Principal Component Analysis of anatomical data (matrix of 41 x 30). The first ten principal 
components are listed with the percentage of variance loading on each component. 



Component 



Eigenvalue 



Percent 



Cumulative 



1 
2 
3 
4 
5 
6 
7 
8 
9 
10 



8.87575 
6.86082 
5.58656 
3.54706 
2.70292 
2.60035 
2.12427 
1.83148 
1.61804 
1 .05972 



21.65 
16.73 
13.63 
8.65 
6.59 
6.34 
5.18 
4.47 
3.95 
2.58 



21.65 
38.38 
52.01 
60.66 
67.25 
73.59 
78.77 
83.24 
87.19 
89.77 



proximal section of the duct of the bursa (as 
seen in Tricula) would become the sperm 
duct, linking the new bursa complex to the 
oviduct to facilitate fertilization. In no case is a 
species of Pachydrobiini seen with the semi- 



nal receptacle arising from the oviduct! The 
most probable sequence is the move of the 
seminal receptacle from the oviduct to the 
duct of the bursa (situation seen in Tricula 
species b). Then the spermathecal duct mi- 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 329 



TABLE 83. Characters with high loadings, that define each of ten components. "High loading on more 
than one axis (above). 



Characters 


Loadings 


COMPONENT 






I. GENERIC ORGANIZERS*, N = 18 


17* 


+ 0.934 


oviduct 360° circle 


18* 


+ 0.838 


spermathecal duct enters pericardium 


24* 


+ 0.734 


origin of seminal receptacle 


27 


-0.678 


male gonad overlaps stomach 


23 


+ 0.658 


seminal receptacle duct length 


21* 


+ 0.649 


seminal receptacle lost 


37 


-0.644 


ejaculatory duct prominence 


36 


-0.623 


position of ejaculatory duct 


6 


-0.616 


gill filament number 


28 


+ 0.610 


male gonad length 


29 


+ 0.610 


presence of vas efferens 


46 


+ 0.610 


position of lateral tooth major cusp 


35 


+ 0.599 


white muscular zone on penis 


13 


-0.580 


bursa size 


38 


+ 0.579 


digestive gland covers Pst 


30 


-0.546 


poisition of seminal vesicle 


41 


-0.460 


radula length 


26 


+ 0.519 


albumen gland length 

II. SPERMATHECAL DUCT, RADULA, PENIS, STOMACH, 
N = 12 


20 


-0.871 


orientation spermathecal duct 


31 


+ 0.847 


where vas efferens leaves prostate 


45 


-0.784 


major cusp of lateral tooth 


19 


-0.723 


length spermathecal duct 


15 


+ 0.696 


bursa shape 


22 


+ 0.665 


configuration duct of seminal receptacle 


2 


-0.601 


operculum shape 


34 


-0.575 


position of penial opening 


40 


-0.575 


color of Ast 


42 


+ 0.522 


number of rows of teeth 


39 


-0.449 


melanin streak on Ast 


33 


-0.471 


penis highly extensible 

III. MALE REPRODUCTIVE, RADULA, OPERCULUM, 
STOMACH, N = 14 


46** 


+ 0.632 


poisition major cusp lateral tooth 


28** 


+ 0.632 


male gonad length 


29** 


+ 0.632 


has/has not: vas efferens 


2** 


+ 0.596 


operculum shape 


40** 


+ 0.589 


Ast color 


34** 


+ 0.589 


position penial opening 


10 


+ 0.573 


gland mass anterior to osphradium 


25 


+ 0.570 


pericardial bursa 


19** 


+ 0.514 


length spermathecal duct 


30** 


+ 0.485 


position serminal receptacle 


5 


+ 0.469 


ridge on operculum 


45** 


+ 0.447 


lateral tooth major cusp 


3 


-0.44G 


opercular layers 


32 


+ 0.446 


penial tip 

IV. RADULA, OPERCULUM, REPRODUCTIVE, N = 5 


44 


-0.631 


central cusp, central tooth 


5** 


+ 0.545 


opercular ridge 


30** 


+ 0.529 


position, seminal vesicle 


14 


-0.502 


bursa position 


26** 


+ 0.470 


albumen gland length 

V. OPERCULUM, FEMALE REPRODUCTIVE, MANTLE 
CAVITY, N = 4 


3** 


+ 0.646 


opercular layers 


12 


-0.484 


female gonad length 

(continued) 



330 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



TABLE 83. (Continued). 



Characters 


Loadings 


COMPONENT 


23" 


+ 0.424 


length duct seminal receptacle 


10* 


-0.421 


gland mass anterior to osphradium 

VI. STOMACH, PENIS, FEMALE REPRODUCTIVE, N = 6 


39" 


+ 0.583 


melanin streak on Ast 


33" 


+ 0.530 


highly extensible penis 


26" 


+ 0.482 


albumen bland length 


38" 


+ 0.475 


digestive bland covers Pst 


32" 


-0.455 


penis tip 


16 


-0.435 


duct of bursa 

VII. CENTRAL TOOTH, OSPHRADIUM, OPERCULUM, 

N = 4 
osphradium length 


9 


+ 0.617 


43 


+ 0.465 


swelling on face of central tooth 


5" 


-0.443 


opercular ridge 


44* 


-0.417 


central cusp of central tooth 
VIII. CENTRAL TOOTH, N = 1 


43** 


-0.610 


swellings on face of central tooth 
IX. BURSA, TEETH, N = 2 


16" 


-0.611 


duct of bursa 


42« 


+ 0.523 


rows of teeth: number 
X. BURSA, N = 1 


14" 


+ 0.538 


position of bursa 



grates along the oviduct to the duct of the 
bursa, with subsequent shift of the sperma- 
thecal duct from opening into the pericardium 
to opening into the posterior end of the mantle 
cavity next to the pericardium, and the uncoil- 
ing of the oviduct (as seen in Neotricula spe- 
cies of type a, and b). Subsequently, either 
the duct of the seminal receptacle migrates 
onto the sperm duct, or the point where the 
spermathecal duct joins the duct of the bursa 
moves distally towards the bursa leaving an- 
terior to it that proximal piece of duct that once 
was the duct of the bursa, with the seminal 
receptacle attached to it (as seen in Pachy- 
drobia). In this scenario, both Halewisia and 
Pachydrobia evolved from progenators re- 
sembling Neotricula species of type a (e.g. N. 
aperta, N. cristella). In Halewisia the sper- 
mathecal duct migrated to open into the bursa 
copulatrix. The shell is a derived type (Davis, 
1979: 111). Pachydrobia has evolved a large 
first-order adaptive radiation (Davis, 1992) of 
greater than 14 species in which the shell has 
modified from the generalized Tricula type 
variously: larger, thick shells with thick lips; 
tendency to have pronounced macrosculp- 
ture (nodes, spines); tendency to shell as- 
symetry. 

There is parallel evolution in the Triculini 
and Pachydrobiini involving loss of the semi- 
nal receptacle and duct with new structures 



arising in different locations to accomodate 
the functions of the seminal receptacle. The 
solution to this loss is quite different in the two 
tribes, hence, no difficulty arises regarding 
correct tribal classification or assessment of 
intergeneric relationships. The tendency to- 
wards loss is minor in the Triculini, involving 
only one genus, Lacunopsis; it is major devel- 
opment in the Pachydrobiini involving the ma- 
jority of genera (five of eight). In Lacunopsis, 
two or more accessary seminal receptacles 
arise to the exterior of the juncture of the ovi- 
duct and spermathecal duct. In the Pachydro- 
biini, solutions to the loss of the seminal re- 
ceptacle are all within the relevant ducts (duct 
of the bursa, sperm duct, oviduct). The most 
parsimonious direction of evolution is as fol- 
lows: the duct of the seminal receptacle is 
lost, the seminal receptacle becomes fused to 
the duct of the bursa close to or at the bursa 
(as in N. dianmenensis); — ► The seminal re- 
ceptacle is replaced by an outpocketing within 
the encapsulated duct of the bursa (as in 
Robertsiella); -> the sperm storage area 
shifts anteriorly to a swelling at the juncture of 
the duct of the bursa, spermathecal duct and 
sperm duct (as in Jinhongia); — * the next shift 
is into the sperm duct (Guo/a-a), and finally 
the oviduct (Guoia-b) -* and farther posterior 
within the oviduct (Wuconchona and Gamma- 
tricula). 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 331 
TABLE 84. Characters and character-states used for the cladistic analysis. 

SYNAPOMORPHIES 

1. Spermathecal duct opens to pericardium: (a) (0); (b) to posterior mantle cavity (1). 

2. Oviduct makes: (a) a closed, tight 360° twist posterior to duct of the bursa (0); (b) an open 360° 
circle (1); (c) does not have the 360° twist or circle (2). 

3. Sperm duct: (a) absent (0); (b) present and short (1); (c) present and elongated (2). 

4. Spermathecal duct: (a) has primitive position between pericardium and oviduct as seen in 
Pomatiopsis (Pomatiopsinae) (0); (b) joins oviduct before oviduct twist (1); (c) joins oviduct 180° or 
more into oviduct circle (2); (d) joins duct of bursa, or common sperm duct (3); (e) joins bursa (4). 

5. Seminal receptacle: (a) arises from primitive condition from oviduct posterior to the duct of the bursa 
(as in Pomatiopsis) (0); (b) from left side of the oviduct and pressed to the contour of the oviduct 
circle complex (1); (c) joins duct of bursa or common sperm duct (2); (d) joins the sperm duct (3); is 
lost; function replaced by: (e) an outpocketing of the encapsulated duct of the bursa (4); (f) an 
internal duct swelling at the juncture of bursa and spermathecal duct (5); (g) an outpocketing of 
sperm duct or swelling of sperm duct at juncture of sperm duct and oviduct (6); (h) within the oviduct 
(7); (i) two or more seminal receptacles arise from the oviduct (outside the duct) at the juncture of 
the oviduct with the spermathecal duct (8). 

6. Duct of serminal receptacle(s): (a) short (0); (b) long (1); (c) lost due to fusion of the seminal 
receptacle to the duct of the bursa (2); (d) lost as the usual seminal receptacle is lost (3). 

7. Gonad 1 or 2, or 3 finger-like tubes: (a) (0); (b) (1). 

8. Vas deferens runs to style sac, turns posteriorly to form seminal vesicle: (a) (0); (b) (1). 

9. Central tooth: (a) anterior cusps 5 or more (0); (b) only one large triangular blade (1) 

10. Foot shape, power: (a) slender, weak (0); (b) wide, powerful (1) 

11. Head shape: (a) standard (0); (b) wide, squat head; eyes in pronounced lobes (1) 

12. Shell shape: (a) ovate-conic (0); (b) globose (1); (c) trochoid (2) 

13. Shell size (a) small, thin (0); (b) increased length, thickness (1) 

14. Penis with chitenous stylet: (a) (0); (b) (1). 

15. Spermathecal duct with vaginal section: (a) (0); (b) (1). 

16. Bursa copulatrix elongated relative to length of albumen gland: (a) (0); (b) (1). 

17. Penis has a strongly developed muscular white zone at concave edge: (a) (0); (b) (1). 

AUTAPOMORPHIES 
Fenou/7/a 

18. Gill filament section Gf, extremely elongated (1) 

Lithoglyphopsis 

19. Radular sac extremely elongated (1). 

20. Posterior palliai oviduct bends 180° around (1). 

21. Extremely long cerebral commissure (1). 

Jinhongia 
5f (above). Function of the seminal receptacle within an internal swelling at the juncture of the bursa 
and spermathecal duct (1). 

Robertsiella 
5e (above). Function of the seminal receptacle within a cavitation of the duct of the bursa (1). 

22. Common sperm duct, duct of the bursa encapsulated (1). 

Guoia 

23. The penis has a glandular lobe (1). 

24. The sperm duct is not only much elongated, it coils (1). 

Wuconchona 

25. The columella of the shell within the body whorl has a raised spiral ridge (1). 

26. The spermathecal duct is short (1). 

27. The male gonad is a wide sac, few lobes, floor of sac opens to a vas deferens, i.e. no vas efferens 

0). 

28. The dominant cusp of the lateral tooth is displaced towards the outside edge of the tooth (1). 

29. The oviduct makes a U-shaped bend (1). 

(continued) 



332 DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 

TABLE 84. (Continued). 

Gammatricula 

30. The posterior half or third of the prostate is smooth (1). 

Pachydrobia 

31. The shell lip is especially thickened (1). 

5d (above). The seminal receptacle arises from the sperm duct. 

Halewisia 
4e (above). The spermathecal duct joins the bursa copulatrix. 



TABLE 85. Scoring 18 OTUs for synapomorphies listed in Table 84. Hub, Hubendickia (outgroup of 
Jullieniini); Tr-a, Tricula species a, Tr-b, species b; Delavaya; Fen, Fenouilia; Lac, Lacunopsis; Lit, 
Lithoglyphopsis; Ne-a, Neotricula species a, Ne-b, species b, Ne-c, species c; Hal, Halewisia; Рас, 
Pachydrobia; Gu-a, Guoia species a, Gu-b, species b; Rob, Robertsiella; Jin, Jinhongia; Wuc, 
Wuconchona; Gam, Gammatricula. 





1 


2 


3 


4 


5 


6 


7 


8 


9 


10 


11 


12 


13 


14 


15 


16 


17 


18 




Hub 


Tr-a 


Tr-b 


Del 


Fen 


Lac 


Lit 


Ne-a 


Ne-b 


Ne-c 


Hal 


Рас 


Gu-a 


Gu-b 


Rob 


Jin 


Wuc 


Gam 


1 























1 


1 


1 


1 


1 


1 


1 


1 


1 


1 


1 


2 


1 




















2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


2 


3 























1 


1 


1 


2 


1 


2 


2 


1 


1 





1 


4 


2 


1 


1 


1 


1 


1 


1 


3 


3 


3 


4 


3 


3 


3 


3 


3 





3 


5 


1 





2 








8 





2 


2 


2 


2 


3 


6 


7 


4 


5 


7 


7 


6 


1 























1 


2 


1 


1 


3 


3 


3 


3 


3 


3 


7 


1 





















































8 


1 





















































9 











1 


1 




1 



































10 














1 




1 



































11 














1 




1 



































12 














2 




1 



































13 











1 


1 




1 














1 




















14 






































1 


1 


1 











15 






































1 


1 


1 











16 























1 








1 

















1 


1 


17 


















































1 


1 



There is clear-cut parallelism in the intra- 
ductal shifting of sperm storage. The Robert- 
siella-Guoia clade has highly derived charac- 
ter-states: the chitenous sheath on the penial 
papilla, the vaginal section of the spermathe- 
cal duct, the thick, straight and relatively short 
continuous duct running from the end of the 
mantle cavity into the bursa, and the loss of 
the seminal receptacle. The clade is relatively 
old within the 1 2 ± 4 million years in which the 
triculine macro adaptive radiation (Davis, 
1992) has been evolving. The presence of 
Robertsiella in Malaysia and Guoia in Hunan, 
China, indicate both the age of the clade in 
and its origin in northern Burma or Yunnan, 
China. Three character-states involving the 
sperm storage location are found within this 
one clade: (1) within an outpocketing of the 
encapsulated duct of the bursa (Robertsiella), 



(2) within the sperm duct next to the oviduct 
(Guoia-a), (3) within the proximal oviduct 
(Guoia-b). The parallel situation is seen in the 
genera Wuconchona and Gammatricula that 
are found only (thus far) in southeastern 
China. In these genera, the simplified gonad, 
penial-characters (but lacking the chitenous 
penial stylet), and sperm storage in a more 
distal section of oviduct indicate a clade of 
derived taxa clearly divergent from the Rob- 
ertsiella-Guoia clade. 

Other parallelisms are simply partitioned 
and understood. In each second order radia- 
tion (Davis, 1980, 1992) there are trends for 
increasing size (seen in shell length), increas- 
ing complexity in sculpture (from smooth -» 
ribs -* reticulate sculpture, spiral cords, 
spines or nodes), and changes in shape 
(ovate-conic to turreted, globose, assymetry 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 333 



-HUBENDICKIA 

I — TRICULA-b 

-TRICULA-a 

-DEL AVAYA 

-FENOUILIA 

-LACUNOPSIS 

-LITHOGLYPHOPSIS 



С 



с 



£í 



, — NEOTRICULA-b 
-NEOTRICULA-c 

NEOTRICULA-a 

HALEWISIA 

PACHYDROBIA 

i — JINGHONGIA 



r: 



Г 



Г 



-HUBENDICKIA 

TRICULA-b 

(—TRICULA- 



£ 



GAMMATRICULA 
WUCONCHONA 
ROBERTSIELLA 
GUOIA-a 
GUOIA-b 



Г 



— DEL AVAYA 

(—FENOUILIA 
LACUNOPSIS 



1 — LITHOGLYPHOPSIS 



— NEOTRICULA-b 
NEOTRICULA- с 

NEOTRICULA-a 



Г 



HALEWISIA 



I — PACHYDROBIA 

JINGHONGIA 

WUCONCHONA 

GAMMATRICULA 

r- ROBERTSIELLA 

■ — GUOIA-a 

' — GUOIA- b 



a 



HUBENDICKIA 

-TRICULA-b 

-TRICULA-a 

-DEL AVAYA 

-FENOUILIA 

-LACUNOPSIS 



r 



г 



с 



г 



г; 



ITHOGLYPHOPSIS 



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-1— H 



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JINGHONGIA 



Г 



Г 



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WUCONCHONA 

ROBERTSIELLA 

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1 — GUOIA-b 



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-TRICULA-b 

-TRICULA- a 

-DEL AVAYA 

-FENOUILIA 

-LACUNOPSIS 



С 



с 



с 



с 



г 



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WUCONCHONA 

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FIGS. 159-162 Four representatives of the 20 computer-generated cladograms following the Hennig-86 
treatment of data given in Tables 85 and 86. See text for details. 



(see Davis, 1979). The primitive small ovate- 
conic shell seen in Tricula, Neotricula, Jinhon- 
gia and Gammatricula is the basic platform 
from which splendid first order adaptive radi- 
ations arose, most with distinctive patterns of 
shell development (e.g. Pachydrobia, Jullie- 
nia, Hubendickia, Lacunopsis. However, 
while one can discern the shells of Lacunop- 
sis from those of Hubendickia or Pachydro- 
bia, there is convergence in the globose shell 
types such that the shell character-states of 
Guoia, Lithoglyphopsis and Lacunopsis 
merge. Fortunately, there is extreme anatom- 
ical divergence between Guoia, Lacunopsis 
and Lithoglyphopsis; accordingly these gen- 
era are readily classified in one or the other of 
the relevant second order radiations. 

The length of the duct of the seminal recep- 
tacle increases with increasing anatomical 
complexity in the Jullieniini and Triculini. The 
trend is particularly pronounced in the Jullie- 



niini (Davis, 1979; 1991). There is a parallel 
increase in the length of the sperm duct in- 
volving some derived genera of the Triculini 
and Pachydrobiini; it is particularly evident in 
Guoia. 



OLD PROBLEMS RESOLVED 

Two unsolved questions of a decade ago 
can now be answered. (1) What is Lithogly- 
phopsis and its relationship to Lithoglyphop- 
sis aperta Temcharoen, 1971? (2) Given the 
impact of the Himalayan orogeny as a driving 
force of the spectacular triculine radiation, too 
few species of Tricula or species morpholog- 
ically close to Tricula had been found a de- 
cade ago (Davis, 1980). There were numer- 
ous species of Pachydrobia, Lacunopsis, 
Hubendickia, etc. known a decade ago, but 
only three species of Tricula, sensu Davis, 



334 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



— HUBENDICKIA 

i — NEOTRICULA-a 
NEOTRICULA-b 
NEOTRICULA-c 

— HALEWISIA 

— PACHYDROBIA 

— JINGHONGIA 
i— WUCONCHONA 
' — GAMMATRICULA 

ROBERTSIELLA 
GUOIA-a 
GUOIA-b 



Г 



-TRICULA-b 

i — TRICULA-a 

DELAVAYA 



€ 



FENOUILIA 

LACUNOPSIS 

LITHOGLYPHOPSIS 



FIG. 163. The nelsen consensus tree resolving the 
20 trees generated following the Hennig-88 treat- 
ment of data given in Tables 85 and 86. 



1979, had been found in Southeast Asia. A 
few species from Burma had been described 
as Tricula on the basis of shells (reviewed in 
Davis, 1980). It was predicted that more 
would be found (Davis, 1980). How extensive 
was the Tricula radiation? 

Concerning the first question, it is evident 
that Lithoglyphopsis is most closely related to 
Fenouilia and Delavaya anatomically and to 
Lacunopsis conchologically. Lithoglyphopsis 
is a derived genus in the tribe Triculini. Litho- 
glyphopsis aperta, the snail host for Schisto- 
sosma mekongi, is the type species for the 
genus Neotricula (Davis et al., 1986a). Neo- 
tricula is an anatomically generalized genus 
in the tribe Pachydrobiini. Thus, while Litho- 
glyphopsis and Neotricula are members of 
the same subfamily, they are highly divergent 
genera within the Triculinae. 

The second issue is now resolved. With re- 
search initiated in Yunnan, China, in 1983 
(Davis et al., 1986b) and with this and other 
ongoing research, it is clear that Tricula, 
sensu Davis, 1979, comprises an extremely 
large radiation primarily located in southern 
China. The concept of Tricula sensu Davis, 
1979, has changed. The anatomical data sup- 
port four genera with a Tr/cu/a-type shell: Tric- 
ula, Neotricula, Gammatricula, and Jinhongia. 
Anatomical data have been published for 
seven species of Neotricula, eleven species 
of Tricula and one each for Jinhongia and 
Gammatricula. These 20 species are one 
third of the estimated 60 species (conserva- 
tive estimate) with Tricula-Wke shells that 



we think occur throughout southern China, 
Burma and northern Vietnam (Davis, 1992). 



BIOGEOGRAPHY 

Biogeography at a glance is provided in 
Figure 165. An area cladogram for river evo- 
lution is given with the number of species of 
each genus for which we have anatomical 
data listed for each region of river. Overall 
Triculinae biogeography has been reviewed 
recently (Davis, 1992). Only a few points 
need to be emphasized here. (1) Thus far, 
Neotricula has its greatest deployment in the 
mid to lower Yangtze River, whereas Tricula 
has more species in the upper Yangtze River. 
(2) The distribution of Guoia, Robertsiella, 
Tricula and Neotricula show that the major 
innovative anatomical developments in the 
Triculini and Pachydrobiini had occurred be- 
fore the Yangtze-Mekong river drainages 
were completely separated. 

(3) The tribes Triculini and the derivative 
Pachydrobiini dominate the Yangtze River 
drainage; no Jullieniini have been found, thus 
far, in the mid or lower Yangtze River drain- 
ages. Conversely, the Jullieniini dominate the 
Mekong River drainage with their greatest ra- 
diation in the lower Mekong River. (4) Only 
Pacydrobia of the Pachydrobiini and La- 
cunopsis of the Triculini have undergone ex- 
plosive adaptive radiations (considering all 
genera of the Triculini and Pachydrobiini); this 
has occurred in the lower Mekong River. (5) 
The most derived anatomical character- 
states are found in the lower river systems. 

(6) There are considerable ecological dif- 
ferences among tribes. With the exception of 
Kunmingia living in the shallow basin of a 
limestone spring, the other genera of the Jul- 
lieniini live in a large river environment. Con- 
sidering the Triculini and Pachydrobiini, only 
Lacunopsis, Lithoglyphopsis, Guoia, Halewi- 
sia and Pachydrobia live in large rivers. 
Fenouilia lives on the bottom of a large lake, 
as did Parapyrgula (presumed extinct). Tric- 
ula, Neotricula, Jinhongia and Gammatricula 
live in upland shaded small streamlets of 
pure, cold water. Only Neotricula aperta has 
adjusted to living in a large river environment. 

There are numerous species of Neotricula 
and Tricula in Hubei Province and provinces 
to the east, species that are in the process of 
being studied. We have seen at least one ad- 
ditional species of Gammatricula. The proven 
distribution of Tricula is from northern India to 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 335 



«HÜ} 

2b 4c 5b 7b 8b 6b 



* 



Hf4 



1b 2c 3b,c 4d 

19 SYNAPOMORPHIES 
15 HOMOPLASIES 
4 LOSSES 

— X 

38 TREE LENGTH 
AUTAPOMORPHIES О 



-Kb 

9b 13b 



{} 



Ü 



10M1b 



rO-O 

• r-o-o- 

19 20, 21 

КО 



* 



г-ШЬ 



^9: 



fOOo 



-X- 



rO 



6d 5e+ 



If 



rOO— — 

5e 22 l— O" 



4x>a 



5g 



{h 



HUBENDICKIA 



TRICULA-a 



TRICULA-b 

DEL AVAYA 

FENOUILIA 
LITHOGLYPHOPSIS 

LACUNOPSIS 

HALEWISIA 

NEOTRICULA-b 
NEOTRICULA-a 

NEOTRICULA-c 
• PACHYDROBIA 

JINHONGIA 

ROBERTSIELLA 

GUOIA-a 

GUOIA-b 



ЧНД- 



rX-QQOOOOOwUCONCHONA 



3a 4a 3a 24 25 26 27 28, 29 

I О GAP 



FIG. 164. The cladogram resulting from weighting one character in particular, the seminal receptacle, and 
the direction of evolution of character states with the primitive condition being 5a ( = score 0) (see Table 85) 
(all Triculini except Lacunopsis); the shift of the duct of the seminal receptacle to the duct of the bursa (5c 
= score 2); the loss of the seminal receptacle, its function transferred into the duct of the bursa (5e + and 
5e on the cladogram); the move within the duct system to the juncture of duct of the bursa, sperm duct and 
spermathecal duct (5f ) -» to the sperm duct (5g) — > to the oviduct (5h). Parallelisms are accounted for; see 
text for details. 



the East China Sea. As discussed in Davis 
(1992), there are numerous diverse Triculinae 
in Yunnan that have not yet been studied. It is 



highly probable that species of Neotricula will 
be found among them in the upper Yangtze 
River drainage. While this study has done 



336 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



GÜOIÄ-2 
LITHOGLYPHOPSIS-1 



DELAVAYA-1 

KUNMINGIA-1* 

LACUNOPSIS-? 


TRICOLA-3 
WUCONCHONA- 1 








NEOTRICDLA-? 
TRICÜLA-3 




HUBEI 
HUNNAN 


GAMMATRICÜLA- 1 




UPPER YANGTZE R 




LOWER 


YANGTZE R. 




FENOÜILIA-1 
PARAPROSOSTHENIA*- 
PARAPYRGÜLA*-E? 
TRICULA-3 


E? 






TRICULA- 1 




LAKE 
ERHAI 




























HALEWISLA 

HOBENDICKIA* 

HYDRORISSOIA* 






MEKONG R. 




TRICÜLA-1 




HUBENDICKIA-1 
JINHONGIA-1 
LACÜN0PSIS>1 
TRICÓLA- 1 


THAI DRAINAGE 

| 


KARELAINIA* 
LACUNOPSIS 
NEOPROS06THENIA* 
NEOTRICÜLA-1 




UPPER MEKONG R. 
YUNNAN 


PACHYDROBIA 
PACHYDROB I E L LA * 












ROBERTSIELLA-2 


1 LOWER MEKONG R. 



MALAYSIA 

FIG. 165. An area cladogram for rivers showing the location of triculine genera. Those genera with the 
asterisk belong to the tribe Jullieniini. The question mark indicates the possibility that the genus might be 
found there. E? indicates probable extinction. With the exception of Pachydrobia and Lacunopsis in the lower 
Mekong River, each with more than ten species, the number of species of each genus of the Triculini and 
Pachydrobiini studied anatomically is given at each location along the drainage systems. Tr/cu/a-1 is Tricula 
montana of northern India. See text for further details. 



much to reduce the probability that new gen- 
era of Triculinae will be found (we would ex- 
pect one or two more, especially in Yunnan), 
there is considerably more to be learned con- 
cerning species of Tricula and Neotricula and 
their patterns of distribution. 



mell. Dr. John Hendrickson is thanked for his 
help in executing the NTSYS programs. The 
work was supported by the National Natural 
Science Foundation of China award No. 
3860607 to Dr. Chen, and N.I.H. award A1 
11 373 to Dr. Davis. 



ACKNOWLEDGEMENTS 

Use of the facilities of Hunan Medical Col- 
lege and the Zhejiang Academy of Medical 
Sciences is gratefully acknowledged. The 
drawings were made by Davis with render- 
ings by Elizabeth Carrossa and Susan Tram- 
mell. Scanning electron microscope work was 
done by Dr. Chen Cui-E and Caryl Hester- 
man. Photographs of the shells were taken 
and printed by Dr. Chen and Caryl Hester- 
man. Graphics were done by Susan Tram- 



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TRYON, G. W., 1862, Notes on American fresh wa- 
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Proceedings of the Academy of Natural Sciences 
of Philadelphia, 14: 451-452. 

THIELE, J., 1928, Revision des systems der Hydro- 
biiden und Melaniiden. Abteilung für Systematik, 
Ökologie und Geographie der Tiere, 55: 351- 
402. 

VOGE, M., D. BRUCKNER & J. I. BRUCE, 1978, 
Schistosoma mekongi sp. n. from man and ani- 
mals, compared with four geographic strains of 
Schistosoma japonicum. Journal of Parasitology, 
64(4): 577-584. 

WENZ, W., 1939, Gastropoda, Part 3, Prosobran- 
chia. Pp. 555-581 in О. H. schindewolf, ed., 
Handbuch der Paläozoologie, Berlin (Borntrae- 
ger). 

YEN, T. C, 1939, Die chinesischen Land-und 
Süsswasser — Gastropoden des Naturmuseums 
Senckenberg. Abhandlungen der Senckenber- 
gischen Naturforschenden Gesellschaft, 444: 1- 
234. 

ZILCH, A., 1974, Vinzenz Gredler und die Er- 
forchung der Weichtiere Chinas durch Franzi- 
kaner aus Tirol. Archiv für Molluskenkunde, 1 04: 
171-228. 



Revised Ms. accepted 16 December 1991 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 339 

APPENDIX 1. Sources of data for species of Triculinae used in the multivariate analyses 
(excluding species treated in this paper). 

Neotricula apena (Temcharoen, 1971): in Davis et al., 1976 [Type species of genus; designated 

in Davis et al., 1986a]. 

Neotricula burchi (Davis, 1968) 

Tricula montana Benson, 1843: in Davis et al., 1986a [Type species of genus] 

T. bambooensis Davis & Zheng, 1986: in Davis et al. 1986b 

T. bollingi Davis, 1 968 

T. gregoriana Annandale, 1924: in Davis et al., 1986b 

T. hudiequanensis Davis & Guo, 1986: in Davis et al., 1986b 

Г. ludongdini Davis & Guo, 1986: in Davis et al., 1986b 

T. xianfengensis Davis & Guo, 1986: in Davis et al., 1986b 

T. xiaolongmenensis Davis & Guo, 1986: in Davis et al., 1986b 

Jinhongia jinhongensis (Guo & Gu, 1985): in Davis et al., 1986b [Type species of genus] 

Gammatricula chinensis Davis, Liu & Chen, 1990: in Davis et al., 1990 [Type species of genus] 

Wuconchona niuzhuangensis Kang, 1983: in Davis & Kang, 1990a [Type species of genus] 



340 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



APPENDIX 2. Factor loading of characters for the ten principal components that collectively account for 
85% of the variation in this study of shell characters. 









PRINCIPLE COMPONENTS 








I. 


II. 


III. 


IV. 


V. 


1. 


-0.0410545 


-0.4614740 


0.1951133 


0.1102495 


-0.1964896 


2. 


0.3371738 


-0.2702949 


0.6452890 


-0.2870722 


-0.2273258 


3. 


-0.5820084 


0.0153342 


0.2508085 


0.0428066 


0.5737585 


4. 


-0.4883084 


-0.0937884 


-0.3041671 


0.3050418 


-0.0443432 


5. 


0.1598786 


0.6532829 


-0.1058710 


-0.0463305 


-0.4989291 


6. 


-0.2746159 


0.8122425 


0.0071554 


-0.0500111 


0.1822269 


7. 


0.2803561 


0.2844659 


0.2436223 


-0.6108634 


-0.2804339 


8. 


0.1706873 


-0.4612060 


0.0912007 


0.4000807 


-0.1060740 


9. 


-0.6064097 


0.0550309 


0.4268830 


0.0825438 


0.4504842 


10. 


-0.0511060 


-0.0086817 


0.7289513 


0.3538858 


0.0138379 


11. 


-0.8494929 


0.0619407 


0.1342359 


-0.1052565 


0.2478488 


12. 


-0.6526540 


0.4972618 


0.2145493 


-0.2189934 


-0.0605389 


13. 


-0.7146944 


0.2136463 


0.0176418 


-0.0641302 


0.0094302 


14. 


0.1056108 


0.2477635 


-0.3376958 


0.0983843 


-0.0247723 


15. 


-0.1938874 


0.2239324 


0.2971711 


0.4426535 


-0.3136139 


16. 


0.3325563 


-0.1555008 


0.1835824 


-0.7154313 


0.0046761 


17. 


-0.6648433 


-0.3846053 


0.0730909 


-0.3156229 


-0.4205736 


18. 


-0.6955758 


0.3660246 


0.1990413 


-0.2286217 


0.2015936 


19. 


-0.6319250 


-0.3369728 


-0.1161682 


0.0574202 


-0.4591451 


20. 


-0.6360441 


-0.1127735 


-0.2589680 


0.1512705 


-0.3323932 


21. 


-0.4095072 


-0.3166545 


-0.2127306 


-0.1982544 


-0.2723410 


22. 


0.3875425 


-0.1664845 


0.1605595 


-0.5852705 


0.3093103 


23. 


-0.0710536 


0.4584002 


-0.2259200 


0.1122301 


-0.2893657 


24. 


-0.7326382 


-0.1977977 


0.0873077 


-0.2158382 


0.1143209 


25. 


-0.0983867 


0.3013054 


-0.2336317 


0.1312115 


-0.1079600 


26. 


0.0980229 


-0.0845837 


0.8104326 


0.3794045 


-0.1045079 


27. 


0.1178917 


0.2651106 


0.7510733 


0.1976523 


-0.3326711 


28. 


0.1214556 


0.5956626 


0.1896647 


-0.1906758 


-0.4167448 


29. 


-0.6648433 


-0.3846053 


0.0730909 


-0.3156229 


-0.4205736 




VI. 


VII. 


VIM. 


IX. 


X. 


1. 


0.3384070 


-0.5698774 


0.0460096 


-0.1038296 


-0.2015172 


2. 


-0.1341500 


-0.1682082 


0.1296563 


0.2262901 


0.1179613 


3. 


0.1338242 


0.0179137 


0.0716831 


0.1584757 


0.0337647 


4. 


0.1847772 


-0.0621649 


-0.3912900 


0.3915480 


-0.0423710 


5. 


0.1626572 


-0.0789925 


0.2185103 


0.1225756 


-0.0940029 


6. 


-0.1021466 


0.0043270 


0.0444586 


-0.0901887 


0.2147945 


7. 


-0.0002180 


0.1959912 


-0.1031375 


0.0608091 


-0.2179681 


8. 


0.0615272 


0.1587772 


-0.2981737 


-0.4940681 


-0.0164338 


9. 


-0.2213138 


-0.0213113 


0.1639645 


-0.1460116 


-0.2374873 


10. 


-0.2431751 


0.0326452 


0.1561172 


0.0089133 


-0.0482282 


11. 


-0.1235609 


0.0485087 


-0.0126241 


0.0552378 


-0.3203997 


12. 


0.1602039 


0.0045610 


-0.0563521 


-0.0245823 


-0.0996069 


13. 


0.2771966 


-0.1739794 


-0.0983610 


0.0695070 


0.4416130 


14. 


0.2559430 


-0.1571185 


0.7337301 


0.0273879 


0.1133017 


15. 


-0.0909669 


0.2889332 


-0.2158527 


0.4116663 


0.2163310 


16. 


-0.3295022 


0.0224967 


-0.0264935 


0.2984503 


0.0080422 


17. 


0.1965575 


-0.1292952 


-0.0028319 


-0.0987587 


0.1400116 


18. 


-0.0673200 


-0.1059933 


-0.1987022 


-0.2522537 


0.2059356 


19. 


-0.1305618 


0.1619921 


-0.0220588 


0.1714101 


-0.1204920 


20. 


-0.4444652 


0.1209360 


0.1189897 


-0.0587002 


-0.0058012 


21. 


-0.4959528 


0.1709711 


0.2976215 


-0.1487671 


0.1337223 


22. 


-0.1349609 


-0.2227013 


-0.2268552 


-0.0828126 


0.1662128 


23. 


-0.5534215 


-0.2637156 


-0.0907283 


-0.2368980 


0.0500118 


24. 


0.1807102 


0.1188745 


0.2030360 


0.0449544 


-0.2547120 


25. 


-0.3337661 


-0.6962163 


-0.1929729 


0.1613878 


-0.2710739 


26. 


-0.0871736 


-0.1570683 


0.1745502 


0.0772054 


0.1175377 


27. 


0.0513585 


-0.0302295 


-0.0577648 


-0.1412964 


-0.0056331 


28. 


0.3292032 


0.2276822 


-0.1024080 


-0.2274468 


-0.2240125 


29. 


0.1965575 


-0.1292952 


-0.0028319 


-0.0987587 


0.1400116 



THE POMATIOPSIDAE OF HUNAN, CHINA (GASTROPODA: RISSOACEA) 341 



APPENDIX 3. Factor loading of characters for the ten principal components that collectively account for 
90% of the variation in this study of anatomical characters. Only 41 characters are used as no missing 
data were allowed in the PCA analysis. 



PRINCIPLE COMPONENTS 




I. 


II. 


III. 


IV. 


V. 


2. 


-0.0971430 


-0.6013535 


0.5959546 


0.0668697 


0.2552416 


3. 


0.2857560 


-0.0616901 


-0.4464561 


0.1288039 


0.6460538 


5. 


0.2334109 


0.0746406 


0.4689891 


0.5445427 


0.1455881 


6. 


-0.6159816 


0.2368112 


-0.1616069 


0.3824686 


0.3790016 


9. 


-0.1030642 


0.2758602 


0.2265553 


0.1676757 


0.3476608 


10. 


0.3783801 


0.2220442 


0.5730512 


0.1711390 


-0.4213342 


12. 


0.2808698 


-0.3578114 


0.3977414 


0.2035502 


-0.4838175 


13. 


-0.5800852 


0.2057583 


0.3564111 


0.3719478 


0.3222399 


14. 


-0.2141655 


-0.0802545 


0.1442952 


-0.5024069 


-0.1107264 


15. 


-0.0625617 


0.6955844 


0.1884190 


-0.0751765 


0.0585363 


16. 


0.1441796 


-0.2877685 


-0.1940611 


0.0395566 


0.0007327 


17. 


0.9340978 


-0.0764409 


-0.0112246 


-0.1052855 


0.1197626 


18. 


0.8379920 


-0.1581357 


-0.2415173 


-0.2205638 


0.2052664 


19. 


0.0948649 


-0.7225091 


0.5135236 


-0.0776931 


0.1336098 


20. 


-0.0477885 


-0.8711195 


0.2605822 


-0.1511693 


0.1926126 


21. 


0.6492946 


0.4411572 


0.2990977 


-0.2707346 


0.3700605 


22. 


0.2164818 


0.6650421 


0.3564631 


-0.3670552 


0.2019860 


23. 


0.6580347 


0.2659786 


0.1374653 


-0.1748385 


0.4240264 


24. 


0.7339792 


0.1832732 


-0.1162168 


-0.3264529 


0.1854340 


25. 


-0.3732890 


-0.4181728 


0.5697897 


-0.3883381 


-0.1400759 


26. 


0.5188465 


-0.1851264 


0.0220450 


0.4697214 


-0.1053183 


27. 


-0.6784778 


0.1793504 


0.0918947 


-0.1980585 


-0.2374376 


28. 


0.6098325 


0.1973294 


0.6315968 


0.2793996 


-0.1918658 


29. 


0.6098325 


0.1973294 


0.6315968 


0.2793996 


-0.1918658 


30. 


-0.5456913 


0.1716460 


0.4846009 


0.5285202 


0.0935103 


31. 


-0.0988049 


0.8467538 


-0.0251339 


-0.2078032 


-0.2906712 


32. 


0.4414210 


0.0242099 


0.4463780 


-0.3216799 


-0.0195030 


33. 


0.3388348 


-0.4707434 


-0.3449156 


0.2567195 


-0.0462704 


34. 


-0.3332398 


-0.5745382 


0.5893458 


-0.3239226 


0.1852541 


35. 


0.5994937 


0.3691855 


0.3543632 


-0.0498222 


0.3475240 


36. 


-0.6232941 


0.2722790 


0.2957349 


-0.2475745 


0.0005819 


37. 


-0.6437625 


0.1028952 


0.3961242 


-0.3618655 


0.0745969 


38. 


0.5793707 


-0.1093442 


-0.1040323 


-0.1968725 


-0.3085475 


39. 


0.2811599 


-0.4485988 


-0.3590495 


0.0503694 


-0.3279842 


40. 


-0.3332398 


-0.5745382 


0.5893458 


-0.3239226 


0.1852541 


41. 


-0.4597675 


0.4187379 


0.0153485 


0.3484048 


-0.0427419 


42. 


-0.1782957 


0.5215056 


0.2159982 


-0.1292289 


0.0143310 


43. 


-0.0840292 


0.1112269 


0.1647247 


-0.0415819 


-0.3936273 


44. 


0.1616456 


0.1293033 


-0.1539454 


-0.6310220 


-0.2087496 


45. 


0.0864995 


-0.7839667 


0.0446516 


0.1787427 


0.0927740 


46. 


0.6098325 


0.1973294 


0.6315968 


0.2793996 


-0.1918658 




VI. 


VII. 


VIII. 


IX. 


X. 



2. 

3. 

5. 

6. 

9. 
10. 
12. 
13. 
14. 
15. 
16. 
17. 
18. 



-0.1394164 
0.0093270 
0.0449117 
0.1163917 
0.1417184 
-0.1248724 
-0.1854555 
0.2550922 
0.2836786 
-0.0788765 
-0.4354456 
-0.1534960 
-0.1997098 



-0.3474175 
-0.0592813 
-0.4425233 
0.0395406 
0.6167990 
0.2536923 
-0.0913738 
-0.1352908 
-0.1617582 
0.2099368 
-0.3648156 
0.0609385 
0.1097419 



-0.0792670 
-0.2851638 
-0.2218537 
-0.1044403 
0.2681562 
-0.3529955 
0.0851108 
0.1456703 
0.3841972 
0.2237864 
-0.2176404 
0.1527720 
0.1298337 



0.1056858 
0.1587237 
0.3159829 

-0.0334291 
0.4167362 

-0.0161008 
0.1708288 

-0.1650157 

-0.0299036 
0.0080864 

-0.6109018 
0.0182737 
0.0507008 



0.0854577 
-0.0517160 
0.1471501 
0.2700133 
0.1386042 
0.1129820 
0.1926282 
-0.1249281 
0.5384613 
0.3954614 
0.1593562 
0.1012241 
0.1254707 



(continued) 



342 



DAVIS, CHEN, WU, KUANG, XING, LI, LIU & YAN 



APPENDIX 3. (Continued) 



PRINCIPLE COMPONENTS 




VI. 


Vil. 


VIII. 


IX. 


X. 


19. 


0.1677881 


0.2533934 


0.1785646 


0.0104447 


-0.0195058 


20. 


0.0126496 


0.1760192 


0.0752291 


-0.1513470 


0.0383810 


21. 


0.1104381 


-0.0205072 


-0.0538776 


-0.1350167 


-0.0573354 


22. 


0.2026985 


0.0578036 


-0.3427233 


-0.0726498 


-0.0024815 


23. 


0.3357801 


-0.0910891 


-0.2196621 


0.1255442 


0.1376799 


24. 


-0.1652149 


0.2716005 


-0.1927697 


-0.0061319 


0.0910426 


25. 


0.0315530 


0.1971248 


-0.2439671 


-0.0636229 


-0.0579249 


26. 


0.4822483 


0.2524238 


0.0512107 


-0.095791 1 


-0.0476433 


27. 


-0.0037041 


0.0253443 


0.1257655 


0.2416408 


-0.0378403 


28. 


0.0709616 


-0.1122485 


0.1196700 


-0.0827769 


-0.0295994 


29. 


0.0709616 


-0.1122485 


0.1196700 


-0.0827769 


-0.0295994 


30. 


0.1328322 


-0.1671411 


-0.2060764 


0.1016164 


0.0785346 


31. 


0.1569631 


0.0579261 


0.1100781 


-0.2140089 


-0.0485999 


32. 


-0.4554683 


0.1161356 


0.0894265 


0.0458263 


-0.2058747 


33. 


0.5295037 


0.3717229 


-0.1185204 


-0.0160033 


0.0198906 


34. 


0.1115931 


0.0793780 


0.1069966 


-0.0967904 


-0.0698220 


35. 


0.0258687 


0.0397086 


-0.0296042 


-0.2666091 


-0.2045914 


36. 


0.1449843 


0.1298960 


-0.2864864 


-0.1526312 


-0.0600707 


37. 


0.2668363 


-0.1405495 


-0.3199402 


0.0507502 


0.0056072 


38. 


0.4748601 


-0.1419334 


0.0072793 


0.2376274 


-0.0643782 


39. 


0.5829850 


0.0145123 


-0.1912881 


0.0057201 


-0.1940589 


40. 


0.1115931 


0.0793780 


0.1069966 


-0.0967904 


-0.0698220 


41. 


-0.0477399 


0.3526877 


0.0829260 


-0.2145180 


-0.0505431 


42. 


-0.3470802 


-0.0603708 


0.1326352 


0.5233748 


-0.3710022 


43. 


-0.2444863 


0.4647836 


-0.6101984 


0.0604030 


0.1866010 


44. 


0.3225032 


-0.4169769 


-0.1464836 


0.2691902 


0.0783850 


45. 


-0.3467677 


0.1343245 


-0.2636901 


0.3034776 


0.0343297 


46. 


0.0709616 


-0.1122485 


0.1196700 


-0.0827769 


-0.0295994 



MALACOLOGIA, 1992,34(1-2): 343-354 

CROP EPITHELIUM OF NORMAL FED, STARVED AND HIBERNATED SNAILS 
HELIX LUCORUM: A FINE STRUCTURAL-CYTOCHEMICAL STUDY 

V. K. Dimitriadis, D. Hondros & A. Pirpasopoulou 

Department of Genetics, Development and Molecular Biology, School of Biology, 
Aristotle University of Thessaloniki, Thessaloniki, 54006, Greece 

ABSTRACT 

The crop epithelium of the snail Helix lucorum consists of four cell types: ciliated and unciliated 
columnar cells, mucous cells, and basal cells. The morphologic features of the epithelium show 
that the digestive activities in the crop lumen are probably mediated by digestive enzymes 
regurgitated from the digestive gland and the stomach, rather than secreted by the crop cells 
themselves. On the other hand, the crop epithelium is probably responsible for the absorption of 
nutrients from the crop lumen. The mucous cells secrete a periodate-reactive, non-sulfated and 
non-carboxylated material, whereas the dense bodies observed in the columnar cells show a 
positive reaction for acid phosphatases, as well as for periodate-reactive, sulfated and carbox- 
ylated glycoconjugates and possibly are of lysosomal origin. Thirty seven days of hibernation 
increased the number of mucous cells and the number of the dense bodies in the columnar cells, 
while the amount of glycogen particles and lipid inclusion drastically decreased in non-mucous 
cells compared to controls. Hibernation did not alter the carbohydrate content of mucous cells. 
In addition, hibernation caused increased appearance of extrusion of cytoplasmic regions into 
the crop lumen, as well as lysis of certain columnar cells compared to controls. Forty days of 
starvation induced similar but less intense phenomena in the crop epithelium. 

Key words: Helix lucorum, snail, crop, fine structure, hibernation, starvation, cytochemistry. 



INTRODUCTION 

The crop of Pulmonata consists of a thin 
tube connected to the oesophagus and fol- 
lowed by the main stomach. There are few 
data about the structure and function of the 
cells that constitute the pulmonate crop, 
which is regarded as a region where food is 
stored for certain periods (Runham, 1975). 
Furthermore, there is little available informa- 
tion on the morphology and function of these 
cells at the ultrastructural level (Roldan, 1987; 
Roldan & Garcia-Corrales, 1988). As far as 
the digestive enzymes of the digestive tube of 
molluscs are concerned, there are contradic- 
tory views about their origin, for example 
whether they are of exogenous or endoge- 
nous origin or both (Fretter. 1952; Jeuniaux, 
1954; Owen, 1966; Charrier, 1990). 

In addition, little is known about the bio- 
chemistry, physiology and fine structural cell 
morphology of snails exposed to such condi- 
tions as hibernation and starvation, and that 
literature mainly refers to the digestive gland 
epithelium (Sumner, 1965; Oxford & Fish, 
1979; Janssen, 1985; Dimitriadis & Hondros, 
in press). To our knowledge, the only related 
information at the ultrastructural level comes 



from the snail Theba pisana, in which starva- 
tion resulted in the disappearance of lipid 
droplets and glycogen particles in the colum- 
nar crop cells (Roldan, 1987). 

The present study was designed: (1 ) to ex- 
amine the fine structural features of the crop 
epithelium of the snail Helix lucorum; (2) to 
cytochemically characterize the crop cells; 
and (3) to study the effect of starvation and 
hibernation on crop cell morphology. One fur- 
ther question was also addressed: is the car- 
bohydrate content of the mucous cells altered 
in starved and hibernated snails compared to 
controls? 



MATERIALS AND METHODS 

Adult Helix lucorum (Gastropoda, Pulmo- 
nata, Helicidae) were collected from Edessa, 
northern Greece. The largest shell diameter 
of the snails used in the present study varied 
from 41 to 43 mm, and the body weight was 
between 19 and 21.5 g. 

The effect of hibernation on crop cells was 
examined in snails kept in a cold room at 
4±1°C under a photopehod of 9 h light : 15 h 
dark (Lazaridou-Dimitriadou & Saunders, 



343 



344 



DIMITRIADIS ET AL. 



1986), for 37 days. Starved snails were kept 
at 1 9± 1 °C and a photopehod of 1 3L : 1 1 D for 
40 days. Control fed snails were kept at 19 ± 
1°C and a photopehod 13L : 11D. 

The tissues were fixed in Karnovsky's fixa- 
tive (Karnovsky, 1965), postfixed in 2% os- 
mium tetroxide, dehydrated, and embedded 
in Spurr's resin. Sections were cut using a 
Reichert OmU3 ultramicrotome, post-stained 
with uranyl acetate and lead citrate, and ex- 
amined under a JEOL 100B electron micro- 
scope operating at 80 KV. For light micro- 
scopic observations, thick sections were 
stained with 1% toluidine blue. 

For carbohydrate cytochemistry, finely 
minced pieces of crop were incubated over- 
night in low iron diamine (LID) (Takagi et al., 
1982) or in high iron diamine (HID) (Spicer et 
al., 1978; Sannes et al., 1979), treated with 
osmium tetroxide and embedded in Spurr's 
resin. Thin sections of these specimens were 
stained with the thiocarbohydrazide-silver 
proteinate (TCH-SP) sequence. Control tis- 
sues were exposed in 1 M MgCI 2 in place of 
LID or HID. Specimens without osmium 
tetroxide treatment were used for the postern- 
bedding periodate-thiocarbohydrazide-silver 
proteinate (PA-TCH-SP) method (Thiéry, 
1967). Control sections were stained without 
the periodate treatment. Thin sections of 
buffer- and osmium tetroxide-treated tissues 
post-stained with uranyl acetate and lead cit- 
rate were used to characterize normal crop 
epithelial ultrastructure. 

To demonstrate acid phosphatase activity, 
a modified method similar to that proposed by 
Barka & Anderson (1962) was applied. Tis- 
sues sections approximately 1 mm in thick- 
ness were fixed in glutaraldehyde and then 
incubated in a medium containing 0.2 M tris/ 
maléate (pH 5) as a buffer an 0.1 M ß-glyc- 
erophosphate as substrate at 37° for 15-30 
min. Control sections were incubated in the 
absence of substrate. 

Morphometric evaluation was performed 
according to the methods described by Wei- 
bel (1979) and Steer (1981). Samples of five 
cells were analysed from each crop section of 
four different snails. The volume density of 
the dense bodies and of the lipid inclusions in 
the supranuclear cytoplasm of the columnar 
cells was determined from point counting ste- 
reology, using a test square lattice with a pe- 
riod d = 10 mm, equivalent to 1 (xm on the 
specimen. A minimum of 480 points were 
counted per cell. The distribution of columnar 
and mucous cells within the crop epithelium 



were determined by measuring nucleated 
cells of both types on light microscope micro- 
graphs at a final magnification of x 500. Mean 
values and standard deviations of the mor- 
phometric parameters were calculated and 
statistically compared using Student's t-test, 
significant level P<0.05. 



RESULTS 
Morphology of the Crop Epithelium 

The light and electron microscope exami- 
nations of the crop epithelium of Helix luco- 
rum revealed that it is surrounded by connec- 
tive tissue, muscular layers and a system of 
haemocoelic spaces surrounded by amoebo- 
cytes. The crop epithelium consists of four 
cell types: ciliated and unciliated columnar 
cells, mucous cells, and basal cells (Fig. 1). 

The ciliated and unciliated cells are colum- 
nar epithelial cells displaying similar morphol- 
ogy to each other. Ciliated cells are the most 
frequent cell type observed in the crop epithe- 
lium. As far as the electron microscope tech- 
niques permit, it is found that the ciliated and 
unciliated cells are gathered in groups or ir- 
regularly dispersed in the crop epithelium. 

The nucleus of both ciliated and unciliated 
cell types is usually in the middle portion of 
the cells (Figs. 1 , 2). Their cytoplasm contains 
small quantities of rough endoplasmic reticu- 
lum and a small number of Golgi complexes 
usually located in the supranuclear cyto- 
plasm. Numerous mitochondria are predomi- 
nantly in the apex of the cells (Fig. 2). The 
apical plasma membrane of the ciliated cells 
forms well-developed microvilli, and long cilia, 
displaying the "9 + 2" system of microtubules, 
are regularly dispersed among the microvilli 
(Figs. 2, 6). A well-developed microvillar bor- 
der, similar to that observed in the ciliated 
cells is also present in the unciliated cells 
(Figs. 1, 2). Both ciliated and unciliated cells 
possess moderate quantities of glycogen par- 
ticles, lipid inclusions, and electron-dense 
bodies in cytoplasmic areas adjacent to the 
nuclei. 

In certain cases, columnar cells showing 
similar morphologic characteristics but pre- 
senting a cytoplasm with a lower electron 
density than the adjacent ciliated and uncili- 
ated cells are apparent in crop epithelium. 

A characteristic feature of columnar cells is 
the presence of many electron-dense bodies 
in cytoplasmic areas adjacent to their nuclei 



CROP EPITHELIUM OF HELIX LUCORUM 



345 




MC 2 



FIG. 1. Drawing of the various cell types observed in the crop epithelium of Helix lucorum. ВС, basal cell; 
CC, ciliated cell; CT, connective tissue; MC^, mucous cell in various stages of their development; MS, 
muscles; UC, unciliated cell. 



(Figs. 2, 17). The dense bodies react posi- 
tively for periodate-reactive, sulfated and 
carboxylated glycoconjugates when the 
PA.TCH.SP, HID.TCH.SP and LID.TCH.SP 
cytochemical sequences are applied, respec- 
tively (Figs. 3, 4). Columnar cells display large 
numbers of glycogen particles that react 
strongly to periodate-reactive polysaccha- 
rides by the PA.TCH.SP technique (Fig. 3); 
they also contain lipid inclusions, usually lo- 
cated in the middle and basal portion of the 
cells, as well as a few infoldings of the basal 
plasma membrane. 

The third cell type, mucous cells, show a 
cytoplasm that is largely composed of mu- 
cous secretory granules (Figs. 1, 7). Mito- 
chondria and rough endoplasmic reticulum 
are dispersed throughout the remaining cyto- 
plasm, and numerous Golgi complexes (Fig. 
9) are frequently observed with their trans 
face in opposition to a mucous granule. The 
nucleus of these cells is usually situated in 
their middle portions. In many cases, the 
rough endoplasmic reticulum appears swol- 
len and contains crystalline-like material (Fig. 
8). Mucous granules are usually spherical in 
shape, are 1-2 u,m in diameter, and contain a 
fibrillar matrix without cores (Figs. 7, 8). 

Mucous cells are irregularly located across 
the epithelium. Mucous cells of different sizes 
and epithelial orientations from each other are 
observed in transverse sections of the crop 
epithelium (Fig. 1). The development of mu- 
cous cells seems to occur in following stages: 
(1) Spheroid or ovoidal mucous cell appear in 



the base of the epithelium via differentiation of 
basal cells. (2) The mucous cells increase in 
size and extend their dorsal side towards the 
apex of the epithelium. (3) The mucous cells 
further increase in size; their dorsal side is in 
close attachment to the luminal surface of the 
epithelium. (4) The mucous cells are trans- 
epithelially oriented. Their luminal surface is 
limited and their apical region is narrow. (5) 
The mucous cells are finally secreted into the 
crop lumen. 

At the same time, mucous cells showing 
all the above-mentioned morphologic stages 
are present in transverse sections of the crop 
epithelium. Mucous cells displaying a non- 
transepithelial orientation are also presented 
in the apex of crop epithelium. 

Mucous cells react positively to the 
PA.TCH.SP cytochemical method indicating a 
content rich in periodate-reactive polysaccha- 
rides (Figs. 9, 10). Most of the reaction prod- 
ucts are observed in a weblike structure within 
the matrix of the secretory granules. A strong 
PA.TCH.SP positive reaction is also found in 
the Golgi complexes and glycogen particles of 
the mucous cells (Fig. 9). The crystalline-like 
material inside the rough endoplasmic reticu- 
lum lack PA.TCH.SP reaction product (Fig. 
1 0). Often, mucous cells located near the apex 
of the epithelium display a large cisterna in 
their apical portion (Fig. 11); this cisterna dis- 
plays a negative reaction for carbohydrate and 
acid phosphatase (Fig. 11). 

Mucous granules react negatively to the 
HID.TCH.SP and LID.TCH.SP sequences 



346 



DIMITRIADIS ET AL. 







-■■-*»" 



Db 



Mi 



MG 



CROP EPITHELIUM OF HELIX LUCORUM 



347 



(Fig. 6), indicating absence of sulfated and 
carboxylated glycoconjugates. 

Finally, the fourth cell type, basal cells, are 
elongated to oval, and their ventral surfaces 
are attached to the basement membrane. 
Few mitochondria and little rough endoplas- 
mic reticulum are found in these cells. 

Control sections of all the cytochemical 
techniques used showed negative reactivity. 

Morphology of the Crop Epithelium of 
Starved and Hibernated Snails 

Snails starved for 40 days or hibernated for 
37 days presented large regions of crop epi- 
thelium where ciliated and unciliated cells 
were morphologically similar to those of con- 
trols. In the latter cells, however, a significant 
increase in the volume density of dense bod- 
ies in the hibernated snails (Fig. 13), as well 
as a significant decrease in the volume den- 
sity of lipid inclusions in both starved and hi- 
bernated snails was noted. The presence 
also of glycogen particles was less apparent 
in the experimental snails than in controls. 

The application of the PATCH. SP, 
HID.TCH.SP and LID.TCH.SP techniques in 
the starved and hibernated snails indicated a 
carbohydrate content of mucous cells similar 
to that identified in controls, that is, the pres- 
ence of periodate-reactive, non-sulfated and 
non-carboxylated polysaccharides was ap- 
parent in both experimental and control 
snails. 

Mucous cells in starved and hibernated 
snails significantly increased in numbers (Fig. 
12). In the experimental snails, the number of 
the luminal surface-oriented mucous cells 
was increased, while the mucous cells lo- 
cated at the base of the epithelium was de- 
creased compared to controls. In many 



cases, mucous cells were smaller in size 
compared to controls and were positioned in 
the apex of the epithelium, displaying a non- 
transepithelial orientation. The latter was 
more apparent in the hibernated snails. 

In the starved and hibernated snails, there 
were also other crop epithelial regions where 
increased lytic phenomena compared to con- 
trols were observed. The latter phenomena 
included lysis of certain cells (Fig. 15), as well 
as disorganization of apical cell regions (Fig. 
16) and extrusion of cytoplasmic regions into 
the crop lumen (Fig. 17). The lytic phenom- 
ena were more intense in the hibernated than 
in the starved snails. 



DISCUSSION 

The available data concerning the role of 
the crop epithelium in Pulmonata is inconsis- 
tent. According to Owen (1966), the crop is 
implicated in food storage and enzyme secre- 
tion. In addition, van Weel (1961) suggested 
the existence of extracellular digestion in the 
crop lumen by enzyme activity of variable or- 
igin. 

It is not clear, however, whether the en- 
zymes are secreted by the crop wall cells 
(Owen, 1966), transferred from the digestive 
gland (Hirsch, 1917; Fretter, 1952), or pro- 
duced by bacteria in the crop lumen (Jeu- 
niaux, 1954). In the present study, the colum- 
nar cells of the snail Helix lucorum did not 
show morphological features that support 
their extensive secretory function. Such char- 
acteristics would be, for example, large 
amounts of rough endoplasmic reticulum, as 
well as large number of Golgi complexes and 
secretory granules produced by the Golgi 
complexes. Therefore, the results of the 



FIG. 2. Transverse view of crop epithelium. In the apical portion of the ciliated (CC) and unciliated cells (UC) 
there are numerous mitochondria (Mi), while in their middle portion there are abundant lipid inclusions (Li). 
Note the similarity in the morphology of ciliated and unciliated cell. The asterisk shows a columnar cell 
presented less electron dense cytoplasm compared to the adjacent cells. Ci, cilia; Dbs, dense bodies; Lu, 
crop lumen; MC, mucous cell; N, nucleus. Bar = 3 |xm. 

FIG. 3. Detection of periodate-reactive polysaccharides (PATCH. SP technique). A positive PATCH. SP 
reaction is shown in the dense bodies (Db) and glycogen particles (arrow) in a columnar cell. Un-counter- 
stained section. Bar = 0.4 ц.т. 

FIG. 4. Detection of sulfated polysaccharides (HID.TCH.SP technique). A positive HID.TCH.SP reaction is 
shown in a dense body (Db) in a columnar cell. Un-counterstained section. Bar = 0.3 jxm. 
FIG. 5. Detection of acid phosphatase. A positive reaction for acid phosphatase indicates a dense body (Db) 
in a columnar cell. Un-counterstained section. Bar = 0.35 ц.т. 

FIG. 6. Detection of carboxylated polysaccharides (LID.TCH.SP technique). A negative LID.TCH.SP reac- 
tion is noticed in the mucous granules (MG) of a mucous cell. Note the adequate ability of the technique to 
stain the mitochondria (Mi) and the cilia (Ci) of a ciliated cell. Un-counterstained section. Bar = 1.5 ц.т. 



348 



DIMITRIADIS ET AL 




CROP EPITHELIUM OF HELIX LUCORUM 



349 



1.2 _ 



CD 
О 



00 

a 
С 

В 

CU 




Columnar cells 



Control 
^ Starved 
Щ Hibernated 



Mucous cells 



FIG. 12. Percentage (%) of the columnar (ciliated and unciliated cells) and mucous cells in the crop 
epithelium of H. lucorum. Significantly different values (P<0.05) between control and starved or control and 
hibernated snails are indicated by an asterisk. 



present study are consistent with the notion 
(see, for example, Oxford, 1977) that diges- 
tive activities in the crop of snails are probably 
mediated by digestive enzymes regurgitated 
from the digestive gland or the stomach, 
rather than by enzymes secreted by the crop 
cells. 

The presence of a large number of glyco- 
gen particles in the columnar cells indicates 
that the cytoplasm of these cells is a site of 
carbohydrate storage. Because other studies 
show cellulase and chitinase activity in the 
crop lumen of Helix (Jeuniaux, 1955; Stras- 
dine & Whittaker, 1963; Koopmans, 1967; 
Flari & Charrier, in press), it is possible that 
the columnar cells of crop epithelium of H. 
lucorum absorb oligosaccharides from crop 
lumen, part of which are stored as glycogen 
particles. In addition, the presence of a very 



well-developed microvillar border and numer- 
ous mitochondria in the apex of crop colum- 
nar cells of H. lucorum, as well as the pres- 
ence of moderate quantities of lipid inclusion 
in their cytoplasm, possibly indicates an ex- 
tensive absorptive function coexisting with 
their function as cells that propel mucus. In 
the snail Theba pisana, an absorptive func- 
tion is also suggested for the crop epithelial 
cells (Roldan & Garcia-Corrales, 1988). 

The dense bodies observed in the digestive 
cells of H. lucorum crop have been also found 
in the crop or intestinal columnar and mucous 
cells in different gastropods (pulmonates and 
prosobranchs) (Carhker & Blistad, 1946; 
Bowen, 1970; Boquist et al., 1971; Lufty & 
Demian, 1976; Ángulo et al., 1986). In the 
present study, the positive acid phosphatase 
reaction established the lysosomal origin of 



FIG. 7. Mucous granules (MG) containing a fibrillar matrix without cores inside a mucous cell. Bar = 2 ^m. 

FIG. 8. Crystalline-like material (asterisk) appears inside the swelled rough endoplasmic reticulum of a 

mucous cell. MG, mucous granule. Bar = 0.3 (im. 

FIG. 9. An intense PATCH. SP positive reaction show the saccules of the Golgi complexes (Go), the mucous 

granules (MG) and the glycogen particles (arrow) in a mucous cell. Un-counterstained section. Bar = 0.6 

p.m. 

FIG. 10. The granules of a mucous cell show a reticulated PATCH. SP positive reaction. Note the negative 

reaction on the crystaline-like inclusions inside the rough endoplasmic reticulum (arrow). Un-counterstained 

section. Bar = 0, 35 (xm. 

FIG. 1 1 . A mucous cell located near the apex of the epithelium displays PATCH. SP positive mucous 

granules and a large cisterna showing a negative reaction (asterisk). Mv, microvilli; N, nucleus. Bar = 3 p.m. 



350 



DIMITRIADIS ET AL 



С 

•ä 

в 

о 

> 




0.00 



Control 
Starved 

Hibernated 



Dense bodies 



Lipid inclusions 



FIG. 13. Volume density of dense bodies and lipid inclusions in the supranuclear cytoplasm of columnar cells 
(ciliated and unciliated cells). Significant different values (P<0.05) between control and starved or control 
and hibernated snails are indicated by an asterisk. 



these structures. A positive acid phosphatase 
reaction was also observed in apical vacuoles 
and granules of the crop and intestinal colum- 
nar cells, and in all the intestinal gland cells of 
the slug Arion ater (Bowen, 1970; Ángulo et 
al., 1986). The lysosomal origin of the dense 
bodies of crop digestive cells of H. lucorum is 
additionally supported by the positive 
PA.TCH.SP, HID.TCH.SP and LID.TCH.SP 
reactions, indicating the presence of period- 
ate-reactive, sulfated and carboxylated glyco- 
conjugates, respectively. The association of 
the latter reactions with lysosomal activities is 
also referred to in studies on mammalian leu- 
kocytes (Bruyn et al., 1975, Spicer et al., 
1978), as well as in studies on H. lucorum 



digestive gland cells (Dimitriadis & Liosi, in 
press). 

There is little information about carbohy- 
drate histochemistry and cytochemistry of the 
molluscan digestive epithelium. In the slug 
Arion ater (Ángulo et al., 1986), intestinal 
gland cells showed either a positive reaction 
for acid and sulphated mucosubstances or a 
positive one for neutral polysaccharides. In 
the molluscs Semerula macúlala (Varute & 
Patil, 1971) and Venus mercenaria (Zacks, 
1965), sulfated acid and neutral polysaccha- 
rides, respectively, were found in their intes- 
tinal mucocytes. The mucous cells of H. luco- 
rum crop epithelium, as the present study 
indicates, were positive to the PA.TCH.SP re- 



FIG. 14. 25 days of starvation. Numerous dense bodies (Dbs) are located in the apical portion of certain 

columnar cells. Bar = 1.9 p.m. 

FIG 15. 40 days of starvation. A crop epithelial cell in a stage of its lysis. N, nucleus Bar = 1 p.m. 

FIG. 16. 37 days of hibernation. Cisternae with membranic materials are located in the apex of a ciliated 

cells. Bar = 1 p.m. 

FIG. 17. 37 days of hibernation. A columnar cell (asterisk) is extruded into the crop lumen. This phenomenon 

is more apparent in the experimental snails than in controls. Note the numerous dense bodies (arrows) in 

the supranuclear cytoplasm of two columnar cells. Mi, mitochondria; Mv, microvilli; N, nucleus. Bar = 1.5 

p.m. 



CROP EPITHELIUM OF HELIX LUCORUM 



351 



* 







*5i 




w%AObs 



.4Hr5jp* 



^rt 



• 




' ■ л 



% . • г S 



Mv 



f 



■ 




352 



DIMITRIADIS ET AL 



action and negative to both HID.TCH.SP and 
LID.TCH.SP sequences, indicating periodate- 
reactive glycoconjugates, which lack sulfated 
and carboxylated esters. However, it is pos- 
sible that the polysaccharides of mucous cells 
in H. lucorum crop epithelium contain hyalu- 
ronic and sialic acid, which are stained by al- 
cian blue (pH = 2.5), as was demonstrated in 
mucocytes in the intestinal gland cells of the 
slug Arion ater (Ángulo et al., 1986). 

Morphology of the Crop Epithelium of 
Starved and Hibernated Snails 

The existing data on morphological 
changes in the fine structure of the crop epi- 
thelial cells of Pulmonata under starvation or 
hibernation is not sufficient to give a clear in- 
sight into the physiology of the cells under 
these conditions. There are reports indicating 
biochemical and physiological differences in 
enzyme secretion, changes in the concentra- 
tion of the components of extracellular body 
fluids (Meenakshi, 1956), changes in the os- 
motic pressure and respiratory rate (Ghiretti, 
1966), and changes in major metabolic prod- 
ucts (Florkin, 1966) due to hibernation or star- 
vation. 

The light and electron microscope observa- 
tions of the crop cells of H. lucorum after 40 
days of starvation and 37 days of hibernation 
showed a significant increase in number of 
the dense bodies located near the nuclei of 
the columnar cells compared to controls. This 
increase probably reflects increased lysoso- 
mal activity and/or increased intracellular di- 
gestion of nutrients by these cells during star- 
vation and hibernation. 

The increased lytic phenomena and extru- 
sions of cytoplasmic regions into the crop lu- 
men of the snails H. lucorum during starvation 
and hibernation compared to controls are 
probably related to the reduction of the en- 
ergy requirements and the maintenance of 
low function activity by the crop epithelial cells 
during the experimental periods. In contrast to 
our observations, autophagic vacuoles were 
formed in tissues of other organisms under 
starvation (Bowen, 1968; Bauer et al., 1977) 
that were related to cell autophagy. The non- 
appearance of autophagic vacuoles in the 
crop cells of H. lucorum during starvation and 
hibernation probably indicates the snail's ad- 
aptation to these conditions. 

Another effect that starvation and hiberna- 
tion induced in the crop epithelium of H. luco- 
rum was the increase in the number of mu- 



cous cells in the starved and hibernated 
snails compared to controls. It is uncertain 
whether the increase in the number of mu- 
cous cells in the starved and hibernated 
snails is a result of increased mucous cell de- 
velopment or of the accumulation of mucous 
cells in the crop epithelium, reflecting de- 
creased mucous cell secretion due to starva- 
tion and hibernation. 

In conclusion, by using light and electron 
microscopic observations, in combination 
with cytochemical characterization and mor- 
phometric evaluation, the present study pro- 
vides information about the effect of starva- 
tion and hibernation on the cells of the crop 
epithelium of the snail H. lucorum. The results 
of the present study should be considered as 
preliminary. Additional information, especially 
by the use of cytochemical techniques, is 
needed in order to clarify the physiology of 
snails during such conditions as hibernation 
and starvation. 



ACKNOWLEDGMENTS 

This work was financially supported by the 
Greek Ministry of Agriculture. We are grateful 
to Dr. M. Lazaridou-Dimitriadou for the provi- 
sion of the snails and their keep under nor- 
mal, starvation and hibernation conditions. 



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LUFTY, R. G. & E. S. DEMIAN, 1976, The histology 
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354 



DIMITRIADIS ET AL. 



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Revised Ms. accepted 9 March 1992 



MALACOLOGIA, 1992,34(1-2): 355-369 

GENETIC SIMILARITIES AMONG CERTAIN FRESHWATER MUSSEL 
POPULATIONS OF THE LAMPSIUS GENUS IN NORTH CAROLINA 

Alan E. Stiven 1 and John Alderman 2 



ABSTRACT 

The objective of this research was to explore ecological and genetic similarities among several 
southeastern freshwater mussel populations, focusing especially on two rare and potentially 
endangered "subspecies" of Lampsilis radiata residing in river systems in the Piedmont and 
coastal plain of North Carolina. Conchological analyses and the examination of genetic identity 
patterns derived from 1 1 electrophoretic loci in six different populations or species formed the 
basis of our interpretations. 

Two currently recognized subspecies of Lampsilis radiata (radiata and conspicua) are now 
quite rare in North Carolina waters, and one, L. r. conspicua, may be endemic to the state. 
However, we find that in spite of the difference in unadjustable sample mean shell length, they 
do not differ in general shell morphometry. For example, covariance analyses indicated homo- 
geneity of shell length-height regression coefficients, and no difference in adjusted mean 
lengths. A similar conchological comparison of these two "subspecies" with the smaller sized 
recently described Lampsilis fullerkati of Lake Waccamaw also failed to find any significant 
conchological differences among these populations. 

Nei's unbiased genetic distance and Rogers modified genetic distance derived from electro- 
phoretic analyses consistently separated on established Ellipito, Leptodea and Lampsilis spe- 
cies. However, these genetic distance measures did not distinguish between the two rare Lamp- 
silis radiata "subspecies" nor between these two groups and Lampsilis fullerkati: Nei's unbiased 
identity ranged from 0.945 to 0.978 among these three populations. Phenograms and a Wagner 
tree derived from Rogers distance supported evolutionary similarities among these three pop- 
ulations. 

We also found evidence of site effects (conchological differences in different habitats) in 
Lampsilis cariosa and Leptodea ochracea, the two species examined for such a phenomenon. 
Genetic identity values among these intraspecific but allopatric populations ranged from 0.922- 
0.982. We suggest that until contrasting genetic and distinctive biological data are forthcoming, 
the two "subspecies" of L. radiata and possibly L. fullerkati simply be considered as a multiple 
population complex of Lampsilis radiata. 

Key words: freshwater mussel, Lampsilis radiata, ecology, genetics, North Carolina, genetic 
distance, phenogram, Wagner tree. 



INTRODUCTION 

Freshwater bivalves constitute a large and 
very diverse assemblage of species in North 
Carolina and other southeastern states (R.I. 
Johnson, 1970; Burch, 1975; Davis, 1984). 
Many of these mussels and clams are highly 
vulnerable to pollutants, and changes in their 
abundance, survival and reproductive status 
can be a warning signal of the declining 
health of an estuary or river system (Goldberg 
et al., 1978). Because many freshwater mus- 
sels in North Carolina are already in a state of 
decline (Clarke, 1983), it is imperative that 
work focus on the assessment of potentially 
rare and endangered species of freshwater 
mussels, especially in the genera Lampsilis, 



Lasmigona, Elliptic*, and Alasmidonta (Clarke, 
1983; Keferl & Shelley, 1988; Alderman, 
1988). The development and use of the "rare 
and endangered" classification, however, has 
been hampered by insufficient ecological, ge- 
netic and systematic information, as well as 
confusion even over the species concept in 
this group. Many early mussel investigators 
relied on shell morphology for distinguishing 
species, but this approach is often suspect, 
especially in light of the extensive concholog- 
ical divergence in this group (Davis, 1984). 

The freshwater mussel genus Lampsilis is 
probably one of the most successful and spe- 
cialized groups of the Unionidae (Davis, 
1984), with more than 20 living species in 
North America. At least five of these species 



department of Biology, University of North Carolina, Chapel Hill, North Carolina 27599-3280, U.S.A. 

2 Nongame Program, North Carolina Wildlife Resources Commission, Raleigh, North Carolina 27604-1188, U.S.A. 



355 



356 



STIVEN & ALDERMAN 



are found in North Carolina waters and some 
may be endemic to the state. Currently, the 
American Malacological Union recognizes 
two subspecies of Lampsilis radiata (Gmelin, 
1791), L r. radiata (eastern lampmussel) and 
L r. conspicua (I. Lea, 1872) (Carolina fat- 
mucket) (Turgeon et al., 1988). The eastern 
lampmussel is found largely in coastal plain 
waters and occupies a broad geographic 
range from the Pee Dee drainage north to the 
St. Lawrence River. Collections of this "sub- 
species" in North Carolina waters during the 
past four years include sites in the Chowan 
River, Swift Creek, Fishing Creek and the 
lower main channel of the Tar River (Clarke, 
1983; Alderman, 1988). Shelley (1987) lists 
its occurrence in the Tar, Neuse, and Cape 
Fear rivers. The second "subspecies," the 
very rare Carolina fatmucket, is apparently 
found only in North Carolina and may be en- 
demic to this state. It has been collected in the 
tributaries of the Pee Dee River drainage (e.g. 
the Little and Uwharrie rivers), and recently in 
the Flat River of the upper Neuse River drain- 
age basin (Alderman, 1988, 1989). In prior 
collections of these two "subspecies," L r. 
radiata often exhibited smaller shells (Al- 
derman, 1989). Lampsilis r. radiata is also 
found in waters containing anadromous fish, 
and its larval glochidia may be parasitic on 
such fish. If these two populations are suffi- 
ciently different to warrant "species" status, 
then the rarer L r. conspicua is a prime can- 
didate for federal listing as "endangered." 

This study utilized biochemical and popula- 
tion genetic methods (Rogers, 1972; Davis et 
al., 1981 ; Davis, 1984) to probe several ques- 
tions about the genetic relatedness among 
certain North Carolina mussel populations. 
Our prime question dealt with ecological and 
genetic differences in the two Lampsilis radi- 
ata "subspecies." In addition, we explored 
three other questions within this genus. The 
first focused on the recent discovery of a 
small population of a Lampsilis-Wke species in 
the Deep River near its confluence with the 
Cape Fear River. This population is com- 
prised of only large old specimens and is 
thought to be declining. Clarification of its ge- 
netic structure and possible systematic status 
is required (Alderman, 1990). The second 
question related to the genetic and ecological 
status of Lampsilis cariosa (Say, 1817). 
Mostly large L. cariosa are now found in sites 
formerly containing both young and adults 
(e.g. Tar and upper Neuse drainages) (Al- 
derman, personal observation), and its level 



of genetic heterozygosity is unknown. Finally, 
Lampsilis fullerkati R. I. Johnson, 1984, a 
Lake Waccamaw species, was first discussed 
by Kat (1983) and then described as a new 
species by R. I. Johnson (1984). Kat (1983) 
noted that the genetic structure of this species 
was similar to Lampsilis radiata from New 
Brunswick and Nova Scotia. To help clarify 
the systematic status of Lampsilis fullerkati, 
we compared its genetic structure with North 
Carolina Lampsilis radiata specimens as well 
as other Lampsilis species. 



METHODS 
Study Sites and Collections 

The description of all collection sites, as 
well as the species/populations and numbers 
of mussels collected, along with their ANSP 
catalog number (except the two rare Lampsi- 
lis radiata subspecies as noted below), are 
given in Figure 1 and Appendix I. 

Collections of mussels were made from 
shallow water by hand and from deeper areas 
by snorkeling and SCUBA. Mussel species of 
prime interest to this study were generally too 
sparse to use quadrat sampling to estimate 
relative abundance. Specimens of the two 
rare "subspecies," L. r. radiata and L. r. con- 
spicua, were returned to their original sites 
following shell measurements and the re- 
moval of a small tissue sample from the man- 
tle for use in genetic analysis. Individuals of 
all other mussel species collected were sac- 
rificed, tissue samples removed, shell mea- 
surements obtained, and shell and remaining 
soft body tissue preserved. Shell measure- 
ments included maximum length, height, and 
width. Leptodea ochracea (Say, 1817), for- 
merly placed in Lampsilis, was identified as 
such by the lack of a distinct mantle flap (this 
flap is located ventral to the incurrent aperture 
in Lampsilis). 

Genetic Methods 

All animals were processed within 10 days 
of collection. Shells were pried open and tis- 
sue samples of mantle and liver (hepatopan- 
creas) were removed, combined and pro- 
cessed, except in the case of the rare 
"subspecies" as noted above. Tissues were 
chopped on cold glass with a razor blade, 
then homogenized (sonified) in an equal vol- 
ume of grinding buffer (0.1 M Tris, 0.001 M 



LAMPSILIS IN NORTH CAROLINA 



357 




FIG. 1. Map of eastern North Carolina showing major river systems and the location of the collection sites 
(A-G) for this study. See Appendix I for details of each site. 



EDTA, and 5 x 10 5 M NADP, pH 7.0) for 
about 20-50 sec. The sample was then cen- 
trifugea" for 30 min at 12,000 g. Sample ex- 
tracts were stored at -70°C. Protein extracts 
were then analyzed by horizontal starch gel 
electrophoresis (Electrostarch, Lot 89) follow- 
ing procedures outlined by Shaw & Prasdad 
(1970), Selander et al. (1971), Davis et al. 
(1981), Kat (1983), Davis (1984), and Stiven 
(1989). The 11 enzyme loci used (Table 1) 
correspond to most of the loci utilized by oth- 
ers in molluscan genetic research (e.g. Davis 
et al., 1 981 ; Kat, 1 983). The numbering of loci 
and lettering of alleles were from the most 
anodal region back to the origin on the gel. 
Sample tissues of Elliptio complanata (Light- 
foot, 1 786) and Leptodea ochracea were run 
with the other species and subspecies to pro- 
vide allelic consistency. Aminopeptidase 1 
and 2 {Ala-1 and Ala-2) are associated with 
the substrate leucyl-alanine. Leucine ami- 
nopeptidase (Lap) or a-aminoacyl-peptide hy- 
drolase (cytosol) was resolved using L-leucyl- 
ß-naphthylamide and fast black К salt. The 
position of this locus differed from the Ala loci 
on the same gel. 

The genetic data were analyzed using 
Swofford & Selander's (1981) BIOSYS-1 
computer program. P, the proportion of loci 



that are polymorphic under the 0.95 criterion, 
and H, the direct-count heterozygosity, were 
measures of genetic variability. Levels of het- 
erozygote deficiency were also estimated by 
D = (H -H E )/H E , where H and H E are the 
observed and expected frequencies of het- 
erozygotes respectively (Koehn et al., 1973). 
Departure of genetic frequencies from Hardy- 
Weinberg expectations were assessed by 
chi-square analysis adjusted for small sam- 
ple sizes (Levene, 1949). All statistical anal- 
yses were carried out using SYSTAT (Wilkin- 
son, 1990). 

Genetic distance/similarity among species- 
populations were measured by Nei's nonmet- 
ric unbiased genetic distance and genetic 
identity (Nei, 1972, 1978) and Rogers metric 
modified distance (Rogers, 1972; Wright, 
1978). Each has its specific assumptions and 
limitations, but all are related to F ST (Wright's 
measure of genetic divergence-Wright, 1978; 
Hartl & Clark, 1989). Nei's unbiased mea- 
sures are less sensitive to small sample sizes 
than are Nei's biased measures (Nei, 1978). 
There is no upper bound on the magnitude of 
Nei's genetic distance measure. We note also 
that when distance measures are relatively 
large between pairs of species, and heterozy- 
gosity is low, the construction of phenograms 



358 



STIVEN & ALDERMAN 



TABLE 1. Isoenzymes and buffers utilized in the genetic analysis of freshwater mussels from North 
Carolina waters. 



Locus/lsoenzyme (No. Loci) 



Abbrev. 



E.C. No. 



Buffer 



Phosphoglucose isomerase (1) 
Phosphoglucomutase (2) 
Peptidase (2) 

Leucine aminopeptidase (1) 
Malate dehydrogenase (1) 
Phosphogluconate dehydrogenase (1) 
Mannose 6-phosphatase isomerase (1) 
a-glycerophosphate dehydrogenase (1) 
Superoxide dismutase (1) 



Gpi 


5.3.1.9 


LiOH 1 


Pgm 


2.7.5.1 


LiOH 


Ala 


3.4.1.2 


LiOH 


Lap 


3.4.11.1 


LiOH 


Mdh 


1.1.1.37 


CM-6 2 


6-Pgd 


1.1.1.43 


CM-6 


Mpi 


5.3.1.8 


CM-6 


a-Gpdh 


1.1.1.8 


TEB-9 3 


Sod 


1.15.11 


TEB-9 



1 Selander et al. (1971) 

2 8.4 g citric acid per liter, to pH 6 with N-(3aminopropyl) morpholine. Gel buffer 1 part electrode: 19 parts water. 

3 Davisetal. (1981) 



can be carried out fairly reliably with only a 
few representative individuals for a species 
(Nei, 1978). Phenograms were constructed 
for Nei's unbiased genetic distance (Nei, 
1978) and for modified Rogers genetic dis- 
tance (Rogers, 1972; Wright, 1978) using the 
unweighted pair group method of construction 
(Sneath & Sokal, 1973). A Wagner tree phe- 
nogram (Farris, 1972; Felsenstein, 1983) was 
also constructed using Rogers modified met- 
ric distance coefficient (Wright, 1978), with 
the tree rooted at the midpoint of the greatest 
patristic distance. Roger's metric measure 
satisfies the Wagner requirement of triangle 
inequality. This method does not assume 
constant evolutionary rates (Felsenstein, 
1983). 



RESULTS 

Ecological and Shell Parameters 

Only nine individuals of the rare L. r. radiata 
were obtained during many searches (by 
hand during wading, and SCUBA in deeper 
waters), some lasting many hours. Most were 
found in fine to medium sand and organic silt 
substrates. In the lower Chowan River site, 
Leptodea ochracea was about three times as 
abundant as L. r. radiata. In contrast, a 40- 
min search on 5-17-90 by two persons in the 
South Flat River yielded 40 Elliptio compla- 
nata and seven individuals of other species. 
Lampsilis r. conspicua, also rare, was ob- 
tained from the Flat River in Durham County. 
This subspecies was found primarily in 
coarse sand especially along sand bars. 

Figure 2 illustrates the shell length vs. 



height regressions for L r. conspicua and L. r. 
radiata. Although these "subspecies" differed 
significantly in mean shell length (F., 30 = 
29.5, P<0.0001) (Table 2), covariance anal- 
ysis indicated that the length-height regres- 
sion coefficients did not differ (F, 28 = 0.61 , P 
= 0.44), suggesting similar relative concholo- 
gies. Shell length, adjusted for differences in 
shell height, was 105.7 mm and 104.1 mm for 
L r. conspicua and L. r. radiata respectively, 
and this difference is not significant (F., 28 = 
0.63, P = 0.43). Shells of L. fullerkati from 
Lake Waccamaw were considerably smaller 
than the two subspecies (Table 2). However, 
covariance analysis of length-height regres- 
sions among L. r. conspicua, L. r. radiata, and 
L. fullerkati indicated no significant differ- 
ences (F 2 61 = 0.32, P = 0.49), and adjusted 
mean lengths also did not differ among these 
three Lampsilis populations (F 2 63 = 2.72, P 
= 0.073). In contrast, shell length-height re- 
gression comparisons among all combina- 
tions of Elliptio complanata, Lampsilis cari- 
osa, and Leptodea ochracea were significant. 
Leptodea ochracea and Lampsilis cariosa 
showed evidence of the "site effects" that 
arise from conchological differences in popu- 
lations of the same species drawn from differ- 
ent sites (Kat, 1982; Hinch et al., 1986), or 
conversely, conchological similarities among 
different species found in the same site (Horn 
& Porter, 1981). In L. cariosa, mean shell 
length was larger (F 2 31 = 46, P<0.001 ; and 
Tukey test) at the Deep River site (1 1 6.4 mm) 
than at the Cape Fear (60.6 mm) or Tar River 
(61 .1 mm) sites. Similarly, Leptodea ochracea 
was much larger in the Chowan River (92.5 
mm) than in Lake Waccamaw (37.6 mm) (F., 72 
= 847, P<0.0001 ; and Tukey test). 



LAMPSILIS IN NORTH CAROLINA 



359 



150 



130 - 



110 



90 - 



70 - 



50 - 



30 

10 20 30 40 50 60 70 80 90 100 

HEIGHT (mm) 

FIG. 2. Plot of shell length against shell height (mm) for samples of the freshwater mussels, Lampsilis radiata 
conspicua (open circles) and L. r. radiata (closed circles) from North Carolina waters. Refer to Appendix I for 
collecting sites. 



E 
E 



O 

z 

Ш 



г 


I 


I I 




I 


о 
о 


i 


- 


- 










&> 


© 




- 








fr 








- 




o° 








- 






• 














• 


• 






Lr.c. 


о 




I 


I 


I I 




I 


L.r.r. 

i i 


• 

I 





TABLE 2. Summary of shell measurements for the freshwater mussels Lampsilis radiata radiata, 
Lampsilis radiata conspicua, and Lampsilis fullerkati from collections taken in North Carolina waters, 
1990. Values are means ± SE. 



Shell Parameters 



L. r. conspicua 



L. r. radiata 



L. fullerkati 



Sample Size 
Length 
Height 
Width 



23 

113.0 ± 2.88 

64.6 ± 1 .65 

40.2 ± 1 .67 



83.1 ± 4.87 
46.4 ± 2.57 
30.0 ± 2.27 



35 
47.1 ± 0.89 
23.5 ± 0.44 
15.5 ±0.25 



Genetic Analyses 

Diversity and Heterozygosity: Alleleic fre- 
quencies and measures of genetic diversity 
derived from the 11 loci for all populations/ 



species/subspecies are given in Table 3. Av- 
erage direct-count heterozygosity and P- 
values were high in the more common and 
wide-spread species, Elliptio complanata and 



360 



STIVEN & ALDERMAN 



TABLE 3. Alleleic frequencies, sample sizes, and locus-specific direct-count heterozygosity, along with 
percentage of loci polymorphic (P at the 0.95 criterion), and mean direct-count heterozygosity (H ± 1 
S.E.) for the nine populations/species of freshwater mussels from North Carolina waters. 



Population 1 



Locus 



PGI 

A 

В 

С 

D 

E 

F 

G 

H 

(N) 

H (de) 

PGM-1 

A 

В 

С 

D 

E 

F 

(N) 

H (de) 

PGM-2 

A 

В 

С 

D 

E 

F 

(N) 

H (de) 

ALA-1 

A 

В 

С 

D 

E 

F 

(N) 

H (de) 

ALA-2 

A 

В 

С 

(N) 

H (de) 

LAP 

A 

В 

С 

D 

E 

F 

G 

H 

I 

J 

(N) 

H (de) 



0.000 
0.000 
0.000 
0.000 
0.067 
0.767 
0.078 
0.089 
45 
0.133 

0.011 
0.744 
0.044 
0.189 
0.011 
0.000 
45 
0.356 

0.000 
0.044 
0.944 
0.000 
0.011 
0.000 
45 
0.067 

0.000 
0.011 
0.067 
0.744 
0.122 
0.056 
45 
0.244 

0.000 
0.274 
0.726 
42 
0.244 

0.022 
0.678 
0.011 
0.133 
0.011 
0.000 
0.011 
0.067 
0.000 
0.067 
45 
0.333 



0.125 
0.833 
0.000 
0.000 
0.042 
0.000 
0.000 
0.000 
12 
0.333 

0.000 
0.000 
0.167 
0.625 
0.167 
0.042 
12 
0.417 

0.042 
0.000 
0.792 
0.000 
0.167 
0.000 
12 
0.250 

0.000 
0.792 
0.208 
0.000 
0.000 
0.000 
12 
0.417 

0.000 
0.625 
0.375 
12 
0.583 

0.000 
0.000 
0.000 
0.000 
0.000 
0.375 
0.292 
0.083 
0.000 
0.250 
12 



0.000 0.294 

0.929 0.706 

0.000 0.000 

0.000 0.000 

0.071 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

7 17 
0.143 .0.235 

0.000 0.000 

0.000 0.000 

0.214 0.147 

0.571 0.559 

0.000 0.294 

0.214 0.000 

7 17 

0.286 0.471 

0.000 0.029 

0.000 0.000 

0.857 0.706 

0.000 0.000 

0.071 0.265 

0.071 0.000 

7 17 

0.286 0.471 

0.000 0.000 

0.714 0.676 

0.214 0.324 

0.071 0.000 

0.000 0.000 

0.000 0.000 

7 17 

0.429 0.529 

0.000 0.059 

0.643 0.647 

0.357 0.294 

7 17 

0.714 0.294 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.250 

0.071 0.313 

0.571 0.250 

0.214 0.125 

0.071 0.063 

0.071 0.000 

0.000 0.000 

7 16 



0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.316 0.718 

0.000 0.000 

0.000 0.000 

0.684 0.282 



19 
0.526 



55 
0.309 



0.667 0.857 



0.375 



0.000 0.000 

0.000 0.000 

0.000 0.000 

0.974 1 .000 

0.026 0.000 

0.000 0.000 

19 55 

0.053 

0.000 0.000 

1 .000 0.964 

0.000 0.027 

0.000 0.000 

0.000 0.000 

0.000 0.009 

19 55 

0.073 

0.053 0.055 

0.947 0.936 

0.000 0.000 

0.000 0.009 

0.000 0.000 

0.000 0.000 

19 55 

0.018 

0.000 0.036 

1 .000 0.964 

0.000 0.000 

19 55 



0.000 0.000 

0.079 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.921 1.000 

19 54 

0.158 



0.086 
0.000 
0.729 
0.000 
0.186 
0.000 
0.000 
0.000 
35 
0.314 

0.000 
0.000 
0.000 
0.814 
0.171 
0.014 
35 
0.314 

0.000 
0.000 
1.000 
0.000 
0.000 
0.000 
35 


0.000 
0.229 
0.029 
0.714 
0.000 
0.029 
35 
0.286 

0.000 
1.000 
0.000 
35 


0.000 
0.000 
0.000 
0.000 
0.000 
0.000 
1.000 
0.000 
0.000 
0.000 

1 





0.000 0.000 

0.000 0.000 

0.935 1 .000 

0.000 0.000 

0.065 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

23 9 

0.130 

0.000 0.000 

0.000 0.000 

0.000 0.000 

1 .000 1 .000 

0.000 0.000 

0.000 0.000 

23 9 

о 

0.043 0.000 

0.022 0.000 

0.935 1 .000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

23 9 

0.130 

0.000 0.000 

0.457 0.722 

0.043 0.000 

0.500 0.278 

0.000 0.000 

0.000 0.000 

23 9 

0.391 0.333 

0.348 0.000 

0.652 1 .000 

0.000 0.000 

23 9 



0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.000 0.000 

0.109 0.000 

0.761 1.000 

0.022 0.000 

0.000 0.000 

0.109 0.000 

23 9 

0.087 



LAMPSIUS IN NORTH CAROLINA 



361 



TABLE 3. (Continued) 













Population 1 










Locus 


1 


2 


3 


4 


5 


6 


7 


8 


9 


MDH 




















A 


0.000 


0.000 


0.000 


0.000 


0.158 


0.009 


0.000 


0.000 


0.000 


В 


0.012 


0.167 


0.000 


0.000 


0.842 


0.991 


0.214 


0.000 


0.167 


С 


0.628 


0.833 


1.000 


1.000 


0.000 


0.000 


0.000 


0.000 


0.000 


D 


0.360 


0.000 


0.000 


0.000 


0.000 


0.000 


0.786 


1.000 


0.833 


(N) 


43 


12 


7 


17 


19 


55 


35 


23 


9 


H(dc) 


0.023 











0.316 


0.018 


0.429 





0.333 


6-PGD 




















A 


0.000 


0.042 


0.000 


0.059 


0.000 


0.000 


0.000 


0.000 


0.000 


В 


0.000 


0.000 


0.143 


0.000 


0.000 


0.009 


0.000 


0.000 


0.000 


С 


0.000 


0.917 


0.786 


0.941 


0.000 


0.000 


0.686 


0.413 


0.500 


D 


0.000 


0.000 


0.000 


0.000 


1.000 


0.973 


0.000 


0.000 


0.000 


E 


0.326 


0.042 


0.071 


0.000 


0.000 


0.018 


0.314 


0.587 


0.500 


F 


0.674 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


(N) 


43 


12 


7 


17 


19 


55 


35 


23 


9 


H (de) 


0.372 


0.167 


0.143 


0.118 





0.018 


0.514 


0.391 


0.556 


MPI 




















A 


0.023 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


0.000 


В 


0.682 


0.000 


0.000 


0.000 


0.000 


0.018 


0.000 


0.000 


0.056 


С 


0.295 


0.792 


0.000 


0.000 


0.974 


0.900 


0.029 


0.348 


0.389 


D 


0.000 


0.000 


0.000 


0.324 


0.000 


0.000 


0.000 


0.022 


0.000 


E 


0.000 


0.208 


1.000 


0.676 


0.026 


0.082 


0.971 


0.630 


0.556 


(N) 


44 


12 


7 


17 


19 


55 


35 


23 


9 


H (de) 


0.068 


0.250 





0.647 


0.053 


0.127 





0.304 


0.556 


a-PGD 




















A 


0.000 


1.000 


1.000 


0.765 


1.000 


1.000 


1.000 


0.870 


1.000 


В 


1.000 


0.000 


0.000 


0.235 


0.000 


0.000 


0.000 


0.130 


0.000 


(N) 


45 


12 


7 


17 


19 


55 


35 


23 


9 


H (de) 











0.353 











0.087 





SOD 




















A 


0.000 


0.000 


0.000 


0.000 


1.000 


1.000 


0.000 


0.000 


0.000 


В 


1.000 


1.000 


1.000 


1.000 


0.000 


0.000 


1.000 


1.000 


1.000 


(N) 


45 


12 


7 


17 


19 


55 


35 


23 


9 


H (de) 





























P 


81.8 


81.8 


63.6 


81.8 


36.4 


27.3 


45.5 


72.7 


36.4 


H ± 


0.169 


0.280 


0.260 


0.318 


0.100 


0.051 


0.169 


0.138 


0.162 


S.E. 


0.044 


0.070 


0.090 


0.064 


0.052 


0.028 


0.061 


0.046 


0.071 



U-Elliptio complanata, S. Fiat River, Durham Co. 
2-Lampsilis cariosa, Cape Fear River, Cumberland Co. 
3-Lampsilis cariosa, Tar River, Nash Co. 
A-Lampsilis cariosa, Deep River, Moore Co. 
5-Leptodea ochracea, Chowan River, Gates Co. 
6-Leptodea ochracea, Lake Waccamaw, Columbus Co. 
7-Lampsilis fullerkati, Lake Waccamaw, Columbus Co. 
8-Lampsilis radiata conspicua, Flat River, Durham Co. 
9-Lampsilis radiata radiata, Chowan River, Gates Co. 



Lampsilis cariosa. Lower heterozygosity esti- 
mates are associated with the two rare Lamp- 
silis radiata "subspecies," Lampsilis fullerkati 
and Leptodea ochracea, which exhibited a 
very low heterozygosity level. 

Elliptio complanata from the South Flat 
River showed the largest number of polymor- 
phic loci deviating from Hardy-Weinberg ex- 
pectation (six of nine) (0.95 criterion for poly- 



morphic loci), whereas Leptodea ochracea 
from Lake Waccamaw had three of four. 
Lampsilis r. conspicua had three of eight, and 
L. r. radiata none. All significant departures 
came from deficiencies in heterozygotes. For 
example, in E. complanata, "D" ranged from 
-0.36 to -0.95 among loci, Leptodea ochra- 
cea from -0.66 to - 1 .0, and in L. r. conspicua 
from -0.21 to -1.0. 



362 



STIVEN & ALDERMAN 



TABLE 4. Matrix of Nei (1978) unbiased genetic identity (below diagonal) and unbiased genetic distance 
(above diagonal) for the nine populations/species of freshwater mussels from North Carolina waters. 



Population/Species 


1 


2 


3 


4 


5 


6 


7 


8 


9 


1 . Elliptio complanata 


***** 


0.853 


0.902 


0.825 


2.138 


2.179 


0.892 


0.809 


0.927 


2. Lampsilis cariosa-Cape Fear 


0.426 


***** 


0.082 


0.090 


0.726 


0.729 


0.420 


0.414 


0.345 


3. Lampsilis cariosa-Tar R. 


0.406 


0.921 


***** 


0.041 


1.111 


1.086 


0.350 


0.422 


0.386 


4. Lampsilis cariosa-Deep R. 


0.438 


0.914 


0.959 


***** 


1.146 


1.140 


0.404 


0.477 


0.435 


5. Leptodea ochracea-Chowan R. 


0.118 


0.484 


0.329 


0.318 


***** 


0.018 


1.049 


0.963 


0.823 


6. Leptodea ochracea-L. Wacc. 


0.113 


0.483 


0.338 


0.320 


0.982 


***** 


1.004 


0.943 


0.819 


7. Lampsilis fullerkati 


0.410 


0.657 


0.704 


0.668 


0.350 


0.366 


***** 


0.056 


0.049 


8. L. radiata conspicua 


0.445 


0.661 


0.655 


0.621 


0.382 


0.390 


0.945 


***** 


0.022 


9. L. radiata radiata 


0.396 


0.708 


0.680 


0.647 


0.439 


0.441 


0.952 


0.978 


***** 



Genetic Relatedness Among and Within the 
Populations: Genetic relatedness among the 
nine populations was explored through pair- 
wise associations using Nei's (1978) unbi- 
ased genetic distance and identity measures 
(non metric) (Table 4) and Rogers modified 
distance metric measure (Rogers, 1972; 
Wright, 1978) (Table 5). Some of the greatest 
distances were found between species of the 
two genera, Elliptio and Lampsilis, confirming 
prior taxonomic designations based largely 
upon morphological work. Notably, some of 
the smallest distance values occurred be- 
tween L. r. conspicua and L. r. radiata, the two 
rare subspecies. In addition, these rare sub- 
species showed corresponding small dis- 
tance values with another species, L. fuller- 
kati, a mussel first collected from Lake 
Waccamaw. Kat (1983) provided morpholog- 
ically and electrophoretic descriptions of this 
population, and R. I. Johnson (1984) judged it 
to be sufficiently distinct from L. r. radiata to 
warrant separate species status. Rogers 
modified distance (Table 5), which produces 
less compaction at the extremes, yielded cor- 
responding results, with L. fullerkati being al- 
most as genetically close to L. r. radiata as L. 
r. conspicua is to L. r. radiata. 

Two species, Lampsilis cariosa and Lep- 
todea ochracea, were sampled from different 
river systems (Fig. 1 , Appendix I) thus provid- 
ing measures of intraspecific genetic similar- 
ity among sites. Nei's (1978) unbiased ge- 
netic identity among the three poulations of 
Lampsilis cariosa from the Cape Fear, Tar, 
and Deep rivers ranged from 0.914-0.959. 
Heterozygosity values for these populations 
(Table 3) were also comparable (0.26-0.32). 
The populations of Leptodea ochracea from 
the Chowan River and Lake Waccamaw (Fig. 
1, Appendix I) were genetically identical 
(Nei's identity was 0.982; Rogers metric dis- 



tance was 0.134). However, the Lake Wacca- 
maw population exhibited a low heterozygos- 
ity value (0.051, almost half that of the 
population from the Chowan River), and 
these within-species genetic distance com- 
parisons essentially correspond to the levels 
found for the L. r. conspicua-L. r. radiata-L. 
fullerkati association (Tables 4, 5). 

Evolutionary relationships among the nine 
populations were explored by performing a hi- 
erarchical cluster analysis using the un- 
weighed pair-group method with arithmetic 
averaging (Sneath & Sokal, 1973). The result- 
ing phenograms, one based upon Nei's unbi- 
ased genetic distance (Nei, 1978) and the 
other based upon unmodified Rogers genetic 
distance (Rogers, 1972; Wright, 1978) are 
given in Figures 3 and 4 respectively. Both 
confirm the close genetic similarity between 
the two rare "subspecies" L. r. conspicua and 
L. r. radiata, as well as their very close simi- 
larity to the Lake Waccamaw L. fullerkati. 
Lampsilis cariosa lies next in genetic related- 
ness to this group of three "species/subspe- 
cies," with Leptodea ochracea and Elliptio 
complanata exhibiting significant genetic dis- 
tance not only from each other but also from 
the Lampsilis group. The phenogram based 
upon Nei's unbiased distance tended to com- 
pact values at the lower end of the scale (Fig. 
3), whereas the phenogram derived from 
modified Rogers distance, a metric scale, pro- 
vided a less compacted view of the genetic 
relationships among the Lampsilis radiata 
subspecies and L. fullerkati (Fig. 4). Even 
viewed in this manner, the relationships are 
still exceedingly close. 

A possible evolutionary or phylogenetic 
pathway among the nine populations is given 
by the Wagner tree (Fig. 5) (Farris, 1970, 
1972). It is based upon Rogers metric modi- 
fied distance measure. Construction assump- 



LAMPSIUS IN NORTH CAROLINA 



363 



TABLE 5. Matrix of Rogers modified distance for the nine populations/species of freshwater mussels 
from North Carolina waters. 



Population/Species 


1 


2 


3 


4 


5 


6 


7 


8 9 


1 . Elliptic» complánala 


***** 
















2. Lampsilis cariosa-Cape Fear 


0.627 


***** 














3. Lampsilis cariosa-Tar R. 


0.652 


0.269 


***** 












4. Lampsilis cariosa-Deep R. 


0.609 


0.262 


0.211 


***** 










5. Leptodea ochracea-Chowan R. 


0.835 


0.647 


0.747 


0.729 


***** 








6. Leptodea ochracea-L. Waco 


0.840 


0.650 


0.745 


0.731 


0.134 


***** 






7. Lampsilis fullerkati 


0.665 


0.516 


0.490 


0.500 


0.749 


0.742 


***** 




8. L. radiata conspicua 


0.629 


0.499 


0.514 


0.518 


0.716 


0.714 


0.219 


***** 


9. L. radiata radiata 


0.680 


0.484 


0.515 


0.522 


0.702 


0.703 


0.213 


0.162 ***" 



DISTANCE 



1.20 1.00 .80 .60 .40 .20 .00 

+ .... + .... + .... + .... + .... + .... + — - + .... + .... + — - + — - + — - + 



Elliptio complanata-S. Flat R. 



— Lampsilis cariosa-Cape Fear R. 

i — Lampsilis car/osa-Tar R. 

*— Lampsilis cariosa-Deep R. 
I — Lampsilis ful/erkati-L. Waccamaw 

- Lampsilis radiata conspicua-F\at R. 

- Lampsilis radiata radiata-Chowan 

L Leptodea осЛласеа -Chowan R. 
' sptodea ochracea-L. Waccamaw 



+ .... + .... + .... + .... + .... + — . + — - + — - + .... + — + — - + — + 
1.20 1.00 .80 .60 .40 .20 .00 

FIG. 3. Phenogram of nine North Carolina freshwater mussel populations based upon Nei (1978) unbiased 
genetic distance and using the unweighted pair group method of construction. Farris (1972) "f "-value = 
4.672. 



tions are discussed in Felsenstein (1983) and 
Swofford & Olsen (1990). The method of root- 
ing is based upon midpoint location (Farris 
1972, 1981). This method usually leads to 
trees having their roots on the branch leading 
to outgroup species (Felsenstein, 1983). This 
tree placed the two rare Lampsilis subspecies 
and L. fullerkati at approximately the same 
distance from the root, and into the same 
common evolutionary assemblage. Lampsilis 
cariosa lies next in genetic relatedness to the 
three aforementioned species-subspecies. 
Leptodea ochracea and Elliptio complánate, 



as expected, exhibited significant genetic dis- 
tances not only from each other but also from 
the Lampsilis group. Populations within both 
Lampsilis cariosa and Leptodea ochracea 
have relative distances from the root that dif- 
fer little from that among the L. radiata sub- 
species-L. fullerkati complex. 

With respect to specific loci, both L. radiata 
"subspecies" also share the same major 
(most common) allele for all loci, and do so in 
approximately the same frequency (Table 3). 
In six of the loci, slight differences among the 
two subspecies do exist, ranging from certain 



364 



STIVEN & ALDERMAN 



DISTANCE 



.80 
+ — 



.67 .53 .40 .27 .13 .00 

+ — + — - + — - + -— + — + — - + — - + — - + — - + — - + 



EHiptio complanata-S. Flat R. 
Lampsilis cariosa-Cape Fear R. 
Lampsilis сал /osa-Tar R. 
Lampsilis cariosa-Deep R. 
Lampsilis ful/erkati-L. Waccamaw 
Lampsi/is radiata conspicua-F\at R. 
Lampsilis radiata radiata-Chowan 
Leptodea ochracea-Chowan R. 
Leptodea ochracea-L. Waccamaw 



+ .... + .... + .... + .... + — . + — . + — . + .... + — . + — . + — - + — - + 
.80 .67 .53 .40 .27 .13 .00 

FIG. 4. Phenogram of nine North Carolina freshwater mussel populations based upon modified Rogers 
genetic distance (Rogers, 1972; Wright, 1978) and using the unweighted pair group method of construction. 
Farris (1972) "f "-value = 0.780. 



DISTANCE FROM ROOT 

.00 .07 .14 .21 .28 .35 

+ .... + .... + .... + .... + .... + .... + .... + .... + .... + .... + .... + 



.42 

- + 



EHiptio complanata-S. Flat R. 



Lampsilis cariosa-Cape Fear R. 
Lampsilis cariosa-Tar R. 



Lampsilis cariosa-Deep R. 
— Lampsilis ful/erkati-L. Waccamaw 



Lampsilis radiata conspicua-F\at R. 



Lampsilis radiata radiata-Cbowan R. 

Leptodea осЛласеа -Chowan R. 

Leptodea ochracea-L. Waccamaw 



+ —- + -— + - 
.00 .07 



+ — - + - 
.14 



. + -— + -.-. + . ... + .... + . 
.21 .28 



- + -— + —- + 
.35 .42 



FIG. 5. Wagner tree depicting phylogenetic relationships of nine North Carolina freshwater mussel species 
and populations. Coefficient used was modified Rogers distance (Rogers, 1972; Wright, 1978). Tree is 
rooted at midpoint of longest path. Farris (1972) "f "-value is 0.893. 



LAMPSILIS IN NORTH CAROLINA 



365 



loci being polymorphic in one but not the other 
(Pgi, Pgm-1, Pgm-2, a-Pgdh) to one locus 
showing an allele in one subspecies that is 
not present in the other (Mpi). 



DISCUSSION AND CONCLUSIONS 

Genetic Relationships Among Lampsilis, 
Leptodea and Elliptio spp. 

Freshwater mussels are one of the most 
diverse and abundant groups of macroinver- 
tebrates in the United States, but also one of 
the most notorious for lack of systematic 
agreement among investigators (Davis & 
Fuller, 1981). Because conchological differ- 
ences among bivalves are so environmentally 
dependent, the use of shell morphology in 
separating groups, especially above the spe- 
cies level, can often be misleading (Davis & 
Fuller, 1981). Molecular genetic methodolo- 
gies have grown in importance in helping sort 
out differences among the Unionacea (Davis et 
al., 1981 ; Davis & Fuller, 1981 ; Kat, 1983; Kat 
& Davis, 1984). However, opinions differ over 
the association between genetic distance, 
level of systematic divergence and specia- 
tion, as well as the broader question of the 
applicability of molecular genetic tools (G.B. 
Johnson, 1977) and their use in phylogenetic 
interpretations (Felsenstein, 1988; Swofford 
& Olsen, 1990). Avise (1976) suggested in 
general terms that when using Nei's (1972, 
1978) genetic identity (I), values of 0.9-1.0 
would be expected among populations within 
a species, 0.8 to 0.89 among subspecies 
within a species, and a value < 0.8 among 
different species. Thorpe (1982), in a litera- 
ture review, found that 97% of Nei l-values 
used for erecting new species were below 
0.85, and 98% of the l-values associated with 
within species comparisons exceeded 0.85. 
In freshwater mussels, however, systematic 
interpretations based upon genetic distances 
vary, often as a function of allopatry-sympa- 
try. Kat (1983) argued that a value of 0.878 
found among Lampsilis spp. and L. radiata is 
characteristic of distinct species among radi- 
ating clades. This is consistent with Davis et 
al. (1981), who reported values > 0.9 among 
apparent sympatric species of Elliptio in Flor- 
ida and Lake Waccamaw in North Carolina. 
The two "sympatric" Elliptio species in Lake 
Waccamaw had different shell characters 
states, differed in allelic frequencies at three 
loci, and preferred different habitats. Habitat 



differences were not apparent in the two Flor- 
ida sympatric Elliptio species. On the other 
hand, Kat & Davis (1984) interpretated a 
value of 0.954 between two allopatric popu- 
lations of Elliptio complánala in adjacent 
drainages in Nova Scotia as consistent with 
the Avise (1976) scheme. Thus, Davis et al. 
(1981) argue that in established conchologi- 
cally distinct sympatric populations, genetic 
identity values of >0.9 could indicate different 
species, yet recognize that in allopatric situa- 
tions, genetic identity values for different spe- 
cies of the same genus would more likely be 
less than 0.9. 

Our intraspecific work with Lampsilis cari- 
osa and Leptodea ochracea populations pro- 
duced unbiased genetic identities (Nei, 1978) 
among separated sites of 0.922-0.959 and 
0.982 for these species, respectively. These 
values are consistent with those suggested 
by Avise (1976), Davis et al. (1981), and 
Thorpe (1982) to encompass the expected 
variation and range within allopatric popula- 
tions of a single species. Both phenograms 
(Figs. 3, 4) illustrate the identical genetic na- 
ture of these two species groups. Thus, the 
large-shelled L. cariosa type sampled from 
the Deep River is probably L. cariosa. It is 
genetically indistinct from the other two pop- 
ulations and also shows no signs of pro- 
longed bottleneck effects (e.g. diminished 
heterozygosity). 

Among paired associations within the four 
species of Lampsilis, we found identity values 
ranging from 0.357 to 0.863, consistent with 
Avise's (1976) expectations for variation 
among different species, as well as values 
found by Kat (1983) for three distinct species 
of Lampsilis. For example, we found identity 
values of 0.382 and 0.439 between Chowan 
River Leptodea ochracea and L. r. conspicua 
and L. r. radiata respectively, whereas Kat's 
(1983) value for a comparable association 
was 0.31 . Genetic identity measures between 
Leptodea ochracea and Lampsilis fullerkati 
ranged from 0.35 to 0.366, and Kat's (1983) 
corresponding value was about 0.49. We also 
found values from 0.33 to 0.48 between Lep- 
todea ochracea and L. cariosa, clearly imply- 
ing distinct species status for each according 
to guidelines in Avise (1976). Identity values 
among Elliptio, Leptodea and Lampsilis gen- 
era in our study ranged from 0.115 to 0.445 
(Table 4), consistent with Avise (1976) and 
also Davis et al. (1981). Again the two phe- 
nograms (Figures 3, 4) clearly depict these 
patterns. 



366 



STIVEN & ALDERMAN 



Heterozygosity Levels 

In examining relative heterozygosity levels 
among the various populations, we found that 
the two rare Lampsilis "subspecies" had H- 
values that were about 40% lower than that of 
populations of the more abundant and com- 
mon Lampsilis cariosa. They were about the 
same as that of the genetically identical 
Lampsilis fullerkati from Lake Waccamaw and 
the common Elliptio complanata. Thus, we 
cannot associate these low heterozygosity 
levels with a prolonged bottleneck effect (Nei 
et al., 1975). Leptodea ochracea populations 
exhibited even lower heterozygosity levels as 
well as low polymorphism, a finding consis- 
tent with Kat's (1983), especially for Lake 
Waccamaw. We note that reliability of genetic 
identity comparisons among populations by 
genetic distance measures when sample 
sizes are small is greatly improved when het- 
erozygosity levels are low (Nei, 1978), as in 
the case of the rare L. radiata "subspecies." 

One possible explanation for the heterozy- 
gote deficiencies may be that a population in 
any one site is simply a mix of age classes 
and genotypes derived from a variety of pop- 
ulations some distance away that possess dif- 
ferent gene frequencies. Dispersal of mussel 
larvae may be substantial (river currents, 
glochidia on fish hosts), and the Wahlund ef- 
fect may be a partial explanation for heterozy- 
gote deficiencies (Dillon, 1988). 

The Eastern Lampmussel and the Carolina 
Fatmucket: Distinct Subspecies? 

The two rare L. radiata populations (subspe- 
cies) are currently recognized by the Ameri- 
can Malacological Union as L. r. conspicua 
(Carolina fatmucket) and L. r. radiata (eastern 
lampmussel) (Turgeon et al., 1988). Turgeon 
et al. (1988) and Alderman (personal obser- 
vation) have reported that sampled L. r. con- 
spicua shells were larger and heavier than L. 
r. radiata, and that differences existed in their 
current range and preferred habitat. Lamp- 
silis r. radiata resides in the lower parts of 
drainage basins of the eastern North Ameri- 
can coastal plain, discontinuously from South 
Carolina to the St. Lawrence, whereas the 
very rare L. r. conspicua seems endemic to 
North Carolina and is currently restricted to 
the Pee Dee River drainage basin and the 
upper Neuse River basin. Previous work on 
these populations differentiated substrate af- 
finities (Alderman, 1989), with the Carolina 



fatmucket found more often in course sands, 
gravel and cobble substrates, whereas the 
eastern lampmussel occurred more often in 
medium to coarse sands. Our own collecting 
of these two groups was consistent with these 
reported substrate preferences. Sample shell 
size data from this study, even though based 
on relatively small numbers, confirmed prior 
evidence of a larger L. r. conspicua, that is, 
about 1.4 times greater than L. r. radiata 
(Table 2). There is also strong evidence for 
"site effects" in the two species examined 
{Lampsilis cariosa and Leptodea ochracea) 
for this phenomenon, and this process cannot 
be eliminated as a basis of the shell differ- 
ences. In addition, covariance analysis of 
shell regressions clearly indicated strong con- 
cohological similarities between these two 
"subspecies." 

Nei's (1978) unbiased genetic identity be- 
tween these two "subspecies" is 0.978 (and 
an unbiased genetic distance of 0.022) (Table 
4). Rogers metric modified distance (Rogers, 
1972; Wright, 1978) was 0.162, hardly distin- 
guishable from the value of 0.134 between 
the two separated populations of Leptodea 
ochracea, and even smaller than 0.211 to 
0.229 among the allopatric populations of 
Lampsilis cariosa. Clearly, Nei's (1978) iden- 
tity between the two "subspecies" is consis- 
tent with Avise's (1976) categorization of pop- 
ulations within a species (0.9-1 .0). It does not 
approach the subspecies category (0.80- 
0.89), let alone values differentiating species 
(<0.8). It is also in the range of values we 
found among the allopatric populations of 
Lampsilis cariosa and Leptodea ochracea. In 
addition, we note that the distinct Lake Wac- 
camaw species, Lampsilis fullerkati, has Nei 
(1978) unbiased identity values of 0.945 and 
0.952 in association with L. r. conspicua and 
North Carolina L. r. radiata respectively. 
Those are above the 0.88 recorded for the 
same species comparisons by Kat (1983), but 
Kat's L. radiata specimens came from Cana- 
da's maritime provinces. Recall also that 
Davis et al. (1981) argued that values >0.9 
could be found among sympatric freshwater 
mussel species that recently underwent spe- 
ciation. However, these two so-called sub- 
species of L. radiata are currently not sympat- 
ric, and, in fact, are found for the most part in 
different river basins with the exception of the 
Neuse River (R. I. Johnson, 1970). Lampsilis 
r. radiata seems confined to the coastal plain 
tidewater areas of the river systems and does 
not appear to overlap with L. r. conspicua. 



LAMPSILIS IN NORTH CAROLINA 



367 



One may also speculate the most pre-existing 
L. radiata populations above the tidewater ar- 
eas have simply been extirpated. 

Thus, the genetic identity and distance 
measures (Tables 4, 5), and the graphical il- 
lustrations given by the phenograms and 
Wagner tree (Figs. 3-5) point out the almost 
identical genetic similarity between the two 
"subspecies" of L radiata, as well as the neg- 
ligible genetic distance separating these 
"subspecies" from the recently designated 
species, L. fullerkati. Conchological data for 
these three populations are also similar (co- 
variance analysis) and suggest either the be- 
ginnings of evolutionary divergence or simply 
evidence of wide phenotypic plasticity coming 
from different shell growth rates and patterns 
in different sites. The genetic similarity of 
these two L radiata populations differ little 
from that found among poulations of the same 
species living in different sites (e.g. Leptodea 
ochracea and Lampsilis cariosa). Clearly, re- 
search on reciprocal transplant growth of 
young individuals (preferably of both "sub- 
species" and L fullerkati) would be useful in 
clarifying this possible shell phenotypic plas- 
ticity. Answers to questions about reproductive 
isolation and potential hybridization among 
these populations also would shed further 
light on this genetically similar complex. Until 
evidence to the contrary appears, we suggest 
that L. r. conspicua and L r. radiata and per- 
haps L. fullerkati be considered simply as al- 
lopatric "populations" of Lampsilis radiata. 

We are aware that much of the work on the 
application of molecular genetic techniques to 
systematic issues has been in one direction, 
that of defining and erecting new species and 
subspecies. For example, Ayala (1975) insti- 
tuted several subspecies of Drosophila willis- 
toni based on allozyme techniques and Nei's 
(1972) genetic distance. Highton et al. (1989), 
also utilizing allozymes, established 16 spe- 
cies of Plethodon salamanders from the sin- 
gle Plethodon glutinosis complex based on a 
minimum Nei distance of 0.15 or an Nei ge- 
netic identity (I) of 0.85. Such taxonomic split- 
ting is quite common in the systematic litera- 
ture, including that of freshwater mussels. 
Our suggestion, therefore, of regrouping the 
two and possibly three previously recognized 
allopatric subspecies/species into one spe- 
cies complex, based upon very high levels of 
genetic identity as well as similar concholo- 
gies, is probably an uncommon event, yet ap- 
propriate until more distinctive biological spe- 
cies properties become evident. 



ACKNOWLEDGMENTS 

Special thanks go to Christopher McGrath 
for his help with the field work. We are also 
appreciative of the helpful comments of two 
reviewers as well as the editor, George Davis. 
The research was partly supported by a grant 
from the Nongame Program, North Carolina 
Wildlife Resources Commission. 



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Revised MS accepted 24 March 1 992 



APPENDIX I. Species and ANSP catalog numbers, number electrophoresed, water system, site code 
(Fig. 1), and description of collecting sites for populations of North Carolina freshwater mussels used in 
this study. The rate L. r. radiata and L. r. conspicua were returned alive to collecting site. 



Species 



ANSP 



No. 



County 



Water System (Code) Location 



Lampsilis radiata 

radiata^ 
Lampsilis radiata 

conspicua^ 
Elliptio 

complanata 


392038 
392039 


9 
23 

44 


Gates 

Durham 

Person 


Leptodea 
ochracea 2 


392040 
392041 


19 
55 


Gates 
Columbus 


Lampsilis cariosa 


392042 


11 


Cumberland 




392043 


7 


Nash 




392044 


17 


Moore 


Lempsilis fullerkati 


392044 


35 


Columbus 



Chowan River (C). Below HiWay 13/ 

1 58 bridge, Hertford-Gates county line 

Flat River (B). HiWay 501 , east on SR 

1 471 to bridge, 200 m below bridge 

S. Flat River (A). HiWay 501, west on 

SR 1123, south on SR 1125 to bridge, 

200 m below bridge 

Chowan River (C) (see above) 

Lake Waccamaw (D). Collections just 

off shore State Park, 2 m depth 

Cape Fear River (F). HiWay 401, site at 

confluence of Carvers Ck. and Cape 

Fear R. 

Tar River (E). Main channel from Spring 

Hope to HiWay 581 bridge 

Deep River (G). Via HiWay 22 west on 

SR 1456 bridge, upstream 100 m. 

Lake Waccamaw (C) (see above) 



1 Returned alive to collecting site, no ANSP number. 
2 Formerly called Lampsilis ochracea (see Turgeon et al., 1988). 



MALACOLOGIA, 1992, 34(1-2): 371-376 



INDEX 



Taxa in bold are new; page numbers in 
bold indicate pages on which new taxa are 
described; underscored pages indicate 
systematic descriptions of previously named 
taxa; pages in italics indicate figured taxa. 

adrianae, Albinaria 37, 38, 40, 41, 47, 52, 

55-57 
adrianae, Albinaria adrianae 34-36, 39, 41 , 

46, 47-52, 56, 60 
aetnea, Helix 121, 126 
Agriolimax reticulatus 68, 71 
Akiyoshia 143, 154, 155 

uenoi 1 55 
Akiyoshia (Saganoa) chinensis 148, 155- 

156. 158-162 . 166, 297; 157-160, 162- 

165 

kishiiana 1 55 

odonta 150, 152 

yunnanensis 1 55 
Alasmidonta 355 
Albinaria 33-61 

adrianae 37, 38, 40, 41 , 47, 52, 55-57 

adrianae adrianae 34-36, 39, 41 , 46, 
47-52, 56, 60 

adrianae dubia 34-36, 38-41 , 47-52, 
56, 60 

contaminata 34-38, 40, 41, 47, 52-55, 
57 

contaminata contaminata 34-36, 38, 39, 

41, 46-51, 53, 54, 56, 57, 60, 61 
contaminata incommoda 34-36, 39, 41 , 

42, 47-51 , 53, 54, 60, 61 
contaminata liebetruti 34-36, 38, 39, 

41, 42, 46-54, 56, 61 
contaminata muraría 38 
contaminata odysseus 34-36, 38, 39, 

41, 46, 47-51, 53, 54, 57, 61 
jónica 37, 38, 40, 41 , 47, 52, 55, 57 
jónica assicola 34-36, 39, 41 , 46, 47- 

51, 54-56, 61 
jónica jónica 34-36, 38, 39, 41 , 42, 46, 

48-52, 55, 60 
rebeli 34, 39, 40, 47-51 , 61 
senilis 34-38, 40, 41 , 52, 54-55, 57 
senilis flavescens 34-36, 39, 41 , 47-51 , 

55, 56, 60, 61 
senilis kolpomyrtensis 34-36, 38, 39, 

41, 47-51, 54, 56, 61 
senilis senilis 34-36, 38, 39, 41 , 42, 

46-51 , 54-57, 60, 61 
teres 39, 47, 50, 51 
teres nordsiecki 34, 40, 48, 49, 57, 61 
Aligena elevata 10 
Amblema 1 34 
Ammonoida 77 
Amnícola 153, 154 
Amnicolinae 153 

apena, Lithoglypopsis 185, 186, 333, 334 
apena, Neotricula 209, 218, 315-317, 320, 
322, 325-328, 334 



aperta, Tricula 209 

apicina, Xerotricha 111, 120, 126 

Aplacophora 77, 78, 80 

Archaeogastropoda 82 

Arion 4/ 

ater 352 

hortensis 68, 72, 87, 95 
armillata, Helix 109 
armillata, Microxeromagna 117, 123 
Arthritica crassiformis 1 1 
aspersa, Helix 68, 71, 72 
assicola, Albinaria jónica 34-36, 39, 41 , 46, 

47-51 , 54-56, 61 
ater, Arion 352 
bambooensis, Tricula 266, 286, 316-317, 

320, 322, 325-328, 339-342 
Bathymodiolus thermophilus 137 
bathytera, Helix {Pseudoxerophila) 109 
beauii, Cydostremiscus 1, 15 
Biomphalaria glabrata 25-32; 26, 28-31 
Bithynia minutoides 259 
Bivalvia 77, 78, 80 
Blanfordia 1 53 
bollingi, Tricula 286, 314, 315-317, 320, 

322, 325-328, 339-342 
burchi, Neotricula 209, 218, 316-318, 320, 

322, 325-328, 339-342 
burchi, Tricula 186 
Bythinella 154 

chinensis 154, 163, 166 

gongjianguoi 1 63 

nubeiensis 1 63 

/// 163 

watanensis 1 63 

wufenensis 1 63 
Caenogastropoda 83, 84 
Calyptogena 131, 132 

magnifica 1 35 
candicans, Helicella 111 
Candidula 107, 109, 112, 126 

inters ecta 1 09 

spadae 1 09 

unifasciata 1 09 
candidula, Glischrus (Helix) 109 
cariosa, Lampsilis 356, 358, 360-367, 369 
caruanae, Cernuella 110 
carusoi, Helicotricha 109, 112-119, 123, 

126; 113-118 
Caucasigena 109-112, 123 
cema, Ceratobomia 20, 21 
Cepaea 95 

nemoralis 68, 72, 87, 95 
Cephalopoda 77, 78, 80 
Ceratobomia cema 20, 21 

longipes 20 
Cerion 57 
Cernuella 108, 109, 111, 112 

caruanae 1 1 

virgata 1 1 
Certatobomia 20, 21 



371 



372 



INDEX 



chinensis, Akiyoshia (Saganoa) 148, 155- 

156. 158-162 . 166, 297; 157-160, 162- 

165 
chinensis, Bythinella 154, 163, 166 
chinensis, Erhaia 166 
chinensis, Gammatricula 218, 316-318, 

320-322, 325-328, 339-342 
chinensis, Pseudobythinella 148, 166. 169- 

170. 173. 175 ; 157, 167, 168, 170-174 
Chondrina dienta 46 
cinctella, Helix 109 
Ci revi us texanus 1, 15 
dienta, Chondrina 46 
Cochlea virgata 109 
Cohilopinae 186 

complanata, Elliptio 57, 357-367, 369 
conspicua, Lampsilis radiata 356, 358-367, 

369 
conspurcata, Helicella (Xerotricha) 112 
conspurcata, Helix 109, 120 
conspurcata, Xerotricha 107, 111, 120, 126 
contaminata, Albinaria 34-38, 40, 41, 47, 

52-55, 57 
contaminata, Albinaria contaminata 34-36, 

38, 39, 41 , 46-51 , 53, 54, 56, 57, 60, 

61 
corde roi, Helicella 120 
cordiformis, Divariscintilla 11-12, 14, 15- 

21 ; 3, 4, 13 
crassiformis, Arthritica 11 
Crassostrea 1 34 
cristella, Hydrobia 211 
cristella, Neotricula 150, 209, 211-214. 

217-218. 220. 222 . 246, 256, 315-317, 

320, 322, 325-328, 330; 215, 216, 219, 

221, 222-224 
cristella, Tricula 211 
cumúlala, Nanaja 109 
Cydostremiscus beauii 1,15 
daliensis, Erhaia 154, 163 

daliensis, Pseudobythinella 161, 166, 183 

darevskii, Hygrohelicopsis 109, 123 

decollata, Rumina 45, 47 

Delavaya 266, 279, 332-336 

Dentalium 25 

Devonia 19, 20 

dianmenensis, Neotricula 150, 209, 218, 

222, 227-231, 233-234, 246, 256, 315- 
318, 320-322, 325-328, 330; 225-227, 
229-233 

Divariscintilla 2, 19-22 

cordiformis 11-12, 14, 15-21; 3, 4, 13 

luteocrinita 6, 9-11, 15-18, 20; 3, 8, 
11 

maoria 1, 10, 14-21 

octotentacula 2, 4-6, 9, 12, 14-17, 19- 
22; 3, 4, 7, 12, 16 

troglodytes 1, 15-21 

yoyo 1, 5, 6, 15-22 
dubia, Albinaria adrianae 34-36, 38-41, 47- 

52, 56, 60 
duplicata, Neotricula 150, 209, 234-238, 

241, 243-249, 256, 297, 316-318, 320, 

322, 325-328; 225, 234-236, 238-240, 

242-246, 248-252 
Edentiella 109-112, 123 



edentula, Helix 109, 123 

edmundi, Isabellaria 34, 38, 57, 61 

edulis, Mytilus 99-106 

eichwaldi, Helix 1 09, 1 23 

elevata, Aligena 10 

elevatus, Leptodrilus elevatus 129-141; 

130, 131, 134, 138 
Elliptio 355, 362, 365 

complanata 57, 357-367, 369 
Entovalva 1 9 

perrieri 20 
Ephippodonta 1 9 
Erhaia 162 

chinensis 1 66 

daliensis 154, 163 

kunmingensis 154, 163 
Erhaiinl 154 
Erycinidae 22 
Erydninae 22 
explanata, Helix 109 
Fenouilia 266, 277, 279, 282, 284, 324, 

331 , 332-336 

kreitneri 284 
tlavescens, Albinaria senilis 34-36, 39, 41, 

47-51 , 55, 56, 60, 61 
fluviatilis, Nertina 25 
fontinalis, Physa 25 
fuchsianus, Guoia 146, 150, 190, 192, 193, 

195, 204-205. 208-209. 211. 213 ; 188, 

191, 197, 199, 206, 207, 210-212, 214 
fuchsianus, Lithoglyphopsis 204 
fuchsianus, Lithoglyphus 187, 204 
fullerkati, Lampsilis 356, 358-367, 369 
Galeomma polita 10 

takii 10 
Galeommatidae 1-24 
galloprovindalis, Mytilus 99-106 
Gammatricula 184, 320-322, 325-328, 330- 

336 

chinensis 218, 316-318, 320-322, 325- 
328, 339-342 
Gastropoda 77, 78, 80 
gibba, Partula 47 
gittenbergeri, Helicopsis 120 
glabrata, Biomphalaria 25-32; 26, 28-31 
Glischrus (Helix) candidula 109 
gongjianguoi, Bythinella 163 
gonzalei, Helicella 120 
grandis, Lithoglyphopsis 282 
gredleri, Neotricula 318 
gredleri, Tricula 150, 152, 155, 246, 286. 

288-293. 295-297 . 315-318, 320-322, 

325-328; 286-290, 292-296 
gregoriana, Tricula 286, 297, 316-317, 319, 

320, 322, 325-328, 339-342 
Guoia 142, 150, 186-187, 199, 324, 330- 

336 

fuchsianus 146, 150, 190, 192, 193, 
195, 204-205. 208-209. 211. 213 ; 
188, 191, 197, 199, 206, 207, 210- 
212, 214 

viridulus 146, 150, 187. 190-196. 198- 
201 . 203 . 208, 209; 188-191, 196, 
197, 199, 201-208, 210, 272 
Gyraulus 187, 235 



INDEX 



373 



Halewisia 184, 211, 324, 327, 330, 332- 

336 
Helicella 107, 109, 110, 112, 120, 126 

candicans 1 1 1 

cordero] 1 20 

gonzalei 120 

mangae 1 20 

spiruloides 1 1 1 
Helicella (Xerotricha) conspurcata 112 
Helicopsis 107, 109-112, 120, 122; 121, 

122 

gittenbergeri 1 20 

îikharevi 110 

retowskii 110 

striata 110, 117 

subcalcarata neuberû 120 

subcalcarata subcalcarata 120 
Helicotricha 109-112; 123, 124, 126 

carusoi 109, 112-119, 123, 126; 113- 
118 
Helix 349 

aetnea 121, 126 

armillata 109 

aspersa 68, 71, 72 

cinctella 109 

conspurcata 109, 120 

edentula 109, 123 

eichwaldi 109, 123 

explanata 1 09 

hispida 109, 123 

holotricha 109, 123 

/fa/a 109, 120 

lubomirskii 109, 123 

/ucorum 63-73, 343-354; 63, 66, 69, 
70, 345, 346, 34S, 35 Г 

obv/a 109 

pomatia 68 

/г /bens 109, 123 

stolismena 1 09 

sfr/ate 109, 120 

turbinate 109 

turcica 1 20 

unifasciata 1 09 

variabilis 1 09 
He//x (Pseudoxerophila) bathytera 109 
Heterostropha 83, 84 
hispida, Helix 109, 123 
holotricha, Helix 109, 123 
hortensis, Mon 68, 72, 87, 95 
hubeiensis, Bythinella 163 
Hubendickia 146, 332-336 
hudiequanensis, Tricula 218, 286, 316-317, 

320, 322, 325-328, 339-342 
hunanensis, Stenothyra 187, 271 
hupensis, Oncomelania 148, 153, 154 
Hydrobia 1 53, 324 

cristella 211 

minutoides 259 
Hydrobndae 153, 186, 324 
Hygrohelicopsis 109-112, 123 

darevskii 109, 123 
Hygromia 109, 112 
Hygromndae 107-128 
Hygromiinae 126 
Hyolitha 77, 78, 80 



incommoda, Albinaria contaminata 34-36, 
39, 41, 42, 47-51, 53, 54, 60, 61 

intersecta, Candidula 109 

Isabellaria 39, 47-51 , 57 
edmundi 34, 38, 57, 61 

Ischnochiton 25 

/Ya/a, He//x 109, 120 

Jeffrey siana, Vasconiella 19, 20 

jianouensis, Pseudobythinella 162, 163 

jinhongensis, Jinhongia 316-318, 320, 322, 
325-328 339-342 

Jinhongia 184, 320, 322, 324-328, 330-336 

jinhongensis 316-318, 320, 322, 325- 
328, 339-342 
jónica, Albinaria 37, 38, 40, 41 , 47, 52, 55, 

57 
jónica, Albinaria jónica 34-36, 38, 39, 41 , 

42, 46, 48-52, 55, 60 
Jullieniini 324, 332 
kishiiana, Akiyoshia (Saganoa) 155 
Kokotschashvilia 109-112, 123 
kolpomyrtensis, Albinaria senilis 34-36, 38, 

39, 41, 47-51, 54, 56, 61 
kreitneri, Fenouilia 284 
kunmingensis, Erhaia 154, 163 
kunmingensis, Pseudobythinella 155, 161, 

166, 183 
Kunmingia 334 
Lacunopsis 266, 277, 282, 284, 324, 330, 

332-336 

yunnanensis 282 
Lampsilis 355-369 

cariosa 356, 358, 360-367, 369 

fullerkati 356, 358-367, 369 

radiata 356, 365-367 

radiata conspicua 356, 358-367, 369 

radiata radiata 356, 358-367, 369 
Lasaea 20 
Lasmigona 355 
Leptodea 365 

ochracea 356-358, 360-367, 369 
Leptodrilus elevatus elevatus 129-141; 130, 

131, 134, 138 

pustulosus 1 39 
Leucozonella 109-112, 123 
liebetruti, Albinaria contaminata 34-36, 38, 

39, 41 , 42, 46-54, 56, 61 
///, Bythinella 163 
Iikharevi, Helicopsis 110 
//7/7, Neotricula 150, 209, 218, 234, 246, 

248, 252, 254, 256-259, 316-318, 320, 

322, 325-328; 215, 225, 253, 255, 256, 

258-261 
imputan us, Lithoglyphus 187, 282; 272, 273 
Limnaea stagnais 25, 26, 32 
Lithoglypheae 186 
Lrthoglyphinae 186 
Lithogfyphopsis 143, 184-187, 266, 324, 

331 , 332-336 

aperta 185, 186, 333, 334 

fuchsianus 204 

grandis 282 

modesta 150, 185, 187, 196, 205, 266, 
268. 271. 275-277. 279. 282. 284, 
286 ; 270, 272-274, 277-286 



374 



INDEX 



ovatus 282 

viridulus 187, 205 
Lithoglyphus 186 

fuchsianus 187, 204 

liliputanus 187, 282; 272, 273 

modestus 268 

tonkinianus 1 87 

viridulus 186, 187 
littorea, Littorina 68, 71 
Littorina littorea 68, 71 

obtusata 25 
/Atf, Pseudobythinella 163 
longipes, Ceratobornia 20 
lubomirskii, Helix 109, 123 
lucorum, Helix 63-73, 343-354; 63, 66, 69, 

70, 345, 346, 346, 357 
ludongbini, Tricula 286, 315-317, 320, 322, 

325-328, 339-342 
luteocrinita, Divariscintilla 6, 9-11, 15-18, 

20; 3,8, 11 
lysiosquillina, Phlyctaenachlamys 16, 18-21 
maculata, Semerula 352 
magnifica, CaJyptogena 135 
mangae, Helicella 120 
maoria, Divariscintilla 1, 10, 14-21 
maxidens, Tricula 150, 155, 286, 297, 

300-302, 304-305, 316-318, 320, 322, 

325-328; 298, 299, 301, 303-307 
mediterránea, Umbrella 25 
mercenaria, Venus 352 
Mesogastropoda 82 
Microxeromagna 107, 109, 111, 112, 124 

armillata 117, 123 
minima, Neotricula 218 
minutoides, Bithynia 259 
minutoides, Hyarobia 259 
minutoides, Neotricula 150, 209, 259. 262- 

264. 266-268 . 316-317, 320-322, 325- 

328; 215, 263, 265-269 
minutoides, Tricula 259 
modesta, Lithoglyphopsis 150, 185, 187, 

196, 205, 266, 268. 271 . 275-277. 279. 

282. 284. 286 : 270, 272-274, 277-286 
modestus, Lithoglyphus 268 
Mollusca 75-86 
Monoplacophora 77, 78, 80 
montana, Tricula 286, 297, 316-317, 319, 

320, 322, 325-328, 339-342 
muraría, Albinaria contamínala 38 
Mytilus edulis 99-106 

galloprovincialis 99-1 06 
Nanaja 109-112 

cumulata 1 09 
Nautilida 77 
Nautilus 1 34 

nemoralis, Cepaea 68, 72, 87, 95 
Neogastropoda 82 
Neotricula 146, 148, 184, 187, 209. 211. 

217, 218, 230, 286, 315, 318, 320, 322- 

328, 330, 332-336 

aperta 209, 218, 315-317, 320, 322, 
325-328, 334 

burchi 209, 218, 316-313, 320, 322, 
325-328, 339-342 

cristella 150, 209, 211-214. 217-218. 
220. 222. 246, 256, 315-317, 320, 



322, 325-328, 330; 215, 216, 219, 
221, 222-224 
dianmenensis 150, 209, 218, 222, 
227-231, 233-234, 246, 256, 315- 
318, 320-322, 325-328, 330; 225- 
227, 229-233 
duplicata 150, 209, 234-238, 241, 243- 
249, 256, 297, 316-318, 320, 322, 
325-328; 225, 234-236, 238-240, 
242-246, 248-252 
gredleri 318 

lilii 150, 209, 218, 234, 246, 248, 252, 
254, 256-259, 316-318, 320, 322, 
325-328; 215, 225, 253, 255, 256, 
258-261 
minima 218 

minutoides 150, 209, 259. 262-264. 
266-268 . 316-317, 320-322, 325-328; 
215, 263, 265-269 
odonta 318 
Nertina fluviatilis 25 
neuberti, Helicopsis subcalcarata 120 
niuzhuangensis, Wuconchona 316-318, 

320-322, 325-328, 339-342 
nordsiecki, Albinaria teres 34, 40, 48, 49, 

57, 61 
nubivaga, Xerotricha 120 
obtusata, Littorina 25 
obvia, Helix 109 

ochracea, Leptodea 356-358, 360-367, 369 
octotentacula, Divariscintilla 2, 4-6, 9, 12, 

14-17, 19-22; 3, 4, 7, 12, 16 
odonta, Akiyoshia (Sagonoa) 150, 152 
odonta, Neotricula 318 
odonta, Tricula 150, 286, 297, 301 . 305. 
307. 311-312. 314-315 . 316-318, 320, 
322, 325-328; 298, 308-310, 312, 313 
odysseus, Albinaria contaminata 34-36, 38, 

39, 41, 46, 47-51, 53, 54, 57, 61 
Oncomelania 1 53- 1 54 

hupensis 148, 153, 154 
Opisthobranchia 77, 79, 81, 82 
ovatus, Lithoglyphopsis 282 
Pachydrobia 183, 184, 324, 327, 330, 332- 

336 
Pachydrobüni 150, 183, 324, 328, 330, 334 
Parabornia 22 

squill i na 20, 21 
Parapyrgula 334 
Partula 57 

gibba 47 
Pectén 1 9 

perlatoris, Schileykiella 126 
perrieri, Entovalva 20 
Phlyctaenachlamys 20, 21 
lysiosquillina 16, 18-21 
Physa fontinalis 25 
pisana, Theba 87-97, 343, 350 
Planorbidae 271 
Planorbis trivolvis 25 
Plicuteria 109-112, 123 
polita, Galeomma 10 
Polyplacophora 77, 78, 80 
pomatia, Helix 68 
Pomatiopsidae 1 43-342 
Pomatiopsinae 148, 153, 324 



INDEX 



375 



Pomatiopsini 153, 324 
Pomatiopsis 324 
Prosobranchia 77, 79, 81 , 82 
Pseudobythinella 143, 154, 155, 161, 162- 

163. 166. 301 

chinensis 148. 166. 169-170. 173. 175 : 
757, 167, 168, 170-174 

daliensis 161, 166, 183 

j i an ou en sis 162, 163 

kunmingensis 155, 161, 166, 183 

Hui 163 

shimenensis 150, 163, 173. 175. 178. 
180. 182-183. 186 ; 157, 176-179, 
181-185 
Pseudobythinellini 148, 153, 154-155 
Pseudopythina 20 
Pseudoxerophila 1 09- 1 1 2 
Pulmonata 77, 79, 81, 82 
pustulosus, Leptodrilus 139 
radiata, Lampsilis 356, 365-367 
radiata, Lampsilis radiata 356, 358-367, 

369 
Radix 187, 271 

rebeli, Albinaria 34, 39, 40, 47-51 , 61 
Rehderiellinae 186 
reinae, Schi ley kiella 126 
reticulatus, Agriolimax 68, 71 
retifera, Rhamphidonta 21 
retowskii, Helicopsis 110 
Rhamphidonta retifera 21 
Robertsieila 184, 187, 199, 324, 330-336 
Rostroconchia 77, 78, 80 
rubens, Helix 109, 123 
Rumina decollata 45, 47 
Saganoa 155 
Scaphopoda 77, 78, 80 
Schileykiella 126 

perlatoris 1 26 

reinae 1 26 
Scintilla 1, 6 
Scintillona 17, 20 
Semerula maculata 352 
Semisulcospira 187, 271 
senilis, Albinaria 34-38, 40, 41, 52, 54-55, 

57 
senilis, Albinaria senilis 34-36, 38, 39, 41 , 

42, 46-51 , 54-57, 60, 61 
shimenensis, Pseudobythinella 150, 163, 

173. 175. 178. 180. 182-183. 186 ; 157, 

176-179, 181-185 
Sibirobythinella 153 
spadae, Candidula 109 
spiruloides, Helicella 111 
squill i na, Parabornia 20, 21 
stagnalis, Limnaea 25, 26, 32 
Stenothyra 199 

hunanensis 187, 271 
stolismena, Helix 109 
striata, Helicopsis 110, 117 
striata, Helix 109, 120 
subcalcarata, Helicopsis subcalcarata 120 
takii, Galeomma 10 
Tentaculitida 77 
teres, Albinaria 39, 47, 50, 51 
texanus, Circulus 1, 15 
Theba pisana 87-97, 343, 350 



thermophilus, Bathymodiolus 137 
tonkinianus, Lithoglyphus 187 
Trichia 109-112, 123 
Trichünae 1 26 

Tricula 146, 148, 186, 196, 218, 266, 279, 
286 . 301, 315, 320, 322-328, 332-336 
aperta 209 
bambooensis 266, 286, 316-317, 320, 

322, 325-328, 339-342 
bollingi 286, 314, 315-317, 320, 322, 

325-328, 339-342 
burchi 186 
cristella 211 

gredleri 150, 152, 155, 246, 286. 288- 
293. 295-297 . 315-318, 320-322, 
325-328; 286-290, 292-296 
gregoriana 286, 297, 316-317, 319, 

320, 322, 325-328, 339-342 
hudiequanensis 218, 286, 316-317, 

320, 322, 325-328, 339-342 
ludongbini 286, 31 5-31 7, 320, 322, 

325-328, 339-342 
maxidens 150, 155, 286, 297, 300- 
302, 304-305, 316-318, 320, 322, 
325-328; 298, 299, 301, 303-307 
minutoides 259 
montana 286, 297, 316-317, 319, 320, 

322, 325-328, 339-342 
odonta 1 50, 286, 297, 301 . 305. 307. 
311-312. 314-315 . 316-318, 320, 
322, 325-328; 298, 308-310, 312, 
313 
xiangfengensis 201, 286, 315-317, 320, 

322, 325-328, 339-342 
xiaolongmenensis 286, 301, 316-318, 
320, 322, 325-328, 339-342 
Triculidae 1 53 
Triculinae 149, 150, 153, 183, 186, 218, 

266, 324, 334-336, 339-342 
Triculini 150, 266, 279, 324, 327, 330, 333 
trivolvis, Planorbis 25 
troglodytes, Divariscintilla 1, 15-21 
turbinate, Helix 109 
turcica, Helix 120 
uenoi, Akiyoshia 155 
Umbrella mediterránea 25 
unifasciata, Candidula 109 
unifasciata, Helix 109 
Unionidae 355 
variabilis, Helix 109 
Vasconiella 1 9 

jeffreysiana 19, 20 
Venus mercenaria 352 
virgata, Cemuella 110 
virgata, Cochlea 109 
viridulus, Guoia 146, 150, 187. 190-196. 
198-201. 203 . 208, 209; 188-191, 196, 
197, 199, 201-208, 210, 272 
viridulus, Lithoglyphopsis 187, 205 
viridulus, Lithoglyphus 186, 187 
Viviparidae 187, 271 
watanensis, Bythinella 163 
Wuconchona 183, 184, 320-322, 325-328, 
330-336 

niuzhuangensis 316-318, 320-322, 325- 
328, 339-342 



376 INDEX 



wufenensis, Bythinella 163 
Xerolenta 107, 109, 112, 126 
Xeroleuca 120 
Xeromunda 107-112, 126 
Xerosecta 109, 111, 112, 124 
Xerotricha 108, 109, 111, 112, 120 

apiana 111, 120, 126 

conspurcata 107, 111, 120, 126 

nubivaga 1 20 
xiangfengensis, Tricula 201, 286, 315-317, 

320, 322, 325-328, 339-342 
xiaolongmenensis, Tricula 286, 301, 316- 

318, 320, 322, 325-328, 339-342 
yoyo, Divariscintilla 1, 5, 6, 15-22 
yunnanensis, Akiyoshia (Saganoa) 155 
yunnanensis, Lacunopsis 282 



MALACOLOGIA, VOL. 34 
CONTENTS 

PHILIPPE BOUCHET & JEAN-PIERRE ROCROI 

Supraspecific Names of Molluscs: A Quantitative Review 75 

ROBERT H. COWIE 

Shell Pattern Polymorphism in a 13-Year Study of the Land Snail Theba 
pisana (Müller) (Pulmonata: Helicidae) 87 

GEORGE M. DAVIS, CUI-E CHEN, CHUN WU, TIE-FU KUANG, XIN-GUO XING, 
LI LI, WEN-JIAN LIU & YU-LUN YAN 

The Pomatiopsidae of Hunan, China (Gastropoda: Rissoacea) 143 

V. K. DIMITRIADIS, D. HONDROS & A. PIRPASOPOULOU 

Crop Epithelium of Normal Fed, Starved and Hibernated Snails Helix luco- 

rum: A Fine Structural-Cytochemical Study 343 

VASILIS K. DIMITRIADIS & DIMITRIS HONDROS 

Effect of Starvation and Hibernation on the Fine Structural Morphology of 
Digestive Gland Cells of the Snail Helix lucorum 63 

J. P. A. GARDNER 

Null Alleles and Heterozygote Deficiencies Among Mussels (Mytilus edulis 

and M. galloprovincialis) of Two Sympatric Populations 99 

FOLCO GIUSTI, GIUSEPPE MANGANELLI & JORGE V. CRISCI 

A New Problematical Hygromiidae from the Aeolian Islands (Italy) (Pulmo- 
nata: Helicoidea) 1 07 

STEPHEN HUNT 

Structure and Composition of the Shell of the Archaeogastropod Limpet 
Lepetodrilus elevatus elevatus (Mclean, 1 988) 1 29 

TOSHIE KAWANO (CAMEY), KAYO OKAZAKI & LILLANE RÉ 

Embryonic Development of Biomphalaria glabrata (Say, 1818) (Mollusca, 
Gastropoda, Pianorbidae): A Practical Guide to the Main Stages 25 

TH. С M. KEMPERMAN & G. H. DEGENAARS 

Allozyme Frequencies in Albinaria (Gastropoda: Pulmonata: Clausiliidae) 

from the Ionian Islands of Kephallinia and Ithaka 33 

PAULA M. MIKKELSEN & RÜDIGER BIELER 

Biology and Comparative Anatomy of Three New Species of Commensal 
Galeommatidae, with a Possible Case of Mating Behavior in Bivalves 1 

ALAN E. STIVEN AND JOHN ALDERMAN 

Genetic Similarities Among Certain Freshwater Mussel Populations of the 
Lampsilis Genus in North Carolina 355 



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VOL. 34, NO. 1-2 MALACOLOGIA 1992 

CONTENTS 

PAULA M. MIKKELSEN & RÜDIGER BIELER 

Biology and Comparative Anatomy of Three New Species of Commensal 
Galeommatidae, with a Possible Case of Mating Behavior in Bivalves 1 

TOSHIE KAWANO (CAMEY), KAYO OKAZAKI & LILLANE RÉ 

Embryonic Development of Biomphalaria glabrata (Say, 1818) (Mollusca, 
Gastropoda, Planorbidae): A Practical Guide to the Main Stages 25 

TH. С M. KEMPERMAN & G. H. DEGENAARS 

Allozyme Frequencies in Albinaria (Gastropoda: Pulmonata: Clausiliidae) 

from the Ionian Islands of Kephallinia and Ithaka 33 

VASILIS К. DIMITRIADIS & DIMITRIS HONDROS 

Effect of Starvation and Hibernation on the Fine Structural Morphology of 
Digestive Gland Cells of the Snail Helix lucorum 63 

PHILIPPE BOUCHET & JEAN-PIERRE ROCROI 

Supraspecific Names of Molluscs: A Quantitative Review 75 

ROBERT H. COWIE 

Shell Pattern Polymorphism in a 13-Year Study of the Land Snail Theba 
pisana (Müller) (Pulmonata: Helicidae) 87 

J. P. A. GARDNER 

Null Alleles and Heterozygote Deficiencies Among Mussels {Mytilus edulis 

and M. galloprovincialis) of Two Sympatric Populations 99 

FOLCO GIUSTI, GIUSEPPE MANGANELLI & JORGE V. CRISCI 

A New Problematical Hygromiidae from the Aeolian Islands (Italy) (Pulmo- 
nata: Helicoidea) 1 07 

STEPHEN HUNT 

Structure and Composition of the Shell of the Archaeogastropod Limpet 
Lepetodrilus elevatus elevatus (Mclean, 1 988) 1 29 

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V. K. DIMITRIADIS, D. HONDROS & A. PIRPASOPOULOU 

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Genetic Similarities Among Certain Freshwater Mussel Populations of the 
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