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AMERICAN ic:
FERN 1
JOURNAL

PUBLISHED BY THE AMERICAN FERN SOCIETY

EDITORS

David W. Bierhorst
Gerald J. Gastony

David B. Lellinger

John T. Mickel

Terry R. Webster

MERCURY PRESS, ROCKVILLE, MARYLAND 20852

CONTENTS
Volume 73, Number 1, Pages 1-32, Issued March 31, 1983

Edgar T. Wherry and his
Contributions to Pteridology W. H. WAGNER, JR 1

Stone Fort at Fort Totten: Last Habitat
for Woodsia obtusa and Asplenium platyneuron
in Queens County, Long Island, New York?
ANDREW M. GRELLER and DAVID C. LOCKE 6

Starch Gel Electrophoresis of Ferns:
A Compilation of Grinding Buffers, Gel
and Electrode Buffers, and Staining Schedules
DOUGLAS E. SOLTIS, CHRISTOPHER H. HAUFLER,
DAVID C. DARROW, and GERALD J. GASTONY

Shorter Notes: A New Combination in Asplenium;
Adiantum furcatum and Microstaphyla furcata;
Noteworthy Pteridotphyte Records for Nebraska;
Spread of Marsilea quadrifolia in McDonough
County, Illinois; Wood Ferns New to Maryland
and ware; A New Combination for an

Asplenosorus Hybrid 28
American Fern Journal 32
Index to Volume 73 122
Errata 124

Volume 73, Number 2, Pages 33-64, Issued June 16, 1983

Vittaria Gametophytes Discovered
in a New Physiographic Province ALLISON W. CUSICK 33

Chapman’s Quillwort Reconsidered BRIAN M. BOOM 39

C-Glycosylxanthones in Diploid and Tissue
Culture-induced Autotetraploid Davallia fejeensis
P. MICK RICHARDSON and HARINDI.2 K. PALTA 43

The Distribution of Woodwardia areolata R. CRANFILL 46

Two Moonworts of the Rocky Mountains:
Botrychium hesperium and a New Species
Formerly Confused with It W. H. WAGNER, JR. and FLORENCE S. WAGNER 53

Shorter Notes: Lycopodium complanatum and
L. annotinum Found in the Black Hills;
Microfibrils in the Xylem of Blechnum
viviparum; Bi i
Sporophylis in Isoétes from Rajasthan 62

Reviews 12, 45, 52

Volume 73, Number 3, Pages 65-96, Issued September 29, 1983

Observations on the Structure and Function
of Hydathodes in Blechnum Iehmannii JOHN S. SPERRY 65

A Reclassification of the Fern Genus Pyrrosia K. H. SHING 73

Notes on the Ecology and Development

of Plagiogyria fialhoi PAULO G. WINDISCH and MARILIA PEREIRA-NORONHA 79
The Ferns of Elden Mountain, Arizona MICHAEL D. WINDHAM 85
Donovan S. Correll (1908-1983) 94
Reviews 93-96

Volume 73, Number 4, Pages 97-124, Issued December 29, 1983

Polyploidy and Aneuploidy in Hypolepis,

and the Evolution of the Dennstaedtiales P. J. BROWNSEY 97
Pecluma, a New Tropical American Fern Genus MICHAEL G. PRICE 109
The Lady Fern, Athyrium filix-femina,

in Saskatchewan VERNON L. HARMS 117
Shorter Note 121
American Fern Journal 122
Index to Volume 73 122

Errata 124

AMERICAN ee
FERN 1
JOURNAL

PUBLISHED BY THE AMERICAN FERN SOCIETY

EDITORS

David W. Bierhorst
Gerald J. Gastony
David B. Lellinger
John T. Mickel

Terry R. Webster

MERCURY PRESS, ROCKVILLE, MARYLAND 20852

CONTENTS

Volume 74, Number 1, Pages 1-32, Issued April 16, 1984

Promotion of Apogamy in Matteuccia struthiopteris,

the Ostrich Fern P. VON ADERKAS
A New Filmy Fern from Puerto Rico GEORGE R. PROCTOR
Chromosome Numbers of Neotropical Isoétes R. JAMES HICKEY
Two New Phenolic Glycosides in Asplenium septentrionale FILIPPO IMPERATO
A Remarkable Cyathea Hybrid R. E. HOLTTUM
The Ligule of Isoétes B. D. SHARMA and R. SINGH

Shorter Notes: Two Species of — Newly
Escaped in Florida; A New Statio
Trichomanes petersii in ralecie ee geen
Gametophytes at Bartholomew’s Cobble

Reviews 6, 18,

Volume 74, Number 2, Pages 33-64, Issued July 18, 1984

A Western Holly Fern, Polystichum x scopulinum
in Newfoundland WARREN H. WAGNER, JR. and ERNEST ROULEAU

Drymoglossum Under Water Stress C. S. HEW

Problems in Asplenium, with Some New Species
from Ecuador ROBERT G. STOLZE
Frequency of Cyanogenesis in Bracken
in Relation to Shading and volute Severity
I. SCHREINER, D. NAFUS and D. PIMENTEL

New Combinations and Some New Names in Ferns DAVID B. LELLINGER
Gualterio Looser (1898-1982)
Shorter Notes: Equisetum ramosissimum in Louisiana;

Three New Combinations in Lox oxogramme; Notes on
North American Ferns, II; Graves’ Spleenwort
in Ohio

36, 50,

Suggestions to Contributors

Volume 74, Number 3, Pages 65-96, Issued December 6, 1984

The Habitat Characteristics and Abundance
of Equisetum x ferrissii and its Parent Species,

Equisetum hyemale and Equisetum laevigatum, in Iow

LORENZ M. RUTZ and DONALD R. FARRAR 65

The Organic Nutrition of Botrychium Gametophytes

Anti-microbial Activity ne Phenolic Acids
in Pteridium aquilinu

DEAN P. WHITTIER 77

MICHAEL SAN FRANCISCO and GILLIAN COOPER-DRIVER 87

Reviews

86, 96

Volume 74, Number 4, Pages 97-124, Issued December 28, 1984

The Identification of Hawaiian Tree Ferns
of the Genus Cibotium

Two New Tree Ferns from Panama

Trunk Length and Frond Size in a Population
of Nephelea tryoniana from El Salvador

An Unusual New Elaphoglossum from Peru
New Tropical American Ferns

Shorter Note: aE Urban Locality for
Asplenium euron

1985 A.1.B.S. Meeting—Call for Papers
American Fern Journal

Index to Volume 74

Errata

RICH BECKER 97

ROBERT G. STOLZE

RALPH L. SEILER

ROLLA TRYON

JOHN T. MICKEL

101

AMERICAN ae
FERN _
JOURNAL |

QUARTERLY JOURNAL OF THE AMERICAN FERN SOCIETY

Edgar T. Wherry and his
Contributions to Pteridology W. H. WAGNER, JR. 1

Stone Fort at Fort Totten: Last Habitat
for Woodsia obtusa and Asplenium platyneuron
in Queens County, Long Island, New York?
ANDREW M. GRELLER and DAVID C. LOCKE 6

Starch Gel Electrophoresis of Ferns
A Compilation of Grinding Buffers, Gel
and Electrode Buffers, and Staining Schedules
DOUGLAS E. SOLTIS, CHRISTOPHER H. HAUFLER,
DAVID C. DARROW, and GERALD J. GASTONY 9

Shorter Notes: A New Combination in Asplenium;
Adiantum furcatum and Microstaphyla furcata;
*eoaaegeng Pteridophyte Records for Nebraska;

Marsilea quadrifolia in McDonough
Cea Illinois; Wood Ferns New to Maryland
and Delaware; A New Combination for an

Asplenosorus Hybrid 28
American Fern Journal 32
MissouR! BOTA

APRIL4:

The American Fern Society

DAVID B. LELLINGER, Smithsonian Institution, Washington, DC 20560. Journal Editor
ALAN R. SMITH, Dept. of Botany, University of California, Berkeley, CA 94720. Memoir Editor
JOHN T. MICKEL, New York Botanical Garden, Bronx, NY 10458. Newsletter Editor

setae Mire Journal

DAVID B. LELLINGER U = cat Herbarium NHB-166, Smithsonian Institution,
Washington, DC 20560.

TERRY R. WEBSTER ....... Biological Sciences Group, University of Connecticut. Storis. CT 06268.

The “American Fern Journal” (ISSN 0002-8444) is an illustrated quarterly devoted to the
study of ferns. It is owned by the American Fern Society, and — at the Smithsonian instieutile
Washington, DC 20560. Second-class postage paid at Washin
Claims for missing issues, made 6 months (domestic) to 12 month (foreign) after the date of issue,
and the matters for publication should be addressed to the Edito
Changes of address, dues, and applications for membership aia be sent to Dr. Judith. E. Skog.
Dept. of Biology, George Mason University, Fairfax, VA 22030. :
Orders for back issues should be addressed to Dr. James D. Montgomery, Ichthyological Associates, —
R.D. 1, Berwick, PA 18603.
General inquiries concerning ferns should be addressed to the Secreta
Subscriptions $9.00 gross, $8.50 net if paid through an = ( agency fee $0.50); sent free to —
members of the American Fern Society (annual dues, $8.00 + $4.00 mailing surcharge beyond U.S.A.,
Canada, and Mexico; life membership, $160.00). A
Back volumes 1910-1978 $5.00 to $6.25 each: single back numbers of 64 pages or less, $1.25; 65-80
pages, $2.00 each; over 80 pages, $2.50 each, plus shipping. Back volumes 1979 et seq. $8.00 each, —
single back numbers $2.00 each, plus shipping. Ten percent discount on orders of six volumes or more. f
Library :
Dr. John T. Mickel, New York Botanical Garden, Bronx, NY 10458. is Librarian. Members .
_ may borrow books at any time, the borrower paying all shipping costs. 4

Newsletter y

Dr. John T. Mickel, New York Botanical Garden, Bronx, NY 10458. is editor of the newsletter.
“Fiddlehead Forum.” The editor welcomes contributions from members and non-members. including —
miscellaneous notes, offers to — or purchase materials, personalia. horticultural notes, and
reviews, of non-technical books on

Mr} Neill D. Hall,
a 1230 None 8 Se le, W. a
oe collection lists Sem, A 98115, is = Spores exchanges

: in fers Botanical books, cs, back a OF the Journal, and cash or other ek i ways wel
a are tax-deductible. - and her gifts are al < wekonel all a
oe ni should be addressed to the Secretary. a ;

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 1 (1983) l

Edgar T. Wherry
and his Contributions to Pteridology'
W. H. WAGNER, JR.*

When he first became interested in ferns, Dr. Wherry was thirty years old. His
background had been in chemistry and geology, and at the time he was an assistant
curator in mineralogy at the U.S. National Museum. Some amateur botanists took
him to see a patch of the walking fern, Camptosorus rhizophyllus, where it was
growing on gneiss, and he became curious about its soil requirements. This led to
his first publication on ferns “A Chemical Study of the Habitat of the Walking Fern”
in 1916. After that Wherry became an expert on hydrogen-ion concentrations in the
soils of eastern United States ferns, and he also broadened out into questions of their
geography, ecology, and systematics. By the end of his career he had become an
authority on North American pteridophytes north of Mexico.

In 1917, Wherry transferred to the Department of Agriculture, where he came
under the influence of F. V. Coville, the Chief Botanist, who encouraged him in the
study of plants. By 1930, Wherry was prominent enough in his new profession to be
invited to join the Botany Department of the University of Pennsylvania. He
accepted this opportunity, which was ideal for him because it represented a return to
his alma mater and to his home city of Philadelphia. Wherry taught at the University
until his retirement in 1955. He stayed active for many years longer than most
botanists: he continued to take field trips well into his 80’s and was still writing
letters as late as his 90’s. Only in the last few years before his death at 97 on 19
May 1982 did he lose touch.

Wherry was very active in the American Fern Society, and three-fourths of his
pteridophyte publications were in the “American Fern Journal.” He was President of
the Society from 1934 to 1939, and he also took charge of preparing a cumulative
25-year index to the “Journal.” When Maurice Broun wrote his “Index to the Ferns
and Fern Allies of North America” (1938), Wherry supplied the habitat and range
data for most of the species. After becoming discouraged by the poor quality of the
manuals then available for identifying pteridophytes, he published one himself
entitled “Guide to Eastern Ferns” (1937), which, together with the second edition,
sold more than 6000 copies and did much to stimulate the popularity of ferns among
naturalists and botanists. Later he wrote another valuable guide, this one for the
southeastern United States. The royalties from his field manuals were donated to the
American Fern Society.

Wherry did not become a major botanist of his time: he was not involved in any of
the “big ideas” such as biosystematics, cytotaxonomy, and phylogeny that burgeoned
during his lifetime. His approach to the study of plants was deficient in such

*Department of Botany and Herbarium, University of Michigan, Ann Arbor, MI 48109.

'A series of papers in tribute to Edgar T. Wherry (19 Sep 1885-19 May 1982) was published in
Amer. Fern J. 66:33-80. 1976. Biographical notes have been published in Fiddlehead Forum 9:32-33.
1982 and by Prof. Wagner in Bull. Torrey Bot. Club 109:545-548. 1982. An obituary for Dr. Wherry
has been published in Bartonia, 49:1—5. 1983, the journal of the Philadelphia Botanical Club.

2 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

subjects as morphology, anatomy, cytology, and physiology, and he used the
laboratory and the microscope very little in his research. He functioned primarily
as a “field naturalist,” as evidenced by his teaching and his publications. Over 40%
of his fern bibliography dealt with geographical distributions, especially range
extensions and reports of what he encountered on his numerous exploring trips.
Among the many individuals who accompanied him on his pteridological field

expeditions were Harry W. Trudell, J. E. Benedict, and myself. For over a third of a ©

century, from high school student to university professor, I had the good fortune to
go exploring from time to time with Wherry and to absorb his ideas of pteridophyte
ecology.

In the “Fern Guide” he wrote as follows: “Sometimes .. . a fern student may
have the ‘thrill that comes once in a lifetime’ by discovering a species in an area
where it had not previously been found.” Wherry passed his love for finding rare
pteridophytes to others, and when the University of Virginia Mountain Lake
Biological Station began its pteridology course in 1962, Wherry wrote to the class
these inspiring words: “To the Mountain Lake Chapter, F. F. A. (Future Ferners of
America) Greetings: At last a real study of the ferns of Mountain Lake is under
way... . Many range extensions are to be expected there. Congratulations on
yours!” He was so right—that first class and the subsequent ones (including the
latest in 1982) made numerous range extensions of species in eight genera. The
success of field pteridology courses everywhere in the United States, especially at
Lake Itasca, MN, Highlands, NC, and at Flathead Lake, MT, owes a lot to Wherry’s
teaching about the roles of soil requirements and indicator species.

Wherry’s most significant contributions to the study of fern distribution and
ecology are those concerned with pH requirements, the subject which originally

stimulated his interest in ferns. He helped to design simple kits for making field —

tests of soil acidity, and he showed, for example, that certain traditional reports of
species growing on particular rock types were wrong. One author copied another

that Asplenium bradleyi grows on limestone, and it took Wherry years to straighten

this out and to convince pteridologists that this species is confined to acidic rocks.

Thanks to his efforts to interest botanists in pH, it has become standard today for
field workers to specify rock and soil types upon which plants grow. And if we wish
to find a particular species, we first locate the proper substrate. We know now that it —
would be practically impossible for Asplenium montanum to be found growing —

beside A. ruta-muraria; their edaphic preferences are too different.

Oddly, Wherry paid little attention to the positive effects of disturbance and —
succession On pteridophytes, even though his viewpoint in general was broadly —
ecological. He did not seem to appreciate how influential upsets such as floods,
fires, and grazing may be in enhancing the abundance and spread of many species, —

including botrychiums, certain dryopterids, and lycopods. Often, when a certain

species disappeared from its habitat, Wherry would attribute it to vandalism or a

over-collecting when it was more probably a matter of simple succession.

When he tried, in his greenhouse, to duplicate the natural growth conditions of |
saxicolous ferns by faithfully assembling the appropriate rocks, he almost invariably _
failed. Had he only known what we now know, namely that most rock ferns can

f
E

‘

W. H. WAGNER, JR.: EDGAR T. WHERRY 3

grow merely in soil-in pots, controlling the pH by simply mixing in limestone chips
or sphagnum peat, he could have learned much more about the characters of rock
ferns by culturing them side-by-side under uniform conditions. Practically everything
he knew about the species he studied came from field observations.

Wherry’s main contribution to pteridophyte taxonomy involved detecting hybrids
in the wild. He was opposed to the wanton designation of trivial forms and varieties
as illustrated by Fernald in “Gray’s Manual” (1950), and he was effective in steering
serious fern students away from this. His argument in “The Fern Guide” (1961) was
that “since these mostly occur in association with typical plants—indeed often
arising from the same rootstocks—they are not named.” He distinguished between
geographical varieties (what some botanists today call subspecies) and true species,
as illustrated by his treatment of the decompound species of Dryopteris, such as D.
spinulosa (= D. carthusiana) and D. intermedia, which by Fernald and his
associates had been interpreted as varieties, D. spinulosa, typical variety, and D.
spinulosa var. intermedia. He developed his concepts on the basis of studies of
natural populations, long before modern experimental and biochemical techniques
were available to confirm them.

Together with W. D. Gray, Wherry presented illustrations of intermediates, which
he interpreted as hybrids, between long recognized species of spleenworts. Appar-
ently, however, neither Gray nor Wherry was aware, or even suspected, that two of
the most important of the species then recognized, A. Xbradleyi and A.
x pinnatifidum, were hybrids themselves (Wagner, W. H., Jr., Evolution 8:103—118.
1954). Both of these men, on the contrary, accepted the hybridity of the fertile A.
x ebenoides from Alabama. The most interesting of the hybrids first recognized by
Wherry is A. Xtrudellii (A. montanum X pinnatifidum, a beautiful example of
hybrid vigor in a “sterile hybrid” which evidently can reproduce by unreduced
spores. Wherry’s work on Asplenium is honored in the name A. Xwherryi D.
Smith, the backcross of A. X bradleyi to A. montanum.

Under Dryopteris he distinguished and/or named a number of hybrids which since
have become important in the genome analysis of the basic species of eastern
American wood ferns. Some of these are common ferns widespread in forests and
swamps. They include D. x slossonae (D. cristata X marginalis), D. X triploidea
(D. intermedia X carthusiana), and D. X benedictii (D. clintoniana X carthusiana).
What he named D. /eedsii and interpreted to be D. goldiana X marginalis has
since turned out to be D. celsa X marginalis (Wagner, W. H., Jr., Virginia
Polytech. Inst. State Univ. Res. Div. Monogr. 2:147-192. 1970). His greatest
difficulties with wood ferns involved the taxa resembling D. x clintoniana, the
same ones that cause us trouble today. He nonetheless had a sharp eye for them and
was the first person to recognize as distinct D. x australis (D. celsa x ludoviciana),
although for good reasons at the time he initially interpreted it as a southern variety
of D. clintoniana. His contributions to our knowledge of Dryopteris hybrids are
commemorated in the name D Xneo-wherryi Wagner, which is the true hybrid of
Goldie’s Wood Fern and the Marginal Wood Fern.

Scientifically, Wherry’s influence on American pteridology was substantial. Many
of today’s pteridologists are indebted to his teachings as well as his inspiration. His

major contributions involved his investigations of the pH reactions of soils, distribu-
tion patterns, and his studies of various taxa, especially spleenworts and wood ferns.

The

than three decades. For the general public, Wherry’s field manuals and his many
lectures and guided trips for amateurs helped engender a widespread enthusiasm for
the natural history of pteridophytes.

1916.

1917.
1920.
1920.

published j

. The soil reactions of certain rock ferns—II. Amer. Fern J. pasar
: ag gravesii in Pennsylvania. Amer. Fern J. 10:119-121.

: Wall ferns in Wilmington, North Carolina. Amer. Fern J.

. Ferns of eastern West Virginia, I. Amer. Fern J. Pay ake

. Some fern finds in Virginia. Amer. Fern J. 15:1-7.

. The Appalachian aspleniums. Amer. Fern J. 15:46—54.

. Soil reaction preferences of three adder’s-tongues. Amer. Fern J. 16:1—3.

. The West Virginia locality of the southeastern relative of Woodsia scopulina. Amer. Fern J.
)

ray
. A fernless area. Amer. Fern J. 17:63-64.
. Observations on the woolly lipfern. Amer. Fern J. 16:107-109.
. Ferns of Dripping Spring, Oklahoma. Amer. Fern J. 18:61-63.
. Notes on Asplenium Trudelli. Amer. Fern J. 17:135-138.
. Further occurrence of the Alleghany cliff-fern. Amer. Fern J. 19:101—102.
. The Asplenium ebenoides locality near Havana, Alabama. Amer. Fern J. 11:30-33. (with H. W.
ll

- Asplenium bradleyi erroneously reported on limestone again. Amer. Fern J. 21:111—113. 4
. Range-extensions and other observations, 1931-32. Amer. Fern J. 22:79-86. ;
- Fern field notes, 1933. Amer. Fern J. 23:109-112.

. Fern field notes, 1934. Amer. Fern J. 24 97-104
. Fern field notes, 1935. Amer. Fern J. 25: 123-126.

. The northern limits of Polypodium polypodioides in the east. Claytonia 2:32.
. Fern field notes, 1936. Amer. ren, 26:127- ae

: Cand eae Aner Fem J. 28: 125-140, ot. Hz.
. Recent fern finds in West Virginia. Castanea 4:14.

:
:
4
fs a
7A full aden Dr. Wherry, including his papers on flowering plants and —— has been a

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

American Fern Society is indebted to him for his help in many ways over more

ee ee eS eS Ne

PTERIDOPHYTE BIBLIOGRAPHY OF EDGAR T. WHERRY’

A chemical study of the habitat of the walking fern, Camptosorus rhizophyllus (L.) Link. J.
Washington Acad. Sci. 6:672-679.

Observation on the habitat of certain ferns. Amer. Fern J. 7:110—-112.

A fruitless search for Asplenium fontanum in ingore Amer. Fern J. 10:90-91.

The soil reactions of certain rock ferns—I. Amer. Fern J. 10:15—22.

a
alia a clean

soil reactions of the ferns of woods and swamps. a al J. 11:5-16.

shel a al hale Ae le Ls el gbila

Wood-ferns on Mt. Desert Island, Maine. Amer. Fern J. 16:3-7.

16:92-95. (with F. W. Gr.

Be oP ot i acre tae de

Trudell)

eee ee pes ee

ESR SRG eee Ce REE 3

in Bartonia, ‘9: 6-14. 1983, the journal of the Philadelphia Botanical Club

|
‘
F
3
4

Ree ne AE LE Oe TT RR TT TO eA TMS PON Ne ETE CRM I Ge eR SON Ns PEE REE TAT eR EO CRE a ET IN NO CE ST EO A BE CE EN eT Te See ee

W. H. WAGNER, JR.: EDGAR T. WHERRY

1940.
1941.
. Two Virginia fern records. Virginia J. a —

. A woodfern hybrid deserves a name. Bartonia 21:1—2, pl. 1. fig. 1

. The ferns and lycosphens of paras ‘Bertie 21:11-63, pl. 1, fig. 2.

. Lycopodium sabinaefolium in Pennsylvania. — nip 9 32:111-113.

. Observations on Florida ferns. Amer. Fern J. 32:139-145.

. The discoveries of new Pennsylvania ferns. oo ae J. 32:148-149.

. Guide to Eastern Ferns, 2nd ed. Science Press Printing Co.. Lancaster, PA.

. Note on the southeastern relatives of oi iia inundatum. Amer. Fern J. 34:24.
. Cystopteris bluff. Amer. Fern J. 34:92-9

. Osmunda cinnamomea f. ile Amer. Fern J. 34:94—95.

. The indument of Cystopteris fragilis. Amer. Fern J. 35:54—55.

. Notes on Illinois pteridophytes. Amer. Fern J. 35:92.

. Our most-renamed native fern. Amer. Fern J. 35:128.

. Notes on Muhlenberg’s ferns. Amer. Fern J. 36:54-58.

. Selaginella rupestris in Pennsylvania. Amer. Fern J. 37:24-25.

Five recent books on ferns. Bartonia 20:38.
Asplenium adiantum-nigrum in Arizona. Amer. Fern J. 31:97—100.

Our easternmost Cheilanthes species. Amer. Fern J. 37:77-79.

. A crispate variant of the Christmas ne Amer. Fern J. 37:121.
. Remarks on the American lady ferns. Amer. Fern J. 38:155—158.
. Guide to Eastern Ferns. Univ. of Hasta Press, Philadelphia.
. Geographic notes on the bog fern. Amer. Fern 9:18-19.
0. A new interpretation of the Dryopteris clintoniana ~ Amer. Fern J. 40:118—120.
. Observations at Bartholomew’s Cobble. Amer. Fern J. 41:13-14.
. Lowland lycopodiums in the New Jersey ay ore ai Torrey Bot. Club 81:364.
4. Nomenclature of the oak-ferns. Amer. Fern J. 44:85—86.

How I became interested in ferns. Amer. Fern J. 50:225—228.

. The Fern Guide. Doubleday, Garden Ci oe

. Supplementary note on Dryopteris hybrids. Amer. Fern J. 51:33-35.

. Native ferns in the garden. Plants and Garden 18:

. The Southern Fern Guide. Doubleday, Garden City, NY,

. Some new name-combinations for southeastern ferns. Amer. Fern J. 54: 143-146.
. Southern records of Ophioglossum vulgatum. Amer. Fern J. 58:182-183.

6 AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 1 (1983)

Stone Fort at Fort Totten: Last Habitat
for Woodsia obtusa and Asplenium platyneuron
in Queens County, Long Island, New York?
ANDREW M. GRELLER* and DAVID C. LOCKE**

In the course of documenting vegetation on the northeastern coast of Queens .
County, Long Island, New York, the senior author discovered two ferns, Asplenium —

platyneuron (L.) B.S.P. and Woodsia obtusa (Spreng.) Torrey, in what may well be —

their last remaining habitat in Queens County (Borough of Queens, City of New

York). They were found in the weathered cement between large blocks of granite ©
used to construct the walls of the “Stone Fort” at Fort Totten, a 19th Century coastal —

fortification on the Willets Point peninsula (40°47'30" N, 73°46’40” W). The Stone

Fort now forms Willets Point, rising from the shore level to a height of thirty feet. It
was built in 1862-66 as part of the seacoast defenses of New York City; photographs
and a brief history of the Stone Fort were published by Alperstein (1977).

Reconnaissances of the site were undertaken in the spring of 1973, in the late spring —
and early summer of 1978, and during the winter of 1981-82. Occasionally a few |

plants were measured, but a census was not attempted.

Soil samples were obtained adjacent to fern roots at two sites on the landward side —
of the granite wall, for elemental analysis. Stones, sticks, and obvious plant matter —

were separated, and a portion of well-mixed soil was ground in a mortar to a fine

powder. A 1.00 g sample of each was dry-ashed at 450°C in a muffle furnace, anda —

semi-quantitative elemental analysis conducted with a Baird Atomics Emission

Spectrograph. A just-saturated slurry in distilled water was made of each raw soil —
sample, and the pH measured with an Orion combination electrode and a Corning —

pH meter.

Further tests for sodium and chloride, indicators of saline spray, were made on
samples taken near Woodsia roots, one from the seaward side of the granite wall, —
and one from the landward side. Soil samples, 1.00 g, were extracted with 10.0 ml —
of distilled, deionized water. The sodium content was determined by atomic —
absorption spectroscopy using a Perkin-Elmer 303, and chloride content by ion- —
selective electrode using an Orion electrode and a Beckman expanded scale pH ~

meter.

Voucher specimens were deposited at the Herbarium of the Brooklyn Botanic |

Garden (BKL).
RESULTS

Fewer than twenty-five individuals of A. platyneuron were observed during the —
most recent visit (30 March 1982). These were concentrated around the central —
stairway connecting the three levels of the fort, between the main and upper levels. —
in a sheltered, moderately well lighted location. A few individuals were scattered —
Over the rest of the main level. The plants grew from partially decomposed cement —

“Department of Biology, Queens College, City University of New York, Flushing, NY 11367.
Department of Chemistry, Queens College, City University of New York, Flushing, NY | 1367.

—.

GRELLER & LOCKE: STONE FORT AT FORT TOTTEN 7

in joints between granite blocks. They were well rooted in the joints, to perhaps 10
cm deep. Fronds seldom exceeded 10 cm in length.

The population of W. obtusa numbered several hundred individuals. Plants
occurred in the horizontal joints of the seaward granite wall of the fort, where sea
spray and salt-laden air are features of the environment. More commonly, individu-
als occurred in both vertical and horizontal joints in the complex of granite walls on
the main, landward side. Plants of Woodsia appeared to be less deeply and securely
rooted than those of Asplenium. Commonly, young sporophytes were seen attached
to gametophytes. Woodsia plants in winter bear a rosette of sterile fronds approxi-
mately 3-4 cm long. Fertile fronds are produced during the growing season; these
regularly reach 30 cm or longer.

The substrate of both ferns consisted of a mixture of black humus and light gray
fragments of cement and sands. The soil evidently arises from the mortar decomposed
through weathering and from organic materials deposited by the ferns and other
organisms which have gained a foothold (Segal, 1969). This is supported by our
chemical analyses. The results of the dry-ashing test show that organic materials
account for nearly 40% by weight of the soil sample. In addition, the emission
spectrographic elemental analyses of the inorganic portions of the wall soil samples
reveal silicon, calcium, and aluminum to be the major constituents and an overall
composition quite similar to that of Portland cement. Portland cement has been in
widescale use since at least 1850 (Lea, 1971), and apparently was used as the
mortar for the Stone Fort. Although raw, wet cement is highly alkaline (pH 11-12),
acidic agents in rainwater and in humic substances from decayed plant materials act
to neutralize the alkalinity. Our soil samples are neutral (pH 6.96—7.00); these pH
values are typical of well-aged wall soils (Segal, 1969).

Results of the sodium and chloride analyses, in wg/g of soil (ppm) are: landward,
250 ppm Na and 110 ppm Cl; seaward, 1200 ppm Na and 280 ppm Cl. The seaward
wall soil has nearly five times as much sodium and more than twice as much
chloride as the soil from the landward wall, confirming the influence of salt spray on
the seaward side.

DISCUSSION

It is uncertain when A. platyneuron and W. obtusa first appeared in the mortar of
the Stone Fort. Little of the mortar remains unweathered, and extensive sections are
missing. It is possible that the latter sections once supported populations of the
ferns. It cannot be decided on the basis of the available data whether it is the high
calcium content or the neutral pH of the soil that accounts for the presence of the
ferns; both conditions are uncommon on Long Island.

We inspected herbaria at New York Botanical Garden, Brooklyn Botanic Garden,
and Planting Fields Arboretum, and consulted with staff of the New York State
Herbarium, Albany. Woodsia obtusa has been collected at Ridgewood, in Queens
County (G. B. Brainerd in 1866, BKL 022985); on rocks near Greenport, in Suffolk
County, Long Island (BKL 1862); and in “dry woods” there (R. Latham in 1918,
NY 824). The Fort Totten collection (Greller in 1973, BKL) is the first Woodsia
from stone walls on Long Island. Edward Frankel (pers. comm.) reports W. obtusa

8 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

as common on old walls in Westchester County, north of Long Island. Woodsia was —
collected “on - of oe ice chute, south side” at Rockland Lake, Clarkstown
Township, Rocklan , New York (J. H. Lehr in 1954, NY), also north of —
Long Island. ee mee listed A. platyneuron as “frequent throughout Long —
Island.” Nevertheless, Greller (1977, 1979) did not encounter it in the only
remaining extensively forested sites in Queens County: Cunningham, Forest, and —
Alley Parks. Until they are reported elsewhere, the Fort Totten populations of W. —
obtusa and A. platyneuron are the only ones documented in Queens County. Their ©
absence from former terrestrial habitats may be caused by a decrease in soil pH,
perhaps as a result of acid rain. Precipitation pH in Queens County since 1970 has —
averaged about 4.0, monthly averages ranging from 3.5 (summer) to 4.5 (spring and
fall) (D. C. Locke, unpubl.). i
Occurrence of W. obtusa on the seaward wall is noteworthy because of the
relatively high levels of sodium and chloride which presumably result from salt
spray. Based on salinity measurements made in Hempstead Harbor, a similar bay a
few miles east of Fort Totten, the salinity of Long Island Sound at Fort Totten is
about 25%e (D. C. Locke, unpubl.), less than that of ocean water (35%c) but more
than sufficient to produce saline spray. The relatively higher level of sodium than |
chloride in both samples presumably reflects differential leaching of the two —
elements; according to Lindsay (1979), the level of sodium in soils is typically —
higher than that of chloride.
The authors gratefully acknowledge the assistance of CWO-4 Jacob M. Fein U. S.
Army (ret.), who acted as our guide and gave us historical information, reprints, and
photographs of the Stone Fort; Thomas J. Delendick, Asst. Taxonomist and Curator —
of the Herbarium, Brooklyn Botanic Garden, who helped us locate specimens; —
Richard Posner, U. S. Testing Company, for emission spectrographic analyses; and
Scott Marcus, for field assistance.

LITERATURE CITED

eS D. M. 1977. Fort Totten at Willets Point. Periodical (Council on Abandoned Military

ee Arizona) 9(2):41—-52. :

GRELLER, 5 M. 1977. A vascular Ha of the forested portion of Cunningham at Pie County,

New York, with notes on the vegetation. Bull. Torrey Bot. Club 104:170- 7

- 1979. A vascular flora of the forested portion of Cunningham Park, oaecis County, be a

York: Corrections and additions. Bull. Torrey Bot. Club 106:45. a
JELLIFFE, W. E. 1899. The Flora of Long Island. Privately Printed, Lancaster, PA.

LEA, F. M. 1971. The Chemistry of Cement and Concrete, 3rd ed. Chemical Publishing Company. —

New York.
LINDSAY, W. L. 1979. Chemical Equilibrium in Soil. Wiley, New York.
SEGAL, S. 1969. Ecological Notes on Wall Vegetation. W. Junk, The Hague, Netherlands.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 1 (1983) 9

Starch Gel Electrophoresis of Ferns:
A Compilation of Grinding Buffers,
Gel and Electrode Buffers, and Staining Schedules

DOUGLAS E. SOLTIS*, CHRISTOPHER H. HAUFLER**,
DAVID C. DARROW***, and GERALD J. GASTONY***

The homosporous pteridophytes have been largely uninvestigated by electrophore-
sis, despite the fact that they offer many exciting research possibilities (Soltis et al.,
1980). The paucity of electrophoretic studies of ferns and fern allies may be due in
large part to the high concentrations of condensed tannins that many species contain
(Cooper-Driver, 1976 and pers. comm.). These compounds render enzymes inactive
by binding with them following cellular disruption, thereby frustrating researchers
who have attempted electrophoretic analysis utilizing standard methods of sample
preparation.

The method of sample preparation developed by Kelley and Adams (1977a, b) in
their analysis of enzyme variation in Juniperus was an important procedural
breakthrough in overcoming the difficulties that result from the liberation of large
amounts of phenolic compounds during tissue preparation. Recently, a simplified
version of that method was applied by Soltis et al. (1980) to fern leaf tissue,
facilitating rapid preparation of active enzyme samples and thereby making electro-
phoretic analyses of large numbers of individuals more feasible.

In an attempt to improve methods of analysis of fern enzymes in starch gel
electrophoresis, we have experimented with modifications of the method of sample
preparation outlined by Soltis et al. (1980). We also have examined several different
methods of sample preparation such as those of Gottlieb (1981a), Mitton et al.
(1979), and Werth et al. (1982), and have evaluated the relative merits of each with
fern tissue. Finally, during the course of our electrophoretic investigations of ferns
we found that standard gel and electrode buffers and staining schedules, such as
those of Brewer (1970) and Shaw and Prasad (1970), often provided unsatisfactory
results when applied to ferns. We have determined gel and electrode buffers, as well
as staining schedules, that provide clear starch gel enzyme banding for 22 enzyme
systems in ferns. Requests for advice resulting from the recent surge of interest in
fern enzyme electrophoresis have prompted us to compile our procedural data so that
other researchers can take advantage of our experimentation. We hope that these data
will stimulate more extensive electrophoretic investigation of pteridophytes and other
electrophoretically difficult taxa.

Gottlieb (1981b) recently reviewed aspects of enzyme electrophoresis primarily in
gymnosperms and angiosperms. His discussion is equally relevant to understanding
the potential applications and limitations of electrophoretic evidence in pterido-
phytes. Since homosporous pteridophytes have high chromosome numbers, it is
tempting to invoke polyploidy in interpreting their enzyme band patterns. It is well

*Department of Biology, University of North Carolina, Greensboro, NC 27412.
**Department of Botany, University of Kansas, Lawrence, KS
***D oy

partment of Biology. Indiana University, Bloomington, IN 47405.

TABLE |. ELECTRODE AND GEL BUFFER RECIPES USED SUCCESSFULLY IN ELECTROPHORETIC ANALYSIS OF FERNS (Gram
ted).

amounts are given for one liter final volume of buffer except where note

Electrode Buffer
1. 0.400 M Citric Acid, trisodium salt;
117.64 g Citric Acid, trisodium salt
dihydrate, 1.0 M HCI to pH 7.0

2, 0.135 M Tris, 0.032 M Citric Acid;
16.35 g Tris*, 6.10 g Citric Acid?

3. 0.135 M Tris, 0.017 M Citric Acid;
16.35 g Tris*, 3.35 g Citric Acid?

4. 0.223 M Tris, 0.086 M Citric Acid;
27.00 g Tris’, 4 52 g Citric Acid”,
NaOH to pH 7

5. 0.223 M Tris, 0.069 M Citric Acid;
27.00 g Tris*, 13.33 g Citric Acid”

6. 0.100 M NaOH, 0.300 M Boric Acid;
4.00 g NaOH, 18.55 g Boric Acid?

7. 0.038 M LiOH, 0.188 M Boric Acid;
1.60 g LiOH-H,O, 11.60 g Boric Acid”
Adjust to pH 8.3 with dry components

pH
@ 22°C
7.0

Gel Buffer
0.020 M Histidine-HCl; 4.19 g L-Histidine- HCI
monohydrate, 1.0 M NaOH to pH 7.0

0.009 M Tris, 0.002 M Citric Acid;
dilute 67 ml of electrode buffer to 1 liter

0.009 M Tris, 0.001 M Citric Acid;
dilute 67 ml of electrode buffer to | liter

0.008 M Tris, 0.003 M Citric Acid;
dilute 35 ml of electrode buffer to | liter

0.008 M Tris, 0.002 M Citric Acid;
dilute 35 ml of electrode buffer to | liter

0.015 M Tris, 0.004 M Citric Acid;
1.84 g Tris*, 0.69 g Citric Acid”

0.045 M Tris, 0.007 M Citric Acid,
0.004 M LiOH, 0.019 M Boric Acid;
Tris- yee: buffer (5.45 g Tris’,
ric Acid”, bring volume to
900 ml), aad 100 ml electrode buffer
to give 9:1 ratio, 1.0 M NaOH to pH 8.3

pH
22°C
7.0

(€861) £2 SWATOA “TWNYNO! NYA NVOININY

8. 0.039 M LiOH, 0.263 M Boric Acid; 8.0
1.64 g LiOH:H,O, 16.23 g Boric Acid”

9, 0.065 M L-Histidine free pis ca. Sc]
0.015 to 0.016 M Citric A
10.09 g L-Histidine free . Citric Acid? to
pH 5.7 (= ca. 2.9 to 3.1

10. 0.180 M Tris, 0.004 M EDTA, 0.100 M 8.6
Boric Acid; 21.80 g Tris*, 1.52 g
EDTA tetrasodium salt dihydrate,
6.18 g Boric Acid”, Boric Acid to
pH 8.6

11. 0.400 M Citric Acid, trisodium salt; 7.0
117.64 g Citric Acid, trisodium salt
2 Syiree. 1.0 M HCI to pH 7.0

0.042 M Tris, 0.007 M Citric Acid;
0.004 M LiOH, 0.025 M Boric acid;
5.04 g Tris*, 1.25 g Citric Acid”,

0.16 g LiOH:H,O, 1.56 g Boric Acid”,
1.0 M HCI to pH 7.6

0.009 M L-Histidine, 0.002 M Citric Acid;
dilute 140 ml of electrode buffer to | liter

0.045 M Tris, 0.001 M EDTA, 0.025 M Boric Acid;

dilute 250 ml of electrode buffer to | liter

0.005 M Histidine-HCI; 1.05 g L-Histidine-HC!
monohydrate, 1.0 M NaOH to pH 7.0

“Trizma-base (Sigma T1503; Sigma eh works rfl Ss oe is much less expens
PA

mounts of Citric Acid and Boric A re give

ve).
anhydrous free acid per liter.

“Check pH of s sion after mixing; at of Tris buffers ty chatae with dilutio

dit are from hal (198 1a).

SNY34 40 SISINOHdOYLIIT3 139 HOYWIS “IW 13 SLNOS 3G

12 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

known, however, that multiple forms of a given enzyme may be coded by different
alleles at a single locus (allozymes) or by genes at more than one locus (isozymes).
Furthermore, many standardly assayed plant enzymes have isozymes located in two
or more subcellular compartments (e.g., the cytosolic and chloroplastic isozymes of
PGI and PGM discussed by Gottlieb, 1981b, 1982—for interpretation of enzyme
symbols see Table 2). When found in pteridophytes, multiple, subcellularly com-
partmentalized isozymes should not be misinterpreted as products of duplicated loci
resulting from polyploidy.

According to Chapman et al. (1979), attempts to determine the amount of
heterozygosity per locus in homosporous pteridophytes are complicated by recombi-
nation between homoeologous duplicated loci. An obvious prerequisite to recombina-
tion of this kind is the actual presence of such loci. Gastony and Gottlieb (1982)
developed a means of demonstrating whether duplicated loci are, in fact, present in
pteridophytes. By electrophoretically analyzing sporophytes taken from nature and
individual gametophytes grown from spores of these sporophytes, segregational
analysis of sporophytic enzyme banding patterns can be conducted. This permits
genetic interpretation of parental sporophytic enzyme phenotypes without the time-
consuming crossing programs required by total reliance on sporophytic tissue. Use
of this methodology enables the investigator to determine whether apparent
heterozygosity of sporophytes is coded by alleles at a single locus or by genes at
duplicated (homoeologous) loci. Our application of this methodology to several fern
genera (e.g., Athyrium, Bommeria, and Pellaea) has demonstrated that the genetic
variability observed in these taxa results from allelic diversity and segregation at
single, not duplicated, loci.

MATERIALS

Living sporophytes of Athyrium filix-femina, Bommeria ehrenbergiana, B. hispida,
B. pedata, B. subpaleacea, Botrychium virginianum, Ceratopteris thalictroides,
Cystopteris bulbifera, C. dickieana, C. fragilis, C. laurentiana, C. protrusa, C.
reevesiana, C. tennesseensis, C. tenuis, Isoétes butleri, I. engelmannii, Lygodium
japonicum, Nephrolepis exaltata, Ophioglossum engelmannii, Pellaea andromedi-
folia, P. atropurpurea, P. glabella, Polypodium polypodioides, P. virginianum,
Polystichum acrostichoides, Pteridium aquilinum, Woodsia obtusa, and W. oregana
were maintained in greenhouse culture and utilized in this investigation. Living
gametophytes representing the species of Athyrium, Bommeria, Cystopteris, Pellaea
and Preridium listed above were cultured on nutrient agar (Gastony & Haufler, 1976)
and also provided material for this study.
GRINDING BUFFER SOLUTIONS
We routinely utilize modifications of either the phosphate grinding buffer—poly-
vinylpyrrolidone (PVP) solution employed by Mitton et al. (1979), the Tris—maleate
grinding buffer-PVP solution of Soltis et al. ( 1980), or the Tris-HCl grinding
buffer-PVP solution of Gottlieb (1981a) with PVP substituted for polyvinylpoly-
pyrrolidone (PVPP), as described below. Recipes for preparing these grinding buffer
solutions are provided below; molarity or percent volume values are provided and

D. E. SOLTIS ET AL.: STARCH GEL ELECTROPHORESIS OF FERNS 13

gram or milliliter amounts required to prepare 25 ml of buffer solution are given in
parentheses.

Phosphate grinding buffer-PVP solution.—0.029 M (0.28 g) sodium tetraborate, 0.017 M (0.08 g)
sodium metabisulfite, 0.20 M (1.0 g) L-ascorbic acid sodium salt, 0.016 M (0.07 g) diethyldithiocar-
bamic acid sodium salt, 4% w/v (1.0 g) PVP average molecular weight 40,000 (Sigma PVP 40T). or
36-40% w/v (9.0-10.0 g) PVP average molecular weight 10,000. Dissolve gram amounts in 25 ml of
0.10 M phosphate buffer pH 7.5 (to make 100 ml of phosphate buffer dissolve 1.36 g KH»PO, in H,O,
add 9.0 ml IM NaOH, and bring volume to 100 ml with H,O) and then add 0.25 ml (1%)
2-mercaptoethanol.

Tris—maleate grinding buffer-PVP solution.—0.20 M (1.91 g) sodium tetraborate, 0.02 M (0.095

sodium metabisulfite, 0.25 M (1.24 g) L-ascorbic acid sodium salt, 0.026 M (0.113 g)
diethyldithiocarbamic acid sodium salt, 0.10 M (0.29 g) maleic acid, 0.10 M (0.30 g) Tris. 4% w/v (1.0
g) PVP average molecular weight 40,000 or 32-40% w/v (8.0-10.0 g) PVP average molecular weight
10,000. For 25 ml of buffer, dissolve amounts indicated in 19 ml distilled water; mix thoroughly and
crush out lumps; adjust to pH 7.5 with 1.0 M HCI; add 0.025 ml (0.1%) 2-mercaptoethanol: add H,O to
25 ml. Originally (when we utilized PVP 10,000) it was necessary to allow the PVP to hydrate overnight
before using the grinding buffer solution. When employing PVP 40,000, it is possible to prepare the
grinding buffer solution immediately prior : sample preparation i stirring the PVP into solution.

Tris-HCl grinding buffer-PVP solution.—0.1% v/v (0.025 ml) 2-mercaptoethanol, 0.001 M
(0.010 g) EDTA (tetrasodium salt), jae M (0.019 g) potassium chloride, 0.010 M (0.050 g)
magnesium chloride hexahydrate, 4 or 20% w/v (1 or 5 g) PVP 40,000, 25 ml 0.10 M Tris-HCl! buffer,
pH 7.5. Stir the PVP into solution or allow it to hydrate in the buffer overnight.

TABLE 2. GEL AND ELECTRODE BUFFER SYSTEMS THAT WE HAVE FOUND TO YIELD THE
BEST BANDING IN FERNS WE HAVE ASSAYED.

Enzyme Symbol Gel and Electrode Buffer System
from Table |

Acid phosphatase APH 6,7

onitase ACN 15,9
Aldolase ALD 1-5, 7, 10, 11
Aspartate aminotransferase AAT (or GOT) 6, 7,8
Catalase CAT 4, 7, $,.10
Esterase (Colorimetric) EST 6.7
Esterase (Fluorescent) FE 8
Fructose- 1 ,6-diphosphatas Fl ,6DP 1, 11
Glucose-6-phosphate deyropenae G6PDH 4, 5, 6,7
Glutamate dehydro GDH 3, 5,7
Glyceraldehyde-3- tl

drogenase G3PDH 1, 11

Hexokinase HK 2, 35, 31 8
Isocitrate dehydrogenase IDH 1, 2,3, 455
Leucine aminopeptidase LAP 3, 7,8
Malate dehy n MDH 1, 4,5, 9
Malic enzyme ME 7,1
Peroxidase PER re
Phosphoglucoisomerase PGI 5, 6,7
Phosphoglucomutase PGM 7.0. 1.5

6-Phosphogluconate dehydrogenase -6-PGD 1, 2, 4,5
Shikimate dehydrogenase SkDH 12, 4,3, 0
Triosephosphate isomerase TPI 2,5, 6, 7,8

14 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Comparison of these three grinding buffer solutions indicates that for most
enzymes the staining results are highly comparable. For some enzymes, however,
the Tris-HCI-PVP solution seems to improve enzyme band clarity (e.g., PGI,
PGM, LAP), while for other enzymes the reverse is true. In Pellaea andromedifolia,
for example, PGI banding was sharp with the Tris-HCI-PVP solution but was
inhibited by the presence of ascorbic acid and sodium tetraborate in the
Tris—maleate-PVP buffer. In Athyrium filix-femina, however, the Tris—maleate—PVP
solution gives superior banding for PGI and one more observable EST locus when
compared to the Tris-HCI-PVP solution. The phosphate-PVP solution often result-
ed in reduced APH activity when compared to the other two grinding buffer
solutions.

In working with species of Asplenium, Werth et al. (1982) used a method of
sample preparation that uses caffeine, but not PVP. Following this technique, equal
weights of tissue and caffeine were ground in a 0.1 M HEPES pH 7.0 buffer with
0.2% 2-mercaptoethanol and 0.5% sodium metabisulfite to produce a slurry. We found
that for virtually all enzyme systems investigated, the method of Werth et al.
provided results roughly comparable to those obtained with grinding buffers contain-
ing PVP. The presence of 0.5% sodium metabisulfite in the grinding buffer appears
to be of great importance. When this ingredient is omitted from the buffer, activity
is noticeably reduced or almost totally lost for many enzymes. Significantly, the full
complement of PGI enzyme bands obtained with PVP was not expressed in most of
the fern taxa investigated when sodium metabifsulfite was omitted. These results are
in agreement with similar observations of Werth et al. (1982).

It should be emphasized that to obtain the best results, the appropriate amount of
PVP depends upon the taxon under investigation and in part upon the molecular
weight of the PVP employed. Soltis et al. (1980) utilized a grade of PVP with an
average molecular weight of 10,000 (although this is not stated in their report) and
incorporated 40% w/v PVP (4 g PVP in 10 ml of Tris—maleate grinding buffer). At
present, we routinely employ a grade of PVP with an average molecular weight of
40,000. Both molecular weights of PVP are suitable, but less PVP 40,000 is
required than PVP 10,000. The amount of PVP used is an important consideration
because, as reported by Soltis et al. (1980), use of excessive amounts of PVP in the
preparation of grinding buffer-PVP solutions frequently results in a decrease or
complete loss of enzyme activity. Although PVPP is effective with a wide range of —
angiosperms, its substitution for PVP failed to produce banding for PGI in Pellaea

andromedifolia whether the PVPP was hydrated in the grinding buffer or added —

directly to the leaf tissue during grinding.
This discussion stresses the importance of selecting optimal grinding buffer
components and procedures for the taxon under investigation. As noted above,

certain components of these buffers may inhibit band expression, whereas in other — |

—. the full complement of components may be required to obtain clear banding. — :
bt ave found that LAP activity is greater if sodium ascorbate, sodium tetraborate,
sodium metabisulfite are eliminated from the grinding buffer. On the other —

be very noticeably reduced when a buffer solution incorporating only PVP is

hand, activity for some enzymes, (e.g., SDH, G6PDH. MDH. CAT. 6PGD) may

D. £. SOLTIS ET AL: STARCH GEL ELECTROPHORESIS OF FERNS 15

utilized. A further complication in some taxa is that when a grinding buffer lacking
2-mercaptoethanol is employed, artifactual (“ghost”) bands proliferate for some
enzymes and can hinder accurate interpretation of the band patterns. Given these
examples, we strongly encourage comparative experimentation with grinding buffer
solutions when initiating electrophoretic studies.

SAMPLE PREPARATION AND ELECTROPHORESIS

Leaf samples to be analyzed electrophoretically were taken from mature sporo-
phytes. The methods discussed herein, however, are applicable to gametophytic as
well as sporophytic tissue. In our original electrophoretic analyses of ferns, leaf
samples were prepared according to the method of Soltis et al. (1980) in which
small amounts of leaf tissue are ground under liquid nitrogen with a porcelain
mortar and pestle until a fine dry powder is obtained. The powder is quickly mixed
with grinding buffer to form a thick slurry. This slurry is absorbed directly into thick
paper wicks (made from Whatmann 3 MM chromatography paper or some other
suitable wick paper).

It should be emphasized that the use of the grinding buffer-PVP solutions alone
(outlined above), without the use of liquid nitrogen in the preparation of samples,
provide clear enzyme banding in all of the fern taxa that we have examined so far.
Therefore, at present we utilize a simplified method of sample preparation in which
liquid nitrogen has been eliminated. In standardizing our grinding procedure by
consistently homogenizing 100 mg of leaf material in 0.5 ml of grinding buffer, we
obtained roughly equivalent staining intensity for all samples, which facilitates
comparing individuals and scoring gels. If small amounts of tissue are being ground,
it may be possible to substitute 43 mm7? plastic weigh boats for mortars and glass or
plexiglass rods for pestles. As discussed by Gastony and Gottlieb (1982), it is
possible to obtain enzyme banding from single gametophytes. Their technique
involves smearing individual gametophytes onto wicks pre-moistened with extraction
buffer-PVP solution. With all of the described grinding/extraction procedures, the
amount of grinding buffer and the type and size of wicks should be tailored to the
taxon and tissues under investigation. Once the plant material has been absorbed
into wicks, the wicks are inserted into a vertical slit in the starch gel and subjected
to horizontal electrophoresis at 4°C. Band definition may be improved by removing
wicks from the gel after the first 10-20 minutes of electrophoresis. It is important to
maintain electrical contact across the slit in the gel. We routinely place some sort of
spacer (e.g., strips of Whatmann paper, a plastic straw, a thin piece of plexiglass) at
the cathodal end of the gel to compress the gel slightly and to insure that the slit
does not open.

To permit comparison of banding patterns between gels, it is important to
maintain a constant starch concentration throughout a study. We have experimented
with a variety of starch concentrations throughout the practical range of 11.5 to 15%
and have found concentrations of 12 to 13.2% most suitable, although in some
instances alternative concentrations may improve band resolution. Connaught, Fish-
er, and Sigma starch have been used and all provide clear enzyme banding, provided
appropriate gel and electrode buffer systems and staining schedules are utilized.

16 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

In order to facilitate determining the location of the anodal front, it is useful
to insert a wick bearing the marker dye Bromphenol Blue (Sulfone Form, Sigma
BO126; 0.04% w/v in 95% ethanol). Depending on the gel and electrode buffer
system employed, as well as the taxon under investigation, the enzyme bands will
migrate varying distances behind the Bromphenol Blue marker. Optimal running
times for each enzyme system will therefore have to be determined empirically.

GEL AND ELECTRODE BUFFER RECIPES

We have found that standard gel and electrode buffer systems, such as those of
Shaw and Prasad (1970), and Brewer (1970), often yield unsatisfactory results with
fern tissue. We have determined gel and electrode buffer systems that provide clear
banding for 22 enzymes in ferns. Recipes for the gel and electrode buffers we most
commonly employ are provided in Table /.

Several of the gel and electrode buffer systems that we employ are modifications
of those of Shaw and Prasad (1970). It should be noted, therefore, that errors are
present in several of the recipes they provided. For example, in Shaw and Prasad
buffer system I (see page 299, Table | of their report), utilization of 16.35 g of Tris
yields an electrode buffer with a molarity of 0.135 (see present report, Table 1,
electrode buffers 2 and 3), rather than 0.155 as reported by Shaw and Prasad.
Similarly, use of 27.0 g of Tris in the preparation of the electrode buffer of Shaw
and Prasad system XII yields a molarity of not 0.233 but 0.223 (see present report,
Table 1, electrode buffers 4 and 5). Furthermore, use of the electrode buffer recipe

of Shaw and Prasad system XII yields a buffer of approximately pH 5.5, which

differs significantly from the estimate of pH 7.0 listed by Shaw and Prasad. Use of
the amounts of Tris and citric acid given for system 4 in Table / of the present report
will also yield an electrode buffer of pH 5.5; sodium hydroxide is then added to
adjust the buffer to pH 7.5.

Of the 11 gel and electrode buffer systems for which recipes are provided in Table
/, some clearly are preferable to others for certain of the 22 enzymes for which we
have obtained sharp banding. For example, best enzyme banding for MDH is
obtained when system 1, 4, 5, or 9 is used. Which system works best is dependent,
in part, on the taxon under investigation. We have found. for example, that the best
systems for Athyrium and Cystopteris differ somewhat from those that supply clear

enzyme banding for Bommeria. The systems provided here, however, should provide —
an excellent framework for conducting electrophoretic analyses of most ferns. The —
gel and electrode buffer systems that we have found to yield the clearest banding for

each of the 22 enzymes with which we have experimented are listed in Table 2.
STANDARD STAINING SCHEDULES

During the course of our electrophoretic investigations of ferns, we found that —
many published staining schedules, such as those of Shaw and Prasad (1970), —
yielded unsatisfactory results. Enzyme banding often can be improved dramatically
when these standard recipes are modified. One very important modification that we
employ involves increasing the pH of the buffer in staining schedules for ALD,
GDH, G6PDH, HK, IDH, MDH, ME, PGI, PGM, and 6PGD. Tris-HCI staining

buffers of pH 7.0 or 7.1 typically are used in staining for these enzymes (Shaw &

D. E. SOLTIS ET AL.: STARCH GEL ELECTROPHORESIS OF FERNS 17

Prasad, 1970). We have found, however, that in ferns the pH optima for these
enzymes are in the 8.0-8.5 range, and we therefore use a Tris—HC] staining buffer
of pH 8.0-8.5 for them. For some enzymes, this simple modification improves
Staining dramatically. For example, standard MDH staining recipes use a buffer of
pH 7.0. Application of such a protocol to ferns, however, often results in very little
or inconsistent staining. Well stained bands are obtained consistently for MDH when
a Tris-HCl staining buffer of pH 8.0-8.5 is used (provided an appropriate gel and
electrode buffer system is employed; see Table 2).

It also should be noted that we have consistently been unable to obtain observable
activity with fern leaf tissue for some enzymes, despite considerable experimenta-
tion. These include Alcohol dehydrogenase, Alkaline phosphatase, a-Glycerophos-
phate dehydrogenase and Lactate dehydrogenase. There are several enzymes for
which we have obtained observable activity with ferns, but have not yet obtained
clear banding, such as Diaphorase and Peptidase, and these are therefore not
included in this report. In addition, there are a number of enzyme staining schedules
that we have not yet attempted. We hope that the staining schedules provided below
will stimulate additional experimentation in this regard.

The staining schedules that we utilize for the 22 enzymes for which we obtain
clear banding are provided below in alphabetical order. The final volume for all stain
recipes is 100 ml. Depending on gel size and staining container capacity, it may be
possible to assay two gel slices simultaneously with each of the following 22
solutions. Alternatively, when assaying single gel slices, the recipes may be halved
to 50 ml final volume. Readers are encouraged to consult Enzyme Nomenclature
/978 (International Union of Biochemistry, Nomenclature Committee, 1979 [here-
after referred to as I.U.B., 1979]) for details of enzyme specificity and Gottlieb
(1981b) for a recent review of the electrophoretic technique and its application to
plant populations. It should be noted that enzymes such as peroxidases, esterases,
and acid phosphatases operate on a large number of substrates in vitro: therefore,
standard staining protocols for these enzymes result in the staining of various
numbers of isozymes whose homologies are not apparent. Although the utility of
these isozymes in between-species comparisons is therefore limited, their banding
patterns can be useful in assessing variability within populations.

Several different organisms serve as the source for commercial preparations of
Glucose-6-phosphate dehydrogenase. This enzyme typically is specific for NADP,
but some forms are capable of utilizing either NADP or NAD. It should be
emphasized that NADP is much more expensive than NAD, hence selecting a
Glucose-6-phosphate dehydrogenase type that can use NAD will save considerable
money. Therefore, in those recipes that call for added Glucose-6-phosphate dehydro-
genase, the notation NAD(P)* reflects this option.

Acid phosphatase (EC 3.1.3.2)

0.05 M sodium acetate buffer, pH 5.0 100 ml
1.0 M MgCl, 0.5 mi
@-naphthyl acid phosphate, sodium salt 100 mg
Fast garnet GBC salt aan

We have obtained results with buffer molarities ranging from 0.05 M to 0.2 M
and within a pH range of 5.0 to 6.0. Stain at room temperature; a modification of

18 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Scandalios (1969). Gottlieb (1981b) noted that plants may have a dozen or more
acid phosphatases. We consistently have observed one very intense zone of acid
phosphatase activity, with several fainter bands occasionally evident.

Aconitase (EC 4.2.1.3.; = Aconitate hydratase in I.U.B., 1979)

1.0 M Tris-HCI buffer, pH 8.5 10 ml
H,0 90 ml
cis-aconitic acid 70 mg
Isocitrate (= Isocitric) dehydrogenase 7 units
1.0 M MgCl, 1 ml
NADP 10 mg
MTT 5 mg

PMS
Stain in the dark at 30°C; a modification of Shaw and Prasad (1970).

Aldolase (EC 4.1.2.13; = Fructose-biphosphate aldolase in I.U.B., 1979)
1.0 M Tris-HCI buffer, pH 8.5 10m

HO 90 ml
Fructose-1 ,6-diphosphate,

tetra(cyclohexylammonium) salt 500 mg

trisodium salt 200 mg
1.0 M arsenic acid, sodium salt 1 ml
Glyceraldehyde-3-phosphate dehydrogenase 200 units
NAD
MTT 20 mg
PMS

Stain at room temperature; a modification of Shaw and Prasad (1970).

Aspartate aminotransferase (EC 2.6.1.1.; = Glutamate oxaloacetate trans-
aminase
1.0 M Tris-HCI buffer, pH 8.0 10 ml
HO 90 ml
L-aspartic acid 100 mg
a-ketoglutaric acid 100 mg
Adjust pH to 8.0 with 1.0 M NaOH as necessary, then add:
Pyridoxal-5’-phosphate 5 mg
Fast blue BB salt 100 mg

Stain in the dark at room temperature; a modification of Gottlieb (1973a) and
Selander et al. (1971, appendix). Our results show that this enzyme is very sensitive
to the pH of the staining buffer; very little staining is observed at pH 7.0 or at pH
9.0. The number of loci coding for proteins that are capable of this aminotransferase
reaction is known to vary in plants. Although we typically encounter a single locus,
Gottlieb (1981b) has reported the likelihood of specific subcellular localizations for
the several dimeric isozymes.

Catalase (EC 1.11.1.6)
3%

D. E. SOLIS ET AL.: STARCH GEL ELECTROPHORESIS OF FERNS 19

Incubate in this solution at room temperature for 1-2(30) minutes: pour off the
solution, rinse several times with distilled water, then add:

0.09 M KI 50 ml

H,O ml

Catalase activity will appear as white bands on a dark blue background: a
modification of Shaw and Prasad (1970). For some of the buffer systems in Table /,
it may be necessary to add several drops (approximately 2 ml) of glacial acetic acid
in order to induce staining. An alternative method is to substitute 50 ml 0.05 M
sodium acetate buffer (pH 5.0) for the 50 ml H;0. The gel may turn completely blue
in a very short time. Be prepared either to photograph the gel or score it while
staining. One locus has been observed in ferns. The protein is reported to be
tetrameric (Scandalios, 1969).

Esterase (Colorimetric; EC 3.1.1.-)
a-naphthy! acetate

40 mg
8-naphthy! acetate 40 mg
dissolved in acetone 2 mi
1.0M phosphate buffer, pH 6.0 10 ml
H,0 90 ml
Fast blue RR salt 100 mg
Stain at room temperature; a modification of Gottlieb (1974).
Esterase (Fluorescent; EC 3.1.1.-)
4-methylumbelliferyl acetate 42 mg
dissolved in acetone 25m
1.0 M sodium acetate buffer, pH 5.0 18 ml
H,0

Stain in the dark at room temperature and observe under long-wave ultraviolet
light; Staining schedule of Mitton et al. (1979). Observe immediately after staining
(bands fade quickly). Caution should be exercised in scoring gels because flavonoid

bands” may also be visualized under UV light. To determine whether flavonoid

“bands” are present, the gel slice should be observed under UV light prior to
Staining,

Fructose-1,6-diphosphatase (EC 3.1.3.11; =Fructose-biphosphatase in LUB.,
1979)

1.0 M Tris-HCI buffer, pH 8.0 Aer
H,0 90 ml
Fructose- | ,6-diphosphate,

tetra(cyclohexylammonium) salt 250 mg

trisodium salt in ae
10M MgCl, ss
Grosphoglucoisomerase paced

UCose-6-phosphate dehydrogenase seat
NAD(P)* “i i ie
MTT 5 mg
PMs ime

Stain in the dark at 30° C; a modification of Shaw and Prasad (1970).

20 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Glucose-6-phosphate eee serene (EC 1.1.1.49)

1.0 M Tris-HCI! buffer, pH 8.0 10 ml
H,0 90 ml
Glucose-6-phosphate, disodium salt 100 mg
NADP 20 mg
MTT (or NBT) 10 mg
PMS mg

Stain in the dark at 37° C; a modification of Shaw and Prasad (1970).
Glutamate pe lscceroe! (EC 1.4.1.2)

1.0 M Tris-HC1 buffer, pH 8 10 ml
HO 70 ml
1.0 M L-glutamic acid, pH 8.0 (use free acid

or monosodium salt and add NaOH to pH 8.0) 20 ml
NAD 20 mg
MTT (or NBT) 10 mg
PMS

2 mg
Stain in the dark at room temperature; a modification of Gottlieb (1973b) and of
Shaw and Prasad (1970).

Glyceraldehyde-3-phosphate ee (EC 1.2.1.12;: = Glyceraldehyde-
phosphate dehydrogenase in I.U. B., 1979)

1.0 M Tris-HCI buffer, pH 8.0 10 ml
H,0 90 ml
Fructose-1,6-diphosphate, trisodium salt 100 mg
Aldolase 10 units
Incubate above mixture approx. 30 min. at 30-37° C, then add:
1.0 M arsenic acid, sodium salt 1 ml
NAD 10 mg
MTT 5 mg
PMS | mg

Stain in the dark at 30° C; a modification of Shaw and Prasad (1970).
Hexokinase (EC 2.7.1 1)

1.0 M Tris-HCI buffer, pH 8.5 10 ml
HO 90 ml
Glucose 90 mg
1.0 M MgCl, 5 ml
EDTA, tetrasodium salt, dihydrate 40 mg
NAD(P)* 10 mg
MTT (or NBT) 15 mg
Pee Phosphate dehydrogenase 40 units
ATP sa

Stain in the dark at room temperature; a modification of Shaw and Prasad (1970).

Isocitrate dehydrogenase [NADP*
ne M Tris-HC] buffer, pH 8 i ie a <
30

10 ml

ee |

Isocitric acid, trisodium salt a wd

10M MgCl, pa
NADP

D. E. SOLTIS ET AL.: STARCH GEL ELECTROPHORESIS OF FERNS 21

MTT (or NBT) 15 mg
PMS 2 mg
Depending on the taxon being analyzed, results have been obtained by using
buffers ranging from pH 7.2 to pH 8.5. Stain in the dark at room temperature: a
modification of Shaw and Prasad (1970).

Leucine aminopeptidase (EC 3.4.11.-)

L-leucine-B-naphthylamide (free base or

acid salt) 20 mg
dissolved in dimethyl formamide 5 ml
1.0 M phosphate buffer, pH 6.0 10 ml
H,0 90 ml

Black K salt or fast black K salt 50 mg

Stain in the dark at room temperature; a modification of Gottlieb (1973c). This
enzyme has broad activity; it can cleave a number of N-terminal amino acids.
Although commonly referred to as Leucine aminopeptidase (“LAP”), it is more
precisely referred to as Aminopeptidase. There are a number of aminopeptidases,
some of which are specific for certain N-terminal amino acids. Aminopeptidase
(cytosol; EC 3.4.11.1) is activated by heavy metals, whereas Aminopeptidase
(microsomal; EC 3.4.11.2) is not activated by heavy metals (I.U.B., 1979). It may
be necessary to modify the pH of the stain in order to increase staining intensity.
One isozyme is usually observed and a second is occasionally apparent; both are
monomeric.

Malate dehydrogenase (EC 1:4:1.37)
8.5

1.0 M Tris-HC1 buffer, pH 8.0 or 10 ml
2.0 M DL-malic acid, pH 8.0 (add NaOH to pH 8.0) 10 ml
HO 80 ml
NAD 10 mg
MTT (or NBT) 10 mg
PMS 2

Stain in the dark at room temperature; a modification of Shaw and Prasad (1970).
Some investigators add 38 mg EDTA (tetrasodium salt, dihydrate) to this recipe. We
use 2.0 M DL-malic acid rather than 1.0 M L-malic acid as most standard staining
schedules require because DL-malic acid is much less expensive than purified
L-malic acid, which is the actual substrate. There are at least four malate NAD*
dehydrogenases reported (I.U.B., 1979). The one most frequently reported in
routine electrophoresis is L-Malate: NAD+ oxidoreductase (EC 1.1.1.37), of which
at least two putative isozymes have been observed. The protein is dimeric; subcellu-
lar compartmentalization of the isozymes is discussed by Gottlieb (1981b).

Malic enzyme (EC 1.1.1.40; = Malate dehydrogenase [Oxaloacetate-decar-
boxylating, NADP*] in I.U.B., 1979)
8.5

1.0 M Tris-HCI buffer, pH 8.0 or 8. 10 mi

20 80 ml
2.0 M DL-malic acid, pH 8.0 (prepare as in Malate dehydrogenase) 10 ml
10M MgCl, 2 ml
NADP 20 mg

MTT (or NBT) 20 mg
PMS 2 mg

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Stain in the dark at room temperature; a modification of Richmond (1972 and
pers. comm.).
Peroxidase (EC 1.11.1.7)

3-amino-9-ethy! carbazole 65 mg
dissolved in dimethyl formamide 5 ml
0.05 M sodium acetate buffer, pH 5.0 95 ml
0.1 M CaCl, 2 ml
2 ml

Incubate the gel in a refrigerator until the bands appear (30-60 min); a modification
of Shaw and Prasad (1970) and of Gottlieb (1973b).
Phosphoglucoisomerase (EC 5.3.1.9; = Glucosephosphate isomerase in I.U.B.,

1.0 M Tris-HC1 buffer, pH 8.0 10 ml

> 90 ml
1.0 M MgCl, 1 ml
Fructose-6-phosphate, disodium salt 30 mg
Glucose-6-phosphate dehydrogenase 40 units
NAD(P)*
MTT (or NBT) 20 mg
PMS 2 mg

Stain in the dark at 37°C or at room temperature; a modification of Shaw and
Prasad (1970). This dimeric enzyme has numerous synonyms (I.U.B., 1979)
including: Phosphohexose isomerase, Phosphohexomutase, Oxoisomerase, Hexose
phosphate isomerase, Glucose-6-phosphate isomerase, Phosphosaccharomutase, and
Phosphohexoisomerase. Cytosolic and chloroplastic isozymes should be expected
(Gottlieb, 1981b, 1982).

Phosphoglucomutase (EC eit)

1.0 M Tris-HCI buffer, pH 8.0 or 8.5 10 ml
H,0 90 ml
1.0 M MgCl, 2 ml
Glucose-1-phosphate, disodium (Sigma G7000)

or dipotassium (Sigma G6875) salt 100 mg
1.7 x 104M «-D-glucose-1,6-diphosphate,

tetra(cyclohexylammonium) salt 5 mi
Glucose-6-phosphate dehydrogenase 40 units
NAD(P)* 10 mg
MTT (or NBT) 20 mg
PMS

; : 2 mg
A modification of Shaw and Prasad (1970). An equally suitable alternate recipe
which eliminates the need to purchase a-D-glucose- 1 ,6-diphosphate is:

- M Tris-HC1 buffer, pH 8.0 oe
; l
Glucose- |-phosphate, disodium salt
Sj
Se€-6-phosphate dehydrogenase peste
NAD(P)*
aD 10 mg
in 10 mg
2 mg

Stain in the dark at 37°C or at room temperature.

TG EL PADS SE

D. E. SOLTIS ET AL.: STARCH GEL ELECTROPHORESIS OF FERNS 23

6-Phosphogluconate dehydrogenase (EC 1.1.1.44)

1.0 M Tris-HC1 buffer, pH 8.0 10 ml

2 90 ml
6-phosphogluconic acid, barium salt 40 mg
1.0 M MgCl, 2 ml
NADP 10 mg
MTT 10 mg
PMS 2 mg

Stain in the dark at room temperature or at 37°C: a modification of Shaw and
Prasad (1970).

Shikimate dehydrogenase (EC 1.1.1.25)

1.0 M Tris—HC1 buffer, pH 8.5 10 ml
H,0 90 ml
Shikimic acid 100 mg
NADP 10 mg
MTT (or NBT) 20 mg
PMS 2 mg

Stain in the dark at room temperature or at 37°C.
Triosephosphate isomerase (EC 5.3.1.1)

1.0 M Tris-HCI buffer, pH 8.0 10 ml
H,0 90 ml
Dihydroxyacetone phosphate, lithium salt 10 mg
EDTA, tetrasodium salt, dihydrate 38 mg
NAD 30 mg
MTT 10 mg
PMS 2 mg
Arsenic acid, sodium salt 460 mg
Glyceraldehyde-3-phosphate dehydrogenase 300 units

Stain in the dark at 37°C; Gottlieb (pers. comm.). An alternative and much less

expensive method of staining for TPI follows (also see agarose recipe):

1.0 Tris-HCI buffer, pH 8.0 ——
H,0 90 ml
DL-a-glycerophosphate 200mg
Pyruvic acid, sodium salt 100 mg
a-Glycerophosphate dehydrogenase 100 units
Lactate dehydrogenase 100 ‘units

Incubate the above solution for 2 hours at 30-37°C. At the end of the incubation
period, inactivate the enzymes by adjusting the solution to pH 2.0 with 1.0 M HCI
(taking care not to go below pH 2.0) and then re-adjust to pH 8.0 with 1.0 M
NaOH. Add:

1.0 M arsenic acid, sodium salt | ml
Glyceraldehyde-3-phosphate dehydrogenase 100 units
NAD 20 m
MTT 5 mg
PMS

mg
Stain in the dark at 30°C. The bands will be dark blue on a light blue background.
Readers who follow this protocol must first show the lack of a-Glycerophosphate
dehydrogenase activity in their taxa. This can be done simply by staining a gel slice
with:

24 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

1.0 M Tris-HC1 buffer, pH 8.0 10 ml

90 ml
DL-a-glycerophosphate 200 mg
NAD 20 mg
MTT 5 mg
PMS | mg

AGAROSE STAINING SCHEDULES

Several of the reagents commonly required in enzyme electrophoresis, such as
NADP, are expensive. Costs may be reduced by using staining schedules employing
agarose (modified from Mitton et al., 1979 and Gaines, pers. comm.), which
require much smaller quantities of most ingredients than do standard staining
schedules but yield comparable results.

Agarose staining schedules are provided below for ALD, G6PDH, GDH, IDH,
MDH, PGI, PGM, 6-PGD, SkDH, and TPI. The following general procedure is
employed to stain one starch gel slice using the agarose technique: dissolve all
ingredients required for enzyme staining (see below) in 6 ml of Tris-HC1 buffer; in
a second flask mix 0.06 g agarose (Agarose type II, Sigma A6877) with 6 ml
distilled water (a 1% agarose solution) and bring this solution to a boil while stirring
constantly; combine the two solutions and very quickly apply to surface of one gel
slice (it may be necessary to increase ml amounts of buffer and 1% agarose
depending on the size of the gel slice); stain all slices in the dark at room
temperature.

Aldolase (EC 4.1.2.13)
PMS

1 mg
MTT 5 mg
NAD 10 mg
Fructose-1 ,6-diphosphate, trisodium salt
or tetra(cyclohexylammonium) salt 75 mg
Arsenic aci 25 mg
Glyceraldehyde-3-phosphate dehydrogenase 40 units
1 M Tris-HCI buffer, pH 8.5 6 ml
1% agarose 6 ml
Glucose-6-phosphate dehydrogenase (EC 1.1.1.49)
PMS I mg
MTT 4 mg
NADP 4 mg
EDTA, tetrasodium salt, dihydrate 6 mg
Glucose-6-phosphate, disodium salt 5 mg
0.1 M Tris-HCI buffer, pH 8.0 6 ml
1% agarose 6 ml
Glutamate dehydrogenase (EC 1.4.1.2)
PMS 1 mg
MTT 4 mg
NAD 3 mg
L-glutamic acid, monosodium salt 100 mg
0.1 M Tris-HCI buffer, pH 8.0 6 ml

1% agarose

D. E. SOLTIS ET AL.; STARCH GEL ELECTROPHORESIS OF FERNS 25

Isocitrate dehydrogenase [NADP* ] (EC 1.1.1.42)

I mg
MTT 4 mg
NADP 4 mg
2 40 mg
Isocitric acid, trisodium salt 10 mg
0.1 M Tris-HCl! buffer, pH 8.0
1% agarose 6 ml
Malate dehydrogenase (EC 1.1.1.37)
PMS 2 mg
MTT 7 mg
NAD 7 mg
L-malic acid 16 mg
0.25 M Tris-HC1 buffer, pH 8.6 6 ml
1% agarose 6 ml
Keep PMS and L-malic acid powders separate until adding the buffer.
Phosphoglucoisomerase (EC 5.3.1.9)
PMS | mg
MTT 4 mg
NAD(P)* | mg
0.1 M Tris-HCI buffer, pH 8.0 4 ml
10% MgCl, 1 ml
0.018 M fructose-6-phosphate, eine salt
Glucose- _ = toe dehydrogena 10 units
1% agaro 6 ml
Phosphoglucomutase (EC 2.7.5.1)
PMS | mg
MTT 5 mg
NAD(P)* 4 mg
0.1 M Tris-HCI buffer, pH 8.0 iy
0.05 M glucose-1-phosphate, sala salt (Sigma G7000) 2.5 ml
Glucose-6-phosphate dehydrogen 10 units
0.001 M fructose-1 ,6- diphaghae: "eounies salt 1 ml
10% MgCl, 1 ml
1% agarose 6 ml
6-Phosphogluconate dehydrogenase (EC 1.1.1.4.4)
PMS I mg
MTT 4 mg
NADP 4 mg
6-phosphogluconic acid, barium salt 10 mg
10% MgCl, 0.5 ml
0.1M Te HCl buffer, pH 8.5 5 ml
1% agar 6 ml

Dissolve the 6- -phosphogluconic acid barium salt in 1.0 ml of 0.1 M Tris—HC1
buffer (pH 8.5) approximately 15 minutes before mixing in other ingredients.

6 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Shikimate dehydrogenase (EC 1.1.1.25)
PMS

I mg
MTT 4 mg

4 mg
Shikimic acid 10 mg
0.1 M Tris-HC1 buffer, pH 8.0 6 ml
1% agarose 6 ml

Triosephosphate isomerase (EC 5.3.1.1)

PMS I mg
MTT 4 mg
NAD 10 mg
EDTA, tetrasodium salt, dihydrate 10 mg
Arsenic acid 150 mg
Dihydroxyacetone phosphate 1.5 mg
Glyceraldehyde-3-phosphate dehydrogenase 75 units
0.1 M Tris-HC1 buffer, pH 8.0 6 ml
1% agarose 6 ml

GEL FIXATION AND DOCUMENTATION

AAT and PER gel slices are fixed in 50% glycerol; all other gel slices are fixed in
50% ethanol.

In order to keep a permanent record of results, we routinely photograph gel slices
using Kodak Technical Pan Film 2415 following the exposure index and developing
procedure recommended for high contrast.

ACKNOWLEDGEMENTS

D.E.S. and C.H.H. thank Jeff Atwood, Harriet Blanton, Leslie Gottlieb, James
Hamrick, Terry Lastovicka, and Andrew Torres for laboratory assistance and helpful
comments and Charles Werth for providing details of his sample preparation

University of Kansas, and to D.C.D. from the Department of Biology of Indiana
University, NSF Doctoral Dissertation Improvement Grant DEB-8117239, and the
Society of the Sigma Xi are gratefully acknowledged.

LITERATURE CITED
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aa G. J. 1970. An Introduction to Isozyme Techniques. Academic Press, New York.
AN, R. H., E. J. KLEKOWSKI, Jr. and R. K. SELANDER. 1979. Homoeologous
Cone eee and recombination in the fern Pteridium aquilinum. Science 204:1207—1209.
mIVER, G. 1976. Chemotaxonomy and phytochemical ecology of bracken. J. Linn. Soc.,

Bot. 73:3
GASTONY, G. J. and L. D. GOTTLIEB. 1982. Evidence for genetic heterozygosity in a homosporous
fern. Amer. J. Bot. 69:634-637.
» and C. H. HAUFLER. 1976. Chromosomes and apomixis in the fern genus Bommeria
(Gymnogrammaceae). Biotropica 8:1—1]

D. E. SOLTIS ET AL.:; STARCH GEL ELECTROPHORESIS OF FERNS 27

GOTTLIEB, L. D. 1973a. Genetic control of Glutamate oxaloacetate transaminase isozymes in the
diploid plant Stephanomeria exigua and its allotetraploid derivative. Biochem. Genet. 9:97—107.
73b. Genetic differentiation, sympatric speciation, and the origin of a diploid species of
Fass Amer. J. Bot. 60:545-553.
. 1973c. Enzyme differentiation and phylogeny in Clarkia franciscana, C. rubicunda and C.
amoena. Evolution 27:205—214.
. 1974. Genetic confirmation of the origin of Clarkia lingulata. Evolution 28:244-250.
. 1981a. Gene numbers in species of Astereae that have different chromosome numbers. Proc.
Natl. Acad. Sci. USA 78:3726—3729.
. 1981b. Electrophoretic evidence and plant populations. Progress in Si beaticphae sh 7:1-45.
——. 1982. Conservation and duplication of isozymes in plants. Science 216 380.
INTERNATIONAL UNION OF BIOCHEMISTRY, NOMENCLATURE he: 1979. Enzyme
Nomenclature 1978. Academic Press, New York.
KELLEY, W. A. and R. P. ADAMS. 1977a. Preparation of extracts from Juniper leaves for
electrophoresis. Anbeeaser reel 16:513-516
and R. P. AD 77b. Seasonal variation of isozymes in Juniperus scopulorum:
systematic signa ied J. Bot. 64:1092—10
MITTON, J. B., Y. B. LINHART, J. L. HAMRICK, and J. + BECKMAN. 1977. Observations on the
genetic structure kee mating system of ponderosa pine in the Colorado front range. Theor
Appl. Genet. 51:5-13.
INHART, K. B. STURGEON, and J. L. HAMRICK. 1979. Allozyme polymorphism
detected 1 in mature needle tissue of ponderosa pine. J. Hered. 70:86-89.
RICHMOND, R. C. 1972. Enzyme variability in the Drosophila willistoni group. I]. Amounts of
variability in the superspecies, D. paulistorum. Genetics 70:87-112.
SCANDALIOS, J. G. 1969. Genetic control of multiple molecular forms of enzymes in plants: a
review. eae Genet. 3:37-79.
SELANDER, R. K., M. H. SMITH, S. Y. YANG. W. E. JOHNSON, and J. B. GENTRY. 1971. IV.
Biochemical polymorphism and systematics in the genus Peromyscus. I. Variation in the
old-field mouse (Peromyscus polionotus). Studies in as VI. Univ. Texas Publ.

7103:49-90.
SHAW, “ R. and R. PRASAD. 1970. Starch gel electrophoresis of enzymes—a compilation of recipes.

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SOLTIS, 3 a ap HAUFLER, and G. J. GASTONY. 1980. gi ie enzyme variation in the fern

genus Bonnet an analysis of nie Syst. Bot. 5:30-
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esata i tissues using ae Isozyme Bull. 15:139.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 1 (1983)

SHORTER NOTES

A NEW COMBINATION IN ASPLENIUM.—A new fern was recently added
to the flora of Alabama with the naming of the backcross hybrid x Asplenosorus
boydstonae Walter (Amer. Fern J. 72:62. 1982.). The type material was collected in
Hale County, Alabama, in a locality famous for its fertile alloploid population of
Scott’s Spleenwort, which arose through hybridization of the Walking Fern and the
Ebony Spleenwort. The new hybrid has Scott’s Spleenwort and the Ebony Spleen-
wort as its parents. If the view is held that the Walking Fern belongs in the genus
Asplenium rather than Camptosorus, the “hybrid genus” X Asplenosorus is taxonom-
ically superfluous. The new hybrid then needs to be placed in Asplenium and the
epithet corrected in conformity with Art. 73.10 of the “International Code of
Botanical Nomenclature”:

Asplenium x boydstoniae (Walter) Short, comb. nov.

* Asplenosorus boydstonae K. §. Walter, Amer. Fern J. 72:62. 1982. TYPE: Havana Glen, Hale Co.,
Alabama, Wagner & Walter 70011 (MICH not seen).

John W. Short, 217 Cook St., Auburn, AL 36830.

ADIANTUM FURCATUM AND MICROSTAPHYLA F URCATA.—The is-
land of St. Helena, outside the Gulf of Guinea off the west coast of Africa, is not
only famous because it was the residence in exile of Napoleon Bonaparte. It has also
produced a fern, Microstaphyla bifurcata, which stands out of proportion when its
diminutive size (ca. 10-20 cm) is compared with the long list of names it has been
called. Since the establishment of the genus Microstaphyla in 1851, it has been
placed in no fewer than nine genera! The last revisionary treatment of Microstaphyla
was by Maxon (J. Washington Acad. Sci. 13:28-31. 1923), who included among
the synonyms the name Adiantum furcatum L. f. In the “Index Fil icum,” Christensen
(1905) accepted that epithet as Elaphoglossum furcatum (L. f.) Christ. That
basionym also accounts for the citation Microstaphyla furcata (L. f.) Presl, which
has been used by some authors, including Maxon, who have overlooked Presl’s
explicit citation of Osmunda bifurcata Jacq. as the basionym of his combination.

In any event, accepting A. furcatum as a synonym for the sole species of
Microstaphyla is an error which originated when the younger Linnaeus included a

Sously interpreted by the younger Linnaeus to be the same as his A. furcatum (Sp.
nmeaenals 1881), a plant he reported as “in Cap. Bonae Spei et insula
so : hunberg. Thouin.” Although Thunberg collected in the Cape and Thouin
in Reunion, neither of them set foot on St. Helena, where Microstaphyla is
Pe. Furthermore, A. furcatum is not an Adiantum at all, but is Asplenium
ee (Lam.) C. Chr. (syn. A. rutaefolium (Berg.) Kunze). For this reason,
. innaean epithet should be deleted as a synonym of M. bifurcata (Jacq.) Presl,

correct name.—Luis Diego Gémez P., Museo Nacional de Costa Rica, Apartado

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 1 (1983) 29

NOTEWORTHY PTERIDOPHYTE RECORDS FOR NEBRASKA.— Field
studies in the Niobrara River Valley of north-central Nebraska during the summer of
1982 yielded several significant finds. Field work was carried out in connection with
a floristic survey of the 54,000-acre Niobrara Valley Preserve, owned by The Nature
Conservancy, and in areas peripheral to the Preserve.

Botrychium matricariifolium A. Br. was collected for the first time in Nebraska.
In the Great Plains it is otherwise known only from Custer County, South Dakota
(Brooks, R. E. Amer. Fern J. 70:91. 1980) and from Lac Qui Parle County,
Minnesota (Atlas of the Flora of the Great Plains, lowa State Univ. Press. 1977).
Three plants were observed in a juniper-oak woodland on the south side of the
Niobrara River about 15 miles NNE of Johnstown, Brown County. The plants were
growing in a sandy, partially shaded soil along the floodplain of the river close to an
access road to a center-pivot irrigation pump along the river. Much of the area had
been cleared for the road and for an irrigation pipe up a steep, north-facing slope.
No additional plants were found during subsequent searches. The plants were
photographed and, after the fronds senesced, collected (Freeman 15/7, KANU).

Ophioglossum vulgatum var. pseudopodum (Blake) Farw. was first collected in
Nebraska in 1913 by J. M. Bates in the vicinity of Kennedy, Cherry County.
Clausen (Mem. Torrey Bot. Club 19:126. 1938) cited the specimen without
infraspecific reference; Brooks (Amer. Fern J. 70:92. 1980) referred it to var.
pseudopodum. A thriving population was found on the south side of the Niobrara
River ca. 55 yds east of the Berry Bridge, 5 miles west and 3 miles south of Sparks,
Cherry County. The plants covered an area of about 7 x 2 yds at the west end of a
beaver pond. They were growing in wet, loamy soil at the edge of a paper birch
forest along with Cicuta bulbifera, Onoclea sensibilis, and Thelypteris palustris.
The area showed obvious signs of human manipulation, as a dike situated between
the river and the west end of the pond was fitted with a pipe to supplement water in
the pond during dry periods, possibly as part of a fish hatchery built many years
ago. Photographs were taken and voucher specimens (Freeman & Churchill 1672)
were distributed to various herbaria, including KANU, KSC, NEB, and NY.

Dryopteris carthusiana (Villars) H. P. Fuchs was collected around the turn of the
century in Brown, Cass, Cherry, Lancaster, and Thomas counties in Nebraska
(A. J. Petrik-Ott, Beih. Nova Hedw. 61:141. 1979). Field studies conducted by a
number of researchers in 1980 resulted in the discovery of one population along
Little Cedar Creek Canyon in Cherry County (Sutherland, Boner, Lamp & Joern
5053, NEB; Sutherland & Harrison 5314, NEB). During the course of our survey,
we found four populations, all in rich paper birch forests along spring-fed canyons
on the south side of the river. Two populations were in Cherry County. One was
photographed and collected along Little Cedar Creek Canyon, 5.2 miles south of
Sparks (Freeman & Churchill 1200, KSC, NEB). A second was just east of Smith
Falls, 3 miles east and 4 miles south of Sparks (Churchill & Brogie 12424, NEB).
The latter was the largest of the populations. Two populations were found in Brown
County. One was along Dutch Creek Canyon, 16 miles north of Ainsworth
(Churchill 12300, NEB). The second was in Kantak Coulee, 3 miles west of
Meadville (Churchill 12408, NEB).

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

We wish to thank Dr. W. H. Wagner, Jr. and Ralph Brooks for verifying the
specimen of Botrychium and Dr. T. M. Barkley and Ralph Brooks for reviewing the
manuscript.—Craig C. Freeman, Division of Biology, Kansas State University,
Manhattan, KS 66506 and Steven P. Churchill, Division of Biological Sciences,
University of Kansas, Lawrence, KS 66045.

SPREAD OF MARSILEA QUADRIFOLIA IN MCDONOUGH COUNTY,
ILLINOIS.—Thirty-five years ago Myers found Marsilea quadrifolia L. in Spring
Lake, McDonough County, Illinois. It is of interest to record how much this alien,
potentially weedy plant has spread in that time. Myers (Amer. Fern J. 40:256. 1950)
stated that during the years following 1947 Marsilea did not become abundant, but
he did recognize that it “has persisted following partial draining of the lake,
dredging, and a considerable increase in the water level when a higher dam was
built” (Annotated Catalogue and Index for the Illinois Flora. Western Illinois Univ.
Ser. Biol. Sci. 10:58. 1972). At present, the plant has not spread much in the lake
and is still not abundant. Water from the lake flows downstream via Spring Creek for
about 4.3 miles where it enters the LaMoine River, which is a tributary of the
Illinois River. In 35 years, M. quadrifolia has migrated down Spring Creek only one
mile. The quantity and distribution of the plants vary, progressing from many
plants in the upper part of the creek below the lake and dam to a very few, widely
scattered plants beyond one-half mile to finally one plant one mile downstream. The
plants are usually at the edge of the creek, particularly near the lake, but further
downstream are found characteristically in small “cut-offs” at the edge of the creek.
Downstream migration of Marsilea has not been rapid, which seems surprising
considering the numerous times it could have been dispersed by propagules (includ-
ing rafts of vegetation), especially during the rush of high waters in spring and fall.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 1 (1983) 31

WOOD FERNS NEW TO MARYLAND AND DELAWARE.— While collect-
ing specimens for Towson State University, I discovered four plants of the Crested
Log Fern Hybrid, Drvopteris celsa x cristata, growing as a fairy ring near
Creswell, Harford County, Maryland (Redman 3547, BALT. MICH). This discovery
represents the fifth state known for this rare hybrid. the closest known locality being
about 150 miles northeast in northeastern New Jersey. The plants grew on alluvial
soils derived from the James Run gneiss formation in a Beech-Oak woods. The
parents were not growing nearby: D. celsa is known trom 5 miles away and D.
cristata from 7 miles away. Unfortunately, the hybrids’ locality is being destroyed for
development, but the plants have been moved into cultivation. | am indebted to Drs.
Warren and Florence Wagner for verifying the identity of the specimens.

I also discovered three plants of Miss Slosson’s Hybrid Woodfern, D. x slossonae
Wherry (D cristata X marginalis), near Kenton, Kent County, Delaware (Redman
2814, BALT). The habitat is alluvial woods under a canopy of oak trees. Plants of
both parents grow nearby. Apparently this is the first record for this hybrid in
Delaware.—Donn E. Redman, 2615 Harwood Road, Baltimore, MD 21234.

A NEW COMBINATION FOR AN ASPLENOSORUS HYBRID.—Tradi-
tionally, the Walking Fern has been segregated from the spleenworts (Asplenium) as
Camptosorus rhizophyllus (L.) Link. A number of pteridologists, however, question
the distinction, pointing out the morphological similarity of Walking Fern to
Asplenium, its comparable diploid chromosome number (n= 41), its preference for
rock substrates, and its northeastern distribution. A close alliance with the spleen-
worts is further demonstrated by its tendency to cross with representatives of the
genus Asplenium to produce sterile hybrids, some of which have become fertile
through chromosome doubling and now behave as distinct species. In the Appala-
chian Mountains such hybridization and ploidy level changes have led to an
extensive allopolyploid complex which has undergone considerable study. Those
Who maintain the Walking Fern in Camptosorus place hybrids with Asplenium and
their derivatives in the hybrid genus Asplenosorus. Most of the hybrids were
Originally described as spleenworts, and therefore have combinations available under
Asplenium. One new Walking Fern—Spleenwort hybrid, however, was recently de-
scribed as an Asplenosorus. Its incorporation into the genus Asplenium may be
accommodated by the following combination:

Asplenium x shawneense (R. C. Moran) H. E. Ballard, comb. nov.

Asplenosorus x shawneensis R. C. Moran, Amer. Fern J. 71:85, 1981.

This is the hybrid of Asplenium rhizophyllum and A. trichomanes, and is known
So far only from southern Illinois.—Harvey E. Ballard, Jr., Department of Biology,
Western Michigan University, Kalamazoo, MI 49008.

32 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

AMERICAN FERN JOURNAL

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AMERICAN 33
FERN —
JOURNAL |

QUARTERLY JOURNAL OF THE AMERICAN FERN SOCIETY

Vittaria Gametophytes Discovered
in a New Physiographic Province ALLISON W. CUSICK
Chapman’s Quillwort Reconsidered BRIAN M. BOOM

C-Glycosylxanthones in Diploid and Tissue
Culture-induced Autotetraploid Davallia fejeensis
P. MICK RICHARDSON and HARINDER K. PALTA

The Distribution of Woodwardia areolata R. CRANFILL

Two Moonworts of the Rocky Mountains:
Botrychium hesperium and a New Species
Formerly Confused with It W. H. WAGNER, JR. and FLORENCE S. WAGNER

Shorter Notes: Lycopodium complanatum and
- annotinum Found in the Black Hills;
Microfibrils in the Xylem of Blechnum
viviparum; Bisporangiate Anomalous
Sporophylls in Isoétes from Rajasthan

Reviews 42, 45,

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MISSOURI BOTANICAL

The American Fern Society

Council for 1983
DEAN P. WHITTIER, Dept. of Biology, Vanderbilt University, Nashville, TN 37235. President
TERRY R. WEBSTER, Biological Sciences Group, University of Connecticut, Storrs, CT 06268.
Vice-President
MICHAEL I. COUSENS, Faculty of Biology, University of West Florida, Pensacola, FL 32504.
Secretary
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Treasurer
JUDITH E. SKOG, Dept. of Biology, George Mason University, Fairfax, VA 22030.
Records Treasurer
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ALAN R. SMITH, Dept. of Botany, University of California, Berkeley, CA 94720. Memoir Editor
JOHN T. MICKEL, New York Botanical Garden, Bronx, NY 10458. Newsletter Editor
American Fern Journal
EDITO

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TERRY R. WEBSTER ....... Biological Sciences Group, University of Connecticut, Storrs, CT 06268.

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AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 2 (1983) 33

Vittaria Gametophytes Discovered
in a New Physiographic Province
ALLISON W. CUSICK*

Since their recognition over 20 years ago, knowledge of the distribution and
taxonomic affinities of the perennial Vittaria gametophytes found in the eastern
United States has expanded greatly. Once thought to be extremely rare and local,
they now are known to be common in selected habitats over a broad region of the
Appalachians. Farrar (1978) summarized the history of this taxon and presented a
map of its distribution as then understood. Vittaria gametophytes have been discov-
ered in the unglaciated Appalachian Plateau, Blue Ridge, Ridge and Valley, and
Upland Piedmont provinces, all uplifted bedrock areas which presumably have been
in continuous existence since Tertiary times. Farrar et al. (1983) discussed these
recent finds and reported the gametophytes from sites in Ohio and Pennsylvania near
the limits of Pleistocene glaciation. This article describes the presence of Vittaria
gametophytes beyond these limits. This unusual fern now has been discovered on the
glaciated Appalachian Plateau nearly 90 km north of the southern limit of
Wisconsinan glaciation. This also is the first report of Vittaria gametophytes from
the watershed of the Great Lakes.

The Vittaria gametophytes were found 10 June 1982 on Little Mountain, Geauga
and Lake counties, Ohio, on property of the Holden Arboretum, Mentor, Ohio (Fig.
1). Accompanying the author were Brian Parsons and Thomas Yates, field natural-
ists of the Arboretum, and Robert McCance of the Division of Natural Areas and
Preserves, Ohio Department of Natural Resources. To say that we were surprised by
this discovery would be an understatement. Only a small collection was made at this
time (Cusick 21673, ISC). The author revisited the site on 24 August, searching
more thoroughly for gametophytes, and again preparing a voucher (C usick 22000,
OS). Colonies of Vittaria were found to occur discontinuously in a linear band over
approximately 0.5 km of the slope of Little Mountain. Identity of the gametophytes
was verified by Dr. Donald R. Farrar of Iowa State University.

The site of this discovery is an extraordinary area not at all typical of glaciated
Ohio and a habitat highly suitable for the growth of Vittaria. Little Mountain is at
the extreme edge of the glaciated Appalachian Plateau, being one of the localized
bedrock exposures that mark the escarpment between the elevated Plateau and the
Lake Plains of the Central Interior Lowlands province. The mountain is shaped
roughly like a dumbbell, oriented north to south, with knobs at either end. The
northern knob rises to 380 m, the southern knob to 386 m. Between these summits
is a flattened saddle only slightly lower in elevation. The northern knob lies partly in
Concord Township, Lake County, while the bulk of the mountain is in Chardon
Township, Geauga County. The shoreline of Lake Erie is only 6.5 km north of the
mountain. The approximate mean elevation of the lake at that point is 174 m. Thus,

*Division of Natural Areas and Preserves, Ohio Department of Natural Resources, Fountain Square,
Columbus, OH 43224.
Volume 73, number 1, of the JOURNAL, was issued March 31, 1983.

34 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

TauMe@uLL

PORTAGE

ft
MAHOMING

STARK COLUMBAN A

a
ae 2h YH

% +
TUSCARAWAS
A HOPPE RSON
7
-

FIG. 1. The northeastern quarter of Ohio. Solid line indicates the northern limit of the glaciated
Appalachian Plateau; dashed line, the southern limit of Wisconsinan glaciation. Solid dot is location of
Little Mountain.

there is a difference in elevation of 206 m between the mountain and Lake Erie
(U.S. Geological Survey, 1970

This dramatic change in elevation between lake and summit has a significant
effect on local weather patterns. Winds moving south across Lake Erie pick up
moisture which is quickly lost once this escarpment is reached. In the lee of these
summits, then, is the locally-famous “snow belt” of northeastern Ohio. This local
phenomenon is clearly shown on the snowfall map in Gordon (1969) on which the
site of Little Mountain may be seen in the pattern of isopleths. Snowfall in the Little
Mountain region averages nearly 3 m yearly, far in excess of Ohio’s annual average
snowfall of about 1 m.

The summits of Little Mountain and the other elevations at the edge of the
glaciated Appalachian Plateau in Ohio are capped by the Sharon Conglomerate
formation of the Pottsville group of the Pennsylvanian System of sedimentary rocks.
Throughout northeastern Ohio, the Sharon Conglomerate is noted for the formation
of massive cliff faces, rock shelters, and slump blocks (Rau, 1970). The designation

Differential weathering of this cement results often in honeycomb patterns in the
cliff faces (Heimlich et al., 1970).

A. W. CUSICK: VITTARIA GAMETOPHYTES IN A NEW PROVINCE 35

At the base of the Sharon formation lies a major disconformity with the Meadville
Shale of the Cuyahoga group of Mississippian age sediments. This disconformity is
responsible for two phenomena which contribute to the habitat supporting the
populations of Vittaria, namely, the origination of springs and the formation of
slump blocks. The Sharon Conglomerate is highly permeable, whereas the underly-
ing shale is relatively impervious. This disconformity, then, is marked by a zone of
seeps and springs (Heimlich et al., 1970; Rau, 1970). The jointed conglomerate
tends to slip along this lubricated shale surface, splitting the rock, which slowly
moves downslope. On Little Mountain this joint-controlled slippage forms a pictur-
esque and complex system of blocks and chasms ranging from | to 3 m wide,
occasionally narrower, 10 to 15 m deep, and often as long as 50 m (Aronson, 1974).

The springs which flow from the disconformity on Little Mountain are excep-
tional in that they average 8-9°C year-round (Parsons, pers. comm.). This cold
temperature allied with the depth and narrowness of the fissures on the mountain
permits snow and ice to persist long into the warmer months. The writer noted the
remains of ice on 10 June 1982. Even in mid-summer one’s breath condenses before
one’s face while walking through the chasms on the mountainside. The environment
within the crevasses of Little Mountain conforms in every particular to the ecological
requirements of Vittaria as outlined by Farrar (1978, p. 4): “a low light intensity of
100 ft-c or less, relatively high humidity, and protection from temperature extremes

. overhanging rock outcrops, dense forest canopy, and nearness to running
water.”

Floristically, Little Mountain is a part of the White Pine-Hemlock—Northern

ardwoods community (Braun, 1950), a forest association of very limited occur-
rence in Ohio. The first botanist to visit the mountain may have been John L.
Riddell, who collected several species from this “pine-clad, rubblestone knob”
(Riddell, 1836, p. 567). Read (1873a, b) listed the major forest trees of the
mountain, particularly noting the Hemlock (7suga canadensis (L.) Carr.) and White
Pine (Pinus strobus L.) on the northern end of the mountain, the extensive growth of
American Chestnut (Castanea dentata (Marsh.) Borkh.) throughout the summit, and
“Rock Oak” (Quercus prinus L.) on the southern knob. All these species are yet
extant on the mountain, although the chestnut is represented only by fallen, dead
trees and root sprouts. Ferris (1887) presented an extensive list of vascular species
for the mountain. While his list contains some obvious inaccuracies, it also provides
further evidence that most of the herbaceous and woody species found on Little
Mountain in the past still grow there today and that the original vegetation
conformed to Braun’s concept of the White Pine—-Hemlock—Northern Hardwoods
community.

In most of its known range, the Vittaria gametophyte seems associated with the
Mixed Mesophytic forest as defined by Braun (1950). This is the case, for instance,
with other known Ohio populations. However, the Warren County, Pennsylvania
Station (Farrar et al., 1983), like Little Mountain, is within the White Pine-
Hemlock-Northern Hardwoods floristic province. The occurrence of Vittaria ga-
metophytes probably is more closely related to appropriate physical and geological
setting than to floristic province.

36 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Little Mountain was a prominent landmark to the first settlers of northeastern
Ohio. As early as 1831, a hotel was built on the mountaintop, and within the
following 50 years Little Mountain became a celebrated resort. At least three hotels
were built on the summit, together with a complex of summer cottages, gazebos,
churches, schoolhouses, and bowling alleys. The largest and best-known of the
hotels, the Pinecrest, built in the late 1880’s, featured a Western Union telegraph
line and a post office. The cool springs also induced developers to advertise water
cures for “invalids.” Guests clambered through the chasms on sweltering summer
days, enjoyed the cool breezes through the white pines, and gazed at the dramatic
view of Lake Erie from the northern knob. It was fashionable to carve one’s name on
the conglomerate in the “Devil’s Kitchen,” as one large grotto was dubbed. Many of
these graffiti are still legible today, a sort of permanent guest register. The resort era
ended by 1920 and the Pinecrest hotel was torn down in 1941. Little Mountain
ceased to be the playground of the public and instead became the private preserve of
the Little Mountain Club, an elite group of wealthy Clevelanders (Ahlstrom, 1961;
Ferris, 1887). The mountain could then recover from the years of wanton vandalism.
The present composition of the vegetation and the beautiful stand of mature White
Pine are tributes to the ability of plant communities to survive and recover from
severe disturbance. Other than the graffiti on the rocks, now largely cloaked by
bryophytes, the most notable evidence of the former resort is the abundance of Vinca
minor L. on the mountaintop.

Little of the disturbance described above would have had a direct effect on the
Vittaria population. However, it is difficult to be certain how common or widespread
the gametophytes might have been prior to the time of the hotels. Today, the
Vittaria, although occurring over a considerable linear area of the mountainside,
grows in isolated pockets, often only a few square cm in extent, on the walls of the

Ohio. An occasional colony of the gametophytes occurs in the cast left by the fall of
a particularly large quartz pebble. Most often the plants grow in recesses or narrow
clefts formed in zones of finer pebble size. The plants definitely are less frequent in

(1980, 1982) maps thin layers of Hiram Till of Wisconsinan age over even the
highest summits of the limits of the Appalachian Plateau in Ohio. Clearly, Little
Mountain was not a refugium on which individuals of Vittaria could have survived
glaciation.

The means of dispersal of this species, even in the heart of its range, is little
Dies § Although Vittaria gametophytes prolifically produce vegetative gemmae,
there seems to be no obvious mechanical means for transporting these gemmae
across the 90 km gap between Little Mountain and the nearest limit of the

A. W. CUSICK: VITTARIA GAMETOPHYTES IN A NEW PROVINCE 37

Wisconsinan ice. It seems unlikely that the few-celled gemmae could survive
wind-borne dispersal over such a distance without fatal desiccation. Also, it is
difficult to understand how wind currents can lift these gemmae from the sheltered
grottos and canyons in which they occur and convey them into the upper air. A
possibility, which seems highly unlikely at least to this writer, is that the plants were
introduced by an animal or human visitor who had accidentally picked up the
gemmae in southeastern Ohio only a short time before.

Both Farrar and this writer have found Vittaria gametophytes at the limit of
Wisconsinan glaciation in Fairfield County, Ohio, yet have failed to locate the
species in similar habitats and on the same rock strata in areas only a few kilometers
beyond the glacial boundary in other Ohio counties. Farrar’s two western Pennsylva-
nia stations for Vittaria gametophytes are located in Lawrence and Warren counties.
The Lawrence County site is about 80 km southeast of Little Mountain and is
located just north of the Illinoian glacial limit and south of the Wisconsinan
boundary. The Warren County population is about 160 km east-northeast of Little
Mountain and is directly south of the Illinoian limit. At present, the Warren County
Station is the northernmost known occurrence of these gametophytes (Farrar et al.,
1983).

It is probably useless to speculate overmuch on the origin of the Little Mountain
population of Vittaria until other, similar sites on the glaciated Appalachian Plateau
of Ohio and nearby ‘states are examined for this species. Appropriate habitats
elsewhere in glaciated regions may well harbor populations of this gametophyte.
Indeed, the perennial gametophyte of an as yet unknown species of Trichomanes has
been found on the glaciated Appalachian Plateau from central Ohio to New
Hampshire (Farrar et al., 1983; McAlpin et al., 1978). Perhaps this find of Vittaria
should not come as such a surprise after all.

In glaciated Ohio, suitable habitats are very limited in extent and highly
disturbed. If the case of Little Mountain is considered, however, disturbance which
may have extirpated other types of vegetation may not have had so severe an effect
on Vittaria. As of this writing, Vittaria gametophytes have not been found elsewhere
in glaciated Ohio. But much field work remains to be done before any firm
conclusions can be reached as to the true distribution of the gametophytes, either
nationally or statewide. This find opens new realms for fern enthusiasts to explore.

The author is indebted to R. Henry Norweb, Jr., Director of the Holden
Arboretum, for access to the Arboretum property and for permission to collect
vouchers. Brian Parsons and Thomas Yates of the Arboretum staff have provided
field assistance as well as access to unpublished data on Little Mountain and its
history. The Arboretum and its staff should be congratulated for their commitment to
the protection of natural areas. Mr. and Mrs. Hugh Pallister of Willoughby, Ohio,
also have provided the author with information on the history of Little Mountain.
My special thanks are owed to Dr. Donald R. Farrar of Iowa State University, Ames,
lowa, for verifying the identity of the Vittaria gametophytes, for his sharing of data,
and for his encouragement. This research was supported by the Division of Natural
Areas and Preserves, Ohio Department of Natural Resources.

38 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

LITERATURE CITED
AHLSTROM, J. M. 1961. The Little Mountain story. The Historical Society Quarterly, Lake Co., Ohio

3:43-46.

ARONSON, J. L. 1974. Mass wasting on Little es Lake and Geauga counties, Ohio (abstr.).
Geol. Soc. America Abs. with Programs

BRAUN, E. L. 1950. Deciduous Forests of Eastern North America. Blakiston, Philadelphia.

COOGAN, A. H., R. M. FELDMAN, E. J. SZMUC, and J. V. MRAKOVICH. 1974. Sedimentary
environments of the Lower Pennsylvanian Sharon Conglomerate near Akron, Ohio. Pp.
19-41 in: R. A. Heimlich and R. M. Feldman, eds. Guidebook No. 2: Selected Field Trips in
staribediets Ohio. Ohio Geological Survey, Columbus

FARRAR, D. R. 1978. Problems in the identity and origin 0b the Appalachian gametophyte, a
fpornye fern of the eastern United States. Amer. J. Bot. 65:1—12.

PARKS, and B. W. McALPIN. 1983. The fern genera Vittaria and Trichomanes in the
northeastern United States. Rhodora 85:83-92.

FERRIS, 'E. J. 1887. History of the Little Mountain from 1810 to 1887. Painesville Advertiser,
Painesville, OH

GORDON, R. B. 1969. The natural vegetation of Ohio in pioneer days. Ohio Biol. Surv. Bull.,
3(2):xi + 113

HEIMLICH, R. A., J. V. MRAKOVICH, and G. W. FRANK. 1970. The Sharon Conglomerate. Pp.
125-133 in: P. O. Banks and R. M. Feldman, eds. Guide to the Geology of Northeastern
Ohio. North. Ohio Geol. Soc., Cleveland.

MCcALPIN, B., and D. R. FARRAR. 1978. Trichomanes gametophytes in Massachusetts. Amer. Fern J.
68:97-98.

RAU, J. L. pl Pennsylvanian System of northeast Ohio. Pp. 69-124 im: P. O. Banks and R. M.
Fe n, eds. Guide to the geology of northeastern Ohio. North. Ohio Geol. Soc., Cleveland.

READ, M. ee Geology of Geauga County. Rept. Ohio Geol. Surv. 1(1):520—533.

—————. 1873b. Geology of Lake County. Rept. Ohio Geol. Surv. 1(1):510-519.

RIDDELL, J. 1836. Supplementary catalogue of Ohio plants. Western J. Med. Phys. Sci.

35
US. Ss Rerind SURVEY. see Mentor Quadrangle, 7.5 minute series (topographic). U. S.
Dept. Interior, Washingto
WHITE, G. W. 1980. Glacial pelea of Lake County, Ohio. Ohio Geol. Invest. Rept. No. 117.
————.. 1982. Glacial geology of northeastern Ohio. Ohio Geol. Surv. Bull. 68.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 2 (1983) 39

Chapman’s Quillwort Reconsidered
BRIAN M. BOOM*

Certainly one of the least known pteridophytes in the southeastern United States is
Chapman’s Quillwort. Described by Engelmann (1882) as /soétes flaccida Shuttlew.
var. chapmanii Engelm., the taxon has been collected only a few times since its
initial collection by A. W. Chapman in 1848. The collections are always from the
same general locality in Jackson County, Florida. Larger megaspores with smoother
ornamentation were said by Engelmann to distinguish it from the typical variety.

Underwood (1900) and Clute (1928) both recognized the distinctness of the
taxon, and Small (1932) even elevated it to specific status. Lakela and Long (1976)
maintained J. chapmanii (Engelm.) Small, and in their key distinguished it from /.
flaccida on the basis of megaspore ornamentation and sporangium shape. On the
other hand, Pfeiffer (1922) and Boom (1979, 1982) were not convinced of the
distinctness of J. chapmanii, and they synonymized it with I. flaccida. Recently I
was able to study the spores of these plants more carefully by means of scanning
electron microscopy (SEM), and I now regard the evidence as sufficient for
maintaining J. flaccida var. chapmanii as a rare, but distinct element of Florida’s
pteridophyte flora.

Spores were coated by a two-step procedure consisting of the deposition from a
low voltage arc of an initial carbon layer onto which was subsequently sputtered a
60:40 gold-palladium alloy using a 15 watt DC diode coater. Specimens were
examined with a Cambridge S4-10 SEM. Photographs were taken with Polaroid
Type 665 Positive/Negative Land Film. Spores of some thirty specimens of /soétes
flaccida were examined (vouchers cited in Boom, 1979), but the photomicrographs
of only three are presented herein: var. flaccida: Lake Flirt, near Lake Okeechobee,
Florida, Aug 1878, A. P. Garber s. n. (US 240797); cypress pond near Cobb,
Sumter Co., Georgia, R. M. Harper 1046 (US 400120); and var. chapmanii:
Tributary of the Chipola River at Marianna, Jackson Co., Florida, R. K. Godfrey
61963 (US 2424571). :

Both megaspores and microspores of the two varieties show differences In perine
ornamentation which are taxonomically significant (Figs. / -6). Additionally, the
differences in megaspore size provide a useful means of distinguishing the varieties.
The ornamentation of J. flaccida var. chapmanii megaspores is low tuberculate,
densely so on the distal hemisphere and sparsely so on the proximal (Fig. /).
Greater magnification shows the perine to be minutely echinate (Fig. 3). In var.
flaccida, the megaspores are also low tuberculate, but densely so on both hemt-
spheres (Fig. 2). Greater magnification reveals a finely rugulate perine, the muri
anastomosing so as to give a spongy appearance (Fig. 4). The diagnostic perine
characters (extent and distribution of tubercules) can be seen at 40X, and the size
difference detected with a dissecting microsc

Although the microspores of the varietie
perine ornamentation differs: that of var. chapmanii b

ope.
s are essentially the same size, their
eing echinate with an arach-

*New York Botanical Garden, Bronx, NY 10458.

40 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

FIGS. 1-6. Isoétes flaccida spores. FIG. . Megaspore of var. chapmanii (US itll ds scale = 100
um. FIG. 2. Megaspore of var. flaccida ‘ie sieht scale = um. F 3. Megaspore perine of
var. chapmanii (US 2424571); scale = 7 wm. FIG. 4. Meg aspore perine of var. _Placelue 240797);
scale = Hm. FIG. 5. Microspore pe var. chapman (US 2424571); scale = § . FIG. 6.
Microspore of var. flaccida (US 400120): scale = Lm.

8
S

————

B. M. BOOM: CHAPMAN'S QUILLWORT RECONSIDERED 41

noid perine between echinae (Fig. 5), and that of var. flaccida being papillate with a
laevigate perine between papillae (Fig. 6). Given their average size of ca. 30 ym, the
microspores do not provide useful taxonomic characters for the field or herbarium
botanist, but the ornamentation differences help confirm their distinctness.

Aside from the spore characters, there are no other apparent morphological,
chemical, or ecological differences that separate the varieties. The orbicular or
obovoid sporangium ascribed to var. chapmanii by some authors (Engelmann, 1882;
Small, 1932; Lakela & Long, 1976) as a feature distinguishing it from var. flaccida
(with an oval, ellipsoid, or subglobose sporangium) is not a good character, as this
structure varies too much depending on plant size and ecological conditions. The
flavonoid chemistry of the two varieties was examined by Boom (1979), but the
chromatographic profiles were identical. Ecologically, there appears to be nothing
striking about the habitat of var. chapmanii. A topotype collection (Boom 333,
TENN) reports “sandy/peaty substrate, clear water in deep bald cypress swamp;” the
stream is underlain by limestone. This general description could apply to many areas
in Florida where var. flaccida occurs, but perhaps there are significant microhabitat
factors involved which have not yet been detected. A distribution map of I. flaccida
is given in Boom (1982).

Just as Taylor et al. (1975) used SEM to confirm the distinctness of Isoétes butleri
Engelm. from J. melanopoda Gay & Dur. based on spore ornamentation, it has been
possible to show that two entities are encompassed by /. flaccida. | prefer to
recognize these two taxa at the varietal level because, unlike the case of J. butleri
and J. melanopoda, the two taxa involved here are not ecologically or geographically
differentiated. I do not regard I. flaccida var. chapmanii as a hybrid because it meets
none of the criteria used in the detection of several instances of hybridization in
other southeastern U.S. quillworts (Boom, 1982). I consider I. flaccida var. flaccida
to encompass J. alata Small and /. flaccida vat. rigida Engelm. The following key
separates the two recognized varieties of the species:

KEY TO VARIETIES OF ISOETES FLACCIDA
Megaspores 300-400(440) zm diam., densely low tuberculate on both hemispheres,
papillate; common throughout Florida and southern Georgia ........:.ssssseereeeeressee es
Megaspores 440-540 xm diam., densely low tuberculate on distal hemisphere, sparingly so on
proximal; microspores echinate; rare in northwestern Florida (Jackson Co.) ......-- var. chapmani

Appreciation is extended to David Lellinger, Andrea Sessions, and Walter Brown
for assistance with /soétes spore studies at US. The present work was materially
aided by the Botany Department of The University of Tennessee, Knoxville, by a
Grant-In-Aid of Research from Sigma Xi, and by a travel grant from The Smithsonian
Institution.

microspores
var. flaccida

LITERATURE CITED
BOOM, B. M. 1979. Systematic studies of the genus Isoétes in the southeastern United States. M.S.
Thesis, Univ. of Tennessee, Knoxville, TN.
. 1982. Synopsis of Isoétes in the southeast

RS aegy ern United States. Castanea 47:38-5
CLUTE, W. N. 1928. The Fern Allies of North America

9.
North of Mexico. Willard N. Clute & Co.,

- Joliet, IL. : :
ENGELMANN, G. 1882. The genus Isoétes in North America. Trans. St. Louis Acad. Sci. 4:358-390.
LAKELA, O. and R. W. LONG. 1976. Ferns of Florida. Banyan Books, Miami, FL.

42 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

PFEIFFER, N. E. 1922. Monograph of the Isoétaceae. Ann. Missouri Bot. Gard. 9:79-233.

SMALL, J. K. 1932. Ferns of Florida. Science Press, New York, NY.

TAYLOR, W. C., R. H. MOHLENBROCK, and J. A. MURPHY. 1975. The spores and taxonomy of
Isoétes butleri and I. melanopoda. Amer. Fern J. 65:33-38.

UNDERWOOD, L. M. 1900. Our Native Ferns and Their Allies. Henry Holt, New York, NY.

REVIEW

Flora Malesiana, Series II—-Pteridophyta, volume 2, part 5, Thelypteridaceae,
by R. E. Holttum. Pp. 331-599, including index and addenda for entire volume,
plus pp. 1-20, contents and dedication. 1982. Martinus Nijhoff, BV, The Hague,
Netherlands. $63.00.—Historically, no other group of ferns has been subjected to
such varied circumscriptions, both of the family and genera; moreover, species have
been notoriously under-collected, ill-defined and confused, and their relationships
misunderstood. Part of the reason is their mundane (cringe) and superficially similar
look that causes collectors to sample one or a few and pass by the rest. Another
reason is that few taxonomists have bothered to look at them closely. The first who
did look was Carl Christensen, to whom Holttum appropriately dedicates this
volume; but Christensen concerned himself mostly with Neotropical species. In
1963, Ching presented a revised classification of Old World genera that provided a
springboard for Holttum’s many precursory revisions culminating in the present work.

The sheer size and complexity of Thelypteridaceae have made Holttum’s task
formidable. This family is probably the largest in ferns, with almost 1000 species. In
the Flora Malesiana region alone Holttum treats 440 species in 22 genera. The
number of new taxa (75 species, 18 varieties) is staggering and reminds us how
poorly known are the tropical floras and how urgent it is to preserve what little is
left. In Sphaerostephanos, with 152 Malesian species, 43 are known only from the
type and many additional species are not much better represented in herbaria.

But Holttum’s most important contribution is not the number of species named
and redefined; it is the understanding of the large natural groups of species. Here, he
has brought order from chaos. Doubtless there will be taxonomists, including
myself, who will prefer to assign lower rank to some of Holttum’s genera. But his
circumscriptions of natural groups are likely to survive intact or with only minor
redefinition.

At least 13 new chromosome counts are embedded in the work: they may well be
overlooked by compilers of chromosome indices. The most interesting, n= 66 for a
species of Coryphopteris, is a new base number (x= 33) for Thelypteridaceae and
may lend support to Holttum’s contention of a relationship between this family and
Cyatheaceae.

The magnitude, originality, and scholarship shown in this work are truly impres-
sive and culminate a brilliant career. But Holttum is already turning to other
ahi — and one wishes him many more years of taxonomic insight.
—Atan RK. Smith, Herbarium, Departme iversi ifornia,
Berkeley, CX oF Dp nt of Botany, University of Califo

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 2 (1983) 43

C-Glycosylxanthones in Diploid and Tissue Culture-induced
Autotetraploid Davallia fejeensis
P. MICK RICHARDSON and HARINDER K. PALTA*

1,3,6,7-tetrahydroxy-C-glycosylxanthones are phenolic compounds which have
been found relatively infrequently in ferns. A recent survey (Wallace et al., 1982)
reported their occurrence in seven genera: Asplenium, Athyrium, Elaphoglossum,
Cardiomanes, Hymenophyllum, Trichomanes, and Marsilea. They have also been
reported in Ctenitis decomposita (Bohm, 1975). The present communication reports
C-glycosylxanthones in Davallia fejeensis Hooker. Diploid and tetraploid samples of
both the sporophytic and gametophytic generations were examined.

MATERIALS AND METHODS

Rhizome tips (about 2 cm long) of diploid Davallia fejeensis were collected in the
Enid A. Haupt Conservatory of the New York Botanical Garden, cleaned, rinsed in
tap water and further trimmed. Voucher specimens are at NY. Explants (6-8 mm
long) were cultured on 1% agar-gelled sterile nutrient medium of Knudson as
modified by Steeves et al. (1955) and supplemented with 2% sucrose in order to
provide control diploid sporophytes growing in axenic culture. Roots, leaves, and
thizomes of the resulting plants were excised and induced to differentiate into
aposporous diploid gametophytes. Stock cultures of diploid gametophytes were
multiplied and further grown on liquid media supplemented with 0.6% agar to effect
fertilization and raise tetraploid sporophytes. Tetraploid gametophytes were isolated
from this tissue culture-induced tetraploid material by repeating the procedure
followed at the diploid level. In this way, both diploid and tetraploid sporophytes and
gametophytes were made available for this study.

Phenolic compounds were isolated by two-dimensional paper chromatography of
an 80% methanol extract of green material in t-BuOH-HOAc-H,0, 3:1:1 (TBA) and
15% aqueous acetic acid (HOAc), followed by one-dimensional paper chromatogra-
phy in water. C-glycosylxanthones appeared as orange compounds in ultraviolet light
and turned yellow when fumed with ammonia. Both purified compounds were
co-chromatographed with mangiferin and isomangiferin isolated from Asplenium
montanum Willd. (Bozeman & Radford 11552, NY). Rr values in TBA, BAW,
HOAc, and H,O were: mangiferin, 0.31, 0.43, 0.43, 0.12; isomangiferin, 0.19,
0.29, 0.23, 0.04; Asplenium mangiferin, 0.34, 0.42, 0.42, 0.11; Asplenium
isomangiferin, 0.21, 0.32, 0.23, 0.03; and rutin standard, 0.43, 0.45, 0,56; 0:25.
The BAW and HOAc values are very similar to those of Smith and Harborne (1971),
but the TBA and HOAc values differ from those of Markham and Wallace (1980).
Absorption spectra (MeOH, nm) were: mangiferin, 232, 260, 270sh, 310, 364;
isomangiferin, 240, 258, 270sh, 312, 365. The compounds were unaffected by acid
hydrolysis (1 hr, 2N HCI, 100°C).

“Harding Laboratory, New York Botanical Garden, Bronx, NY 10458.

44 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

RESULTS AND DISCUSSION

The C-glycosylxanthones mangiferin and isomangiferin were the only phenolic
compounds appearing on the two-dimensional chromatograms of an 80% methanol
extract of each sample. The diploid and tetraploid sporophytes contained relatively
large amounts of both compounds, but in both cases the mangiferin spot was much
larger than the isomangiferin spot. Gametophyte extracts contained very small
amounts of each compound. Further identification was performed only on com-
pounds from the sporophytes. Spots from the chromatograms were eluted and then
chromatographed in water to purify them sufficiently for ultraviolet spectrophotome-
try and co-chromatographic testing. The purified compounds showed reduced Ry
values in comparison to those measured on the two-dimensional chromatograms,
strongly suggesting that Ry values based on such chromatograms may be misleading.

These results suggest that the same C-glycosylxanthones are produced in both
sporophytes and gametophytes. This is comparable to the only similar published
study of fern flavonoids (Petersen & Fairbrothers, 1980), where the same flavonol
glycosides appeared in both generations. Secondly, it appears that an induced
doubling of the chromosome number in Davallia has no effect on the production of
C-glycosylxanthones in either generation. If we accept that the C-glycosylxanthones
in Davallia are equivalent to the flavonoids in flowering plants, then the effects of
induced chromosome doubling on phenolic production in ferns may be compared
with the effect of a similar chromosome doubling in angiosperms. Mears (1980) has
reviewed the chemistry of polyploids. In some cases of induced autotetraploids in
Phlox (Levin, 1968), no qualitative differences in phenolics were detected; in
another study in the same genus (Levy, 1976), such differences did occur. In Briza
(Murray & Williams, 1976) qualitative differences in phenolic production some-
times occurred in induced tetraploids. A more complex situation occurred in
autotetraploids of Gibasis (del Pero de Martinez & Swain, 1977), where polyploidy
is correlated with Robertsonian Fusion, a phenomenon which may be responsible for
changes in phenolic expression.

This is the first study of the chemistry of induced autoploidy in fern gametophytes
and sporophytes. The tissue culture method followed in this study is an excellent
way to effect such changes in chromosome numbers. A survey for the presence of
C-glycosylxanthones in other fern genera is being undertaken in our laboratory; it is
pees that these compounds may not be as rare as they were previously thought to

LITERATURE CITED

BOHM, B. A. 1975. Xanthones in the fern Ctenitis decomposita. Phytochemistry 14:287—288.

LEVIN, D. A. 1968. The genome constitutions of eastern North American Phlox amphidiploids.
Evolution 22:612-632.

LEVY, M. 1976. Altered piycofiavane expression in induced autotetraploids of Phlox drummondii.
Biochem. Syst. Ecol. 4:249-254.

MARKHAM, K. R. and J. W. WA ALLACE. 1980. C-glycosylxanthone ~ flavonoid variation within
the filmy ferns (Hymenophyllaceae). Phytochemistry 19:415—4

MEARS, J. A. 1980. Chemistry of polyploids: a summary with comments on Parthenium (Asteraceae-
Ambrosiinae). In W. H. Lewis, ed. Polyploidy. Plenum Press, New York and London.

RICHARDSON & PALTA: C-GLYCOSYLXANTHONES IN DAVALLIA FEJEENSIS 45

MURRAY, B. G. and C. A. WILLIAMS. 1976. Chromosome number and flavonoid biosynthesis in
Briza L. (Gramineae). Biochem. Genet. 14:897—904.

PERO de MARTINEZ, M. A. del and T. SWAIN. 1977. Variation in flavonoid patterns in relation to
chromosome changes in Gibasis schiedeana. Biochem. Syst. Ecol. 5:37-43.

PETERSEN, R. L. and D. E. FAIRBROTHERS. 1980. Flavonoid synthesis and antheridium initiation in
Dryopteris gametophytes. Amer. Fern. J. 70:93-95.

SMITH, D. M. and J. B. HARBORNE. 1971. Xanthones in the Appalachian Asplenium complex.
Phytochemistry 10:2117-2119.

STEEVES, T. A., I. M. SUSSEX and C. R. PARTANEN. 1955. In vitro studies on abnormal growth of
prothalli of the bracken fern. Amer. J. Bot. 42:232-245.

WALLACE, J. W., K. R. MARKHAM, D. E. GIANNASI, J. T. MICKEL, D. L. YOPP, L. D.
GOMEZ, J. D. PITTILLO and R. SOEDER. 1982. A survey for 1,3,6,7-tetrahydroxy-C-
glycosylxanthones emphasizing the “primitive” leptosporangiate ferns and their allies. Amer.
J. Bot. 69:356—-362.

REVIEW

THE GENUS SELAGINELLA IN TROPICAL SOUTH AMERICA, by
A. H. G. Alston, A. C. Jermy, and J. M. Rankin. Bull. Brit. Mus. (Nat. Hist.),
bot. ser. 9(4):233-330. 1981. £14.00 postpaid.—A. H. G. Alston was the premier
Selaginella authority in the middle of this century. He published extensively on the
genus in the Old World and in southern South America, Brazil, the West Indies, and
Central America in the New World. At the time of his death, he had begun to study
Selaginella from the Andean countries and the Guyanas. The present paper com-
pletes and extends Alston’s work and includes Brazil, thus accounting for virtually
all of tropical South America. The region contains 133 species and six infraspecific
taxa of Selaginella. A table of species distribution by countries shows that Colombia
has by far the greatest number of species. The bracketed key to the species can be
read forward or backward, a decided advantage because there are no species
descriptions. Each taxon treated has a synonymy, a statement of range, a list of
specimens, and, sometimes, additional notes. The authors have uncovered some
overlooked synonyms in rare literature, always a pleasure to see. However, most
synonyms state only the country where the type was collected, rather than the type
locality, the collector and number, and the herbarium of deposit. A list of references
and an index to accepted names and synonyms concludes the volume. New taxa are
illustrated by a photograph of the herbarium sheet and by SEM photographs of the
leaves, which range in magnification upwards from a mere eight times natural size, a
useful technique for illustrating clearly the cilia and other small details of the leaves.
This paper is indispensible for identifying Selaginellas from tropical America and
Will be the standard reference for many years to come.—D.B.L.

46 AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 2 (1983)

The Distribution of Woodwardia areolata
R. CRANFILL*

Phytogeographical studies in North America, as compared with those in Europe,
are still in their infancy. Lack of detailed atlases and the relative paucity of extensive
paleo- and neoecological studies in North America hamper the botanist who wishes
to investigate the factors affecting the distribution of a particular species. A few
botanical studies of this sort are available (Hocker, 1956; Koevenig, 1976; Salisbury,
1926), although most are limited to local areas or to single factors.

The distribution of a particular plant is determined by several interrelated factors,
including: (1) climate and soils, both present and past, (2) interactions with other
organisms, (3) production and dissemination of propagules, and (4) evolutionary
history, including time and place of origin (Billings, 1952; Cox et al., 1976; Krebs,
1972; Pielou, 1979). In general, physical parameters of the environment, such as
moisture availability or extremes of temperature, place absolute limits on_ the
distribution of an organism. Interactions with other species (including man!), soils,
dispersability, and other historical factors (e.g., hurricanes, stream piracy, and
glaciation) constrain the organism within these bounds (Billings, 1952; Gorham,
1954). As many have pointed out (Cox et al., 1976; Koevenig, 1976), the study of
the interaction of limiting factors is problematic because of our inability to deal
quantitatively with several seemingly inextricably linked variables at once. Analysis
of widespread or narrowly restricted species is often easiest because in these
Situations one or a small number of variables contributes differentially to the
distribution. Even so, the implications of such correlations are not always as clear as
they may seem. In the absence of corroborative experimental evidence, caution
should always be exercised in the interpretation of distributional correlations.

Woodwardia areolata (L.) Moore is ideal for this type of study because it is
widespread and common throughout the southeastern and Atlantic United States and
is well collected. One of the three species in subgenus Lorinseria, it is characteristic
of acidic mucky, sandy, and peaty bogs throughout the southeastern United States
(Wherry, 1921, 1964). The aim of this paper is to make some ecological inferences
from the distribution of W. areolata and to discuss some of the problems inherent in
this sort of induction.

MATERIALS AND METHODS

The distribution of W. areolata was compiled from herbarium specimens depos-
ited at the following institutions: DHL, F, FLAS, FSU, GH, KY, MEM, MICH,
MO, NCU, NY, SMU, TAES, TENN, TEX, TNS, UC, USF, VDB (abbreviations
of Holmgren et al., 1981). Over 700 sheets were examined and plotted accurately on
a base map of the eastern United States and Canada. Climatic and edaphic
parameters were drawn from a variety of sources. Isotherms were composed and
redrawn from individual state maps (National Atmospheric and Oceanographic
Institute, 1980).

*Department of Botany, University of California, Berkeley, CA 94720.

—e

—

R. CRANFILL: DISTRIBUTION OF WOODWARDIA AREOLATA 47

RESULTS AND DISCUSSION

The distribution of W. areolata is shown in Fig. 1. A specimen found at USF
(Cheever s. n., 25 June 1917) verifies the occurrence of this species in Maine
(previously questioned by Ogden et al., 1948). It also seems likely that the species
may occur in southern Indiana and extreme eastern Kansas. Associations of the
distribution with several edaphic and climatic factors were found and are discussed
below.

Climatic Factors.—The climatic feature that appears to have the greatest influ-
ence on W. areolata is minimum winter temperature. Of the three isotherms
examined (0°C, —2.0°C, and —4.5°C), the —4.5°C mean made the closest fit. All
stations, save five, fell to the south of the line, with especially close correspondence
in New England. Of the sites north of the isotherm, three are no longer extant
(Kitfield, 1974; Price, pers. comm.), while another lies close to the Atlantic Ocean
which may have a moderating influence not reflected in the climatic data. It is
interesting to note the absence of this species from the higher Appalachians of
Virginia and West Virginia (Wagner, 1963), even though suitable acidic bogs and
seeps exist in the area.

It is apparent that mean temperature in itself is not a very useful index. As an
indicator of potential climatic severity it can be important, although analysis is
nearly impossible in the absence of values for variance. Thus, survival of W.
areolata may not depend as much on its tolerance of —4.5°C as on its ability to
withstand the occasional much colder winter. Of course the problem may be even
more complex. Winter minima may reflect other associated parameters, such as the
length of the growing season, that more directly affect the ability of the plant to
grow, reproduce, and compete with its neighbors.

Competition can be critical in the distribution of organisms (Hynes, 1954; Jaeger,
1970). Since competition varies as a function of community composition, which
changes over the distribution of any particular species, a perfect or even very close
fit between abiotic influences and distributional limits often does not obtain.
Distributions of weedy species, therefore, probably more closely parallel absolute
climatic limits than do distributions of plants of stable habitats, where interactions
with other organisms are more structured. The recent northward spread of Asplenium
platyneuron (L.) B.S.P., primarily a southern species, is a result of an increase
in the disturbed habitats that provide this species with suitable sites in which
to become established (Wagner and Johnson, 1981) and demonstrates that the
Previous exclusion of this spleenwort was more a result of its competition with
established communities than a result of its tenderness. A similar explanation may
account for the presence of W. areolata on siliceous cliffs and ledges at the northern
and western limits of its range, even though seemingly suitable bogs and seeps exist
in these areas. At some point, the species composition of the typical habitat of this
Species changes sufficiently so that it no longer is competitive in such situations.
Sandstone cliffs and ledges in the same vicinity, which present physiological
demands similar to those of acidic bogs (in the form of nutrient inavailability and
drought stress), possess less plant cover and are characterized by a much lower
diversity of species. It seems likely, therefore, that lithophily in W. areolata may be a

AMERICAN FERN JOURNAL: VOLUME 73 (1983)
48

FIG. 1. Distribution of Woodwardia areolata. Dots represent terrestrial and unspecified neha
while stars indicate epipetric stations. The ragged line represents the Janu —4.5°C isotherm. ert
shows Canadian province of Nova Scotia. X indicates extirpated population. Arrows indicate outlying
populations.

R. CRANFILL: DISTRIBUTION OF WOODWARDIA AREOLATA 49

direct response to competition. Such cases would be parallel to but more subtle than
Billings’ (1950, 1952) “Compensation Effect,” in which edaphic factors of certain
soils and rock types compensated for the negative effects of other environmental
factors. Transplants of individuals of W. areolata from Kentucky to Michigan
survived over two winters (until 1981 when the experiment was discontinued) and
indicate that winter minima alone are insufficient to account for the distribution of
this fern. Further demographic and transplant experiments are needed to adequately
test the hypothesis that competition is the controlling factor.

Edaphic Factors.—The distribution of W. areolata fitted that of certain soil types
even better than it did temperature. My measurements and those of Wherry (1921)
indicate that the sporophyte of this fern is extremely acidophilous. The pH of several
sites checked in Kentucky and Tennessee all fell below 5.5, with one reading of 4.0.
It is not surprising then that the species is entirely absent from calcareous regions
within its range. No stations are known in the limestone regions of the Interior Low
Plateaus or in the calcareous Black Belt region of Alabama and Mississippi.
Absence of the species from glaciated areas in southern Illinois and southeastern
Ohio can be ascribed to the predominance of calcareous glacial drift in these areas
(Fig. 2). The alluvial plain of the lower Mississippi River is also avoided. As a
region of heavy basic to circumneutral clays (Kellogg, 1936; Braun, 1950), the
Mississippi Alluvial Plain is devoid of most species characteristic of the sandy
uplands and acidic seeps to the east and west. Correspondence of the distributions of
W. areolata and Pinus echinata Miller (Little, 1971), another calcifuge, is striking.

The best fit to soils occurs in eastern Texas (see inset, Fig. 2). The absence of the
Chain Fern from the regions of the Blackland Prairie and Coast Prairie soils, both
predominantly calcareous clays (Arbingast et al., 1976), is striking. Because of their
greater matrix potential, clay soils must be hydrated to a higher percentage than
coarser grained soils to exceed the permanent wilting percentage. As a result, clay
soils tend to be droughtier in more arid regions and may become critical for W.
areolata as it reaches its western limit. The patchwork distribution of these soils has
made it impossible, therefore, to assess the relative importance of moisture availabil-
ity (for an index, see Thornthwaite, 1948) and relative humidity as limiting factors;
no good correlation was found between these factors and the western limit of W.
areolata.

As with the consideration of climate, distributional evidence is insufficient to
demonstrate that heavy clay soils or pH are in themselves limiting. The indefinite
Survival of adult sporophytes on the heavy, calcareous, nutrient-rich soils of the
Inner Bluegrass region of Kentucky indicates that competition may actually consti-
tute the limiting factor.

The absence of this fern from much of the Allegheny Plateau cannot be explained
by the climatic or edaphic criteria just discussed. Contrary to its name, the plateau
region is highly dissected and affords little of the seep and bog habitat necessary for
the species. It also lacks extensive deposits of resistant sandstone and conglomerate
that form the secondary habitat of the fern.

Although the previous discussion concerned the sporophyte, other stages 1n the
life history may be important in controlling the distribution. Pteridophytes are like

0 AMERICAN FERN JOURNAL: VOLUME 73 (1983)
bs)

FIG. 2. Distribution of Woodwardia areolata in relation to soils; a = Mississippi Alluvial Plain, b =
region of Interior Low Plateaus with calcareous soils, c = Ridge and Valley Province, d = Black

blackland prairie soils while crosshatched area represents the extent of coastal prairie soils. Hatched line
depicts extent of Wisconsin Glaciation. X indicates extirpated population.

R. CRANFILL: DISTRIBUTION OF WOODWARDIA AREOLATA 5]

lower plants in which the alternation of generations can have drastic consequences
on the distribution of the organism (see the example of Laminaria, Dixon, 1965, pp.
109-115).

In summary, the present distribution of W. areolata is clearly associated with the
mean —4.5°C minimum January temperature to the north. The distribution of
calcareous soils, especially clays, effectively limits the distribution to the south and
may be important, in concert with increasing aridity, to the west. No correlation was
found between the distribution of this fern and the indices of moisture availability
and relative humidity. In all cases it appears likely that competition narrows the
limits imposed by abiotic influences since preliminary experiments demonstrate the
ability of this species to survive in more rigorous environments when competition is
removed. The change from bogs and seeps to cliffs and ledges at the range limit is
also compatible with this hypothesis. This “competitional compensation point” is
probably more important in limiting the distributions of non-weedy species, which
occupy more stable habitats than do weedy species.

Experimental studies of problems of distribution are greatly needed. Through nar-
rowing and refining our investigations, they may aid materially in the study of the
effects of competition on community structure. Ferns make excellent subjects
because they are small and fairly easy to grow and because the production and
dissemination of propagules, critical in many flowering plants, is not so limiting in
pteridophytes.

I am grateful to A. R. Smith and Herbert Baker who provided comments on a
draft of this paper. I am indebted to D. L. Melroy who provided assistance and
criticism in numerous ways.

LITERATURE CITED

ARBINGAST, S. A., L. G. KENNAMER, and R. H. RYAN. 1976. Atlas of Texas. Bureau of Business
Research, Univ. of Texas, Austin, TX.

BILLINGS, W. D. 1950. Vegetation and plant growth as affected by chemically altered rocks in the
western Great Basin. Ecology 31:62—74
. 1952. The environmental complex in
Biol. 27:251-265.

BRAUN, E. L. 1950. Deciduous Forests of Eastern North America. Free Press, New York, NY.

COX, C. B., I. N. HEALEY, and P. D. MOORE. 1976. Biogeography, an Ecological and Evolutionary
Approach. Blackwell Scientific Publications, Oxford, U.K.

DIXON, P. S. 1965. Changing Patterns of Distribution in Marine Algae. Pp. 109-1 15 in C. G. Johnson
and L. P. Smith, eds. The Biological Significance of Climatic Changes in Britain. Academic
Press, London, U.K. :

GORHAM, E. 1954. An early view of the relationship between plant distribution and environmental
factors. Ecology 35:97—98. ae

HOCKER, H. W. 1956. Certain aspects of climate as related to the distribution of Loblolly Pine.
Ecology 37:824-834. :

HOLMGREN, P., W. J. KEUKEN, and E. SCHOFIELD. 1981. Index Herbariorum, Part I. Guide to
the Herbaria of the World, 7th ed. W. Junk, The Hague, Netherlands.

HYNES, H. B. N. 1954. The Ecology of Gammerus duebeni Lilljeborg and its occurrence in fresh

water in western Britain. J. Animal Ecol. 23:38-84.

, R. C. 1970. Potential extinction through compe

salamanders. Evolution 24:632-642.

relation to plant growth and distribution. Quart. Rev.

JAEGER tition between two species of terrestrial

52 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

KELLOGG, C. E. 1936. Development and Significance of the Great Soil Groups in the United States.
. S. Dept. Agr. Misc. Pub. :
KITFIELD, A. 1974. A range extension for Woodwardia areolata. Rhodora 76:312.
KOEVENIG, J. L. 1976. Effect of climate, soil, physiography and seed germination on the distribution
of the River Birch (Betula nigra). Rhodora 78:420-437.

KREBS, C. J. 1972. Ecology, the Experimental Analysis of Distribution and Abundance. Harper and
Row, New York, NY.

LITTLE, E. L. 1971. Atlas of United States Trees, vol. I. Conifers and Important Hardwoods. U. S.
Dept. Agr. Misc. Pub. 1146.

NATIONAL ATMOSPHERIC AND OCEANOGRAPHIC INSTITUTE. 1980. Climate of the States, 2
vols. Dept. of Commerce, Washington, :

OGDEN, E. C., F. H. STEINMETZ, and F. HYLAND. 1948. Checklist of the Vascular Plants of
Maine. Bull. Josselyn Bot. Soc. Maine 8.

PIELOU, E. C. 1979. Biogeography. Wiley and Sons, New York, NY.

SALISBURY, E. J. 1926. The geographical distribution of plants in relation to climatic factors. Geogr.
. 67:312-335.

THORNTHWAITE, C. W. 1948. An approach to a rational classification of climate. Geogr. Rev.

38:55—94.,
WAGNER, W. H., Jr. 1963. Pteridophytes of the Mountain Lake area, Giles County, Virginia,
including notes from Whitetop Mountain. Castanea 28:113-150.
————, and D. JOHNSON. 1981. Natural history of the Ebony Spleenwort, Asplenium platyneuron
(Aspleniaceae) in the Great Lakes area. Canad. Field Nat. 95:156-166.
WHERRY, E. T. 1921. The soil reactions of ferns of woods and swamps. Amer. Fern J. 11:5-16.
—————. 1964. The Southern Fern Guide. Doubleday Nature Guide Series. Doubleday, Garden City,
NY.

REVIEW

FERNS AND FERN ALLIES OF THE DRIFTLESS AREA OF ILLINOIS,
IOWA, MINNESOTA AND WISCONSIN, by James H. Peck. Milwaukee Public
Museum Contributions in Biology and Geology 53:1-140. 1982. $13.50 postpaid.
—The driftless area, a pocket in the upper midwestern landscape which escaped at
least the more recent glaciations, holds, by virtue of its topography and age of its
habitats, a diverse and interesting array of plants. It has been the subject of botanical
Study for many years. The present account is by far the most complete and useful of
any we have had for the pteridophytes, which number 73 species, 13 hybrids, and 6
infraspecific taxa. Peck’s treatment includes a useful introduction, a discussion of
the affinities of the flora with special reference to the interesting disjunct Thelypteris
simulata, a systematic list with Synonyms, statement of habitat, and specimen
citations, a key to the genera and species of pteridophytes found in the driftless area,
and a large and useful section of literature cited. Ninety. pages are devoted to
full-page county distribution maps for the driftless area ferns that include all of
Minnesota, Wisconsin, Iowa, Illinois, and Missouri. The maps not only place the
ferns of the driftless area in the context of the surrounding region, but they are sure

to stimulate searches for pteridophytes in counties where the first records are yet to
be obtained.—D.B.L.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 2 (1983) 53

Two Moonworts of the Rocky Mountains;
Botrychium hesperium and a New Species
Formerly Confused with It
W. H. WAGNER, JR. and FLORENCE S. WAGNER*

The taxonomy of western North American botrychiums still needs much research.
Interpretations of the past were based largely upon scanty and poorly prepared
collections. For over 30 years only two B. matricariifolium-like moonworts have
usually been accepted for this region—B. boreale subsp. obtusilobum (Rupr.)
Clausen and B. matricariifolium subsp. hesperium Maxon and Clausen (Clausen,
1938). As to the former, we conclude that the western North American plant is not
closely related to B. boreale, as will be discussed in a monograph of this genus
currently in preparation. The correct name for taxon obtusilobum is B. pinnatum St.
John (Fig. 1, a—g). Taxon hesperium also proves to be a distinct species, readily
distinguished from B. matricariifolium A. Br. subsp. matricariifolium, which
occurs in North America only east of the Great Plains (Fig. 2, h-n). With our recent
opportunity to investigate large populations of these plants in the field in numerous
localities, we are now confident of the distinctness of not only B. pinnatum and B.
hesperium, but of a third element as well, which is described here for the first time.

It is no surprise to discover a new species related to B. hesperium in western
North America, where the rate of endemism among moonworts is the highest in the
world. With some ten out of 14 of the described and undescribed species known
only there, western North America is clearly the metropolis for this subgenus
(Botrychium subg. Botrychium). Using primarily the sterile lamina as a basis, we
provide the following key to its major groups.

KEY TO THE GROUPS OF BOTRYCHIUM SUBG. BOTRYCHIUM
IN NORTH AMERICA
1. Sterile lamina absent; frond composed of two sporophores........--+.+++++s00sser007 B. paradoxum group
1. Sterile lamina present; frond with a single sporophore.
2. Larger pinnae mostly fan-shaped or wedge-shaped, not pinnatifid; costa absent or poorly developed.
3. Pinnae and lobes, especially the distal ones, commonly irregularly confluent; basal pinnae often

SPONBAY GRammer ated. 22. Sin cccateage ss ste = swe ennennnenderseostengyeossienesstrees®* B. simplex group

3. Pinnae and lobes regularly separated; basal pinnae conform or only slightly exaggerated.
B. lunaria group

2. Larger pinnae mostly oblong to lanceolate, pinnatifid, costa usually present, at least in basal
i B. lanceolatum group

Early in our studies we did not notice that the original collections of B. hesperium
were mixtures of two species. Although certain specimens were different in a

we simply believed that some specimens had narrow, ere
an

*Herbarium and Department of Botany, University of Michigan, Ann Arbor, MI 48109.

54 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

shag 1. Silhouettes of moonworts similar to B. echo and B. hesperium. FIGS. a-g. B. pinnatum
: ae ae MICH). FIGS. h-n. B. matricariifolium (h, j, n, Wagner 81003, MICH; others Wagner

WAGNER & WAGNER: TWO ROCKY MOUNTAIN MOONWORTS 55

and that its apparent variability resulted from this. Our field studies, however, soon
dispelled this interpretation. The type specimen of B. hesperium includes only
individuals with rounded segments. It was taken at Glacier Lake in Rocky Mountain
National Park by a high school teacher, E. Bethel, in 1914. Bethel and others took
numerous specimens there during the period 1911 to 1921. Bethel was aware that
there were actually two elements involved, and he and Ira Clokey separated their
1921 collection at Glacier Lake into two groups—Bethel & Clokey 3987, typical,
and 3987a, with “segments narrow, more acute.” W. R. Maxon was also aware of
these distinctive plants as shown by his annotations of one of the herbarium sheets
(US984959).

Our new interpretation is based upon over 200 specimens of B. hesperium studied
in the field in seven localities, and over 300 of the new species, B. echo, in ten
localities. The two species grow in such similar habitats that one description of the
habitat will suffice for both. They tend to occur together, often side-by-side. Also
associated in genus communities with them in the southern Rockies are B.
lanceolatum, B. lunaria, and B. minganense. In Montana and Alberta, where B.
echo is not known, B. hesperium occurs also with B. paradoxum and B. pinnatum.
Most of the localities that we studied had only a few plants. To illustrate, for B.
echo, ten localities yielded respectively 1, 2, 9233, 9, al, 25, 9, and 150+
plants. The two most productive localities were in Arizona at Mount Baldy and at
San Francisco Peaks. In the former locality, B. hesperium was absent, but at the
latter we recorded 88 individuals of this species. At only one locality in the southern
Rockies did we find only B. hesperium (Wagner 81158, 3 individuals in company
with B. lanceolatum), but in the northern Rockies it apparently regularly occurs
alone.

The two species grow on grassy slopes, roadsides, and at edges of lakes. The soil
is usually rocky, the substrate including decomposed granite as well as other rock
types. In the southern Rockies the plants grow at elevations between 8,500 and
11,500 ft. The easiest way to find them is to drive along roads at proper altitudes
and to seek flat roadside ditches with gravelly soil and scattered shrubby vegetation,
Picea saplings and Salix shrubs dominating. Plants are sometimes found growing
even in the gravel of the road shoulder! In addition to spruces and willows, other
woody associates encountered are Lonicera involucrata, Potentilla fruticosa, and
species of Abies, Juniperus, and Ribes. The herbaceous associates include weeds
and involve such genera as Achillaea, Antennaria, Arenaria, Carex, Cerastium,
Epilobium, Festuca, Fragaria, Frasera, Mertensia, Penstemon, Potentilla, Saxifraga,
Sedum, Selaginella, Setaria, Solidago, Trifolium, Valeriana, and Zygadenus. Seek-
ers of these plants should be warned that only one out of ten or twenty seemingly
appropriate habitats yield these botrychiums. It is therefore necessary to sample
many likely sites. :

Rather than distinguish only B. hespertum and B. echo, all of the North American
taxa of the B. lanceolatum group that might be confused with them are keyed. For
more details of the differences between B. hesperium and B. echo themselves, the
reader is referred to the descriptions and to Figure 4. This key is based upon
medium and large individuals.

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

uw
mn

uch
FIG. 2. Silhouettes of moonworts. FIGS. a—i. B. echo (Wagner 82107b, MICH). FIG. ee wae
divided form of B. echo (Bethel & Clokey 3987a, US; see also Fig. 3, a—e). FIGS. k-o. B. hesp
(k-n, Wagner 82107a, MICH; 0, Bethel in 1914, US—type).

WAGNER & WAGNER: TWO ROCKY MOUNTAIN MOONWORTS 57

KEY TO THE BOTRYCHIUM LANCEOLATUM GROUP IN NORTH AMERICA

1. Sterile lamina broadly deltate, sessile to subsessile; sporophores of full-sized plants usually
composed of several major upright axes; plants of northern North America :.....B. lanceolatum

1. Sterile lamina mostly oblong to oblong-deltate, subsessile to stalked; sporophores of full-sized plants
usually with one major upright axis, sometimes with one or two upright laterals.

2. Sterile segment mostly conspicuously stalked, the stalk 20-30% of the blade length; segment
tips usually serrulate or crenulate; living lamina pale blue-green, dull; sporophore commonly
twice as long as sterile lamina; plants of eastern North America B. matricariifolium

2. Sterile segment short-stalked or subsessile, the stalk 5-20% of the blade length; segment tips
usually entire, repand, or pointed; living lamina color various; sporophore usually only 1.5 times
as long as sterile segment (except in B. hesperium); plants of western North America.

3. Pinnae and lobes well separated, not approximate or overlapping, mostly more or less parallel-
sided, linear to oblanceolate; pinna tips pointed; basal pinnae, except in the smallest and
largest fronds, usually deeply cleft into a single lower projection and larger upper projection;
lamina shiny green in life............--.s:sceecsseeeerersernnenenrerseeeesees ech

3. Pinnae and lobes usually approximate or overlapping, the large ones abruptly contracted at
base, oblong-lanceolate to ovate to deltate; pinna tips blunted or rounded; basal pinnae not
cleft into two projections; color and luster various.

4. Pinnae with few lobes, these mainly on the basal side; lowest pinnae exaggerated, ascending
and subclasping, strongly asymmetrical, the lower side with coarse basiscopic lobe; segments
broadly adnate at base; lamina gray-green, dull in POS Baga gr el een ORES B. hesperium

_ Pinnae with numerous lobes, these roughly equal in number on the upper and basal sides;
lowest pinnae mostly equal to or slightly larger or smaller than next distal pair, not ascending
or clasping, nearly symmetrical, the lower side with small lobes subopposite to those on
upper side; segments narrowly adnate at base; lamina bright green, shiny in life.

B. pinnatum

The following descriptions have been condensed to emphasize the most important
differences between the species.

Botrychium echo W. H. Wagner, sp. nov. Figs. 2-5.

Sporophorum segmento sterili plerumque sesquilongius; segmentum sterile vivum
supra vivide viride nitidumque, fere sessile vel brevissime stipitatum, lamina late
oblonga 2.2 (1—4.5) cm longa; pinnae bene separatae, non imbricatae, oblanceolatae,
, icl asibus subzygomorphis; pinnarum par
infimum longitudine par proximum aequans vel paulo superans, patens, non
amplectens; sporae tenelle verrucatae.

Plants exclusive of their roots 9.5 (3-15) cm tall, the common stalk 6 (2-10) cm

g ,
short-stalked, broadly oblong, 2.2 (I-4.5) cm long; pinnae narrowly attached to a
telatively narrow rachis, remote to approximate, not overlapping, lanceolate

e
aminar margins nearly entire; basal pinna pair not exaggerated in length, equal to or
somewhat longer than the adjacent pair, spreading or only moderately ascending, not
clasping; spores 37 (27-53) pm in maximum diameter, irregularly and finely
verrucate, the warts small, low, and separated by narrow, shallow channels.

TYPE: Glacier Lake, Boulder Co., Colorado, 2800 m. alt, E. Bethel & I. W.
Clokey 3937a (US; isotypes CAS, WTU).

PARATYPES:

ARIZONA: Apache Co.: White Mts., Mt. Baldy, on open bald, Wagner 82101 (MICH). Coconino
Co.: San Francisco Mt., Inner Basin, E. L. Little, Jr. 4741 (US—mixed with B. lunaria), NE slope of

58 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Fey Me

FIG. 3. Silhouettes of moonworts. FIGS. a-—e. Unusually dissected forms of B. echo resembling B.
matricariifolium (Wagner 82101, MICH—see also Fig. 1, j). FIG. f. Probable B. lanceolatum X
minganense hybrid; note the few, long segments (Wagner 82/2], MICH). FIGS. g—i. Probable B. echo
< minganense hybrids; note the relatively more numerous, shorter segments (Wagner 82104, MICH).

Mt. Doyle, Wagner 82107b (MICH). COLORADO: Boulder Co: Glacier Lake, July 1914, E. Bethel
(US), 13 July 1912, E. R. Cross (US), Bethel & Clokey 3987 (UC—2 sheets, both mixed with B.
hesperium); Arapahoe Moraine, E slope, 1 mi S of University Camp, W. A. Weber 3431 (WTU). Clear
Creek Co.: Roadside near Echo Lake, Wagner 80136b (MICH), 8/153b (MICH); Warren Mt. Picnic
Ground, 2.8 mi E of CO-515, Wagner 81158 (MICH); US-6, 0.8 mi S of I-70, Wagner 81160 (MICH).
El Paso Co.: Pikes Peak below moraine, H. L. Shantz 52 (US). Gunnison Co.: Monarch Pass, Wagner
82118 (MICH); 0.5 mi E of Monarch Pass on CO-50, Wagner 82123 (MICH). Lake Co.: Road to
Independence Pass, 3.1 mi E of Hairpin Turn, Wagner 82/27 (MICH). Summit Co.: Near Breckenridge,
K. K. Mackenzie 99 (NY—mixed with B. lanceolatum); CO-91, 2-3 mi S of 1-70, Wagner 81164
(MICH). UTAH: Summit Co.: 1.5 mi S of Spirit Lake, A. H. Holmgren et al. 7130 (UC—mixed with

B. lunaria).

The Greek specific epithet echo is used here in apposition. It was chosen to
reflect the fact that this moonwort seems to repeat the characteristics of other,
similar species. One of the best areas to study it is where we first recognized its

distinctions from B. hesperium, namely Echo Lake on the slopes of Mount Evans,
olorado

WAGNER & WAGNER: TWO ROCKY MOUNTAIN MOONWORTS 59

A 10 MM BR

FIG. 4. Comparison of B. hesperium (A) and B. echo (B) from a mixed population at San Francisco
P eaks, AZ. Tracings of cleared sterile blades. The characters are: 1=ovate, broadly attached vs.
linear-lanceolate, narrowly attached pinnae. 2= Overlapping vs. separated pinnae. 3=Repand vs.

vs. short

In certain respects, especially the shape of the pinnae and the glossy laminar
surfaces, B. echo resembles B. lanceolatum. We have not yet detected any hybrids
between the two species, but they might readily be confused with one or the other of
the parents.

A distinctive but uncommon form of B. echo with more dissected pinnae than
normal is known (Fig. /, j; Fig. 3, a-e). The basal pinnae and sometimes the medial
pinnae possess 3 or 4 lobes on the lower side and 2 or 3 lobes on the upper side.
Comparison of the outline of these with B. matricariifolium (Fig. 1, h—-n) shows
considerable resemblance. Clausen (1938), who combined B. echo and B. hesperium
as a single taxon, surely had such plants as these in mind when he wrote that some
specimens “can be matched almost exactly by material of [typical matricariifolium).”
The dissected form is especially well developed at Mount Baldy, White Mountains,
Arizona. It is connected by intermediates to the normal form.

Botrychium hesperium (Maxon & Clausen) Wagner & Lellinger,
71:92. 1981, pro hybr.

BS bes ny matricariifolium subsp. hesperium Maxon & Clausen, Me
38.

Amer. Fern J.

m. Torrey Bot. Club 19:88.

60 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

FIG. 5. Spores of B. echo (A) and B. hesperium (B). SEM photographs x 1,100. Note the differences
in size and form of the verrucae.

Plants exclusive of their roots 12 (5-20) cm tall, the common stalk 7 (3-13) cm
tall, sporophore relatively tall, 5 (3-10) cm tall, nearly twice as long as sterile
segment, 80 percent with | or more basal branches 4 or more as long as main axis of
sporangial cluster; sterile segment gray-green and dull in life, mostly short-stalked,
subdeltate, 2.5 (1-5) cm long; pinnae broadly attached to a relatively wide rachis,
crowded to commonly overlapping, ovate to lanceolate with rounded apices, the
pinna bases asymmetrical, the lamina margins finely repand; basal pinnae com-

monly exaggerated, up to twice as long as the adjacent ones, often upright and
commonly clasping; spores 37 (29-50) um in maximum diameter, irregularly and
coarsely verrucate, the warts large, prominent, and separated by wide, deep
channels.

TYPE: Glacier Lake, Boulder Co., Colorado, 8500 ft, July 1914, E. Bethel
(US—S isotypes in addition).

OTHER COLLECTIONS EXAMINED:

CANADA: Alberta: Waterton Lakes National Park, W of Red Rock Canyon Parking Area, Wagner
81103a-d (MICH).

ARIZONA: Coconino Co.: San Francisco Peaks. Inner Basin, NE slope of Doyle Mt., Wagner
82107a (MICH). COLORADO: Boulder Co.: Glacier Lake, Bethel & Clokey 3987 (CAN, CAS, US);
Rocky Mt. National Park, Loch Vail Trail, Glacier Gorge, A. E. Porsild & B. E. Willard 23102 (CAN).
Clear Creek Co.: Roadside near Echo Lake, Wagner 80136a (MICH). 8/153a (MICH). Lake Co.:
Road to Independence Pass, 3.1 mi E of Hairpin Turn, Wagner 82128 (MICH). MONTANA: Deer

ge Co.: Flint Ridge Mts., Storm Lake, S of Georgetown Lake, Wagner 80/29a (MICH), 81116b
(MICH).

It is interesting to note that Maxon’s original, unpublished interpretation of this
taxon was as given here. On the type specimen he had written the label to read
“Botrychium hesperium Maxon sp. nov.” The occurrences that we have found so far

WAGNER & WAGNER: TWO ROCKY MOUNTAIN MOONWORTS 6l

in Montana and Alberta contain plants much smaller on the average than those in
Colorado and Arizona. The northern plants resemble young stages of the southern
ones, and they may be dwarfed by the climate. The coarsely sculptured spores that
we observed in B. hesperium (Fig. 5, B) are like those of B. lanceolatum.

For B. echo, our collection records range from 13 July to 20 September, with
most specimens taken in August. Botrychium hesperium may be similar, but there is
some evidence that it may appear earlier and die down earlier. Judging from the
condition of the large sample of plants taken at San Francisco Peaks, B. hesperium
shows more effects of ageing. Seventy percent of the specimens of B. hesperium
collected on 21 August 1982 showed evidence of damage—browned margins and
broken, eaten, or otherwise tattered pinnae—compared to only 30% of B. echo
collected at the same place and time. The leaves of B. echo appeared to be fresher.

s would be expected, sterile interspecific hybrids involving both B. hesperium
and B. echo have been encountered. Their hybrid origin is deduced by their
association with parents, occurrence as usually one or a few plants, morphological
intermediacy, and abortion of spores.

Because only a single leaf is produced per year, it is difficult to obtain cytological
observations as a rule; however, Sahashi (1979) has documented irregular meiosis in
a Japanese hybrid. The most obvious interspecific hybrids are those in which the
parents differ strongly (e.g., the Scandinavian hybrid of B. boreale and B. lunaria).
Botrychium hesperium hybridizes readily with B. paradoxum to produce very
obvious intermediates, striking because one of the two sporophores is “half sterile”
(Wagner & Wagner 1981; Wagner et al., 1982). We have encountered a few definite
hybrids involving B. echo. These involve B. minganense Victorin. Some are from
Mount Baldy, Arizona, where the parents occur together locally in abundance. At
San Francisco Peaks, we found a number of sterile plants involving what appear to
be combinations of B. echo, hesperium, and lunaria, but these will require more
study to separate. Some hybrids involve yet other species, for instance, a specimen
from Monarch Pass, Colorado, we first thought was B. echo X minganense (Fig. 3
f). However, the cutting is somewhat different; also B. echo is rare and sporadic at
this locality, while both B. lanceolatum and minganense are abundant and the latter
species probably are the parents.

This is part of an investigation of the evolution and systematics of Botrychium
made possible by NSF grants DEB 800536 and DEB 8202768. We are indebted to a
number of individuals who have helped us, especially W. R. Anderson, R.
Eccleston, R. H. Hevly, J. Kuijt, J. D. Montgomery, A. Neas, W. A. Weber, M. D.
Windham, and G. Yatskievych. The following herbaria have kindly supplied speci-
mens of the species considered here: ASC, BRY, CAN, CAS, DAO, MO, NY, UC,
US, and WTU.

‘Confirmation of spore abortion is accomplished in Botrychium using only a dissecting microscope at
30-60X magnification. The spores of specimens that were dried while the sporangia still had not
completely discharged tend to be released and become attached (electrostatically?) to the sporophore
axes and opened sporangial walls. If the spores are abortive, the sizes are extremely irregular—very
small, normal, and very large. The largest spores tend to be more or less spherical and not tetrahedral.

62 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

LITERATURE CITED
CLAUSEN, R. T. 1938. A monograph of the Ophioglossaceae. Mem. Torrey Bot. Club 19:1—-177.
SAHASHI, N. 1979. Morphological and taxonomical studies on Ophioglossales in Japan and the
adjacent regions. Identity of Sceptridium Lyon in the Izu Islands. J. Jap. Bot. 54:273—281.
WAGNER, W. H., Jr. and F. S. WAGNER. 1981. New species of moonworts, Botrychium subg.
Botrychium (Ophioglossaceae), from North America. Amer. Fern J. 71:20-30.
, F. S. WAGNER, and C. HAUFLER. 1982. A hybrid population involving the “all-fertile”
Botrychium paradoxum and the hemidimorphic B. heSperium (Ophioglossaceae). (Abstract).
Bot. Soc. Amer. Misc. Publ. 162:77-—78.

SHORTER NOTES

LYCOPODIUM COMPLANATUM AND L. ANNOTINUM FOUND IN THE
BLACK HILLS.—Lycopodium complanatum L. and L. annotinum L. were found
growing together in the Black Hills of Wyoming on July 27, 1982. The location is in
Crook County, Upper Sand Creek at confluence with Spottedtail Gulch, T51N
R60W Section line of 20-21, elevation 5600 feet. The plants were growing under
White Spruce, Picea glauca (Moench) Voss, and Hazelnut, Corylus cornuta
Marsh. along with Low Red Huckleberry, Vaccinium scoparium Leiberg. Lycopo-
dium complanatum was found only at this location, but L. annotinum extended on
down the canyon at several more localities. Specimens are deposited at the Univer-
sity of Wyoming (RM) and the New York Botanical Garden (NY) (Dorn 3793,
3792). The closest known locality for L. complanatum is about 450 miles to the
northwest in Lewis and Clark County, Montana. The closest known locality for L.
annotinum is about 270 miles to the west in Park County, Wyoming.

Several other pteridophytes were collected within two miles of the Lycopodium
location along the same creek: Equisetum sylvaticum L. (Dorn 3777, collected for
the first time in Wyoming one day earlier by E. F. Evert in the Big Horn
Mountains), Athyrium filix-femina (L.) Roth (Dorn 3 778), and Equisetum scirpoides
Michx. (Dorn 3809, another first record for Wyoming).—Robert D. Dorn, Box
1471, Cheyenne, WY 82003.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 2 (1983) 63

MICROFIBRILS IN THE XYLEM OF BLECHNUM VIVIPARUM. —
Cellulose forms the framework of plant cell walls by a system of microfibrils (as
well as other molecular shapes) in which various substances are incrusted. The
presence of microfibrils can be demonstrated by chemical removal of the incrusting
substances or, rarely, by their actual appearance in the lumen of pits, particularly in
relation to tori and plasmodesmata of some angiosperms and conifers. After the
formation of secondary walls in some Gnetaceae and Coniferae (Warchop et al.,
Holzforsch. 13:115-120. 1959), a sort of “tertiary growth” appears in the shape of
warts or of fibrils, either isolated, at random, or in a margo (a marginal reticule that
fuses at or near the center of the torus), and sometimes obliterates the lumen
(Butterfield & Meyland, Three-dimensional Structure of Wood, p. 72. 1980).

FIG. 1. Isolated, sporadic xylem elements in Blechnum viviparum interpreted as secondary vessels. Two
pits showing a loose net of microfibrils, = 10,000.

In the material of Blechnum viviparum (Broadh.) C. Chr. studied and reported by
Montiel and Guevara (Rev. Biol. Trop. 27:171-176. 1979) and reinterpreted by
Gomez (Brenesia 18:253—258. 1980), some of the pits show what in my opinion are
microfibrils (Fig. 1) which, to my knowledge, have either been overlooked or have
not yet been reported from vascular cryptogams where xylem elements or secondary
growth have been detected. It must be pointed out that the microfibrils reported here
seem somewhat thicker than those usually found in higher plants and, unlike those,
ours are sporadic and quite scattered throughout the material.

The presence of microfibrils in ferns is interesting from a phylogenetic point of
view inasmuch as they represent another intermediate character between the tracheid
and the coniferophyte-type element. Furthermore, the presence of loose microfibrils
Partially obliterating the lumina between crassulae, projecting from the middle
lamellae to the secondary walls in a pteridophyte, seems to support the idea of a
coherent, organized constitution of cellulose in the primary membranes as proposed
by Frey-Wyssling et al. (Planta 47:1 15-126. 1956) and Miihlethaler (Biochem.
Biophys. Acta 5:1-9. 1950), at least in these primitive plants.—Luis D. Gomez P.,
Museo Nacional de Costa Rica, Apartado 749, San José, Costa Rica.

64 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

BISPORANGIATE ANOMALOUS SPOROPHYLLS IN ISOETES FROM
RAJASTHAN—The presence of a solitary, adaxial sporangium on or in association
with a sporophyll is the fundamental feature of all lycopods. Only two exceptions
have been reported. The fossil Lycostachys protostelicus was described by Pant and
Walton (Palaeontographica 108:1—10. 1961) as having two subarchesporial pads in a
sporangium; and in /soétes indica, Pant and Srivastava (Proc. Natl. Inst. Sci., India,
B, Biol. 23:242-280. 1962) described sporophylls having two, almost equally
developed sporangia. The discovery of bisporangiate sporophylls in two additional
species of Isoétes, I. coromandelina L. f. collected at Poasa near Jaipur, Rajasthan,
and in an apparently new species of /soétes from Mt. Abu, Rajasthan, India, are
additions to this list.

FIG. 1. Bisporangiate, abnormal sporophylls of /soétes coromandelina, X 1.2. FIG. 2. Longisection
of bisporangiate sporophyll of /soétes sp. from Mt. Abu showing two unequal sporangia, X 20.

A number of abnormal sporophylls were discovered in J. coromandelina which
possess double sporangia (Fig. 1). These seem to be produced by fusion between
adjacent sporophylls during their development. The proximal portions of the
sporophylls are united and bear two sporangia; the distal, acicular portions of the
sporophylls are free. The ligules and upper labia are also double. However, in other
details of morphology and anatomy the sporophylls are normal.

In the Isoétes from Mt. Abu, an abnormal sporophyll bearing two sporangia was
discovered during sectioning of the material. The two sporangia are unequal and are
Separated by a partitioning wall (Fig. 2). The smaller sporangium is empty and
seems to be a bud-like, sterile structure; the bigger one is normal and contains
numerous megaspores.

From the present discovery, it seems that double sporangia are not so rare in
Isoétes. Further investigations are needed to establish any possible phylogenetic
conclusions.—B. D. Sharma, R. Singh, and D. R. Bohra, Department of Botany,
University of Jodhpur, Jodhpur 342001, India.

——— ee

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AMERICAN ae
FERN ee mber, 19
JOURNAL

QUARTERLY JOURNAL OF THE AMERICAN FERN SOCIETY

Observations on the Structure and Function

of Hydathodes in Blechnum lehmannii JOHN S. SPERRY 65
A Reclassification of the Fern Genus Pyrrosia K. H. SHING 73
Notes on the Ecology and Development

of Plagiogyria fialhoi PAULO G. WINDISCH and MARILIA PEREIRA-NORONHA 79
The Ferns of Elden Mountain, Arizona MICHAEL D. WINDHAM 85
Donovan S. Correll (1908-1983) 94
Reviews 93-96

RSSOURT BOTANICAL
OcT 6 1983

GARDEN LIBRARY

The American Fern Society

American Fern Journal
EDITOR

DAVID B. LELLINGER U.S. Nat’l Herbarium NHB-166, Smithsonian Institution,
Washington, DC 20560.

ASSOCIATE EDITORS
DAVID W. BIERHORST Rt. 3, Box 188, Picayune, MS 39466.
GERALD J. GASTONY Dept. of Biology, Indiana University, Bloomington, IN 47401.
JOHN T. MICKEL New York Botanical Garden, Bronx, NY 10458.
TERRY R. WEBSTER ....... Biological Sciences Group, University of Connecticut, Storrs, CT 06268.

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AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 3 (1983) 65

Observations on the Structure and Function of Hydathodes
in Blechnum lehmannii
JOHN S. SPERRY*

The fronds of many ferns in the Polypodiaceae (Ogura, 1972) and Cyatheaceae
(Weiler, cited in Lippmann, 1925) possess swollen vein endings associated with
specialized adaxial epidermal cells. Their structure is similar in all ferns (Gardiner,
1883: Potonié, 1892; Poirault, 1893; Goebel, 1930; Guttenburg, 1934), including
Blechnum lehmannii Hieron. (Figs. 1-3). The depressed epidermis (Fig. 3) sug-
gested the term “Wassergriibchen” for the vein endings to European anatomists of
the nineteenth century (e.g., Potonié, 1892). Vein endings on developing fronds
secrete water (Fig. /) when conditions reduce or stop transpiration (Mettenius,
1856; Haberlandt, 1894a; Goebel, 1926, 1930). In some species, the secreted water
contains salts in solution which form “chalk scales” after the water has evaporated
(Mettenius, 1856; Potonié, 1892; Lippmann, 1925).

Most sources report that water secretion is dependent on the metabolic activity of
the specialized epidermal cells adaxial to the vein ending (Fig. 3, e); it is thus
regarded as an active, or glandular, process (Gardiner, 1883: Haberlandt, 1894a,
1914; Lepeshkin, 1906; Stocking, 1956; Fahn, 1979). This conclusion is reflected in
various references to the vein endings as water glands, salt glands, or chalk glands.
There is, however, very little evidence to support any conclusions on the mechanism
of secretion. All work on the subject was performed in the last century, and the
results obtained are questionable due to the bias in experimental design which
prevented testing of alternative hypotheses and to the possibility of artifacts in
technique (see comments in Spanjer, 1898; Haberlandt, 1898; Lepeshkin, 1923).
Moreover, the results are contradictory, since they have been interpreted as support-
ing either an active (Gardiner, 1883; Haberlandt, 1894a, 1914) or a passive (Spanjer,
1898) mechanism of secretion. Finally, all experiments were performed on a single
species, Polypodium aureum L., and need not represent the situation in other ferns.

Because the mechanism of secretion from vein terminations is unknown, they will
be referred to in this paper by the term “hydathode” which, as originally defined by
Haberlandt (1894b), did not presuppose a mechanism of secretion. Haberlandt used
his term to refer to structures on above-ground portions of the plant, particularly
foliage leaves, which function in water transfer to and from the plant surface.

Previous work on the mechanism of hydathode secretion in ferns neglected
important aspects of the problem. Despite the fact that root pressure could play a
Significant role in the secretion process, the relationship of root pressure to secretion
in intact plants was not studied. In fact, there is no well documented account of root
Pressure in ferns. In addition, investigators could not analyze secretion from a
structural standpoint because anatomical studies were confined to the mature,
non-secreting hydathodes; the structure of the secreting hydathodes on developing
leaves was unknown. In this study, the relationship of root pressure to secretion as
well as developmental changes in hydathode structure were investigated in B.

oe Forest, Petersham, MA 01366.
olume 73, number 2, of the JOURNAL was issued 16 June 1983.

66 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

FIG. 1. Hydathodes secreting water on a developing frond; arrow indicates a droplet of water above a
hydathode. FIG. 2. Epidermal peel of hydathode photographed with dark field illumination. There 1s no
intercellular space in the epidermal layer; the separation of cells marked by the arrow is an artifact of
preparation due to the weak adherence between epidermal cells. FIG. 3. Transverse section of a
hydathode on a mature frond made parallel to the leaf margin and treated with I-KI/H,SO,. e = epidermal
cells of hydathode, en =endodermis; arrows indicate Casparian strip.

lehmannii in its native locality, the Central American cloud forest. Observations
Suggest that one mechanism of secretion from fern hydathodes is the passive
transmission of xylem sap from vein endings to the plant surface through the
apoplast of the hydathode under a pressure gradient induced by root pressure.

MATERIALS AND METHODS

Plants growing in the cloud forests near Xalapa in the state of Veracruz, Mexico,
provided material for both experimental and anatomical work. Individual plants
consist of a single, erect rhizome with short internodes. ;

Root pressure was measured with a bubble manometer attached to the cut stipe
(Fig. 4A) and a thermometer. The manometer was made with surgical tubing (ca.

J. S, SPERRY: OBSERVATIONS ON BLECHNUM HYDATHODES 67

= :

= S

® 2 |

= 0b

2 3

5 |

ep)

Yt |

C5, | hos ee oikece rater, CEL ae

05 10 2.0

B time (hrs.)

FIG. 4A. Manometer attachment for measuring root pressure. B = clear plastic bag enclosing the entire
crown of an individual plant, S=cut stipe to which the manometer is attached, C=clamp, Y = forked
connector, BB = bubble at sealed end of the manometer tube. FIG. 4B. Root pressure vs. time for plants
1, 2, 3. S=onset of hydathode secretion.

0.8 mm inside diameter) partially filled with acid fuchsin and sealed at one end so
as to leave a bubble at the sealed end (Fig. 44, BB). Bubble length was assumed to
be proportional to bubble volume. At the beginning of the experiment, bubble
length at ambient pressure and air temperature was determined by releasing the
clamp (Fig. 4A, C). After the clamp was closed, subsequent readings of bubble
length and air temperature were made, and exudation pressure relative to ambient
was calculated according to Gay-Lussac’s law (Weast & Astle, 1981). Clamping
itself did not affect bubble length. Care was taken when attaching the manometer so
that a continuous column of liquid was present from stipe to bubble, and the
attachment was gently secured with twist-ties to help guard against leakage.

The relationship between root pressure and hydathode secretion was studied in
three plants growing along a shaded stream in the cloud forest on a clear day. The
entire above-ground portion of each individual plant was enclosed in a clear plastic
bag and root pressure was monitored with a manometer attached to a single cut
frond for each plant (Fig. 4A). Observations of xylem pressure and hydathode
secretion were then made.

Anatomical differences between the secreting and non-secreting hydathodes on

alcoholic sodium hydroxide followed by lactic acid. Observations were made on
Preparations left unstained or stained in toluidine blue. The distribution of lignin
was determined from hand sections stained in phloroglucinol and concentrated HCI.
Suberized and cutinized cell walls were identified using the I-KI/H2SO,j test.

68 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

FIGS. 5-13. Development of Casparian strip. All preparations were treated in I-KI/H,SO,. FIGS. 5-8,
10, and 12 are from cleared preparations; FIGS. 9, 11, and 13 are transverse sections made parallel t0
the leaf margin. Arrows = Casparian strip. FIGS. 5~9. Non-secreting hydathodes on mature frond.
FIG. 5. Vein subtending a hydathode and surrounded by endodermis with fully developed Casparian
strip. FIG. 6. Hydathode viewed from the adaxial epidermis and focussed on the Casparian stip
bordering the edge of the vein ending. FIG. 7. Higher magnification of the Casparian strip as viewed 9

J. S. SPERRY: OBSERVATIONS ON BLECHNUM HYDATHODES 69

OBSERVATIONS

Secretion.—Secretion was observed on developing, unenclosed fronds in foggy
and rainy weather in the cloud forest and when individual crowns were enclosed in
plastic bags. In bagged plants, secretion was initiated on successive pinnae in an
acropetal direction so that the last hydathodes to begin secreting on developing
fronds were those on the most distal pinnae. Based on qualitative observations,
distal hydathodes appeared to have a greater rate of secretion than more proximal
hydathodes, which on developing fronds sometimes showed no secretion. Observa-
tions of fronds at different stages of development as judged by relative size, color,
and texture suggested that the permanent loss of secreting capability by hydathodes
occurs acropetally. No salt deposits of any kind were observed to accumulate above
the hydathodes

oot pressure.—Root pressure was determined by examining the bleeding
behavior of cut stipes (Fig. 4A, S) on plants growing in conditions which were
assumed to severely reduce or stop transpiration. Such conditions were either
natural, as on rainy or foggy days, or induced by enclosing the entire above-ground
portion of an individual plant in a clear plastic bag (Fig. 4A, B). Root pressure was
assumed to be present when the attached stipes showed continual bleeding after
being cut. Initially, some of this bleeding was due to mucilage flow as judged by the
viscous nature of the exudate. However, after continual blotting over a period o
several minutes, the viscosity of the bleeding fluid decreased considerably, and with
the aid of a hand lens, xylem sap could be seen issuing from the stele. Plants in
which transpiration was not inhibited did not show this bleeding behavior from cut
stipes; here the only exudation noted was a small amount of mucilaginous material.

Although the secretion of mucilage from the cut end of the stipe was a potential
problem for root pressure measurements, the results indicate that the pressure
produced by the secretion is insignificant. Certain plants (e.g., plant 1, Fig. 4B) did
not show root pressure even when transpiration was reduced by enclosure of the
crown in a plastic bag. The absence of root pressure in these plants was apparently
due to insufficient soil moisture. Despite the fact that the stipes of these plants
showed mucilage secretion when cut, no significant increase in pressure above
ambient was noted from attached manometers.

The relationship between root pressure and hydathode secretion studied in the
three bagged plants (labelled 1, 2, 3) is summarized in the graph of Figure 4B. In
plants 2 and 3, the buildup of root pressure in the xylem correlated positively with
the observation of water secretion from hydathodes of developing fronds. In bot
plants, the measured pressures were sufficient to support a column of water taller
than the height of the plant (1.8 m for plant 2, 1.2 m for plant 3). Plant | developed

directly on the tracheids of the vein ending. FIG. 8. Casparian strip adaxial to the vein ending in surface

Probably phenolic vacuoles.

70 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

no root pressure, and no secretion was observed from its hydathodes.

Variation in root pressure in the three plants is correlated with position relative to
the stream flowing under the bank where they were growing. Plant 2, which showed
the most dramatic rise in pressure, was growing roughly 0.3 m above the stream
level. Plant 1, which showed no root pressure, was 2.5 m above the stream. Plant 3
was intermediate in pressure and bank position. Possibly the explanation for this
correlation is decreasing soil moisture with increasing height above the stream;
greater soil moisture would favor the generation of higher root pressures.

Hydathode development.—The ability of hydathodes to secrete is correlated with
specific features of their development. Hydathode cells reach their mature size and
are capable of secreting water long before the cells of the surrounding leaf tissue
have finished expanding. Hydathodes capable of secreting protrude above the
surrounding epidermal cells (Fig. 13). Only after the leaf cells reach their full size
and the leaf achieves its mature thickness do hydathodes become the depressed
“Wassergriibchen” consistently described by early anatomists (Figs. 3 and 9). In the
recessed stage, hydathodes were observed to be incapable of water secretion.

The structural change that appears to be most closely related to secretory function
is the progressive suberization of the hydathode endodermis. The Casparian strip
surrounding the hydathode (Figs. 3 and 9) is the distal extremity of a network which
surrounds the entire mature vascular system of the plant. The development of this
continuous Casparian strip is acropetal, and lags considerably behind the maturation
of protoxylem and hydathode cells (Figs. /0, 12, and 13). Thus, hydathodes in
developing tissues do not possess a suberized endodermis.

The acropetal extension of the Casparian strip to the hydathode is first evident in
the endodermis abaxial to the vein ending (Fig. //, arrows). The vein subtending the
hydathode is completely surrounded by a Casparian strip (Fig. 5). At this time, the
Casparian strip around the vein ending is like a mitten with the abaxially turned
palm cut out. Development of the Casparian strip proceeds with the first steps
represented by thin bands of suberin extending from the well developed strip below
the vein. Eventually, the abaxial endodermis of the vein ending possesses 4
Casparian strip as well developed as anywhere else in the frond (Fig. 9).

The degree of Casparian strip development at the hydathode correlates closely
with the capability of the hydathode to secrete water. Examination of developing
fronds from plants 2 and 3, which possessed distal secreting hydathodes and
proximal non-secreting hydathodes, indicated that secretion only occurred from
hydathodes where the Casparian strip was either absent or incompletely developed.
In one frond, for example, distal secreting hydathodes which from qualitative
observation showed the highest secretion rate possessed no Casparian strip (Figs. 10
and /3). Hydathodes on pinnae from the middle part of the frond, which appeared to
have a lower rate of secretion, possessed an incompletely developed Casparian strip
adaxial to the vein ending (Fig. //). Hydathodes at the base of the frond showed no
secretion and were found to be completely surrounded by a thick Casparian strip
(Figs. 6-9). In addition, examination of fronds at different stages of Casparian strip
development indicated that the acropetal progression of complete hydathode suber-
ization correlates with the acropetal loss of secretion capability in the hydathodes.

:
{

J. S. SPERRY: OBSERVATIONS ON BLECHNUM HYDATHODES 71

DISCUSSION

The correlation of root pressure with secretion from hydathodes on the one hand,
and the correlation between Casparian strip development and the cessation of
hydathode secretion on the other, suggest a simple explanation for the occurrence of
water secretion from hydathodes in B. lehmannii. Root pressure is directly responsi-
ble for secretion by supplying a pressure gradient sufficient to drive the passive
movement of xylem sap from the vein ending of the hydathode, through the apoplast
of the sub-epidermal and epidermal cells, and through the thin cuticle to the outside
of the plant. When the Casparian strip is completely formed around the vein ending
of the hydathode, the apoplast is sealed and secretion from the hydathode is blocked.
Preliminary results from Polypodium and Nephrolepis species indicate that this
hypothesis can also apply to hydathodes in these genera.

Root pressure is known to be directly responsible for secretion from the epidermal-
pore hydathodes of angiosperms (Fahn, 1979). Although pores are not present in the
epidermis of fern hydathodes, root pressure can move water to the plant surface
through the anticlinal walls of the epidermal cells. According to Meidner (1977),
root pressure causes the secretion of water from the epidermal cell walls of
Gladiolus leaves which lack epidermal-pore hydathodes, and the same explanation
may account for Goebel’s observation (1930) of secretion from the blade margins of
ferns without hydathodes.

The results presented in this paper do not preclude the possibility that water
secretion is partially or wholly active, and more research is required to decide the
question. Hydathode epidermal cells could play a role in the modification of the
solute content of the secreted xylem sap. Thus, they would be analogous to
glandular cells found associated with epidermal-pore hydathodes of the angiosperms
(Dieffenbach, et al., 1980). If the epidermal cells are capable of secreting salts,
there may also be a metabolically dependent osmotic mechanism which contributes
to water secretion, at least for those fern species that characteristically secrete large
amounts of salts and form “scales” (see Lepeshkin, 1906). Regardless of the specific
mechanism of secretion, however, the correlation of root pressure with hydathode
secretion suggests the functional significance of the fern hydathode is to be sought in
its relationship with root pressure.

This project was initiated while I was enrolled in the Harvard course Biology 247,
which included a field trip to Mexico. I would like to thank the instructors, Rolla
and Alice Tryon, for organizing the trip, which was financially supported by the
Atkins Fund of Harvard University. I also thank the Tryons for advice during early
Phases of the project and P. B. Tomlinson and M. H. Zimmerman for their critical
review of the manuscript. Frank Lang provided the photograph in Figure 1; Figure 2
was photographed with the assistance of George Wilder.

LITERATURE CITED

DIEFFENBACH, H., U. LUTTGE, and M. G. PITMAN. 1980. Release of guttation fluid from passive
hydathodes of intact barley plants. Il. The effects of abscisic acid and cytokinins. Ann. Bot.
—712

FAHN, A. 1979. Secretory Tissues in Plants. Academic Press, London.

12 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

GARDINER, W. 1883. On the physiological significance of water glands and nectaries. Proc. Camb.
Phil. Soc. 5:35-50.
GOEBEL, K. 1926. Morphologische und biologische Stunden. IX. Beitrage zur Kenntnis der
Verwandschaftsverhaltnisse einiger Javanischer Farne. Ann. Jard. Bot. Buitenz. 26:107-160.
________. 1930. Archegoniatenstudien. XX. Farne mit punktierten Blattern. Flora 124:410-422.
GUTTENBERG, H. 1934. Studien an Pflanzen der Sunda-Inseln. Ann. Jard. Bot. Buitenz. 34:1-62.
HABERLANDT, G. 1894a. Ueber Bau und Function der Hydathoden. Ber. Deutsch. Bot. Ges.
12:367-378.
_ 1894b. Anatomisch-physiologische Untersuchungen iiber das tropische Laubblatt. II. Uber
wassersecernirende und -absorbirende Organe. Sitzungsber. Akad. Wiss. Wien., Math-Nat.
KI. 103:489-583.
_ 1898. Bemerkungen zur Abhandlung von Otto Spanjer “Untersuchungen tiber die
Wasserapparate der Gefisspflanzen.” Bot. Zeit. 61:177-181.
—_——.. 1914. Physiological Plant Anatomy. (Translated from the 4th German edition by Montague
Drummond.) Macmillan, London.
LEPESHKIN, W. W. 1906. Zur Kenntniss des Mechanismus der aktiven Wasserausscheidung der
Pflanzen. Beih. Bot. Zentralbl. 19:409—452.
_ 1923. Uber active und passive Wasserdrusen und Wasserspalten. Ber. Deutsch. Bot. Ges.

41:298-300.

LIPPMANN, E. 1925. Ueber das Vorkommen der verschiedenen Arten der Guttation und einige
physiologische und 6kologische Beziehungen. Bot. Arch. 11:361—464.

MEIDNER, H. 1977. Sap exudation via the epidermis of leaves. J. Exp. Bot. 28:1408-1416.

METTENIUS, G. H. 1856. Filices Horti Botanici Lipsiensis. Leopold Voss, Leipzig.

OGURA, Y. 1972. Comparative Anatomy of Vegetative Organs of the Pteridophytes. Gebriider Born-
traeger, Berlin.

POIRAULT, G. M. 1893. Recherches anatomiques sur les cryptogames vasculaires. Ann. Sci. Nat.
Bot., VII, 8:113-256.

POTONIE, H. 1892. Uber die den Wasserspalten physiologisch entsprechenden Organe bei fossilen und
recenten Farnarten. Sitzungsber. Ges. Naturf. Freunde Berlin 1892:1 17-124.

SPANJER, O. 1898. Untersuchungen iiber die Wasserapparate der Gefasspflanzen. Bot. Zeit. 61:6-81.

STOCKING, R. C. 1956. Guttation and bleeding. Pp. 489-502 in: W. Ruhland, ed. Encyclopedia of
Plant Physiology, vol. 3. Springer-Verlag, Berlin.

WEAST, R. C. and M. J. ASTLE, eds. 1981. CRC Handbook of Chemistry and Physics. CRC Press,
Inc., Boca Raton, Florida.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 3 (1983) 73

A Reclassification of the Fern Genus Pyrrosia'
K. H. SHING*

The name Pyrrosia, although published by Mirbel in 1803, was overlooked by
pteridologists until relatively recently. Instead, the synonyms Cyclophorus Desv.,
1811, or Niphobolus Kaulf., 1824, were used. Giesenhagen (1901) published the
only monograph, treating 50 species in Niphobolus and describing their venation,
epidermis, and indument. Farwell (1931, pp. 241-246) recognized that Pyrrosia is
the correct name of the genus. R. C. Ching (1935) studied the species of the Asian
mainland, Japan, and Taiwan Province of China. He transferred 49 known species to
Pyrrosia and described five new species. This has been the only extensive system-
atic paper on the genus. Nayar and others (1961, 1965, 1967) did detailed work on
the morphology and anatomy of the Indian species of Pyrrosia in which they divided
the 13 Indian species into six groups, some of which are apparently unnatural and
inconvenient to use. For example, they placed P. lingua (Thunb.) Farw., which has
stellate hairs with only lanceolate arms in the P. heteractis group, although each
stellate hair of P. heteractis (Mett. ex Kuhn) Ching has 1-3 longer, acicular arms in
addition to the lanceolate arms. Van Alderwerelt van Rosenburg (1908, pp. 678-696)
divided 34 Malesian species into two sections under the generic name Cyclophorus.
Section Niphobolus had venation similar to that of Campyloneurum (more or less
regular, subquadrangular areolae with two or more simple or forked, included,

or less irregularly netted with single, free, included, variously oriented veinlets).
This arrangement has something to recommend it, but since the blade of Pyrrosia is
very thick and in most species the veins cannot be seen without clearing, venation is
difficult to use for identifications.

In China, Pyrrosia is an herb of traditional use. Since the laminae are shaped like
a dagger with a leathery texture and usually trail on rocks, it was called “Rocky
Leather” or “Flying Dagger” in old Chinese herbals. It is a diuretic which can be
used to clean the lungs and to alleviate fever. Now in the pharmacy there are two
kinds of medicine on the market, “Shiwei Tablet” and “Instant Shiwei Powder”
(Shiwei in Chinese means Pyrrosia), made from Pyrrosia plants and said to have
curative effects for nephritis and chronic tracheitis. Since special care is unnecessary
for the survival of these plants, except for good drainage, they may be cultivated in

hed i iji hina, but during the writer's visit to the United States, he received
a grant from the “cpap tee e Foundation (020138) and through the kindness of the curators had ae
city to examine more specimens in the ipllowing herbaria: GH, MICH, NY, and US. es woul
like to extend his thanks to Prof. R. C. Ching and Prof. W- H. Wagner, Jr. for their kind guidance, to
Mr. M. G. Price for his helpful discussions, and to Dr. A. F. Tryon for her helpful discussion of spore
characters and for providing SEM photograph for Figure 2.

74 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

PE). FIG. 7. Spore of P. polydactilis, x 1200 (Taiwan, Tanaka 140, PE). FIG. 8. Spore of P.
borneensis, 1100 (Borneo, H. F. Sun 198, PE). FIG. 9. Lower epidermis of P. clavata. FIG. 10.
Upper epidermis of P. clavata. FIG. 11. Lower epidermis of P. angustata.

K. H. SHING: RECLASSIFICATION OF PYRROSIA 75

FIG. 12. Types of stellate hairs in Pyrrosia. a= broad arm type, b=acicular arm type, c, d= dimorphic

arm types, e=crispate-lanose arm type.

a number of Pyrrosia species have been imported into the United States from the
East Indies, the Philippines, Japan, and New Guinea (Hoshizaki, 1981).

Pyrrosia includes about 100 species, mostly in Asia, with a few in Australia,
New Zealand, Oceania, and Africa. Fifty species occur in China. Because of their
more or less uniform appearance, identification may be quite difficult. During the
past several years, the writer has observed their spore morphology, epidermises,
stomata, and indument, and has tried to establish their taxonomic relationships. The

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16 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

present paper will discuss these characters and will propose a new classification
scheme for the genus.
MORPHOLOGICAL CHARACTERISTICS

Spores.—Almost all species of the genus have a uniform, tubercular perine,
which varies only slightly in size and thickness (Figs. 2—8). However, spore
ornamentation of P. angustata (Swartz) Ching is sharply different from most other
members of the genus, and appears ribbed under the SEM (Fig. /). Since the fronds
are dimorphic and firmly coriaceous, the sori very large, round, deeply sunk, and
uniseriate on each side of the costa, J. Smith (1857, p. 6) considered this species to
be a separate genus, Niphopsis. But in my opinion, considering all important
features, it is really a member of Pyrrosia. Another similar species, P. samarensis
(Mett. ex Presl) Ching, has the same spore ornamentation. I propose to use the rank
of subgenus to indicate their advanced position in Pyrrosia.

Rhizome scales.—The rhizome scales of Pyrrosia are either entire or are ciliolate
with the hairs spread along the sides or sometimes with a tuft of hairs at the apex.
Since the scales are easily broken and their tips lost on most specimens, it 1s
difficult to use this feature for identification. However, since the scales of different
species may also be very different in form, size, and texture, they still are useful for
distinguishing some species.

Epidermis and stomata.—The arrangement of the upper epidermal cells in the
genus is uniform (Fig. /0). There are, however, mainly two types of stomata in the
lower epidermis. Most are pericytic, with the stomata completely surrounded by a
single subsidiary cell with guard cells and subsidiary cell not linked together by any
anticlinal walls (Figs. 9 and //). Some are polycytic, having the stomata in large
part surrounded by a single, U-shaped subsidiary cell with the anticlinal cell walls of
the guard cells and the subsidiary cell linked together toward the distal end. A few
are desmocytic or copericytic (Sen & Hennipman, 1981).

Indument.—There are four kinds of stellate hairs in different groups of Pyrrosia:
the broad arm type has 6-9 lanceolate arms (Fig. /2, a), the acicular arm type has
8-12 acicular arms (Fig. 12, b), the dimorphic arm type has 6—9 lanceolate arms
and I-3 acicular arms (Figs. 12, c and d), and the crispate-lanose arm type has the
arms crisped and intertwined like cotton velvet, mostly on the bottom layer of the
dense, thick indument (Fig. /2, e). Hair types are a constant and reliable character
for subdividing the genus.

CONSPECTUS OF THE GENUS PYRROSIA

Pyrrosia subg. Pyrrosia

TYPE: Acrostichum lingua Thunb. [= Pyrrosia lingua (Thunb.) Farw.]. 4!
Pyrrosia sect. Pyrrosia

YPE: Acrostichum lingua Thunb. | =Pyrrosia lingua (Thunb.) Farw.].

Pyrrosia ser. Pyrrosia

bes Acrostichum lingua Thunb. [= Pyrrosia lingua (Thunb.) Farw.].
: is is the largest series in the genus, with nearly 40 species, and is widespread 10
the Asian tropics and subtropics. Two species occur in Australia and one in Africa.

K. H. SHING: RECLASSIFICATION OF PYRROSIA 77
204 \3

Pyrrosia ser. Heteractides Ching & Shing, ser. nov.
Pilis stellatis e brachiis lanceolatis necnon setis paucioribus (1-3) aciculatibus
longioribusque compositis.
TYPE: Polypodium heteractis Mett. ex Kuhn [=Pyrrosia heteractis (Mett. ex
Kuhn) Ching].
This series includes only P. heteractis, which is found in southwestern China,
Vietnam, and the Himalayan area, and P. eberhardtii (Christ) Ching, distributed in
r southern China and Vietnam.
0° Pyrrosia ser. Drakeanae Ching & Shing, ser. nov.
Pilis stellatis e brachiis 8-12 aciculatis compositis.
TYPE: Polypodium drakeanum Franch. [ = Pyrrosia drakeana (Franch. ) Ching].
This series includes about 20 species from eastern Asia to southeastern Asia, with
one species in Africa.
,oo\4 Pyrrosia sect. Dichlamys Ching & Shing, sect. nov.
Frons subtus indumento bistrato e pilis stellatis stipitatis dimorphis laxe composito
praedita, superne e pilis aciformibus, et inferne e pilis lanosis crispatis compositis.
TYPE: Niphobolus mollis Kunze [= Pyrrosia mollis (Kunze) Ching].
90° Pyrrosia ser. Costatae Ching & Shing, ser. nov.
Pilis strati superi e brachiis lanceolatis, inferi e filamentis lanosis crispatisque
compositis.
TYPE: Apalophlebia costata Pres| [= Pyrrosia costata (Presl) Tag. & Iwats.].
This series includes, besides the type species, P. nummularifolia (Swartz) Ching
and P. strigosa (Swartz) Ching; the species are limited to the Himalayas and
southeastern Asia.
1o\-© Pyrrosia ser. Molles Ching & Shing, ser. nov.
Pilis strati superi e brachiis longe aciculatis compositis, inferi crispatis lanosisque.
TYPE: Niphobolus mollis Kunze | = Pyrrosia mollis (Kunze) Ching].
This series contains more than 20 species, with one in Africa.
+o Pyrrosia subg. Niphopsis (J. Smith) Shing, comb. & stat. nov.

Niphopsis J. Smith, Cat. Cult. Ferns 6. 1857. TYPE: Polypodium angustatum Swartz [= Pyrrosia
angustata (Swartz) Ching].

Rhizome thick, long-creeping, the scales caudate-lanceolate, grayish brown to-
ward the apex and broken off in age, dark brown toward the persistent base, leaves
fully dimorphic, the sterile blades shorter, lanceolate or elliptic-lanceolate, the
fertile blades longer, linear-lanceolate or often narrowed to a linear apex; sori large,
round or oblong, in a single series along the costae; spores bilateral, with many
linear ridges, contracted at each end to an abrupt beak.

This subgenus includes two species distributed in southeastern Asia, Singapore,
Malaya, Indonesia, and the Philippines to New Guinea and New Zealand. Morpho-
logically this subgenus is advanced in the genus. Its perispore is similar to that of
some species of Dryopteris. Pyrrosia samarensis (Mett. ex Presl) Ching has the
same spore ornamentation as P. angustata, but its sterile leaf is narrower and its sor!
are confluent at maturity. Its laminae bear many, dark brown, stellate hairs with
aciculate arms covering the hairs of the basal layer of indument, which are of the
crispate-lanose type. This species is endemic to the Philippines.

78 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

NEW COMBINATIONS IN PYRROSIA
According to my studies, the following species require names in Pyrrosia:

Pyrrosia dispar (Christ) Shing, comb. nov.

Cyclophorus dispar Christ, Nova Guinea 8:155. 1909.—New Guinea.
Pyrrosia intermedia (Goy) Shing, comb. nov.

Cyclophorus intermedia Goy, Queensl. Nat. 10:48, t. 6. 1937.—Queensland.

Pyrrosia macrocarpa (Copel.) Shing, comb. nov.

pee gion macrocarpa Copel. Univ. Calif. Publ. Bot. 12:381. 1931.—Pacific Islands.

Thi cies is somewhat similar to P. angustata in outline, and Christensen (Ind.
Fil. mht 3:65. 1934) reduced it to a variety. It differs from P. angustata in that
the plants are smaller, the spore ornamentation is closely tubercular (Fig. 2), and the
sori are never impressed. It is found only in several islands of the Pacific, such as
the Cook Islands, Pitcairn Island, and the Austral Islands.

Pyrrosia distichocarpa (Mett.) Shing, comb. nov.

Polypodium distichocarpum Mett. Ann. Lugd. Bat. 2:231. 1866.—Sumatra.

Cyclophorus winckleri Rosenst. Repert. Sp. Nov. Fedde 7:149. 1909.—Sumatra.

Pyrrosia winckleri (Rosenst.) Tagawa, Acta Phytotax Geobot. 25:180. 1973.

Pyrrosia winckleri and P. distichocarpa are here united for the first time. I have
seen several specimens of the latter species, but no type material.

Pyrrosia borneensis (Copel.) Shing, comb. nov.
Cyclophorus borneensis Copel. Phil. J. Sci. 12C:64. 1917.—Borneo.

Pyrrosia rasamalae (Racib.) Shing, comb. nov.
Polypodium rasamalae Racib. Pterid. Buit. 99. 1899.—Java.

LITERATURE CITED

ALDERWERELT van ROSENBURGH, C. R. W. K. van 1908. Malayan Ferns. Dept. Agr. Netherl.
Indies, Batavia.

CHING, R. C. 1935. On the genus Pyrrosia Mirbel from the mainland of Asia including Japan and
Formosa. Bull. Chin. Bot. Soc. 1:1-72.

FARWELL, O. A. 1931. Fern notes II. Amer. Midl. Nat. 12:233- oe

GIESENHAGEN, K. 1901. Die Farngattung Niphobolus. Fischer, Jen

HOSHIZAKI, B. J. 1981. The genus Pyrrosia in cultivation Pobjnediseaet Baileya 21:53-76.

NAYAR, B. K. 1961. Studies in Polypodiaceae. VII. Pyrrosia Mirbel. J. Indian Bot. Soc. 40: 164-183.

————, and S. CHANDRA. 1965. Ferns of India~XV, Pyrrosia Mirbel. Bull. Natl. Bot. Gard.
Lucknow 117:1-98.

———, and S. CHANDRA. 1967. Morphological series within the genus Pyrrosia, and their phyloge-
netic interpretation. Canad. J. Bot. 45:615—634.

SEN, U. and E. HENNIPMAN. 1981. Structure and ontogeny of stomata in Polypodiaceae. Blumea

SMITH, J. 1857. Cultivated Ferns. Wm. Pamplin, London.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 3 (1983) 79

Notes on the Ecology and Development of Plagiogyria fialhoi
PAULO G. WINDISCH* and MARILIA PEREIRA-NORONHA**

Plagiogyria fialhoi (Fée & Glaziou) Copel. is a shade-loving fern that can be
found in cloud forests of elevated regions in the Brazilian states of Rio de Janeiro,
Espirito Santo, Minas Gerais, S40 Paulo, Santa Catarina, Rio Grande do Sul, and
probably also Parana (Brade, 1956; Sehnem, 1967). Some authors (e.g., Tryon &
Tryon, 1982) consider Plagiogyria in America to be a single species, P. semicordata
(Presl) Christ, but in this study the separation into six species by Lellinger (1971)
was adopted.

Some aspects of the ecology and development of the gametophytes and sporo-
phytes of a population of P. fialhoi occurring in a cloud forest in the Serra da
Mantiqueira in southeastern Brazil (45°25'S, 22°40’W, ca. 2,000 m alt.) were
studied. The locality is on a small ridge next to the Campos do Jordao State Park, in
the state of Sado Paulo. Cloud forests occur mainly on the eastern slopes of the
mountains, above 1,600 and up to 2,000 m alt., and are composed of low (ca. 6 m)
trees covered by a great quantity of epiphytes. Frequent drizzle, fog, and low clouds
keep the humidity extremely high. The rainfall at the locality is thought to be
about 2,000 mm per year.

Climatological data collected at considerably lower altitudes in the nearby state
park (Seibert et al., 1975) indicate July as the driest month (ca. 30 mm rainfall) and
January as the wettest (more than 300 mm). The warmest month is February (17.7°C
average, absolute maximum 27.2°C); the coldest is July (9.5°C average, absolute
minimum —4.4°C). A short description of the floristic composition of these cloud
forests is presented by Seibert et al. (1975), together with the detailed climatic data
for the region.

MATERIALS AND METHODS

Gametophytes and ca. 60 young sporophytes with fronds up to 3 cm long were
collected and preserved in weak chromo-acetic fixative (Sass, 1951); 36 larger
specimens were pressed (voucher specimens at HRCB). Frond longevity was studied
in 15 plants by marking their youngest fronds and following their development during
visits 240 and 374 days later (February and June, 1982). The complete underground
parts (rhizome, roots, and stipe bases) of ten well developed specimens were
carefully removed and the shape and position of the rhizome correlated with the
topography.

The average number of fronds per adult plant was calculated from data from the
ten removed specimens (April, 1981) and from the 15 marked plants (June, 1981).
Average size of the fronds was obtained based on those of the 10 removed specimens
and confirmed later (March, 1983) by measuring 128 adult fronds. The distance
between the elements of 55 pairs (nearest neighbors) of plants was measured.
*Instituto de Biociéncias, Letras e Ciéncias Exatas, Universidade Estadual Paulista UNESP, Caixa
Postal 136, 15100 S. José do Rio Preto - SP, Brazil.

**Curso de Pés-graduacao em Ciéncias Bioldgicas, Universi
Postal 178, 13500 Rio Claro - SP, Brazil.

dade Estadual Paulista—UNESP, Caixa

80 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

FIGS. 1-14. Gametophytes and heteroblastic development of the sporophyte of Plagiogyria fialhoi.
Figs. 1-4. Gametophytes attached to young sporophytes. FIGS. 5-12. Sporophytes in diverse stages of
development. FIG. 13. Lamina of a frond 15.5 cm long. FIG. 14. Pinna from the middle of a frond 40
cm long. y

a > Se —

i oe ar i

ee ee ee ee

WINDISCH & PEREIRA-NORONHA: NOTES ON PLAGIOGYRIA FIALHOI 81

Humus and soil samples were taken from ten points (March, 1983), keeping the
top material (O—5 cm depth) separated from the humus-soil mixture below (S—10 cm
depth). Spores were collected (June, 1981 and February, 1982) and germinated in
the laboratory after storage at 3-5°C for three and 120 days, using a solid agar
medium (technique described by Dyer, 1979). Gametophyte cultures were kept for
100 days growing under 850 lux (fluorescent plus incandescent bulbs, 12 hours/day)
at ca. 24°C.

RESULTS AND DISCUSSION

Plants of P. fialhoi were found in great numbers on the steep, humid slopes
(30-50% declivity) of the southeastern side of the ridge, which rises ca. 70 m above
a small stream. The density of plants increases closer to the water. The distance
(nearest neighbor) between the elements of 21 pairs of plants growing more than ca.
10 m from the stream was 45-520 (ave. 189.4) cm, whereas the distance between
the elements of 34 pairs growing within 10 m of the stream was 18-300 (ave. 95.9)
cm. The plants grew in the rich humus layer that covers the ground. Humus samples
from the top layer (5 cm) had a pH of 3.2 (in CaCly); 48g/cm? P (resin method)
and K, Ca, Mg, H+Al values (m. eq. 100 cm’) of 0.5, 0.8, 0.6, and 24.4. Samples
from the humus and soil mixture (19.6% organic matter) from the layer below (5-10
cm) had a pH of 3.1; 43 pg/cm? P; and K, Ca, Mg, H+Al values of 0.29, 0.2, 0.3,
and 46.0, respectively.

In a few places on the slope, associations of gametophytes and juvenile sporo-
phytes of Plagiogyria in diverse stages of heteroblastic development, together with
fern gametophytes and young sporophytes of other species, mosses, liverworts, and
angiosperm seedlings were found. In contrast with the surrounding area, where the
adult fern sporophytes occurred, the amount of organic matter on the ground at those
spots was very small, indicating that these associations were of a pioneer nature,
probably covering soil exposed by running water or fallen trees.

In nature, the first frond of the young sporophytes (still attached to the gameto-
phytes) measured 5.5—8.0 (ave. 7.0) mm long, with stipes 3.0-5.2 (ave. 3.9) mm
long and laminae 1.3—4.3 (ave. 3.1) mm long. These laminae were symmetric and
bilobed or asymmetric with one of the segments once more lobed (Figs. 1-5). The
margin of the laminae in plants of all developmental stages presented a characteristic
row of more or less rectangular cells (Fig. 15), which was of basic importance In the
identification of juvenile specimens. Fronds ca. 14-35 mm long had a central
vascular strand and tended to form pinnatifid laminae (Figs. 6-7). Pinnatisect to
pinnate laminae were observed on plants with fronds longer than 35 mm (Figs.
8-13). These fronds (up to 80 mm long) had segments with serrate margins and
simple veins. The lower pinnae of fronds ca. 85-150 mm long had biserrate margins
(Fig. 13) and bifurcate proximal veins. Free veins that reach the lamina margins
Was a consistent character in all stages of development. The sterile laminae of adult
sporophytes had biserrate margins (Fig. /4), and most of the veins were bifurcate, so
that a veinlet ended in each tooth. In well developed specimens (with rhizomes
longer than 6 cm) the fronds were 43-117 (ave. 75-68) cm long, the stipe
representing about 1/3 of the total length.

82 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

FIGS. 15-23. Detail of the lamina and rhizome of the sporophyte and earl ages of gametophyte
Placi

arly st
development of Plagiogyria fialhoi. FIG. 15. Laminar margin of a young sporophyte. FIG. 16. pee
curved rhizome (old stipe bases removed). FIGS. 17-23. Early developmental stages of gametophy
in the laboratory.

WINDISCH & PEREIRA-NORONHA: NOTES ON PLAGIOGYRIA FIALHOI 83

Fertile fronds were observed from February to June. It seems that they are formed
around January and probably persist until July. Viable spores were collected in June,
1981. A visit in February, 1982 revealed many young fronds, but the spores
collected from the most developed of these did not germinate. These data indicate a
definite periodicity of fertility. Fertile fronds measured 70-117.5 (ave. 103.0) cm
long (the stipe representing about 1/2 of the total length) and were usually longer
than the sterile fronds of a given plant. The minimum ratio observed for numbers of
sterile to fertile fronds observed was 10:4, the maximum 21:1.

The rhizome, covered with the bases of fallen fronds and adventitious roots, was
usually erect if no longer than 4 cm, but curved in larger specimens so that the
vegetative parts usually were level with or barely above the litter on the ground. The
older parts remained buried. The apex of each rhizome bore the fronds in a rosette
protecting frond primordia and young fronds. Litter carried down the slope accumu-
lated against the frond-covered rhizome apices. The resulting pressure on the uphill
side and perhaps a more intense washing of the ground on the other side, together
with apical rhizome growth, promoted a slow and gradual movement of the whole
plant downhill. The rhizome and its semi-decayed remains are curved, with the main
axis oriented down the slope. In a large specimen, the angle of deviation between the
alignment of the rhizome apex and that of the oldest remains was ca. 155° (Fig. 16),
indicating the change of relative position of the old parts in the ground due to the
downhill movement.

The one year-long observation of marked fronds revealed that fronds persisted for
about one year. The number of fronds on adult plants (of different ages) was 9-21
(ave. 12). Therefore, the minimum age of a plant can be estimated by dividing the
total number of stipe bases by 12. On very old specimens, the semi-decayed parts
still present are of large size, indicating that at the time those were formed the plant
was already well developed. One specimen with a rhizome 21 cm long had a
minimum age of 24 years. The youngest fertile specimen collected had a rhizome 12
cm long; its age was estimated to be more than 15 years, indicating a minimum
Spore-to-spore life cycle of considerable duration. Vegetative reproduction was not
observed, although Bower (1926) cited the occurrence of stolons in P. pycnophylla
(Kunze) Mett. and bifurcation of the rhizome in P. semicordata.

Spores sown on culture media three days after collection did not germinate up to
30 days after sowing, but after 50 days, germinating spores and gametophytes with
3, 4, 9 and 12 cells were seen. Slow germination occurring at irregular intervals
was also observed in cultures of P. glauca (Blume) Mett. and P. semicordata by
Stokey and Atkinson (1956). This kind of gradative germination can explain the
finding in nature of gametophytes together with young sporophytes in diverse stages
of development in a single small area. Spores stored for 120 days showed an
extremely low germination rate; a single spore in about 2,000 germinated after 30
days, reconfirming the short viability of the spores of this genus as discussed by
Stokey and Atkinson (1956) and Nayar and Kazmi (1962).

The first stages of gametophyte development (Figs. 17-20) are very similar to
those described by Nayar and Kazmi (1962) for P. triquetra Mett. No apical
meristematic cell was observed. The development of a heart-shaped apex (Figs.

84 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

21-23) started earlier (65 days after sowing) than in the Old World species studied
by Nayar and Kazmi (1962). Elongated young plate stages and filamentous stages as
cited for P. glauca and P. semicordata by Stokey and Atkinson (1956) were not
observed; perhaps they were a consequence of the physical conditions under which
Stokey and Atkinson’s material was grown. They considered the probability of
competition with algae as being the cause of these elongated plate stages, but our
cultures were also heavily contaminated. Unfortunately, the algae were so numerous
that further development of the gametophytes could not be followed in the laboratory.
In nature, we found that mature gametophytes were cordiform (up to 4.4 mm wide,
5.3 mm long at the larger wing), generally with one of the wings overlapping the
other (Figs. 1-4), and had a thick central region.

is study was made possible by the cooperation of the officers of the Forestry
Institute of the State of Sao Paulo, Campos do Jordao State Park and by support
rom the Conselho Nacional de Desenvolvimento Cientifico e Tecnolégico—CNPq
(Proc. 30.1339/77 and Proc. 10.6254/79), the Universidade Estadual Paulista Julio
de Mesquita Filho—UNESP, and the Fundagao de Amparo a Pesquisa do Estado de
Sao Paulo—FAPESP (Proc. 81/1369-5).

LITERATURE CITED

BOWER, F. O. 1926. The Ferns (Filicales), vol. Il. Cambridge Univ. Press, Cambridge.

BRADE, A. C. 1956. A flora do Parque Nacional do Itatiaia. Bol. Parque Nac. Itatiaia 5. Ministério da
Agricultura, Rio de Janeiro.

DYER, A. F. 1979. The culture of fern gametophytes for experimental investigation. /n Dyer, A. F, ed.
The Experimental Biology of Ferns. Academic Press, London.

are D. B. 1971. The American species of Plagiogyria sect. Carinatae. Amer. Fern. J.

7110-118.

NAYAR, B. K. and F. KAZMI. 1962. Morphology of the spores and prothalli of five species of
Plagiogyria. Bull. Soc. Bot. Bengal 16:3-8.

SASS, J. E. 1951. Botanical Microtechnique. Iowa State Coll. Press, Ames.

SEHNEM, A. 1967. Plagiogiridceas. Jn R. Reitz, ed. Flora Ilustrada Catarinense, part I. Itajat.

SEIBERT, P. et al. 1975. Plano de manejo do Parque Estadual de Campos do Jordao. Bol. Tec. Inst.
Flor. Sao Paulo. 19:1-153.

STOKEY, A. G. and L. R. ATKINSON. 1956. The gametophytes of Plagiogyria glauca (BI.) Mett. and
P. semicordata (Pr.) Christ. Phytomorphology 6:239-249.

TRYON, R. M., Jr. and A. F. TRYON. 1982. Ferns and Allied Plants. Springer Verlag, New York.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 3 (1983) 85

The Ferns of Elden Mountain, Arizona
MICHAEL D. WINDHAM*

Elden Mountain, located immediately northeast of Flagstaff in north-central
Arizona, is an unusual volcanic peak with a diverse fern flora. The earliest
pteridophyte collections from the area are those taken by L. N. Goodding in 1913.
Whiting and Bradey collected extensively on the mountain during the 1930’s and
were apparently the first to encounter Asplenium adiantum-nigrum in 1935. In a short
article discussing the occurrence of this species in Arizona, Wherry (1941) men-
tioned five other ferns casually observed during a visit to the locality. Phillips (1946,
1947) listed five additional taxa for the mountain, bringing the number of reported
species to 11. The flora of Elden Mountain received little attention during subse-
quent years, and much of the area remained unexplored when a forest fire inflicted
heavy damage in June, 1976. Concerned about the fate of several rare species, I
began a comprehensive survey of surviving fern populations in 1978.

Elden Mountain is a massive dacite dome located near the center of the San
Francisco volcanic field. Although precise dates are not yet available, geologists
agree that the dome was formed sometime during the last million years (Kluth &
Kluth, 1974). The dacitic magma was quite viscous, and well-defined lava flows are
evident on all slopes. Juxtaposition of lava flows created a highly complex topogra-
phy, and erosion has produced innumerable cracks and crevices that provide
favorable habitats for a variety of unusual plants. The elevation at the base of the
mountain averages 7000 feet; the summit falls just short of 9300 feet.

A lengthy meteorological record is available for nearby Flagstaff at 6900 feet
elevation (Sellers & Hill, 1974); much of the general climatic data is applicable to
Elden Mountain. At Flagstaff, the lowest average monthly temperature (21,3: ¥)
occurs in January, the highest (65.5° F) in July. The mean frost-free period extends
from June 8 to September 26, a total of 110 days. Annual precipitation averages
18.31 inches, and seasonal rainfall patterns are distinctly bimodal. The greatest
amount of rainfall occurs in July and August, when storm systems enter the region
from the Gulf of Mexico. Pacific-based storms produce a second precipitation peak
in January and February, but the months of May, June, and November are character-
ized by drought, which was a major factor in the disastrous fire of 1976.

The distinct vegetation zones observed on the San Francisco Peaks (Merriam,
1890) are all but absent on Elden Mountain. Abrupt topography brings diverse
habitats into close proximity, and plant associations
complex. The areas surrounding the base of the mountain are i ’
ponderosa, which is frequently associated with Quercus gambelii. South-facing
s, Berberis fremontit, Juniperus
deppeana, J. osteosperma, Pinus edulis, Opuntia phaeacantha, and Yucca baccata.
The northern slope and crest of the mountain support a mixed conifer forest in
which Pseudotsuga menziesii, Abies concolor, and Pinus flexilis dominate. Small
meadows and stands of Populus tremuloides occupy 4 limited area near the summit.
Some areas show a haphazard mixture of all the species mentioned above.

*Department of Biological Sciences, Northern Arizona University. Flagstaff, AZ 86011.

86 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

METHODS AND MATERIALS

The data presented herein result from intensive fieldwork undertaken between
February, 1978 and March, 1983. Considering the size and complexity of Elden
Mountain, it would be unwise to suggest that the pteridological survey is complete.
However, most suitable habitats have been examined, and recent visits to the area
have failed to yield additional, unreported species. Detailed field notes were kept
concerning the distribution and habitat preference of each taxon, and population
estimates were derived through a census of individual adult plants. Ten mature
sporangia were gathered from each species, and their contents were examined in
glycerol to determine spore number. Most cytological materials were field fixed in
May and June using Farmer’s solution (3 parts ethanol: | part glacial acetic acid).
Plants not demonstrating the proper meiotic condition were transferred to a green-
house to stimulate the production of fertile fronds. Fixed sporangia were hydrolyzed
for approximately 15 minutes in IN HCI prior to staining with aceto-orcein. Hoyer’s
solution was used as the squashing and mounting medium in all chromosome
preparations. Counts were derived from cells at late diplotene or diakinesis and were
documented using a camera lucida. Voucher specimens for all phases of this study
have been deposited at the Deaver Herbarium, Northern Arizona University (ASC).

RESULTS AND DISCUSSION

All fern species previously reported for Elden Mountain were relocated during
this survey, proving that none was lost as a direct result of the 1976 fire. In fact, the
number of recognized species was nearly doubled by the discovery of two unreported
herbarium collections and seven taxa new to the mountain. None of the additions
were found in fire-damaged areas, and it is apparent that they have existed on the
mountain for quite some time. Nearly all are characterized by low population
densities, which may explain why they were overlooked by previous collectors. The
survey has revealed that the Elden Mountain fern flora comprises 20 species
representing 11 genera. Chromosome counts have been obtained for 15, and spore
count data are available for all but one.

Adiantum pedatum L.—Discovered in 1979, the colony consists of ca. 25 plants
growing on the western slope at 8100 ft elevation. The plants are confined to a small
area of permanent seepage on the back wall of a narrow cliff recess. The
length/width ratio of the laminae is unusually high compared to other Arizona
collections, but elongate fronds are occasionally observed throughout the range of
the species. Mature sporangia consistently produced 64 normally developed spores.

Asplenium adiantum-nigrum L.—The Elden Mountain population comprises at
least 100 individuals scattered over the southern and eastern slopes at 7100-8100 ft
elevation. The plants favor shaded cracks, crevices, and ledges on relatively dry.
south-facing cliffs. All sampled sporangia yielded 64 normal spores, and a chromo-
some count of n=72 II (Fig. 1A) agrees with previous determinations from Europe
and Colorado (Shivas, 1969) Colorado specimens were originally described as
Asplenium andrewsii A. Nels., and Chihuahuan collections provided the type
materials of A. chihuahuense J. G. Baker and A. dubiosum Davenp. (Knobloch &
Correll, 1962). ‘The proliferation of names in the New World resulted from the

M. D. WINDHAM: FERNS OF ELDEN MOUNTAIN 87

sae hes
eid Bie
LY & Pak ] ‘ a,
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ite Woy
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AN B C
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= + oo +, x0
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ty ot eH LS hehe an , Cy
+, '° 2
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oF sf ~ hee,
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ay * > gy" 4 *
me eal a = ig i ie aay % 44 4
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- oe is = |
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a | » = % <A i
4 ~ opty tn we x *
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FIG. 1. Camera lucida drawings of meiotic chromosomes. A. Asplenium adiantum-nigrum, Windham

351. B. A. platyneuron, Windham 394. C. A. resiliens, Windham 400. D. A. septentrionale, Windham
393. E. A. trichomanes, Windham 397. F. Cheilanthes fendleri, Windham 26. G. C. wootonii,
Windham & Yatskievych 266. H. Cystopteris cf. tennesseensis, Windham & Windham 319. I.
Dryopteris filix-mas, Windham 395. J. Pellaea truncata, Windham 149. K. P. wrightiana, Windham
396. L. P olypodium hesperium, Windham 392. M. Polystichum scopulinum, Windham 398. N. Woodsia
mexicana, Windham 402. O. W. oregana, Windham & Windham 253.

88 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

failure of early investigators to recognize the Eurasian affinities of these highly
disjunct populations. Shivas (1969) stated that meiotic pairing in A. onopteris X A.
adiantum-nigrum was identical whether the latter parent came from Europe or
Colorado. A recent analysis of polyphenolic constituents (Richardson & Lorenz-
Liburnau, 1982) provides further evidence that A. andrewsii (including all collec-
tions from Elden Mountain) is conspecific with A. adiantum-nigrum.

Asplenium platyneuron (L.) B.S.P.—The Elden Mountain colony is located ca.
600 km WSW of the westernmost record in Colfax County, New Mexico. My wife
and I first encountered the plants in February, 1978, but there is some indication
that the species was originally discovered in the 1940's. In a letter to William
Maxon dated July 15, 1947 (on file at ARIZ), Dr. Phillips mentioned that an
associate in California had collected an unusual Asplenium in northern Arizona. The
precise collection locality was unknown, but Phillips tentatively identified the
specimen as A. platyneuron. In a reply dated July 21, Maxon requested a frond from
the plant in order to verify its identity. However, Maxon passed away shortly
thereafter, and the species was never reported for Arizona. The local population
comprises at least 150 plants on the eastern, southern, and western slopes at
7000-7600 ft elevation. For the most part, the plants are restricted to deep cracks
and crevices with semi-permanent shade and an ample supply of moisture. Individu-
als occupying more exposed habitats tend to develop coriaceous pinnae with
prominent marginal serrations. The colony appears to be actively reproducing, and
all sampled sporangia contained 64 normal spores. Local plants are morphologically
similar to those of the eastern United States, and a chromosome count of n= 36 Il
pk 1B) agrees with all previous determinations (Love, Léve & Pichi Sermolli,

Asplenium resiliens Kunze—Discovered in 1978, this species is scattered over
the southern and western slopes of the mountain at 7000-7800 ft elevation. Its
preferred habitat is nearly identical to that of A. adiantum-nigrum, but the two
species were never observed growing in close proximity. A sample of ten sporangia
yielded nine with 32 normal spores and one with 64 malformed, presumably aborted
spores. Meiotic squashes revealed the presence of two types of cells with different
chromosome complements. Sporangia following an apogamous pathway contained
eight large spore mother cells that showed n= 108 II. This mechanism of sporogene-
Sis, resulting in 32 viable spores, is dominant judging from the spore count data.
However, occasional sporangia follow the normal sexual pathway, and they contain
16 smaller spore mother cells at the beginning of meiosis. Manton (1950) suggested
that the latter cells provide evidence of genome homologies in hybrid taxa reproduc-
ing by apogamy. In the Elden Mountain collections, all cells of this type showed
n= 108 I (Fig. 1C). These data confirm previous observations that the species is an
apogamous triploid and suggest that the putative diploid and tetraploid parents were
not closely related.

Asplenium septentrionale (L.) Hoffm.—Widely distributed on all slopes of the
mountain at 7100-8700 ft elevation. The plants occupy a variety of habitats, but
most are found in shaded cracks on exposed boulders and cliff faces. They appear to
be morphologically indistinguishable from European specimens, and all sampled

M. D. WINDHAM: FERNS OF ELDEN MOUNTAIN 89

sporangia contained 64 normal spores. A chromosome count of n=72 Il (Fig. 1D)
agrees with previous determinations in Eurasia (Love, Love & Pichi Sermolli, 1977)
and the disjunct population in Monroe County, West Virginia (Emory, 1970).

Asplenium trichomanes L. subsp. trichomanes.—Found in deep cracks and
crevices on all slopes of Elden Mountain at 7000-8300 ft elevation. Plants are
closely associated with A. platyneuron in several places, but there is no evidence of
local hybridization. Mature sporangia consistently produced 64 normal spores with a
mean diameter of 28 zm. Moran (1982) assigned Arizona collections to the diploid
cytotype on the basis of spore dimensions, and a chromosome count of n=36 Il
(Fig. 1E) substantiates his prediction.

Cheilanthes eatonii Baker—Discovered in 1978, this fern inhabits exposed
ledges and cliffs on the southern and western slopes at 7000-8100 ft elevation.
Considering its abundance on the south side of the mountain, it is surprising that the
species was overlooked by previous collectors. Local materials have traditionally
been assigned to forma eatonii, which exhibits a dense tomentum on the upper
surface of the frond. All sampled sporangia yielded 32 large, well formed spores,
suggesting that the plants are apogamous.

Cheilanthes feei Moore—Common on all slopes of the mountain, this species
favors relatively dry, exposed cliffs at 7000-8500 ft elevation. The presence of 32
spores in each sporangium suggests that local plants are apogamous triploids similar
to those studied by Knobloch (1967).

Cheilanthes fendleri Hook.—Discovered in 1978, the Elden Mountain popula-
tion comprises at least 50 plants on the eastern, southern, and western slopes at
7200-8100 ft elevation. Most plants inhabit well shaded ledges on the eastern side
of the mountain. All sampled sporangia contained 64 normal spores, and a chromo-
some count of n= 30 II (Fig. /F) is apparently the first report for the species.

Cheilanthes wootonii Maxon—Favoring shaded ledges on south-facing cliffs,
this species is scattered over the eastern, southern, and western slopes at 7100-7700
ft elevation. Mature sporangia consistently produced 32 large, well formed spores.
Meiotic squashes revealed that both apogamous and sexual pathways operate during
sporogenesis, although the latter was rarely observed. Sporangia following the
apogamous pathway contained eight large spore mother cells that showed n=90 Il
(Fig. 1G). Sporangia utilizing the sexual pathway contained 16 smaller spore mother
cells at the beginning of meiosis. It was not possible to obtain an exact chromosome
count on cells of this type, but several preparations suggest that n=ca. 30 II + 301.
Most univalents do not converge on the plate during metaphase, and the resultant
unequal distribution of chromosomes leads to Spore malformation. These data
indicate that local plants of C. wootonii are apogamous triploids of hybrid origin. A
Texas plant studied by Knobloch (1967) showed 2n= 116, suggesting that this
polymorphic species includes at least two cytotypes-

Cystopteris cf. tennesseensis Shaver—Not previously reported for Arizona,
although a number of pteridologists have been aware of its occurrence In the state
(W. H. Wagner, pers. comm., 1982). The Elden Mountain colony is located ca. 700
km NW of the westernmost station in the Guadalupe Mountains of Texas (Blasdell,
1963). The species was originally collected by L. N. Goodding in 1913 and

90 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

identified as C. fragilis. Goodding’s collection was overlooked by Phillips (1947)
and omitted from his checklist of Arizona ferns. The population consists of ca.
plants growing on the western slope at 8000-8100 ft elevation. The plants are
confined to seepage zones in deep crevices and cliff recesses; several grow in close
proximity to the Adiantum pedatum colony. A sample of 20 sporangia was removed
from two different plants, and 19 of them contained 64 normal spores. The
remaining sporangium yielded 32 exceptionally large, well formed spores. Further
study will be necessary to determine the viability and ploidy level of these
unreduced products of sporogenesis. Chromosome counts of n= 84 II (Fig. 1H)
were obtained from sporangia with the normal complement of 16 spore mother cells.
The presence of minute bulblets on some plants suggests that the species is an
allotetraploid whose parentage includes C. bulbifera. The plants resemble typical C.
tennesseensis in many respects, but the two taxa are probably not identical.

Dryopteris filix-mas (L.) Schott— This conspicuous fern is frequently collected
and occurs on all slopes at 7100-8600 ft elevation. Its preferred habitat consists of
shaded cracks and crevices with an ample supply of moisture. All sampled
sporangia contained 64 normal spores, and a chromosome count of n= 82 II (Fig.
11) agrees with an earlier report from western North America (Reisender, 1974). The
morphology of local specimens is highly variable, and fronds range from early
deciduous to evergreen.

Pellaea truncata Goodding—Discovered in 1979, the Elden Mountain colony
consists of ca. 10 plants growing on the eastern and southern slopes at 7400-7700 ft
elevation. The species is rarely collected on elevated sections of the Colorado
Plateau, and plants are confined to dry, south-facing cliffs and ledges. A sample of
ten sporangia yielded nine with 64 spores and one with 32 exceptionally large spores
that may be unreduced products of sporogenesis. A chromosome count of n=29 I
(Fig. 1J) agrees with previous reports (as P. longimucronata) by Knobloch and
Britton (1963). Most fronds tend to be tripinnate at the base, but local specimens are
otherwise similar to their low elevation counterparts.

Pellaea wrightiana Hook.—Common on the eastern, southern, and western
slopes of the mountain at 7000-8100 ft elevation. The habitat requirements of this
species are nearly identical to those of Cheilanthes eatonii, and the two often grow
together on the low cliffs of the southern slope. A sample of ten sporangia yielded
nine with 64 spores and one with 32 exceptionally large spores. This situation is not
unusual in P. wrightiana, and all Arizona collections thus far examined have
produced similar ratios. A chromosome count of n=58 II (Fig. 1K) provides further
support for Wagner’s (1965) assertion that this taxon is typically a fertile tetraploid.
Sterile triploid hybrids between this and the preceding species have been found on
the southern slope at 7100 ft elevation.

Pityrogramma triangularis (Kaulf.) Maxon—Discovered in 1979, the Elden
Mountain colony provides the first record for Coconino County and indicates that
the species has a much greater elevation range than was previously suspected. The
colony consists of three small plants growing from a permanently shaded crack on
the southern slope at 7125 ft elevation. Mature sporangia consistently produced 64

M. D. WINDHAM: FERNS OF ELDEN MOUNTAIN 9]

normally-developed spores. Atypical morphology and a paucity of material preclude
identification at the varietal level.

Polypodium hesperium Maxon—Found in shaded cracks and crevices on all
slopes of the mountain at 7600-8700 ft elevation. All sample sporangia yielded 64
normal spores, and a chromosome count of n=74 II (Fig. /L) agrees with previous
Arizona reports by Knobloch (1962) and Lloyd (1963). Local plants have acrid,
pruinose rhizomes that distinguish them from typical populations in the Pacific
Northwest.

Polystichum scopulinum (D. C. Eaton) Maxon—Arizona lies at the southern
distributional limit of this species, and most populations are small and sporadically
distributed. The Elden Mountain colony is the largest in the state, including at least
250 individuals on the western, southern, and eastern slopes at 7100-8200 ft
elevation. The plants favor shaded cracks and crevices on south-facing cliffs. All
sampled sporangia contained 64 normal spores, and a chromosome count of n= 82
Il (Fig. 1M) agrees with previous determinations by Wagner (1973).

Pteridium aquilinum (L.) Kuhn var. pubescens Underw.—This widespread
taxon is rarely observed on Elden Mountain and was not collected until 1979. It
favors rocky soils in open coniferous forests, occurring on all slopes at 7000-7700 ft
elevation. Chromosome and spore counts are not available because the population
showed no evidence of sexual reproduction from 1979 to 1983.

Woodsia mexicana Fée—Common on the eastern, southern, and western slopes
of the mountain at 7000-7600 ft elevation. The plants grow on partially shaded
cliffs and ledges and are often associated with Pellaea wrightiana. All sampled
sporangia yielded 64 normal spores, and chromosome counts of n=76 Il (Fig. IN)
were obtained from five different plants. These counts conflict with Knobloch and
Correll’s (1962) report of n= 82 and substantiate Brown’s (1964) hypothesis that the
species is a tetraploid based on x= 38. Specimens from northern Arizona and New
Mexico are somewhat atypical, showing substantial reductions in the length of both
indusial filaments and marginal processes. ;

Woodsia oregana D. C. Eaton—All Elden Mountain collections of this species
have previously been referred to W. plummerae (Phillips, 1947). The specimens are
unusually glandular and their indusial segments tend to be united at the base.
However, they bear little resemblance to typical W. plummerae of southern Arizona,
and preliminary flavonoid data indicate that the two are biochemically distinct.
Scanning electron microscopy has also revealed clear and consistent differences in
spore ornamentation. Seven plants from Elden Mountain were examined cytological-
ly, all of which yielded counts of n= 76 Il (Fig. 10). Similar materials gathered
from four other localities in northern Arizona also proved to be tetraploid. Overall
morphology suggests a close relationship to W. oregana, but typical collections of
that species are reportedly diploid (Brown, 1964). Additional data will be necessary
to resolve the apparent discrepancy between morphology and cytology. The species
is sporadically distributed on all slopes at 7000-8700 ft elevation. The plants are
usually found in shaded cracks and crevices, and all sampled sporangia contained 64
normal spores. Sterile tetraploid hybrids between this and the preceding species have
been found on the southern slopes of Elden Mountain at 7000 ft elevation.

9? AMERICAN FERN JOURNAL: VOLUME 73 (1983)

The fern flora of Elden Mountain includes ca. 20% of the species known to grow
in Arizona. Several are relatively common in the state, but Asplenium adiantum-
nigrum and A. platyneuron are apparently confined to this locality. The number of
xerophytic ferns is unusually high considering the elevation, and 40% of the taxa are
rarely found north or west of Elden Mountain. Included in this category are
Asplenium resiliens, Cheilanthes eatonii, C. fendleri, C. wootonii, Pellaea
wrightiana and Woodsia mexicana. Four species (20%) are known or suspected
apomicts, showing triploid chromosome numbers and 32 spores per sporangium.
Local Pteridium colonies are strongly rhizomatous and they, too, form the bulk of
their populations without sexual processes. Approximately 69% of the sexual
species are tetraploids; the remainder are diploids. Several of the tetraploids are
nothospecies of hybrid origin, including Asplenium adiantum-nigrum, Cystopteris
cf. tennesseensis, Pellaea wrightiana, and Polypodium hesperium. In each case, one
or both of the putative parents are absent from Elden Mountain and there is no
evidence that the species were formed locally. However, interspecific hybridization is
moderately common on the mountain, and sterile hybrids have been detected in both
Pellaea and Woodsia.

I would like to thank the following for their help with various aspects of the study:
Clark Schaack, Dr. Warren Wagner, Jr., Dr. Florence Wagner, George Yatskievych,
Dr. Richard Hevly, Dr. Timothy Reeves, and Theresa Windham. Drs. James
Rominger, Darrel English, and Gerald Gastony read the manuscript and provided
helpful comments. Special thanks to Dr. Dean Blinn for providing access to the
phase microscope used in this study.

LITERATURE CITED
BLASDELL, R. F. 1963. A monographic study of the fern genus Cystopteris. Mem. Torrey Bot. Club.

BROWN, D. F. M. 1964. A monographic study of the fern genus Woodsia. Beih. Nova Hedwigia
16:1-154.

EMORY, D. L. 1970. A major North American range extension for the forked spleenwort, Asplenium
septentrionale. Amer. Fern J. 60:129-134.

KLUTH, C. F. and M. J. KLUTH. 1974. Geology of the Elden Mountain area, Arizona. In T. N. VY:
Karlstrom, G. A. Swann, and R. L. Eastwood (eds.). Geology of Northern Arizona, Part Il:
Area Studies and Field Guides. Northern Arizona Univ., Flagstaff, AZ.

KNOBLOCH, I. W. 1962. Tetraploid Polypodium vulgare var. columbianum from Arizona. Amer. Fern
J. 52:65-68.
. 1967. Chromosome numbers in Cheilanthes, Notholaena, Llavea and Polypodium. Amer. J.
Bot. 54:461—464.
. and D. M. BRITTON. 1963. The chromosome number and possible ancestry of Pea
wrightiana. Amer. J. Bot. 50:52-55.
, and D. S$. CORRELL. 1962. Ferns and Fern Allies of Chihuahua, Mexico. Texas Res.
Found., Renner, TX.

ene R. M. 1963. New chromosome numbers in Polypodium. Amer. Fern J. 53:99-101.

VE. A., Dz LOVE and R. E. G. PICHI SERMOLLI. 1977. Cytotaxonomical Atlas of the

Pteridophyta. Cramer, Vaduz, Liechtenstein.

MANTON, I. 1950. Problems of Cytology and Evolution in the Pteridophyta. Cambridge Univ. Bian
Cambridge, England.

MERRIAM, C. H. 1890. Results of a biological survey of the San Francisco Mountains and the desert
of the Little Colorado in Arizona. U. S. Dept. Agr. N. Amer. Fauna 3:1-136.

M. D. WINDHAM: FERNS OF ELDEN MOUNTAIN 93

MORAN, R. C. 1982. The Asplenium trichomanes complex in the United States and adjacent Canada.
Amer. Fern J. 72:5-11.

PHILLIPS, W. S. 1946. A check-list of the ferns of Arizona. Amer. Fern J. 36:97-108.

______. 1947. A check-list of the ferns of Arizona. Amer. Fern J. 37:13-20, 39-51.

REISENDER, E. A. 1974. Chromosomes of the male fern from the western United States. Amer. Fern
J. 64:

RICHARDSON, P. M. and E. LORENZ-LIBURNAU. 1982. C-glycosylxanthones in the Asplenium
adiantum-nigrum complex. Amer. Fern J. 72:103-106.
SELLERS, W. D. and R. H. HILL. 1974. Arizona Climate: 1931-1972. Univ. of Ariz. Press, Tucson,

AZ.
SHIVAS, M. G. 1969. A cytotaxonomic study of the Asplenium adiantum-nigrum complex. Brit. Fern
Gaz. 10:68—80.
WAGNER, W. H., Jr. 1965. Pellaea wrightiana in North Carolina and the question of its origin. J.
Elisha Mitch. Soc. 81:95-103.
_ 1973. Reticulation of holly ferns (Polystichum) in the western United States and adjacent

Canada. Amer. Fern J. 63:99-115.
WHERRY, E. T. 1941. Asplenium adiantum-nigrum in Arizona. Amer. Fern J. 31:97-100.

REVIEW

hy is diverse and ranges

from 100 to 1990 m in elevation. Vegetation types include evergreen and semi-
evergreen forests, deciduous forests, subtropical pine forests, grasslands, and tem-
perate forests. Terrestrial, lithophilic, and epiphytic ferns are found. The pterido-
phytes number 256 species in 91 genera, a rich and diverse flora. None of the genera
has more than 20 species; the largest are Selaginella and Asplenium. The present
volume includes keys to the families, genera, and species, synonymies, descriptions,
habitat and range notes in the Flora area, an interesting introduction, a bibliography,
and an index. Some of the species are illustrated with photographs or line drawings.
The keys seem well constructed. The book will be of interest to all who deal with
the ferns of India and the surrounding region. The book is available from United
Book Traders, Opposite Police Lines, Ratanada, Jodhpur 342001, India. The U.S.
price of $20.00 far exceeds the rupee price. —D.B.L.

94 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Donovan S. Correll (1908-1983)

Donovan Stewart Correll was born on 13 April 1908 in Wilson, North Carolina.
He grew up in Winston-Salem and attended Duke University, which awarded him an
A.B. in 1934 and an A.M. in 1936. From 1939 to 1943 he did graduate study at the
Botanical Museum, Harvard University. He was awarded a Ph.D. by Duke Univer-
sity in 1943. He joined the U.S. Navy for the balance of World War II, and then
worked in the Division of Plant Exploration and Introduction, U.S. Department of
Agriculture, from 1944 to 1956. He was the Chief Botanist at the Texas Research
Foundation from 1956 until 1972, worked for the National Science Foundation as a
program director in 1972 and 1973, and then “retired” to the Fairchild Tropical
Garden, where he carried out a strenuous research program until shortly before his
death on 28 Mar 1983, not long after completing his monumental “Flora of the
Bahamas Archipelago.” His principal works on ferns were “The Ferns and Fern
Allies of Louisiana” (with Clair A. Brown), the “Ferns and Fern Allies of Texas,”
and “The Ferns and Fern Allies of Chihuahua” (with I. W. Knobloch). Besides the
ferns, his specialties were the Orchidaceae, Solanaceae, and Palmae, and also the
floras of the southwestern United States and the Bahamas. His knowledge of the
floras of the New World was tremendous. He conducted field work in the United
States, Mexico, Central America, the West Indies (especially the Bahamas), South
America, and Hawaii. An obituary and bibliography is to be published in Brittonia.

Vee

REVIEW

FERNS AND ALLIED PLANTS. WITH SPECIAL REFERENCE TO
TROPICAL AMERICA, by Rolla M. Tryon and Alice F. Tryon. xiv + 858 pp.
illustr. Habitat photography principally by Walter F. Hodge. Springer-Verlag, New
York. 1982. ISBN 0-387-90672-X. $148.00—Fern specialists are blessed (some
would say cursed!) with almost as many classifications from which to choose as there
are pteridologists. The Tryons provide us with yet another choice. In a juxurious
format designed to facilitate quick reference and comparison, they give a synonymy,
a description, comments on classification, a key to the genera, and selected
references for each of 29 families. Each generic treatment includes a synonymy, a
description, a statement on classification, a discussion of the tropical American
species, often keys, discussions of ecology, geography, spores, and chromosome
numbers, additional observations (mostly interesting notes on natural history), and
literature. Taxonomic opinions also have been rendered at the species rank, e.g-, in
Schizaea (p. 78), Paesia (p. 396), and Dryopteris (p. 501), usually with a reduction
in species. An index to scientific names completes the work.

: One of the best features of the book is the abundance of good illustrations. These
include habit photographs of nearly all genera, photographic details of diagnostic
features (€.g., venation, indument, and sori), silhouettes of fronds (chosen to show
diversity in a genus), line drawings and pencil sketches, and SEM photographs of

AMERICAN FERN JOURNAL: VOLUME 73 (1983) 95

spores and occasionally other characters. In addition there are dot maps showing
generalized American distributions for each genus; these I find of questionable
value—their place could have been taken by a simple statement of range.

The SEM photographs of spores are one of the most conspicuous features of this
book, represent a primary source of new data in the classification, and deserve
special comment. I was struck by the often large variation in spore ornamentation
within many genera and even within species (e.g., Cystopteris fragilis). Such
diversity leads me to question the utility of spore morphology in arriving at
taxonomic judgments; at least the use of this evidence must be tempered with data
from many other sources. The authors are to be commended for giving vouchers for
spore photographs. However, they fail to mention sample sizes when making
generalizations based on spore morphologies, e.g., the distinction between
Microlepia and Dennstaedtia (p. 375).

Interspersed in the text are a few original chromosome counts (e.g., Platycerium
andinum, Trichipteris microdonta). It is apparent that chromosome numbers also
form a principal body of evidence in the Tryons’ new classification. There is an
inclination to refer to ancestral base numbers for some groups, despite the lack of
convincing evidence for such numbers (e.g., 23 from 69 for the Cyatheaceae).

In general, the authors have taken a moderate view in recognition of families,
somewhere between the narrow circumscriptions of Pichi Sermolli (1977) and Ching
(1940) on the one hand and Wagner (1973) on the other. Genera are circumscribed
mostly in a broad sense, a position with which I generally agree; prominent
examples are Asplenium, Dryopteris, Gleichenia, Grammitis, Tectaria, Thelypteris,
and Trichomanes. Exceptions are the genera of the Cyatheaceae, the subject of
recent monographic work by R. Tryon and students, and of the Polypodiaceae sensu
stricto. I especially like the breakdown of large genera into species clusters—groups
of presumably related species—with a characterization of these clusters (examples
are in Adiantum and Polypodium, in addition to some of those mentioned above).
There are a few surprising generic circumscriptions. For example, Camptosorus and
Ceterach are accorded generic rank even though they readily hybridize with
Asplenium; other equally distinct genera, ©.g., Antigramma, are sunk in Asplenium.
A similar situation is found in Pteridaceae, where Adiantopsis is recognized but
Aspidotis and Mildella merged in Cheilanthes. Therein, Cheilanthes siliquosa has
been placed in a different species group than C. californica, even though they are so
closely related that they hybridize to produce a fertile allopolyploid, and juveniles of
all three are almost indistinguishable. In the same family, the authors postulate a
relationship between Llavea and Lygodium (p. 310). citing as evidence the similarity
in spores and chromosome number; the chromosome number here seems like
spurious evidence and the spores of Llavea actually appear more similar to those of
certain other cheilanthoid genera, e.g., Cryptogramma.

The chief fault of most recent classifications is insufficient evidence. The Tryons
provide more documentation for their views than most have done. This alone will
make their book a standard and indispensable reference for specialists and general-
ists alike.—Alan R. Smith, Department of Botany-Herbarium, University of Cali-
fornia, Berkeley, CA 94720.

96 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

REVIEW

FIELD GUIDE TO MISSOURI FERNS, by J. S. Key. Missouri Dept. of
Conservation, 220 pp. 1982. $3.00.—This book was written primarily for non-
botanists, and so has attractive line drawings rather than keys for identification of 79
species, varieties, and forms in 28 genera found in Missouri. Introductory material
includes fern morphology, collecting, preservation, and habitats. A pictorial guide
using small illustrations and based mostly on frond morphology and habitat leads
readers to genera. Within the genus, each taxon is illustrated with an attractive habit
sketch and detailed drawings of diagnostic characters. The illustrations, plus a brief
description, permit identification of the material. Each taxon has the scientific
name, with an occasional synonym, the common name, and a statement of habitat
and range in Missouri, as well as the description. A glossary, checklist, and an
index conclude the volume. The perfect-bound book is available from the Missouri
Dept. of Conservation, P.O. Box 180, Jefferson City, MO 65102. Missouri
residents should add $0.12 tax.—D. B. L.

REVIEW

AZOLLA AS A GREEN MANURE: USE AND MANAGEMENT IN CROP
PRODUCTION, by T. A. Lumpkin and D. L. Plucknett. Westview Press, Boulder,
CO. xx + 230 pp. 1982. $20.00.—Although the focus of this paperback volume is
agronomic, the chapter on botany and ecology (pp. 15-38) is likely to be of interest
to taxonomic botanists. Illustrations and a tentative key to the species the authors
recognize (A. nilotica, A. pinnata and var. imbricata, A. microphylla, A. filiculoides,
A. rubra, A. mexicana, and A. caroliniana) will be useful in making identifications
in this difficult genus. Most of the chapter concerns the morphology, anatomy, an
life cycle of Azolla, and contains many illustrations. Unfortunately, the reproduction
IS poor, and so not much detail can be seen in the illustrations. A world-wide map
showing the distribution of the Azolla species concludes the chapter. The bulk of the
book concerns the physiology, nursery culture, field cultivation, pests, and uses in
agriculture of Azolla. The book is available from Westview Press, 5500 Central
Ave., Boulder, CO 80301.—D.B.L.

a EEeaIsaneesuar
BRITISH PTERIDOLOGICAL SOCIETY

Open to all who are interested in growing and studying ferns and fern-allies.
Full members receive THE FERN GAZETTE and the BULLETIN OF THE
BRITISH PTERIDOLOGICAL SOCIETY. Membership subscriptions are £7
for full members, £ 5 for ordinary members (not receiving the GAZETTE), and
£5 for student members (under 25 years of age). For particulars, U. S.
residents should apply to Dr. J. Skog, Biology Department, George Mason
University, Fairfax, VA 22030. Non-U. S. residents should apply to Lt. Col. P.
G. Coke, Robin Hill, Stinchcombe, Dursley, Gloucestershire, England.

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FERN BE
JOURNAL

QUARTERLY JOURNAL OF THE AMERICAN FERN SOCIETY

Polyploidy and Aneuploidy in Hypolepis,

and the Evolution of the Dennstaedtiales P. J. BROWNSEY
Pecluma, a New Tropical American Fern Genus MICHAEL G. PRICE
The Lady Fern, Athyrium filix-femina,

in Saskatchewan VERNON L. HARMS
Review

American Fern Journal
Index to Volume 73

Errata

109

117

The American Fern Society

DAVID B. LELLINGER, Smithsonian Institution, Washington, DC 20560. Journal Editor
ALAN R. SMITH, Dept. of Botany, Un niversity of California, Berkeley, CA 94720. Memoir Editor
JOHN T. MICKEL, New York Botanical Garden, Bronx, NY 10458. Newsletter Editor

American Fern Journal
EDITOR

DAVID B. LELLINGER U.S. Nat’] Herbarium NHB-166, Smithsonian Institution,
Washington, DC 20560

ASSOCIATE EDITORS
DAVID W. BIERHORST Rt. 3, Box 188, Picayune, MS 39466.
GERALD J. GASTONY Dept. of Eprey: Indiana ete Bloomington, IN 47401.
JOHN T. MICKEL w York Botanical Garden, Bronx, NY 10458.

TERRY R. WEBSTER ....... Biological Sciences peo University of Connecticut, Storrs, CT 06268.

The “American Fern Journal” (ISSN 0002-8444) is an illustrated quarterly devoted to the general
study of ferns. It is owned by the American Fern Society, and a at the Smithsonian Institution,
Washington, DC 20560. Second-class postage paid at Washington.

Claims for missing issues, made 6 months (domestic) to +9 "geen (foreign) after the date of issue,
and the matters for publication should be addressed to the Edito

Changes of address, dues, and applications for membership should be sent to Dr. Leslie G. Hickok,
—. of Botany, University of Tennessee, Knoxville, TN 37916.

rt back issues should be addressed to Dr. James D. Montgomery, Ichthyological Associates,
5 De Nalin PA 18603.

General i inquiries oe ferns should be addressed to the Secretary.

Subscriptions $9.00 gross, $8.50 net if paid =e an agency (agency fee $0.50); sent free to
members of the American Fem Society (annual dues, $8.00 + $4.00 mailing surcharge beyond U.
Canada, and Mexico; life membership, $160.00).

Library

Dr. John T. Mickel, New York Botanical Garden, Bronx, NY 10458, is Librarian. Members
may borrow books at any time, the borrower paying all shipping costs.

Newsletter
« Dr. John T. Mickel, New York Botanical Garden, Bronx, NY 10458, is sarap of the newsletter
Fiddlehead Forum.” The editor welcomes contributions from members and no mbers, including

miscel notes, offers to exchange or purchase materials, personalia, rete notes, and
reviews of non-technical books on ferns.

Spore Exchange
Mr. Neill D. Hall, 1230 Northeast 88th Street, Seattle, WA 98115, is Director. Spores exchanged and
collection lists sent on re request
Gifts and Bequests
Gifts and bequests to the Society enable it to expand its services to members and to others interested

in ferns. Botanical books, back issues of the Journal, and cash or other gifts are always welcomed, =
are tax-deductible. Inquiries should be addressed to the Secretary.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER f (1983) 97

Polyploidy and Aneuploidy in Hypolepis,
and the Evolution of the Dennstaedtiales
P. J. BROWNSEY*

The fern genus Hypolepis is widespread in tropical and south temperate parts of
the world. It has one center of distribution in southeastern Asia and Australasia
(with a few species spreading as far as west Africa, Korea, the eastern Pacific and
New Zealand) and another in the New World from the southernmost U.S.A. through
Mexico and the Caribbean to Central and South America. Only one or two species
of circumantarctic distribution are possibly common to both centers.

Morphologically the genus is fairly well defined, but the limits of individual
species are much less obvious. Copeland (1947, p. 57) suggested that about 45
species should be recognized, although more recent investigations indicate that this
is probably an underestimate. Particular problems arise in widely distributed,
polymorphic taxa. These difficulties are compounded by the widespread and
persistent misapplication of well known names such as H. punctata (Thunb.) Mett.
and H. tenuifolia (Forst. f.) Bernh. and because many critical taxonomic features,
notably the lamina and rachis hairs, are often poorly preserved or lost from old
specimens, including many type collections.

The present paper is concerned with some of the wider implications resulting
from a recent study of Hypolepis in Australia and New Zealand (Brownsey &
Chinnock, in prep.). Although some of the ideas expressed here are speculative,
they are put forward now in the hope of stimulating further research, particularly
cytological investigation, into critical members of this group of ferns.

MORPHOLOGY

Hypolepis includes species which have the following combination of characters:
rhizomes long-creeping; fronds bipinnate or more compound, often large, free-
veined; hairs, glandular or bristly, on some part of the lamina, rachis, stipe or
rhizome; scales absent; sorus + round, ranging from terminal on a vein at the
lamina margin, protected by a reflexed indusial flap or modified portion of the
lamina, to submarginal, not quite terminating the vein, and totally unprotected,
spores monolete.

These characters distinguish Hypolepis from allied genera, notably Paesia,
Pteridium, and Dennstaedtia. In Paesia, the sori are continuous along the margins
of the ultimate pinnules, borne on a marginal connecting vein, and protected both by
the reflexed edge of the pinnule and by a distinct inner indusium. Pteridium 1s
Similar to Paesia, except that the inner indusium is less well developed and the
Spores are trilete. In Dennstaedtia, the spores are also trilete, but the marginal sorus
is protected by a cup-shaped or slightly bivalvate indusium formed by the fusion of
Inner and outer indusia.

*National Museum of New Zealand, Private Bag, Wellington, New Zealand.

Volume 73, number 3, of the JOURNAL was issued 29 Sept 1983.

98 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

44) INN mt
Th

j
f

MN Ei:
ASN

——
saaaeliias Netting

=| &
WS =
om 7
Te)
fe)

FIG. 1. Frond silhouettes and drawings of pinnules of Hypolepis distans (A) compared with H.
tenuifolia sensu Allan (1961) (B) and H. millefolium Hook. (C).

Some species have been thought to transcend these generic boundaries. Mickel
(1973) noted that an inner indusium was present in Hypolepis bivalvis v.A.v.R.. Dut
this taxon is better treated as Paesia elmeri Copel. (Holttum, 1958). More signifi-
cantly, Bower (1928, fig. 587) reported the occasional presence of a vestigial inner
indusium in H. repens (L.) Presl which he suggested linked Hypolepis with the
Dennstaedtioid ferns, but this observation requires further investigation.

Many Hypolepis species are rather similar and must be distinguished by combina-
tions of a few vegetative characters. In some cases, a well developed indusial flap 1S
characteristic, but mostly their sori are unprotected. The hairs on the underside of
the lamina are a key distinguishing character, varying in length, color, position, and
glandularity. Also important are the degree of pinna dissection and the stipe and
rachis color.

In Australia and New Zealand, H. distans Hook. stands out as being markedly
different from the other ten or eleven species. The laminae are oblong-lanceolate
and somewhat coriaceous, rather than characteristically deltoid and membranaceous,

P. J. BROWNSEY: POLYPLOIDY AND ANEUPLOIDY IN HYPOLEPIS 99

FIG. 2. Scanning electron micrographs of spores of Hypolepis distans (A) with a rather smooth
perispore, and H. rufobarbata (Col.) Wakef. (B) with a perispore bearing numerous flattened projections.

the lower pinnae arise at 90° to the rachis (Fig. JA), not at the usual acute angle
(Fig. 1B); the stipes are very thin and highly polished, rather than generally thicker
and hispid; and there are very few hairs anywhere on fronds of H. distans, which is
unusual for Hypolepis. Most importantly, the veins of the ultimate pinnules end in
emarginations (Fig. JA) rather than in lobes (Fig. /C). Finally, the spores of H.
distans are very distinct, being dark brown and lacking any marked projections, in
contrast to the spores of other species, which are pale and bear quite long, flattened
Projections (Fig. 2).

Species rather similar to H. distans are found in the New World, notably H.
nigrescens Hook., which ranges from southern Mexico through Central America and
the Caribbean to Colombia and Brazil, and the doubtfully distinct 1. hispaniolica
Maxon from the Dominican Republic and Haiti. Hypolepis nigrescens resembles H.
distans in having coriaceous fronds, relatively few hairs on the lamina, pinnae
arising at 90° to the rachis, veins ending in emarginations, and dark spores. On the
other hand, it has rather thick stipes, which are pubescent and armed with sharp
Prickles, and spores which bear very fine spines. In these characters, H. nigrescens
more closely resembles a distinctive group of Hypolepis species from southeast Asia
represented by H. brooksiae v.A.v.R. and H. papuana Bailey. Like H. nigrescens,
these species have a scrambling habit and periodic dormancy of the rachis (Holttum,
1958). Furthermore, their veins have a slight tendency to end in emarginations, but
this is by no means so pronounced as in either H. distans or H. nigrescens. To what
extent these three elements are related is not entirely clear from morphology alone.

100 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

A : HB

FIG. 3. Meiotic chromosome preparations ( x 1000) from Hypolepis distans (A) Lake Kopureherehere,
Levin, New Zealand, showing 28 bivalents, and H. rufobarbata (B) Kaituna Spur, Banks Peninsula,
New Zealand, showing 52 bivalents. Voucher specimens in WELT.

HYBRIDIZATION AND CYTOLOGY

Evidence is accumulating that species of Hypolepis, like those of Asplenium,
Dryopteris and Polystichum, have a considerable capacity for hybridization. There 1s
good cytological evidence for this at different levels of ploidy among New World
species of the genus and also in isolated records from Australia and Japan (Table !).
In New Zealand, hybridization in Hypolepis has long been suspected (Carse, 1929;
Cockayne & Allan, 1934), and my own morphological and cytological investigations
confirm this. Hybrids are recognizable by their intermediate morphology, shrivelled
spores, and by the irregular pairing of their chromosomes at meiosis. Five of the six
species of Hypolepis occurring on the main islands of New Zealand whose
distributions overlap have been found to hybridize. Only H. distans does not appeat
to hybridize, which suggests, in common with the morphological and cytological
evidence, that it is fundamentally distinct from the other native species.

My own unpublished results together with those of Brownlie (1954, 1957, 1958,
1961) indicate that of the six New Zealand species of Hypolepis other than H.
distans, four have n=52 (Fig. 3B) and two n= 104. By contrast, plants from four
populations of H. distans have n=28 (Fig. 3A). Furthermore it is noteworthy that
the chromosomes of H. distans are markedly larger than those of other New Zealand
species (cf. Figs. 3A and 3B).

Previously published cytological reports relating to species other than those 1D
New Zealand are summarized in Table 1. All identifications are those of the original

mat Reports showing irregular meiotic pairing have been segregated as putative
ybrids.

P. J. BROWNSEY: POLYPLOIDY AND ANEUPLOIDY IN HYPOLEPIS 101

TABLE 1. CHROMOSOME COUNTS FOR SPECIES OF HYPOLEPIS FROM AREAS OTHER
THAN NEW ZEALAND

Species Origin n 2n__ Reference
OLD WORLD
H. punctata Japan c. 104 - Kurita, 1962
J 98 - Kurita, 1967, 1972
Japan, Obarano Go9z - Mitui, 1
Japan, Saitama Pref. 98 - Mitui, 1975
Japan, Niigata Pref. 98 - Mitui, 1976
Taiwan 52 - “Teal, 1979
Himalayas, Darjeeling c. 102 - Mehra & Khanna, 1959
Himalayas, Mussoorie 104 - Mehra & Verma, 1960
Nepal, Kathmandu Valley 104 - Royetal., 1971
Ceylon, Hakgala 51-53 - Manton & Sledge, 1954
Malaya, Taiping Hills c. 100 - Manton & Sledge, 1954
Malaya, Taiping Hills c. 104 - Manton in Holttum, 1954
Malaya, Fraser Hills c. 104 - Manton in Holttum, 1954
H. tenuifolia Samoa - 104. Manton & Vida, 1968
H. villoso-viscidum Gough Island c. 100 - Manton & Vida, 1968
H. sp. Australia 98 - Kurita, 1972
Hybrids
eo Australia - c. 150 Manton & Sledge, 1954
“H. punctata x meiosis c. 200 II+I Kurita, 1967, 1972
alte-gracillima” Japan irregular
H NEW WORLD oy
. Tepens i - Wagner en in
pe U.S.A., Florida 104 by & Solbrig, 1964
Costa Rica, Platanillo 52 - Smith & Mickel, 1977
Puerto Rico, El Verde 39 - Sorsa in Fabbri, 1965
H. viscosa Mexico, Oaxaca 52 - Smith & Mickel, 1977
H. nigrescens Mexico, Oaxaca 29 - Mickel et al., 1966
Jamaica, Caledonia Peak 29 - Walker, 1966
Jamaica, Hardwar Gap 29 - Walker, 1966
Hybrids
“H. bogotensis” Mexico, Chiapas 5011 + 54 I - Smith & Mickel, 1977
a = repens” Costa Rica, Platanillo 4911 + 50 I - Smith & Mickel, 1977
- aff. vi ” i
a hoe. ne, Comes 5211+ 52 | - Smith & Mickel, 1977

The majority of Hypolepis species so far investigated have chromosome comple-
ments of either n=52 or 104. The various reported numbers for collections
identified as H. punctata and H. repens strongly suggest that these taxa are
ill-defined. My own morphological investigations suggest that counts for “H.
Ppunctata” may belong to as many as four different species from Japan and northern
India, Taiwan, Ceylon, and Malaya, although without acc —
cannot be verified. In the New World, there is clear evidence of polyploidy in H.
repens, and it is also likely in the H. bogotensis/viscosa complex

102 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

actual counts of n= 104 have yet to be demonstrated. Similarly, in Australia, the
count from the hybrid plant with 2n=c. 150 strongly suggests the presence of
parental species with n=52 and 104.

Of the anomalous numbers, the most improbable is n= 39 for H. repens from
Puerto Rico which must be compared with n= 104 for the same species in Florida
and n=52 from Costa Rica. Such divergent numbers within a single species
complex are hard to accept, notwithstanding their arithmetical relationship based on
13 and the undoubted fact that H. repens in Central and South America is an
ill-defined aggregate (Stolze, 1981, p. 280).

Another curious number, n= 98 in Japanese plants of H. punctata, must be taken
more seriously since it has been consistently reported from several populations and
illustrated at least three times (Kurita, 1967, 1972; Mitui, 1975, 1976). Presumably
it has arisen by aneupoidy, either by direct reduction from a parental species with
n= 104, or, earlier in its evolutionary history, from one with n=52 (i.e. 52 to 49 to
98). The cells illustrated by Kurita (1972, fig. 11) and Mitui (1976, fig. 5) show
some disparity in the size of individual bivalents consistent with the idea of
reduction in chromosome number by fusion. Further investigation is required to
ascertain the geographical distribution of this cytotype and the extent to which the
loss of chromosomes is reflected in its external morphology. From a nomenclatural
point of view, it is significant that the type of H. punctata is from Japan.
The report of n=29 in H. nigrescens cannot be seriously doubted, having been
independently reported by Mickel et al. (1966) from Mexico and by Walker (1966)
from two populations in Jamaica. Furthermore, the number can now be very
Satisfactorily related to the discovery of n=28 in H. distans from New Zealand.

owever, what is not known is whether there is a direct relationship between these
two species, or whether they represent fragments of quite different aneuploid lines.
While the morphological and cytological evidence confirms that H. distans and H.
nigrescens (and probably also the H. brooksiae/papuana aggregate) are significantly
different from most other members of the genus, it does not yet resolve the
relationship between the three elements.

n summary, these cytological observations point to a base number of x=26 in

Hypolepis, despite the fact that such a number has yet to be recorded for the genus.
There is a well established polyploid line of evolution represented by the numbers
n= 104 and 52 that is almost certainly derived from n=26, and there is the more
recently demonstrated evidence of aneuploidy, with n=29 in H. nigrescens and
n= 28 in H. distans, that can also be related to n= 26 (Fig. 4). The former existence
ig n=26 in Hypolepis is indirectly supported by cytological observations from
Paesia, where the numbers n= 26 and 104 have been reported, and from Pteridium,
Where n=26, 52, and 104 are all known (for original references see Love, Love
Pichi Sermolli, 1977, p.189). Although a base as low as 13 is theoretically possible,
it seems highly unlikely unless the count of n=39 from Puerto Rican plants of 17
repens can be confirmed. On the other hand there is a strong possibility that the
PORNaVS number in the genus may actually be 29, from which species having n=2
were derived by aneuploidy, ultimately giving rise to a successful polyploid line of
evolution radiating at the tetraploid and octoploid levels.

P. J. BROWNSEY: POLYPLOIDY AND ANEUPLOIDY IN HYPOLEPIS 103

13 yp fee a
.
%
ot Dennstaedtia
” scandens
\29 7;
N
\
‘\
NX
Mee
7 “*
\28 1
-*
H. nigrescens i
* cae
H.distans eee:
aoe a2 1 o* ae
-” a ! ry ~
r v =
HYPOLEPIS PAESIA PTERIDIUM

oat
/ eo
s 62) c.8 spp 52) (62) 2spp
7 sat
/

>
(98 }
ye >)
H.punctata®
c. 5spp. 150. 1Sp.

FIG. 4, Phyletic scheme deriving the Hypolepidaceae from the Dennstaedtiaceae. Solid lines and
numbers in unbroken circles indicate probable lines of evolution and established chromosome numbers.
roxen lines and numbers in broken circles indicate possible lines of evolution and doubtful or

ke
Postulated cytotypes.

104 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

PHYLOGENETIC CONSIDERATIONS

Mickel (pers. comm.) has pointed out that the spores of H. distans are more
similar in their perispore pattern to those of Paesia than to any species of Hypolepis,
including H. nigrescens. He recognized some similarity in frond form between H.
distans and the species of Paesia and suggested that it could be part of an aneuploid
series in Paesia which has resulted in the loss of the characteristic inner indusium.
Despite the similarity of its spores to Paesia, I do not believe that this alone provides
sufficient evidence for regarding H. distans as a species of Paesia. In all other
characters it is consistent with Hypolepis rather than Paesia: the inner indusium is
absent: the sorus terminates a single vein; the veins are conspicuous (in Paesia they
are difficult to see); the veins end in emarginations (rare in Hypolepis, but unknown
in Paesia); the hairs are scarce and non-glandular (in Paesia they are usually
abundant and glandular); and the rachis is straight, bearing pinnae which arise at
right angles in more or less opposite pairs (in Paesia it is characteristically zig-zag
bearing pinnae which arise alternately at an acute angle). Holttum (1958) related the
shape of the rachis to periodic dormancy of the apex, pointing out that in P. elmeri
the rachis rests while a single pinna develops in turn, but that in H. brooksiae and
other genera showing periodic dormancy, a pair of pinnae develop together while the
rachis rests.

Although, in my opinion, H. distans is closer to the polyploid species of

Hypolepis (with n=52 or 104) than to species of Paesia, there is no doubt that all
three groups are very closely related. Nevertheless, H. distans, with or without H.
nigrescens and H. brooksiae, constitutes a distinctive element. Smith (1981, P. 136)
has already noted the cytological heterogeneity of the genus and suggests that
“Hypolepis as presently circumscribed may be unnatural.” There is certainly a
strong case for distinguishing the aneuploid species from the polyploid, but whether
such taxonomic recognition should be at the generic or subgeneric level, and
whether the aneuploid species themselves constitute a natural grouping, are very
much more difficult questions to answer. For the moment, no change is proposed,
and Hypolepis is interpreted here in a broad sense.
The question arises as to whether the aneuploid species of Hypolepis provide a
link with other allied genera, or whether they are merely the end products of
divergent evolution from an ancestral stock with n= 26. Paesia and Dennstaedtia are
central to any consideration of this possibility. Copeland (1947, p. 57), Holttum
(1949), and Mickel (1973) have pointed to the similarity between Hypolepis and
Dennstaedtia, Copeland in particular remarking “The more primitive element In
[Hypolepis] .. . is hardly distinguishable from a similar element in Dennstaedtia.”
Species of the two genera share many vegetative characters, although in my
experience the indusial and spore characters will always distinguish them. Neverthe-
less, the presence of an aneuploid line in Hypolepis including the numbers 28 and
29 is of considerable interest since it ties in remarkably with the numbers 30-34,
46, and 47 so far known in Dennstaedtia (Lovis 1977, p. 275; Léve et al. 1977, P-
184). These numbers have already been cited by Lovis (1977, p. 303) in postulating
that the evolution of the Dennstaedtiaceae (i.e., Pichi Sermolli’s Dennstaedtiales) !8
the result of a long series of aneuploid reductions from a base in the Cyatheaceae.

P. J. BROWNSEY: POLYPLOIDY AND ANEUPLOIDY IN HYPOLEPIS 105

It may be more than mere coincidence, therefore, that Hypolepis brooksiae bears
a quite extraordinary morphological similarity to Dennstaedtia scandens (Blume)
Moore. In the field both are scrambling, thicket-formers with periodic dormancy of
the rachis (Holttum, 1958); in the herbarium they can be distinguished only by a
very careful scrutiny of the indusium or by the spores. It is highly significant that
Tsai (1973) has reported the chromosome number of D. scandens in Taiwan to be
n= 30. Unfortunately, the chromosome number of H. brooksiae is not yet known,
but if, as its morphological similarity to H. distans and H. nigrescens suggests, the
number proves to be in the high 20’s, this would constitute good evidence of an
evolutionary link between Dennstaedtia and Hypolepis.

Some features of H. distans, notably the pronounced emarginations, the thin,
highly polished stipes, and the virtually glabrous fronds, are not typical of
Dennstaedtia. One possible answer to this paradox may be that H. nigrescens, H.
distans, and perhaps H. brooksiae, represent remnants of unstable karyotype
combinations thrown up in the course of a series of aneuploid reductions from a
source in Dennstaedtia extending back to at least x=34 and possibly as far as
x=47. An unbroken series of base numbers from 34 in Dennstaedtia punctilobula,
through 33 or 32 in D. scabra, 31 in D. wilfordii, 30 in D. hirsuta and D. scandens,
29 in Hypolepis nigrescens, and 28 in H. distans, ultimately giving rise to a
felicitous combination of 26 chromosomes in Paesia, Pteridium, and the polyploid
species of Hypolepis, is an entirely plausible evolutionary pathway (Fig. 4). The
very fact that many of the same characters appear in different combinations in the
four genera reinforces the belief that these genera do share a common origin. For
example, Dennstaedtia, Paesia and Pteridium have an inner indusium, but Hypolepis
does not; Dennstaedtia and Hypolepis have sori terminating a single vein, whereas
Paesia and Pteridium have sori linking several veins; Dennstaedtia and Pteridium
have trilete spores, whereas Hypolepis and Paesia have monolete spores.

LIMITS OF THE HYPOLEPIDACEAE

The possibility that members of the Hypolepidaceae were derived from a source
in the Dennstaedtiaceae necessitates a more careful consideration of the genera
which constitute the former family. There is no doubt in my mind that Hypolepis,
Paesia, and Pteridium form a natural group of genera with a common ancestry based
on x= 26 and that they are related to—and possibly derived from—Dennstaedtia,
but I find it much more difficult to ally them with some of the other genera which
have been referred to the Hypolepidaceae. ae :

The family was first circumscribed and its relationships outlined by Pichi Sermolli
(1970, 1977, p. 431). He included in it Hypolepis, Paesia, Preridium, Histiopteris,
Lonchitis, and Blotiella. To these (as subfamily Hypolepidoideae of the Denn-
Staedtiaceae) Lovis (1977, p. 276) added Monachosorum, Taenitis, and /diopteris,
although most workers would now probably agree that the latter two genera are
better placed with the Gymnogrammeoid and Pteridoid ferns, respectively.

The true affinities of Monachosorum have never been satisfactorily established,
but it fits no better in the Hypolepidaceae than in any of the other families to which
it has been assigned (Christensen, 1938; Holttum, 1947; Copeland, 1947, p. 51;

106 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Mickel, 1973). Lovis’s main reason for putting it with the Hypolepidaceae was the
belief that 7 =56 in M. maximowiczii (Bak.) Hayata might have been derived from
x=28, which relates to known numbers in Hypolepis.

Lonchitis and Blotiella have also been regarded as somewhat discordant elements
in the Hypolepidaceae, both by Mickel (1973) and even by Pichi Sermolli (1977, p.
432). Their chromosome numbers (7 =50 and 38) do not fit at all comfortably.

There remains the genus Histiopteris, which Mickel (1973) and Pichi Sermolli
(1977, p. 432) found no difficulty in allying to Hypolepis, Paesia, and Preridium.
However, Copeland (1947, p. 60) considered it related to Preris, and Holttum (1973)
included it in his list of genera whose relationships need “fresh examination.” My
own experience of Histiopteris in New Zealand suggests that it is not closely allied
to other members of the Hypolepidaceae. The presence of scales on the rhizome,
glaucous and virtually glabrous fronds, anastomosing veins, and sessile pinnae are
all characters alien to Hypolepis, Paesia, and Preridium. Most importantly, the
known chromosome numbers of n=48 and 96 (Love et al. 1977, p. 191) cannot
easily be related to a base of 26 in the other genera.

Assuming that Monachosorum, Lonchitis, Blotiella, and Histiopteris, if not
members of the Hypolepidaceae, are at least Dennstaedtialean, room must obviously
be found for them elsewhere. Crabbe et al. (1975, p. 155) have already proposed for
Monachosorum a monogeneric subfamily, Monachosoroideae, within their
Dennstaedtiaceae (equivalent to Pichi Sermolli’s Dennstaedtiales), and a solution of
this nature may be necessary for the other genera.

EVOLUTION OF THE DENNSTAEDTIALES SENSU PICHI SERMOLLI

Looking beyond the Hypolepidaceae, Wagner (1980) has recently proposed that
the neotropical genus Loxsomopsis might have an “affinity to either the dennstaedtioid
or lindsaeoid ferns” on account of its chromosome number of n=46. This contrasts
with n= 50 reported for Loxsoma (Brownsey, 1975), a genus previously considered
very close to Loxsomopsis but whose relation to other genera has always been very
uncertain. Investigation of its gametophytes by Stokey and Atkinson (1956) and of its
stomata by van Cotthem (1970, 1973) suggested an affinity between Loxsomaceae
and Cyatheaceae, but the cytological evidence indicates that any such relationship
must be fairly remote. However, Lovis’s (1977, p. 303) suggestion that the
Dennstaedtiaceae (i.e., Pichi Sermolli’s Dennstaedtiales) originated by a long series
of aneuploid reductions from a source in the Cyatheaceae provides a means 0
reconciling these differences. If Loxsoma and Loxsomopsis are admitted to be
Dennstaedtialean ferns, as at least some of their morphological characters and
certainly their chromosome numbers suggest, then the distant affinity with the
Cyatheaceae, hinted at by the gametophyte and stomatal characters, lends some
support to Lovis’s hypothesis.

It may be that the troublesome Monachosorum also belongs on the evolutionary
line from the Cyatheaceae to the Dennstaedtiales. Its position here could help to
reconcile the views of Copeland (1947), Crabbe et al. (1975), and Pichi Sermolli
(1977, p. 430), who allied it with the Dennstaedtiaceae, with those of Christensen
(1938), who allied it with the Thelypteridaceae; in both Holttum’s (1973) and

P. J. BROWNSEY: POLYPLOIDY AND ANEUPLOIDY IN HYPOLEPIS 107

Lovis’s (1977, fig. 3) scheme, the latter family is itself an offshoot of the
Cyatheaceae. Certainly the chromosome number of Monachosorum (n=56) fits
better here than in the Hypolepidaceae. Moreover, the numbers 46, 50, and 56 now
_ in Loxsomopsis, Loxsoma, and Monachosorum go some way to bridging

e gap between the highest confirmed number of n=48 in the Dennstaedtiales
. 1977, p. 304) and the range of 56—69 (excepting 95 or 96 in Metaxya) in the
Cyatheaceae sensu Lovis (1977, p. 273)

This leaves an awkward group of genera, Blotiella (n= 38), Histiopteris (n= 48),
and Lonchitis (n=50), whose affinities are far from clear. Blotiella and Histiopteris
have much in common morphologically, but cytologically the latter genus is closer
to Lonchitis. Tryon (1962) suggests that Blotiella is related to Hypolepis, but that
Lonchitis is closer to Pteris. Histiopteris has also been allied with both the
Dennstaedtialean and Pteridoid ferns, but its cytology fits comfortably into neither
group; the Pteridoid ferns are clearly based on x =29 or 30, albeit with an aberration
of n=27 in Idiopteris, while the known range in Dennstaedtialean ferns (26—c. 50)
would require Histiopteris to be placed at the higher end of the cytological spectrum
where its morphology is even more out of place than at the lower end with the
Hypolepidaceae. The affinities of these three genera need to be more thoroughly
investigated; meanwhile, I regard their placement as uncertai

I am extremely grateful to Dr. J. E. Braggins, University of Auckland, New
Zealand, for supplying me with the photograph illustrated in Figure 2, to Dr. J. T.
Mickel, New York Botanical Garden, for his unpublished observations on New
World species of Hypolepis and Paesia, and to Dr. Mickel and Professor J. D.
Lovis, University of Canterbury, New Zealand, for their critical and extensive
comments on the manuscript.

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AMERICAN FERN JOURNAL: VOLUME 73 EER F (1983) 109

Pecluma, a New Tropical American Fern Genus
MICHAEL G. PRICE*

The species of Pecluma were submerged in Polypodium until modern taxonomists
separated Grammitids from Polypods. Then, Copeland (1956) included some
members of Pecluma in a generic treatment of the Grammitid Ctenopteris. That
provided the impetus for three papers which supplied evidence that Pecluma species
indeed belonged with Polypodium (Stokey, 1959; Wilson, 1959; Sota, 1963). A later
revision of the individual species by Evans (1969) has facilitated a new examination
of the relationships of the group, and I have come to the conclusion that those
authors who reaffirmed an alliance with Polypodium are correct, but that Copeland’s
error may be partly excused on the grounds that Pecluma also has many features
reflecting a shared ancestry with Ctenopteris, features that mark it as abundantly
distinct from Polypodium. The most conspicuous of these differences from
Polypodium are: rhizomes short and non-branching, paleae basally attached, axes
terete, and fronds pectinate. From Ctenopteris, diagnostic differences are: spores
neither trilete nor green, hairs never acicular from a broad base and not dark
maroon, and stipes articulate. These and other characters are detailed below.

Pecluma Price, gen. nov.
Polypodium subg. Pectinatum Lellinger, Amer. Fern J. 71:93. 1981.

eve, non ramificans nec glaucescens, radices saepe prolificas
amina

angustaque, plerumque sursumdecurrentia, marginibus non cartilagineis pilis
ascendentibus multiseptatis dispositis, infra pagina pilis multiseptatis glandulosis
appressis praedita. Venae liberae vel fortuito anastomosantes. Stomata polocytica
atque anomocytica vel copolocytica. Sori orbiculati paraphyses trichoideas
continentes. Sporae monolaetae, non chlorophyllosae.

“TYPE: Polypodium pectinatum L.

ETYMOLOGY: Pecluma is a compound of the epithets pectinatum and plumula
which have been used to exemplify the group.

DISTRIBUTION: Tropics and subtropics of the New World, from Bermuda,
Florida, and northern Mexico to southern Brazil and northern Argentina (Evans,
1969). Species about 28, generally epiphytic or epipetric.

The only previous author to accord formal taxonomic rank to Pecluma was
Lellinger (1981), who noted that it was “sharply distinct” from the rest of
Polypodium. As a genus, Pecluma is immediately recognizable and clearly defined
and circumscribed. Separation from Polypodium (sensu stricto or lato) is important
for the portrayal of the evolutionary lines involved, since Pecluma appears to have
Preserved characters that are possibly primitively simple among Polypods. Some of
these character elements, especially of rhizomes, paleae, and axes, are fundamen-
tally different from the usual condition in not only the genus Polypodium but also

i
Division of Biological Sciences and Herbarium, University of Michigan, Ann Arbor, MI 48109.

110 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

the entire subfamily Polypodioideae (the Polypods). The removal of Pecluma thus
leaves Polypodium far more uniform, definable, recognizable, and phylogenetically
meaningful. However, Pecluma does agree with Polypodium in a number of crucia
features that require microscopy for detection (characters of the spores, sporangial
stalks, and gametophytes), as well as the articulate stipes, which taken together are
my basis for maintaining it in the Polypodioideae. In the other direction, Pecluma
agrees with many members of subfamily Grammitidoideae in various characters by
which it differs from the other genera of subfamily Polypodioideae, again characters
of rhizomes, paleae, and axes, and also of frond-form and venation.

Loxogramme exhibits a mosaic of characters otherwise associated with éither the
Polypodioideae or the Grammitidoideae, and therefore is living evidence that those
two are linked evolutionarily. A subfamily Loxogrammoideae may therefore be
appropriate. I believe that Pecluma must be retained in subfamily Polypodioideae,
rather than be placed in a subfamily of its own, but I am convinced that its
numerous character states so similar to those widespread in the Grammitidoideae
cannot be attributed to evolutionary convergence or reversal, but rather indicate
shared ancestry. Pecluma, then, provides further evidence of the affinity between the
Polypod and the Grammitid ferns and strengthens the case for ranking them as
subfamilies of a greater Polypodiaceae, as apparently first done by Wagner (1961).

n the following discussion of the characteristics of Pecluma, I will not attempt to
repeat the morphological observations made in the excellent paper of Evans (1969),
except to place emphasis on certain character states. A summary of major characters
compared with Polypodium and Ctenopteris,' both confused with Pecluma in the
past, is presented in Table 1.

n Polypodium, the rhizome is wide-creeping, even rampant, regularly branching
or producing short, dormant side branches, and is often glaucous beneath the
paleae. Pecluma differs strongly in all these features, with the rhizome invariably
short, whether ascending or creeping, and even when creeping having phyllopodia
close together (exceptionally to 1 cm apart), and never regularly branching. The
absence of rhizome branches is sometimes compensated for by the production of
proliferations from the roots, already confirmed for seven species by Evans (1969,
pp. 195, 196, and 203), and one more first noted here, P. funicula. The rhizome
surface is never glaucous. All these rhizome characters, as they contrast with
Polypodium, are the general condition of Grammitid ferns, which usually have short,
non-branching, non-glaucous rhizomes, although the Malesian Ctenopters
taxodioides and the American C. moniliformis, for example, have slender, long-
creeping rhizomes. Root proliferations are known among Polypods only in the
highly-specialized Platycerium, where they should not be construed as evidence of
affinity to Pecluma. Among Grammitids, root prolifery has only been confirmed for
Adenophorus subg. Oligadenus (Bishop, 1974), perhaps first shown in the careful
illustration of A. pinnatifidus Gaud. by Brackenridge (1855, pl. 2, fig. 3.)- In the Poly-

ne . —- . si here for expedience to refer to miscellaneous pinnate oe pepe
rammitids, although its Malesian type is v ely related to the type ©
described genus Prosaptia. ee ype

M. G. PRICE: PECLUMA, A NEW FERN GENUS

Ill

TABLE 1. COMPARISON OF MAJOR CHARACTERS OF Pecluma WITH Polypodium SENSU
STRICTO AND THE PINNATE OR SUBPINNATE GRAMMITIDS (“Crenopteris”).

Structure
Rhizome

Roots
Paleae

Stipes
Axes

Lamina
segments
Hairs
Venation
Stomates
Paraphyses
Sporangial
Stalks
Spores

Chromosome
numbers

podiaceae as here circumscribed, root prolifery is also exhibite

Polypodium

peltately attached

often clathrate

sometimes with
rhizoids

often marginate

often scattered on
lamina
articul

ate
variously channeled
usually not dark

usually relatively few
often lanceolate :
occasionally
sursumcurrent
various or glabrescent

free to regularly
osing
usually polocytic +
copolocytic and/or
ocytic
scales, hairs, or

monolete
not green
c= SF

a Close ally of Loxogramme.

Basally attached paleae characterize the Gram
tous condition is peltate attachment; even among
attached paleae occur only in a few highly speci
i.e., Platycerium, a few large Pyrrosia species, an
has paleae basally attached both on the rhizomes and t
frond they are situated only along axes, never scattere
clathrate in whole or in part, and not strongly
species of Pecluma as well as Polypodium and Cten

Pecluma
short
unbranched

not glaucous
often proliferous

sometimes with
rhizoids

margin not

differentiated

only along axes or
absent from lamina

articulate

perfectly terete

dark

numerous

usually

sursumcurrent

multiseptate, dense on
rachis above, appressed
on lamina below and
glandular

free or casually
anastomosing

polocytic + anomocytic
or copolocytic

hair-like
2 or 3 cells thick

monolete
not green
x

Ctenopteris
rt

not glaucous

sometimes proliferous

basally attached

usually not clathrate

sometimes with

izoids

margin usually not
differentiated

not on lamina

usually not articulate
usually terete
often dark

various, often acicular,

broad-based, dark
m

free or casually
nastomosing
usually polocytic +

hair-like or apparently
none
uniseriate at base

trilete
chlorophyllose
x=37

d by Anarthropteris,

mitids. In Polypodium, the ubiqui-
the entire Polypodioideae, basally
alized cases unrelated to Pecluma,
d some drynarioid ferns. Pecluma
he fronds, and if borne on the
d on the laminar surface, not
differentiated at the margins. Some
opteris have rhizoids on the

surfaces of the rhizome scales which appear to be homologous with root hairs, and
SO presumably function for absorption. Marginal hairs on the paleae, when they

‘ rp arg wie
occur in any of these three, are morphologically quite different. Surficial hairs on

112 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

the paleae are also found in Tricholepidium (Ching, 1978) of Pleopeltid affinity,
where they resemble the slender, maroon setae of the Grammitids, in Leucostegia of
the Davalliaceae, in Pleurosoriopsis, where they seem to be rhizoidal, and in the
Thelypteridaceae (Holttum, 1971), where they are related to the hairs on the fronds.

Pecluma agrees with the Polypodioideae rather than the Grammitidoideae in the
important character of stipes articulate to the phyllopodia, but this is not an
exclusive feature, as Ctenopteris celebica, for example, has articulate stipes. The
stipe and rachis of Pecluma are dark and virtually perfectly terete; they do not shrink
during drying to become grooved, channeled, or differentially dimensional. No
Polypodium has this character, although it is widespread in the Grammitidoideae,
and occurs in some species of Pleopeltis of the Polypodioideae. In checking for this
character, one must be wary of several deceptive conditions. If immature, still-
developing fronds are dried before the deposition and hardening of the sclerenchy-
matous sheath responsible for the character, the axes will shrink along certain lines
and appear to be channeled, or if unusually heavy or unequal mechanical pressure is
used in drying, portions of the axes may appear slightly compressed. Laminar tissue
is long-decurrent down the stipe, forming a very narrow strip on each side, but the
actual shaft of the stipe is invariably terete.

Laminar segments (pinnae) are very numerous (over 40) and long and narrow, the
ratio of length to breadth ranging from 6:1 to 14:1, giving the fronds a marked
pectinate appearance, and although nearly the same character state occurs in a few
species of Polypodium, the situation is far more frequent in Ctenopteris. In most
species, some or all of the segments are sursumcurrent, the bases extending upwards
along the rachis more markedly than downwards. This character is more common in
Polypodium than in Ctenopteris, but occurs in both. The margins are never
cartilaginously reinforced as in many Polypods.

Fronds are always hairy with multiseptate hairs, copious on the rachis adaxially,
less so abaxially, and at intervals along the margins (except in Pecluma curvans and
P. pectinatiformis); the lamina beneath almost always has appressed, pale,
multiseptate hairs with glandular terminal cells (except P. hygrometrica, which has
erect, non-glandular hairs). The morphology and distribution of these hairs are most
similar to those of some species of Polypodium, and although a few Ctenopteris have
hairs of similar appearance and distribution, the widespread condition in the
Grammitidoideae of dark maroon, broad-based, slender, acicular hairs is never
found in Pecluma.

The combinations of stomatal types are similar to those widespread among the
Grammitids (van Cotthem, 1970) and Polypods (Sen & Hennipman, 1981). Pecluma
plumula has a combination of polocytic and copolocytic stomates, whereas the other
species I have examined (P. pectinata, P. hygrometrica, P. ptilodon, and P.
pectinatiformis) combine polocytic with anomocytic stomates.

orl are orbicular, never elongate as in some Polypodium and Ctenopter's,
surficial (slightly impressed in P. sursumcurrens), and always contain hair-like
paraphyses. Setose sporangia are common in Pecluma and Ctenopteris, but are rare
in Polypodium. Wilson (1959) pointed out that although no fundamental differences
exist between the sporangial capsules of Polypods and Grammitids, P. pectinata and

M. G. PRICE: PECLUMA, A NEW FERN GENUS 113

P. plumula both agree with the Polypodioideae in having two rows of cells at the
base of the sporangial stalk (vs. a single row at the base in the Grammitidoideae).
Spores are normally monolete and fabiform (globose in P. dispersa), finely tubercu-
late, and achlorophyllous. Spore characters are probably most consistently useful for
distinguishing between subfamilies Polypodioideae and Grammitidoideae, as excep-
tions are very rare, and in only one case is a Grammitid known to have monolete
spores (F. S. Wagner, in prep.).

Gametophyte characters agree best with Polypodium as to early appearance of the
first rhizoid, short duration of the filamentous stage, form of the mature thallus, and
distribution and types of hairs (Stokey, 1959), although gametophytes of the
apogamous Pecluma dispersa agree with the Grammitidoideae in several characters
(Evans 1969, 07).

Most similar to Pecluma is the group of species’ around Polypodium
hartwegianum Hook., as pointed out by Evans (1969, p. 217) and Smith (1981, p.
187), but all these species have the usual differences in rhizomes, paleae, and axes.
The Old World Thylacopteris, a segregate of Polypodium, is similarly pectinate, but
has a very slender, long-creeping, branched rhizome, peltate paleae, grooved axes,
and glabrescent laminae.

I am indebted to Prof. W. H. Wagner, Jr. and David M. Johnson for critical
comments and helpful discussion.

ENUMERATION OF SPECIES a

Pecluma absidata (Evans) Price, comb. nov.

Polypodium absidatum Evans, Ann. Missouri Bot. Gard. 55:238, f. 20. 1969.
Pecluma alfredii (Rosenst.) Price, comb. nov.

Polypodium alfredii Rosenst. Fedde Repert. 22:15. 1925.

Pecluma atra (Evans) Price, comb. n
Polypodium atrum Evans, Ann. Missouri Bot. Cad 55:237, f. 18. 1969.

Pecluma bermudiana (Evans) Price, comb. n
Polypodium bermudianum Evans, Ann. Missouri Bot. pied: 55:228, f. 17. 1969.
Pecluma boliviana (Rosenst.) Price, comb. nov.
Polypodium bolivianum Rosenst. Fedde Repert. 5:236. 1908.
Pecluma camptophyllaria (Fée) Price, comb. nov.
Polypodium camptophyllarium Fée, Huit. Mém. 86. 1857.
Pecluma chiapensis (Evans & Smith in Smith) Price, comb. nov
Polypodium chiapense Evans & Smith in Smith, Amer. Fern J. 70:23, f. 16-17.
Pecluma choquetangensis (Rosenst.) Price, comb. nov.
Polypodium choquetangense Rosenst. Meded. Rijks Herb. 19:18. 1913.

Pecluma consimilis (Mett.) Price, comb. n
Polypodium consimile Mett. Ann. Sci. Nat. (Paris) <i ae 1864.

"1980.

Goniophlebium, but the

“These species have often been included in Polypodium subg. an Old World cis

Goniophlebium when used as a subdivision of Polypodium is typ! ified by a
Whose relationship with American species remains to bee arified.

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

| 1 2
| ice ae -
Be top |

FIG. 1. Pecluma funicula showing a fertile frond (left) and an elongate root with a prctic
plantlets produced by prolifery (Oriente, Cuba, “creeping on trees,” von Eggers 5104,
FIG. 2. Plate 37 of Plumier’s “Description des Plantes de |’ Amerique.”

Pecluma cupreolepis (Evans) Price, comb. nov.

Polypodium cupreolepis Evans, Ann. Missouri Bot. Gard. 55: 224, £47. 1969.

This species was reduced to P. alfredii je Stolze (1981, p. 395).
Pecluma curvans (Mett.) Price, comb. n

Polypodium curvans Mett. Ann. Sci. Nat. (Paris) nts 2-253. 1864.
Pecluma dispersa (Evans) Price, comb. no

Polypodium dispersum Evans, Amer. Fern J. 58: 173; ef 27. 1968.
Pecluma eurybasis (C. Chr.) Price, comb. n

Polypodium eurybasis C. Chr. Svensk. Vet. Akad. tae IHL, 16:71, t. 16, f. 12-13, 1937.
Pecluma ferruginea (Mart. & Gal. ) Price, comb. nov.

Polypodium ferrugineum Mart. & Gal. Mém. Acad. Bruxelles 15: 36, t. 6, f. 2. 1842.
Pecluma filicula (Kaulf. ) Price, comb. noy.

Polypodium filiculum Kaulf. Enum. Fil. 275. 1824, as “filicula.”
Pecluma funicula (Fée) Price, comb. n

Polypodium funiculum Fée, Gen. Fil. 241. son ay Mém. 12, t. 8, f. 2. 1854.

Ctenopteris funicula (Fée) J. Smith, Hist. Fil. 185. 1875.

M. G. PRICE: PECLUMA, A NEW FERN GENUS 115

This Cuban species, overlooked by Evans (1969), is one of two members of the
genus Pecluma with deeply pinnatifid segments, the other being P. choquetangensis,
known from only Bolivia. Pecluma funicula differs conspicuously by the rachis +
straight (not sinuous) and the pinnae and lobes only slightly ascending (vs.
ascending at c. 45°). But the most interesting character of P. funicula is the rampant
root system (see Fig. /), freely proliferating and imitating rope-like, long-creeping
thizomes. Fée described these roots as rhizomes, and illustrated an elongate,
interwined clump of roots and fronds.

Pecluma hygrometrica (Splitg.) Price, comb. nov.
Polypodium hygrometricum Splitg. Tijdschr. Nat. Gesch. 7:409. 1840.
Pecluma paradiseae (Langsd. & Fisch.) Price, comb. nov.

Polypodium paradiseae Langsd. & Fisch. Icon. Fil, 114-18 13.
Pecluma pectinatiformis (Lindm.) Price, comb. nov.

Polypodium pectinatiforme Lindm. Hedwigia 43:309. 1904.

Pecluma pectinata (L.) Price, comb. nov.

Polypodium pectinatum L. Sp. Pl. 1085. 1753.

This species provides the type of the genus. Its lectotype, chosen by Evans (1969,
p. 246), is plate 37 of Plumier’s (1693) “Description des Plantes de 1’ Amerique”
(Fig. 2)

Pecluma plumula (Humb. & Bonpl. ex Willd.) Price, comb. nov.

Polypodium plumula Humb. & Bonpl. ex Willd. Sp. Pl. ed. 4, 5:178. 1810.
Pecluma ptilodon (Kunze) Price, comb. nov.

Polypodium ptilodon Kunze, Linnaea 9:42. 1834.

Evans (1969) divided this species into four varieties, two of which are tetraploids.
If the two tetraploids are united into a distinct entity, as they were by Love & Love
(1977), the earliest name for it is Polypodium robustum Fee.

Pecluma recurvata (Kaulf.) Price, comb. nov.

Polypodium recurvatum Kaulf. Enum. Fil. 106. 1824.

Pecluma sicca (Lindm.) Price, comb. nov.

Polypodium siccum Lindm. Ark. Bot. 1:234, t. 11, f. 4. 1903.
Pecluma singeri (de la Sota) Price, comb. nov.

Polypodium singeri de la Sota, Opera Lilloana 5:181. 1960.
Pecluma sursumcurrens (Copel.) Price, comb. nov.

Polypodium sursumcurrens Copel. Univ. Calif. Publ. Bot. 19:291. t. 42. 1941.
Pecluma truncorum (Lindm.) Price, comb. nov.

Polypodium truncorum Lindm. Hedwigia 43:309. 1904.
Pecluma venturii (de la Sota) Price, comb. nov.

Polypodium venturii de la Sota, Opera Lilloana 5:186, f. 31. 1960.

LITERATURE CITED
BISHOP, L. E. 1974. Revision of the genus Adenophorus (Grammitidaceae). Brittonia 26:217-240.
BRACKENRIDGE, W. D. 1855. U. S. Expl. Exped., Botany, Atlas. C. Sherman, Philadelphia.

116 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

CHING,.R.C. ee Tricholepidium Ching, a new genus of the Polypodiaceae in Asia. Acta Phytotax.
Geobot. 19:41—46.

COPELAND, E. B. 1956. Ctenopteris in America. Philip. J. Sci. 84:381—

COTTHEM, W. van 1970. we Be morphological study of the stomata in the Filicopsida. Bull.
Jard. Bot. Nat. Belg. 40:81—239.

EVANS, A. M. 1969. Interspecific relate in the Polypodium pectinatum-plumula complex. Ann.
Missouri Bot. Gard. 55:193—293.

wigs atts " E. 1971. Studies in the family Thelypteridaceae III]. A new system of genera in the Old

. Blumea 19:17-52.

ne ot S B. 1981. Notes on North American ferns. Amer. Fern J. 71:90—94.

LOVE, A. and D. LOVE. 1977. New combinations in ferns. Taxon 26:324—326.

SEN, U. and E. HENNIPMAN. 1981. Structure and ontogeny of stomata in Polypodiaceae. Blumea

SMITH, A. R. 1981. Flora of Chiapas, Part 2, Pteridophytes. California Academy of Sciences, San
Francisco, CA.

SOTA, E. R. de la 1963. Sobre la ubicacion oi oae de “Polypodium truncorum” Lindman
(Polypodiaceae). Bol. Soc. Argentina Bot. 10:117—

STOKEY, A. G. 1959. Polypodium pectinatum and P. a ee or Grammitidaceae?

Amer. Fern J. 49:142—
STOLZE, R. G. 1981. Ferns and Kern Allies of Guatemala. Part Il, Polypodiaceae. Fieldiana, Botany,
n. s., 6:1-522.

— e a Jr. 1961. Problems in the oo of ferns. Pp. 841-844 in Recent Advances in
y, vol. 1. Univ. of Toronto Press.
WIN. x 7 1959. The sporangia of ee problematic species of Polypodium. Amer. Fern J.
49:147-151.

AMERICAN FERN JOURNAL: VOLUME 73 NUMBER 7/1988) 117

The Lady Fern, Athyrium filix-femina, in Saskatchewan
VERNON L. HARMS*

Several years ago, Harms (1978) reported the first Saskatchewan records of the
Lady Fern, Athyrium filix-femina (L.) Roth, from the Wollaston Lake—-Reindeer Lake
region in the northeastern part of the province. These collections partially filled in
an apparent mid-continental distribution gap of over 1300 km from central Manitoba
to westernmost Alberta, between the known eastern and western ranges of A.
filix-femina s.lat. in northern North America (Figs. 1, D and 2).

During the last several years, 13 additional collections (with duplicates, including
84 individual plants) of A. filix-femina have been made from five widely separated
regions of Saskatchewan, plus one collection from northeastern Alberta. These
greatly expand the known range of the species in the province and in west-central
Canada (Figs. J and 2). The six general regions (and in parentheses the more
specific localities within each) for collections of the Lady Fern in Saskatchewan now
include the following: (1) Wollaston Lake—Reindeer Lake (Hidden Bay of Wollaston
Lake, Geikie River, Courtenay Lake, and W of Numabin Bay of Reindeer Lake,
previously reported in Harms, 1978); (2) Cypress Hills (Boiler Creek north of Loch
Leven); (3) Clearwater River (Smoothrock Falls and Descharme River); (4) Lake
Athabasca south shore (Cantara Bay and MacFarlane River); (5) Wapawekka Hills
(north slope above and below abandoned military site); and (6) the Pasquia Hills (S.
Fork White Poplar Creek and N. Fork Stony Creek). The above regions are indicated
by letter-symbols on the Saskatchewan distribution map (Fig. /), and the actual
‘collection sites by dots (to the extent the sites are far enough apart to be
distinguishable as separate dots at the scale of the base map). The new collections
are those of the author with Robert A. Wright et al., except for the Cypress Hills
collection, which was made by W. C. Harris and S. M. Lamont. The single
collection from northeastern Alberta was from along the Clearwater River 24 km
west of the Saskatchewan border (15 June 1979, Wright s. n.), and essentially fits
into our general Clearwater River region (Fig. /, B). Vouchers of all collections are
deposited in The W. P. Fraser Herbarium (SASK). Complete collecting data for
these is available upon request from the Herbarium or the author. is

The local abundance of A. filix-femina at most Saskatchewan sites was surprising,
considering that the species had not previously been reported from the province. Our
floristic studies in the Pasquia Hills and Wapawekka Hills revealed this to be one of
the more frequent ferns at higher elevations, third in abundance only to Matteuccia
struthiopteris (L.) Todaro and Dryopteris spinulosa (O. F. Muell.) Watt. Lady Ferns
appear to have been overlooked previously in Saskatchewan, possibly mistaken in the
field for the Spinulose Shield Fern, D. spinulosa, to which at least the western forms
bear a cursory resemblance. But even vegetatively, Lady Ferns may be distinguished
from the latter by their veinlets extending to tips of the pinnule teeth and their teeth
ips not being truly spinulose. In the Pasquia and Wapawekka Hills, D. spinulosa
tended to be rather more characteristic of upland coniferous woods, in contrast to

Vc
“The W. P. Fraser Herbarium, University of Saskatchewan, Saskatoon, Sask. S7N OWO, Canada.

118 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

— 58°
=
>
Z
A
of
-54°
Prince ~
Albert
F
North Battleford
ss Saskatoon a 52°
» ra
Swift Current, * * Regina 50°
49° -—_
MONT. = ! ' 1 eS DAK.
110 106° 102
FIGURE 1.

Map of Saskatchewan, Canada, showing localities for populations of A. Mia ide fe
sitchense (closed circles), of var. michauxii (open circles), and of those with intermediates a6 a ae!
var. sitchense (half-open circles). Symbols for the regions are: A =Cypress Hills Provincia ge
B=Clearwater River, C=South re of Lake Athabasca, D=Reindeer Lake—Wollaston ;
E = Wapawekka Hills, and F = Pasquia Hills.

V. L. HARMS: THE LADY FERN IN SASKATCHEWAN 119

DISTRIBUTION OF ar ee
ATHYRIUM FILIX-FEMINA : | :
s.iat. IN NORTH AMERICA QQ = va: michauxii

i aati UT =— var, asplenioides

S <“F wd Pa cama =- var. californicum

60 py > oS
LZ {2 SY ( @ -A. filix-femina sg, lat.

Gyn “gy disjunct isolates
Zips R

H Lie O -A. filix-femina sg. lat.
Soe. present Saskatche-
wan records

x NSS
ee ®
thy LA
Le,
(40 fp x \
& sia, es
Ss
: 0 500 1000
km
FIGURE 2.

FIG. 2. Map of North America showing the main ranges of A. filixfemina giana (by gest
location of previously reported disjunct isolates (by closed circles), me aes Fee stars
Saskatchewan collections (by open circles), indicating the geographical aeons sd oO ie iiaied be
to the former. Based mainly on Hultén (1964, pp. ee 155-156) Boivin
information from McGregor et al. (1977, p. 5). Scoggan (1957, p. 63, 1978, -. JJ usr
(1967, pp. 29-30), Gleason and Cronquist (1963, p. 17), and Hitchcock et al. (1907, pp. ®°—)"”-

the usual Lady Fern habitat here and elsewhere in the province aiscggtta ape
wooded or shrub-thicketed to more or less open stream sides, lake shores, an
seepage areas. : :

The Saskatchewan specimens of A. filix-femina are especially a nomen
they occur in the mid-continental distributional gap between the _ asaeeneiote and
American var. michauxii (Spreng.) Farw. [syn. subsp. angusturm ie ~yclosorum
the western var. sitchense Rupr. [syn. var. cyclosorum Rupr. and subsp. ©)

(Rupr.) C. Chr.] (Fig. 2). The distinguishing characteristics between these varieties,

120 AMERICAN FERN JOURNAL: VOLUME 73 (1983)

as compiled from various published keys and descriptions including those of Butters
(1917), Scoggan (1978, pp. 155-156), Hultén (1968, p. 48), Gleason (1952, p. 43),
Fernald (1950, p. 40-41), Boivin (1967, pp. 29-30), and Hitchcock et al. (1969,
pp. 63-64), are summarized in the following key:

1. Indusia more than | mm long, less than half as broad as long, straightish or crescent-shaped, not
crossing the vein, short-ciliate; spores light brownish, smooth to sparingly papillate; pinnules more
narrow, mostly 5 mm wide or less, slightly to only medium-toothed; laminae mostly less than 6
dm long, widest below the middle; stipes relatively longer, the laminae mostly less than 2(2.5)
times as long as the stipes; stipe base scales 7 mm long or less, mostly dark-brown, often with
blackish streaks; rhizomes horizontal to oblique-ascending, with the new frond growth appearing
lateral to the tuft var. michauxii
Indusia mostly less than | mm long, about half to as broad as long, horseshoe-shaped to
suborbicular, often crossing the vein, conspicuously long-ciliate; spores yellowish, more densely
papillate to appearing finely warty; pinnules broader, the best-developed ones over 5 mm wide,
strongly toothed or lobed to pinnatifid; laminae often more than 6 dm long, widest at or somewhat
above the middle; stipes relatively shorter, the laminae mostly more than 2.5 times as long as the
stipes; stipe base scales usually pale brown, mostly over 10 mm long; rhizomes strongly ascending
to erect, with the new frond growth appearing central in the tuft var. sitchense

Most Saskatchewan Lady Fern collections clearly belong to the western var.
sitchense, rather than to the eastern var. michauxii, despite the wider geographical
gap apparently remaining to the west than to the east of the Saskatchewan
populations. All the collections (including 70 individual plants) from the more
western Saskatchewan regions of the Cypress Hills, Clearwater River and Lake
Athabasca south shore, as well as those from the Wapawekka Hills, were consis-
tently identifiable as var. sitchense (Fig. 1, A, B, C, and E). The collections
(including 24 individual plants) from the more eastern Saskatchewan regions 0
Reindeer Lake—Wollaston Lake and the Pasquia Hills were also mostly determined as
nearest to var. sitchense (Fig. 1, D and F), but at least some specimens showed
various degrees of intermediacy to the eastern var. michauxii. The only two
Saskatchewan collections to be determined as nearest to var. michauxii were from
Courtenay Lake and from 20 km west of Numabin Bay of southern Reindeer Lake
(Ternier & Jasieniuk 1420 and 2113, respectively).

_The Lady Fern seems to display a relatively common boreal North American
distributional pattern, with major eastern and western ranges of relatively high plant
frequency, but an intervening central region where the plants are less common to
rare and sporadic, or even absent. Our recent findings of A. filix-femina at widely
spaced localities across Saskatchewan suggest it should be looked for as well across
central and eastern Alberta and in western Manitoba. The formerly apparent
mid-continental gap for the species, from western Alberta to central Manitoba, may
actually be nonexistent. It is interesting that farther south across the Great Plains a
similar series of “stepping stones” seems to be formed by disjunct isolates in
north-central Nebraska, the Black Hills in southwestern South Dakota, and Sheridan
County in northeastern Wyoming (Fig. 2). Most likely all of these apparently
disjunct isolated populations represent relicts of a formerly continuous west-east
distribution.

The author expresses his sincere appreciation to Wayne C. Harris. and Sheila M.
Lamont for their valuable contribution to this paper in first discovering and

—

V. L. HARMS: THE LADY FERN IN SASKATCHEWAN 121

collecting A. filix-femina in the Cypress Hills Provincial Park, and for subsequently
measuring the varietally diagnostic characteristics on a standing population sample
of 21 plants at the collection site. Thanks are also given to field associates, Robert
A. Wright (along on all collecting expeditions), and to John H. Hudson, Donald F.
Hooper, and Les Baker (each accompanying us on one field trip).

LITERATURE CITED

BOIVIN, B. 1967. Flora of the Prairie Provinces. Part I—Pteroids, Ferns, Conifers, and Woody
Dicopsids. Provancheria, Université Laval, Quebec City.

BUTTERS, F. K. 1917. The genus Athyrium and the North American ferns allied to Athyrium
filix-femina. Rhodora 19:170—207.

FERNALD, M. L. 1950. Gray’s Manual, 8th ed. American Book, New York.

GLEASON, H. A. 1952. The New Britton and Brown Flora of the Northeastern United States and
Adjacent Canada. Vol. 1-The Pteridophyta, Gymnospermae, and Monocotyledoneae. New
York Botanical Garden, New York.

—, and A. CRONQUIST. 1963. Manual of Vascular Plants of Northeastern United States and
Adjacent Canada. Van Nostrand, New York.

HARMS, V. L. 1978. Athyrium filix-femina new to Saskatchewan. Amer. Fern J. 68:119—120.

HITCHCOCK, C. L., A. CRONQUIST, M. OWNBEY, and J. W. THOMPSON. 1969. Vascular Plants
of the Pacific Northwest. Part I-Vascular Cryptogams, Gymnosperms, and Monocots. Univer-
sity of Washington Press, Seattle.

HULTEN, E. 1964. The Circumpolar Plants. |-Vascular Cryptogams, Conifers, Monocotyledons.
Almqvist & Wiksell, Stockholm.
. 1968. Flora of Alaska and Neighboring Territories. Stanford University Press, Stanford,
California.

McGREGOR, R. L., R. M. BARKLEY, et al. 1977. Atlas of the Flora of the Great Plains. Iowa State
University Press, Ames.

SCOGGAN, H. J. 1957. Flora of Manitoba. Bull. Natl. Mus. Canada. Ottawa, 140:i-v, 1-619.

————. 1978. The Flora of Canada. Part 2-Pteridophyta, Gymnospermae, Monocotyledoneae.
National Museums of Canada, Ottawa.

/ REVIEW

HYBRIDS IN EUROPEAN ASPLENIACEAE (PTERIDOPHYTA), by T.
Reichstein, Botanica Helevtica 91:89-139. 1981.—Two-thirds of the more than 30
species of European Aspleniaceae are as fully promiscuous as those of the Appala-
chian Asplenium complex in America, and the number of hybrids they produce iS
even greater. The species and their hybrids have been studied intensively since the
1950’s, principally by Bouharmont, Lovis, Meyer, Reichstein, Sleep, and Vida. The
present paper summarizes the accumulated knowledge of past research and presents
it in the form of annotated checklists of the species and of the hydrids. Several new
hybrids and new cytological results are described in two appendices, and many
hybrids are illustrated with line drawings. Extensive introductory material is applica-
ble to fern hybridization in general, as well as to European Asplenium—D.B.L.

122

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

AMERICAN FERN JOURNAL

Manuscripts submitted to the JOURNAL are reviewed for scientific content by
one more of the editors, and, often, by one or more outside reviewers as well.
During the past year we have received the kind assistance of D. S. Barrington, L. E.

Evans, D. R. Farrar, F. R.

em, J.D, Montgomery, Kee

sion
Taylor, to whom we are deeply indebted. We welcome suggestions of other
i

reviewers.—D.

INDEX TO VOLUME 73

perencaee subg. Oligadenus, 110; pinnatifidus, 110
Adian 8, 95; furcatum, 28; pedatum, 86, 90
Adlets hal ii and Saceu a ae 28
Alston, ; A. @.- Jem . M. sie nkin. The genus
Seapine in eae ge ‘men bey. ), 4

Apaloplebia costata, 77

As 85

pac ce 7, 14, 28, 31, 42, 88, 93, 95, 100; Hanmi
28; and 5

nigram x

subsp. trichoetises; 80: x trudellii, 3; X wherry

x Asplenosorus, 31; boydsto: 28; Xshawneensis, 31

Athyrium, 12, 16, 42; fis denies, sas 14, 62, 117, 119-121,
S osorum, 119, var. 8 aceon
Nagel 118-12

Azolla as a green manure: use od management in crop mae

(rev.),

aishya, A. K. & R. R. Rao. Ferns and fern-allies of Meghalaya
State, ning (rev.), 93
Ballard, H. E., Jr. A new combination for an Asplenosorus hybrid,

Bisporangiate a sporophylls in oe from Rajasthan, 64

Blechnum “omg i, 65, 66, 71; viviparum, 63

Blotiella, 105- 7

Bohra, D. R. (se

Ronseieria, £2; 16 hoe 12; hispida, 12; pedata, 12;
subpaleacea

Cha apman’s eget reconsidered, 39

0; subg. Botrychium, 53; boreale,
3; echo e

se, :
+ 57-59, subsp. hesperium

oss . 53, 59; minganense, 55,
; paradoxum, 53, 61;
12

pinnatum, 53, 34, 57; simplex, 53;

rownsey, P. J. Polyploidy and aneuploidy in Hypolepis and the
evolution of the Dennstaedti
Camptosorus, 28, 95; rhizophyllus. i, 3i

Campyloneurum, 73

Cardiomanes, 42

Ceratopteris dhalictndides: 12

Ceterach, 95

— osylxanthones in diploid and tissue culture- induced auto-
paenieidt nee llia fejeensis, 43

nsidered,

Cheilanthes, 95; californica, 95; eatonii, 89-92, f. eatonii, = feei,

89; fendleri, 87, 89, 92; s of osa, 95: wootonii, 87, 89, 9

i ps

Churchill, S P. e C. C. Freeman)

alge eris

Cra yi The ‘aibeus of Woodwardia areolata, 46
Crypto 5

eee 109-112: gene 112; funicula, 114, 115; monili-

eerie 1
Vittaria gametophytes discovered in a new physio-

ike re
graphic aie 33

Cyatheaceae, 65, 104, 106, 107 ; pe

Cyclophorus, 73; borneensis, 78; dispar, 78: interme MB; be
macrocarpa, 78; sect. Nipho bolus, 73; sect. Niphopsis, /:
winckler

12, 90; dickieana, 12; fragilis,

Cystopteris, "16 bulbifera,
: : _ 12; reevesiana, 12; oe

‘ow,
De avallia ens 42,
Davallia in i
Death Notice: gpa * Correll (1908-1983), 94 =
spnenciger 04-106; <— ae aoe punctilobula, F
105; ‘nies. a wilfor

Dense ee subfam. Syinaee 105, subfam.
Monachosoroideae

The stb of “abst pe — 46

Donov: orrell (1908-19

i id in
copodium staan and L, annotinum foun
xa sans: 3; x benedict :
Heer 3: celsa, 31; oe
a x marginalis, 3:

Dryopteris, 3, 77, 94, 95, 100

* margina as, a
marginalis, 3; nee sates 3. Xleedsii, 3; "x ne0 aise
a spinulosa, 3, 117, var. interme

x triploidea, 3
Edgar T. Wherry and his contributions to pteridology, |

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Elaphoglossum, i furcatum, 28

62; sci 62

Fems and allied plants, with ectel ‘reference to tropical America
(rev.), 94

a0 and fern-allies of opiate et India (rev

Ferns and fern allies of t less Area a sg Iowa,
Ginent and Wisconsin ri rev.), age

The ferns bus esi Mountain, Arizona, 85

Field gui i ferns (rev.), 96
ora Ma alesiana, series I[-Pteridophyta, volume 2, part 5, Thely-

Lee

Freeman C. C. & S. P. Churchill. Noteworthy pteridophyte records

raska, 29
Gastony, G. J. (see D. E. Soltis)
The genus — in tropical South America (rev.), 45
roi enia “
. D. Adiantum furcatum and snare furcata,
28: Mii in the xylem of Blechnum viviparum, 63
Grammitis, 9:
Greller, A. M. & D. .. ee Stone fort at Fort A oor Last
Habitat for \
ree a Island, New York?, 6
Harm Lady Fern, Athyrium filix-femina in Saskatche-

in, -
mre C. H. (see D. E. Soltis)
Henry, - Dz Spread of Marsilea quadrifolia in McDonough Coun-
ty, Illinois, 30

Haters, 195-1 107
Holttu . Flora Ma — series II-Pteridophyta, volume 2,
part 5 Th Thlyteidacese (re 42

Hybrids in Europea (Pteridophyta) (rev.), 121

Hypolepis, 104-107; altegracillima _punctata, 101
bivalvis, 98; bogotensis, 101; br , 99, 04, 10
istans, 98-100, 102, 104, 105; hispaniolica, 99; millefolium

i ens, 99, :.. : 105; pa 99, 1
1, 102, rufobarbata, 1

inct 97; repens 10

was 97, 98, 101; na viscidum, 101; viscosa, 101
“Soe
Isoétes 41

23:
fa
eS;

j » 41; ape , 12, 41; chapmanii, 39; coroman-
delina, 64; hese i o a ccida, 39-41, var. chapmanil,
39-41, var. flaccida, aris r. rigida, 41; melanopoda, 41

Jermy, A. C. (see A. H. G. Alst es

Key, J. S. Field Guide to Missouri ferns (rev.), 96

The Lady Fern, Athyrium ‘is tocnien, in percents 117
an 112

Llavea, 95

see A. M. Greller)

, 106
Lumpkin, T. A. & D. L. Plu sie Azolla as a green manure: use
: and management in crop production (rev.), 96
“asians 62; annotinum, 62; sah A , 62
si ape complanatum and L. annotinum found in the Black
ill
re tachys protostelicus, 64
i. , 85; japonicum, 12
ecg a, 42; quadrifolia, 30
atteuccia struthiopteris, 117
sgn . 107
i, in the xylem of Blechnum viviparum, 63
Microstaphyla, 28; bifurcata, 28

123

Mildella, 95

Monachosorum, 105-107; maximowiczii, 106
Nephrolepis, 71, 73; exaltata, 12

A new combination for an eager hybrid, 31
A new combination in Asplenium, 28

Sapa 73; mollis, 77

Ops

Notes on ih ecology and development of Plagiogyria fialhoi, 79

Noteworthy pteridophyte records for Nebraska, 29

Observations on the structure and function of hydathodes in
6

echnum
Onoclea sensibilis,
Ophioglossum ron 12; vulgatum var. pseudopodum, 29
Osmunda os a, 28
Paesia, 94, eo 104-107; st 98, 104
Ita, H . ee see P. M. Ric
Peck, J. H. Ferns and fern allies i the Driftless Area of Illinois,

Ow: Ot
Pecluma, 109-113, ; absidiata 3; alfred . 113; atra, ns

di:
chiapensis, 113, ¢ etange

lep! : curvans, 112, ee ; no

ybasis, 114; ferruginea, 114: filicula, 114; funicula, 110

114; hygrometrica, 112 iseae, 115 ata, 112
115 tinati s, 112, 115; plumula, 112, 115 ptilodon,
142,115; rec ita, H rod ee 1
currens, 112, 115; tru m, 115; venturii,

Pecl W al American genu: oss

Pellaea, 12, 92; andromedifolia, 12, 14, 26; atropurpurea 2:
glabella, 12; longimucronata, 90; truncata, 87, 90; wrightiana,
87,
Pereira-Noronha, M. (see we G. Windisch)

Pityrogramma, 73; triangul s, 90
Nie! ed mic Se 79-82: glauca, 83; pycnophylla, 83;

Plagiogyria
semic watt 79, 83, 84; triquetra,

Platycerium, 110, 111; andinum, 9:

Pleopeltis, 112

Pleurosoriopsis, |

Plucknett, D. L. A. Lumpkin)

Polyploidy and aneuploidy in Hypolepis, and the evolution of the
Dennstaedtial

Polypodiaceae ey 110, subfam. Grammitidoideae, 110, 112,
113, subfam aaa, 110, subfam. Polypodioideae,

110-11
Polypodium, 71, 109-113; ee 113; alfredii, 113;
angustatum, 77 , 113, aureum, 65; bermudianum, 113;

pect ctinatum,

97. 102, 105, 106; aquilinum, 12, var.

Pteris, 106,
Pyrrosia, a s
bor!

aoe . 74; a? 74, 76-78,
74, The . 74; costat r. Costatae
~ Deion

Sar 2 fs

sect. ome ptiree snes 77;
set. Moles, 7. eg took. "Th do ii, 74; dispar, 78:
tichocarpa, 78; drakean el eberhardtii, 74, 77; heteractis,

73, 77; intermedia, 78; can: ae? 4, 78; mollis,

124

TT, subg. oT 77: nummularifolia, 77; polydactilis, 74;
sect. Pyrrosia, 76; ser. Pyrrosia, 76; subg. Pyrrosia, 76;
78: samarensi 7; strigosa, 77; winckleri, 78

i
A reclassification of the fern genus Pyrrosia.

ferns new to Saag ag — aA
Reichstein, T. Hybrids in iso Aspleniaceae (rev.),

; Flora Mal
5. anger , 42; ge’ h
America, 45; Hybrids in European pia eas (Pteridophyta),
121
Richardson, P. M. & H. K. Palta. C-glycosylxanthones in diploid
4% Stare air eign ploid Davallia fejeensis, 43

Schizaea, 94

Selaginella, 8g SS; an

Sharma, B Singh & D. R. Bohra. So anomalous
eee in mes st Rajasthan

Pees H. Are classificat ation of the fe P ia, 73

Asplenium 28

— = ines B. D. Sharma)
Sone, » E., a H. Heuties, D.C, Depron. & O. J. Gertony. ~—

and electrode buffers, and aining —
Sperry, J. S. Observations on the ture and Joni of hyda-
thodes in ae aaa
Sphaerostep
Spread of a pal ap in McDonough County, Illinois, 30

AMERICAN FERN JOURNAL: VOLUME 73 (1983)

Starch gel electrophoresis of ferns: A compilation of grinding
Ts

Stone Fort at Fort Totten: Last Habitat for Woodsia o

eu platyneuron in Queens County, Long Island, New

Teens, 105

Tectaria, 95
Thence e, 42, 106, 112
fags seeatip i simulata, 52

on)

Tryon, R. M. & A. F. ‘i . Ferns and allied plants, with special
reference to tropical Amerie (rev.), 9
wo moonworts of the Roc untains: << hesperium
and a new species ebnes pear with

Vittaria gametophytes discovered in a new prin province,

Vittaria, 32, 35-37
Wagner, F. S. (see W. lan Wagner, Jr.)
gar T. Wherry and his contributions to

agner, W. in Jr.
pteridolo;
oad ah H., . & F. S. Wagner. Two moonworts of the Rocky
M shim hespesions and a new species formerly
ria wit
Windham, M. ” gc ferns | of Elden Mountain, Arizona
Windisch, p ra-Noronha. Notes on the — and
development of Paeioeves fialhoi, 79
ers toM d Delaware, 31
sia, 6, 7, 92; mexicana, 87, 91, 92: oregana, 12, 87, 91;

9
ardia areolata, 46-51; subg. Lorinseria, 46

ERRATA FOR 1982
Page 85, line 11 should read: “(Rousseau, 1974; Boivin, 1966; Hultén, 1958).

Abbe (1936) reporting on the northern range of this species in. . .

”

Page 85, line 29: For “marina” read “mariana”

BRITISH PTERIDOLOGICAL SOCIETY

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Full members receive THE FERN GAZETTE and the BULLETIN OF THE
BRITISH PTERIDOLOGICAL SOCIETY. Membership subscriptions are £7
for full members, £ 5 for ordinary members (not receiving the GAZETTE), and
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residents should apply to Dr. J. Skog, Biology Department, George Mason
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G. Coke, Robin Hill, Stinchcombe, Dursley, Gloucestershire, England.

Ce eee eee re

4 Sy” yf _.. from TRIARCH

Having completed 25 years in the management of
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AMERICAN
FERN
JOURNAL

Volume 74

Number 1

January-March, 1984

QUARTERLY JOURNAL OF THE AMERICAN FERN SOCIETY

Promotion of Apogamy in Matteuccia struthiopteris,
the Ostrich Fern

A New Filmy Fern from Puerto Rico
Chromosome Numbers of Neotropical Isoétes
Two New Phenolic Glycosides in Asplenium septentrionale
A Remarkable Cyathea Hybrid
The Ligule of Isoétes
Shorter Notes: Two Species of Adiantum Newly
caped in Florida; A New Station for
Trichomanes petersii in Alabama; Trichomanes
Gametophytes at Bartholomew’s Cobble

Reviews

‘8

. VON ADERKAS

GEORGE R. PROCTOR

R. JAMES HICKEY

FILIPPO IMPERATO

R. E. HOLTTUM

B. D. SHARMA and R. SINGH

oth NICAL

gourt BO"

The American Fern Society

Council for 1984
TERRY R. WEBSTER, Biological Sciences Group, University of Connecticut, Storrs, CT 06268.
President
FLORENCE S. WAGNER, Dept. of Botany, University of Michigan, Ann Arbor, MI 48109.
Vice-President
MICHAEL I. COUSENS, Faculty of Biology, University of West Florida, Pensacola, FL 32504
Secretary
JAMES D. CAPONETTI, Dept. of Botany, University of Tennessee, Knoxville, TN 37916.
Treasurer
LESLIE G. HICKOK, Dept. of Botany, University of Tennessee, Knoxville, TN 37916.
Records Treasurer
DAVID B. LELLINGER, Smithsonian Institution, Washington, DC 20560. Journal Editor
. SMITH, Dept. of Botany, University of California, Berkeley, CA 94720. Memoir Editor
DENNIS Wm. STEVENSON, Barnard College, Columbia University, New York, NY 10027.
Fiddlehead Forum Editor
— Aba Journal

DAVID B. LELLINGER U = Se 1 Herbarium NHB-166, berger meer
ashington, DC 2
ASSOCIATE EDITORS
DAVID W. BIERHORST Rt. 3, Box 188, Picayune, MS 39466.
GERALD J. GASTONY Dept. of Biology, Indiana University, Bloomington, IN 47401.
JOHN T. MICKEL New York Botanical Garden, Bronx, NY 10458.

TERRY R. WEBSTER ....... Biological Sciences Group, University of Connecticut, Storrs, CT 06268.

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AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984) l

Promotion of Apogamy in Matteuccia struthiopteris,
the Ostrich Fern
P. VON ADERKAS

The facultative development of sporophytes from vegetative gametophyte cells has
been recorded for many species (Steil, 1939). There are also cases in which
individual sporophyte organs such as sporangia develop directly on gametophytes
(Lang, 1898; Lawton, 1932). The appearance of isolated organs or tissues is
considered to be evidence of apomixis if the gametophytes asexually produce
sporophytes (Dépp, 1967). Tracheary tissue in gametophytes of Matteuccia
struthiopteris (L.) Todaro (Ostrich Fern) cultured over a prolonged period was not
interpreted as an apogamous phenomenon, since the gametophytes did not produce
buds or shoots.

Induction and promotion of apogamy in ferns has been investigated by Whittier
and co-workers with an emphasis on factors such as carbohydrates (Whittier &
Steeves, 1960; 1962), ethylene (Elmore & Whittier, 1975a, b), and osmotica
(Whittier, 1975). These studies of Preridium aquilinum have been concerned with
apogamous development of buds. Attempts to induce similar development in
Matteuccia are described in this note.

MATERIALS AND METHODS

Spores used in this study originated from a single fertile frond of a plant collected
by the author (no. 821) from the Five Mile River, near South Maitland, Hants
County, Nova Scotia (voucher in ACAD). Following separation from the sporangia,
spores were stored in glass vials at — 20°C until required. Spores were sterilized for
2 min in a solution of 3% “Clorox,” a commercial bleach, to which a drop of Tween
80 surfactant had been added. Spores were trapped on a Millipore membrane filter
(pore size = 8 xm) and then were washed twice with sterile, deionized water. They
were plated onto Petri plates containing Knudson’s medium supplemented with
Nitsch’s modified trace elements (Whittier, 1964), FeEDTA (1 »M), and 1% agar.
These plates were placed in growth chambers set at 23°C with light level of 67
WE-m~?-s~! and a light/dark cycle of 16:8 h. Plants were subcultured monthly,
and these cultures served as stock for the experiments. oo

Gametophytes were incubated in Erlenmeyer flasks stoppered with silicon bungs
each penetrated by a long intake and a short outlet tube. Each flask had 100 ml of
Knudson’s medium supplemented with varied amounts of either sucrose or sorbitol.
Sorbitol is very poorly metabolized by ferns, but is preferable to mannitol which
may be inhibitory to development (Hirsch, 1975). The osmolarity of the media was
increased by 0, 0.06, 0.12, 0.19 osmolal, (as measured on a Fisons Osmometer
Model 3W), which corresponded to concentrations of 0, 2, 4, and 6% sucrose, or 0,
1.1, 2.3, and 3.4% sorbitol.

* Atlantic Research Laboratory, National Research Council of Canada, 1411 Oxford St., Halifax, Nova
Scotia B3H 3Z1, Canada. N.R.C.C. no. 22710.
olume 71, number 4, of the Journal was issued December 29, 1983.

0 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

FIGS. 1-4. Sporophytic features on Matteuccia struthiopteris gametophytes raised in vitro. All bars =
mm. FIG. 1. Awl-like structure (A) arising from the anterior end of the central cushion (CC). FIG. <
Awl-like projection from meristematic tissue and two leaves, L1 from the marginal meristem and L2
from the central cushion (CC). FIG. 3. Apogamous bud (B) surrounded by numerous glandular scales
(GS). FIG. 4. Dissected juvenile leaves of apogamous sporophyte (L).

P. VON ADERKAS: PROMOTION OF APOGAMY IN MATTEUCCIA 3

FIGS. 5-6. Sporophytic features on Matteuccia struthiopteris gametophytes raised in vitro. All bars=|
mm. FIG. 5. Juvenile leaf (L) of sporophyte of sexual origin. FIG. 6. Cluster of sporangia (S) at base of
awl-like structure (A).

The flasks were connected by sterilized plastic tubing (Tygon) to a glass manifold,
and this assembly was linked to a 2] Erlenmeyer flask containing distilled water that
served as a humidifier. One set of flasks was supplied from the central air supply,
and the other from a cylinder containing | ppm ethylene in air (CanOx). Flow rates
were maintained at 10 ml-min~! with calibrated flowmeters. Gametophytes were
transferred to the flasks in a laminar flow hood. The flask assembly was transferred
to a growth chamber and raised under the same conditions as the stock cultures. The
total weight of all plants in a replicate was recorded at the beginning of the
experiment, and individual plant weights, as well as total replicate weights, were
recorded at the end of a 35-day period. There were 20 plants per treatment. The
experiment was run three times.

RESULTS

Structures that developed on gametophytes fell into three categories: The most
frequent outgrowths were awl-like projections (91.5%) from either the marginal
meristem or sub-meristematic tissue (Figs. / and 2). Less frequent (8.3%) were
reduced leaves (Fig. 2), and observed only once was an apogamous bud ( Fig. 3).
The sporophyte which developed from this bud had dissected leaves (Fig. 4) with an
abundance of ramenta along the stipe, unlike sporophytes of sexual origin (Fig. 5).

ere were noticeable intergradations between awl-like projections and reduced
leaves, but the latter were distinguished by the obvious development of a leat surface
Fig. 2). These leaves developed from either the meristem or sub-meristematic
tissue. The most extreme sporophyte reduction was a cluster of sporangia at the base
of an awl (Fig. 6). No spores were produced, the sporogenous tissue having aborted
Prior to meiosis as revealed by aceto-carmine squash of fixed material. Sporangia
removed from the plant and raised on nutrient media did not develop further.
Sporophytic organs were not restricted to plants of a particular size, as the smallest
Plants of either experiment had awl outgrowths.

4 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

TABLE 1. NUMBER OF GAMETOPHYTES (n=60) RAISED IN DIFFERENT CONCENTRATIONS
OF CARBOHYDRATE AND ETHYLENE, WHICH DEVELOPED SPOROPHYTIC STRUCTURES.

Carbohydrate Ethylene (1 ppm)
4

0% sucrose 6 6
2% sucrose 37 35
4% sucrose 41 24
6% sucrose 38 35
1.1% sorbitol 30 19
2.2% sorbitol 26 20
3.3% sorbitol aT. 13

TABLE 2. MEAN RELATIVE INCREASE (W,/W,) IN FRESH WEIGHT OF PLANTS RAISED
IN DIFFERENT CONCENTRATIONS OF CARBOHYDRATE AND ETHYLENE.

Carbohydrate Ethylene (1 ppm)

+ ae
0% sucrose 2.3 23
2% sucrose 5.6 6.5
4% sucrose 53 4.5
6% sucrose 2.8 4.8
1.1% sorbitol 2.6 3.6
2.2% sorbitol 3.0 3.0
3.3% sorbitol 2.3 mo

Apogamous development occurred most frequently on gametophytes raised on
sucrose and | ppm ethylene (Table ]). Increasing the concentration of sucrose above
2% had no effect on apogamous development. The effect of ethylene was only
marginally promotive, the more pronounced effect being attributable to sucrose.

Sorbitol promoted apogamous growth compared to controls, but at a level much
lower than that of parallel sucrose treatments. Sorbitol- and ethylene-treated plants
had a higher frequency of apogamy than those without.

Increases in plant weight were greatest overall on media supplemented with
sucrose (Table 2), and these were also the treatments which elicited the highest
apogamous response. However, treatment with sorbitol and ethylene elicited an
apogamous response equal to that of sucrose alone, even though the former had only
half the weight increase of the latter. That increase in weight of gametophyte and
apogamous response are not related is made clear from the 4% and 6% sucrose
treatments.

DISCUSSION

apogamy, in spite of their tracheary tissue. Dopp (1967) stated that the mere
presence of a sporophytic organ or tissue on gametophytes which never asexually
developed whole sporophytes was insufficient evidence of apomixis. The appearance
of tracheids in gametophytes of the otherwise non-apomictic fern Todea barbara \s a

P. VON ADERKAS: PROMOTION OF APOGAMY IN MATTEUCCIA 5

similar case (DeMaggio, 1972). However, apogamous buds of M. struthiopteris do
occur, and have also been isolated, albeit very infrequently, from maintained stock
cultures. Apomictic sporophyte development is much less frequent in this species
than in Matteuccia orientalis (Lloyd, 1973). Unlike some other ferns in which
development of apogamous structures was preceded by formation of either a special
cylindrical process (Lang, 1898) or a prothallial cushion (Whittier, 1962), in M.
struthiopteris the reduced leaves and awls developed either directly from the
marginal meristem or from tissue of the central cushion.

Promotion of apogamy has long been known to occur when a carbohydrate is
added to the substrate (Whittier & Steeves, 1962). Initially, only the nutritional
effects of carbohydrate were considered important (Whittier, 1964), but recently the
role of osmotic potential of the media in promotion of apogamy in Pteridium was
shown by Whittier (1975). For the Ostrich Fern there appears to be a threshold
response to sucrose, but much of this sucrose effect is certainly of an osmotic
nature. Similar to experiments with Pteridium in which an osmoticum replaced
some of the requirement for sucrose (Whittier, 1975), organ development on the
Ostrich Fern gametophytes occurred on up to 33% of the plants raised on sorbitol.

Nevertheless, the effects of sucrose and ethylene on apogamy differ markedly
from those of a similar study on Pteridium aquilinum (Elmore & Whittier, 1975a).
In Bracken, induction of apogamy resulted in whole plant formation, but in the
Ostrich Fern, with a notable exception, only reduced sporophytic organs developed.
The general failure to promote bud development in Matteuccia struthiopteris by the
addition of compounds promotive of apogamy in Pteridium does not imply that the
mechanisms for the two species are necessarily different. The short duration of each
experiment, when compared with the 3-5 months required to induce apogamy in
Ampelopteris prolifera (Mehra & Sulklyan, 1969) or 15 weeks for Osmunda
cinnamomea (Whittier & Steeves, 1960), may account for the low level of sporo-
phyte formation observed.

LITERATURE CITED
DeMAGGIO, A. E. 1972. Induced vascular tissue differentiation in fern

i 133:311-317. ;

DOPP, W. 1967. Apomixis bei Archegoniaten. Pp. 531-549 in W. Ruhland, ed. Encyclopedia of Plant
Physiology, vol. 18. Springer, Berlin, Heidelberg, New York.
ELMORE, H. W. and D. P. WHITTIER. 1975a. The involvement of ethylene and sucrose in the
inductive and developmental phases of apogamous bud formation in Pteridium gametophytes.
Canad. J. Bot. 53:375-381. :
, and D. P. WHITTIER. 1975b. Ethylene and carbohydate requirements fo’
induction in Pteridium gametophytes. Canad. J. Bot. 52:2089-2096. :
HIRSCH, A. 1975. The effect of sucrose on the differentiation of excised leaf tissue into either
metoph _ Pl. Physiol. 56:390-393.
LANG, W. H. 1398. On wa se acai of sporangia upon fern prothalli.
Royal Soc., B, 187-239.
LAWTON, re Re aa induced polyploidy in ferns. Amer. J : Bot. 19:303-333.
LLOYD, R. M. 1973. Facultative apomixis and polyploidy in Matteuccia orientalis. Amer. Fern. J.

gametophytes. Bot. Gaz.

r apogamous bud

Phil. Trans.

s on apogamy, apospory, and controlled

ueues
HRA, P.N. and D. S. SULKLYAN. 1969. In vitro studie y
lopteris prolifera (Retz.) Copel. Bot. J.

differentiation of rhizome segments of the fern Ampe
Linn. Soc. 62:431—443

6 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

MOTTIER, D. M. 1927. Behaviour of certain fern prothallia under prolonged cultivation. Bot. Gaz.
1244-266.

STEIL, W. N. 1939. Apogamy, apospory, and parthenogenesis in the pteridophytes. Bot. Rev.
53

WHITTIER, D. P. 1962. The origin and development of apogamous structures in the gametophyte of
Pteridium in sterile culture. Phytomorphology 12:11—20.
: . The influence of cultural conditions on the induction of apogamy in Pteridium
gametophytes. Amer. J. Bot. 51:730-—736.

1975. The influence of osmotic conditions on induced apogamy in Pteridium aquilinum.

Phytomorphology 25:246-249.

——_——., and T. A. STEEVES. 1960. The induction of apogamy in the bracken fern. Canad. J. Bot.
38:925-930.

————, and T. A. STEEVES. 1962. Further studies on induced apogamy in ferns. Canad. J. Bot.
40:1525-1531.

REVIEW

ILLUSTRATIONS OF PTERIDOPHYTES OF JAPAN, VOLUME 3, S.
Kurata and T. Nakaike, eds. x + 730 pp. + folding map. 1983. University of
Tokyo Press, yen 12,000.—This is the third in a continuing series of magnificent
volumes with detailed treatments of 100 Japanese ferns. The third closely follows
the format of the first two volumes which were reviewed in this JOURNAL 72:11.
1982 and 72:48. 1982. The unusually sturdy volumes contain line drawings, photos,
and maps for each species, plus descriptions and lists of localities in Japanese. The
nomenclature is as presented by T. Nakaike (Enum. Pterid. Japon. Filices, 1975)
where synonymies are listed. Volume 3 is mainly devoted to Diplazium, Polystichum
(continued from volume 1) and the thelypterids. Regrettably, the nomenclature of
the thelypterids was not updated to incorporate the latest publications of Prof. R. E.
Holttum. Two other minor points are that Diplazium petri Tard. was reduced to D.
latifrons v.A.v.R. by Iwatsuki & Price (South East Asian Studies 14:565. 1977), and
that Salisbury should not be the parenthetic author for Thelypteris palustris because
his name was illegitimate. Furthermore, the nomenclature of the Marsh Fern in
Japan, as well as in the United States, cannot be settled without a new taxonomic
study. Holub (Taxon 21:331-332. 1972) believes it should be called T. thelypteroides
(Michx.) Holub on the basis of earlier work by Morton, but Tryon and Tryon (Amer.
Fern J. 63:67. 1973) are skeptical.

Fern people in North America will be especially pleased with this third volume,
since it includes several other ferns besides the Marsh Fern, with which they are
familiar. These are: Polystichum braunii, P. lonchitis, P. tsus-simense (cultivated
here), Phegopteris connectilis, Thelypteris torresiana, Cyclosorus dentatus, and
Diplazium esculentum, the most delicious of all ferns, with a photo showing a whole
field of it—M. G. Price, Herbarium, North University Building, University of
Michigan, Ann Arbor, MI 48109.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984) 7

A New Filmy Fern from Puerto Rico
GEORGE R. PROCTOR*

The writer is presently engaged in field work sponsored by the Puerto Rican Dept.
of Natural Resources, which is intended to lead to the production of an up-to-date
volume on the ferns and allied plants of this island. Already, after five months of
collecting and collating other information, it appears that 45 taxa have been added to
the 320 listed by Liogier & Martorell (1982), several of them new to science. At the
end of one year, I intend to publish a complete checklist of Puerto Rican ferns in
order to summarize new additions and describe any new species or varieties, as a
preliminary to final preparation of the book.

The purpose of the present paper is partly to publicize the work being done, and
partly to provide a name for a species of Trichomanes of which fragmentary material
has already been seen from other parts of the Antilles, and which has been confused
in the past with another species known only from Jamaica.

Trichomanes (Subg. Achomanes) padronii Proctor, sp. nov. Fig. 1.
A Trichomanes pinnatifido rhizomatibus crassioribus, stipitibus longioribus, laminis
6-15 cm longis pinnato-pinnatifidis, pilis ramosis vel stellatis effusioribus differt.
Rhizome wide-creeping, finely cordlike, 0.5-0.8 mm in diameter, densely
clothed toward apex with flexuous, reddish, hairlike rhizoids ca. | mm long. Fronds
distant, 9-18 cm long, long-stipitate; stipes 3—4.5 cm long, shorter than the blades,
nonalate except at the extreme apex, loosely and deciduously clothed with branched
or stellate, pluricellular hairs up to 1.3 mm long; similar hairs also on vascular parts
and margins of the blade. Blades oblong-lanceolate, 6-15 cm long, 2.5-3.5(5) cm
broad, pinnate-pinnatifid, acuminate at the apex, subtruncate at the base; rhachis
alate (wings 0.3-0.7 mm wide on either side), pinnae ascending, alternate, 7-13 on
a side below the pinnatifid apex, 1.5—2.5 cm long, short-stalked (the stalks winged
like the rhachis), the expanded portion rhombic-acuminate, 1-1.5 cm broad below

apex subtruncate to slightly flaring; receptacle becoming long-exserted.

TYPE: South of Road 120 ca. 0.5 km ESE of Observation Tower, Maricao State
Forest, Municipio San German, Puerto Rico, elev. 850-860 m, 23 November 1983,
G. R. Proctor 39833 (US). Scandent on trunk of a tree-fern in subtropical wet
forest, serpentine substrate.

Collected with, and named for, Sr. Rubén Padron, diligent and well-informed
Biologist in Charge of the Maricao State Forest.

This species has been recorded from Hispaniola under the name Trichomanes
pinnatifidum v. d. Bosch (Christensen, 1937, P.- 10), but that species as now
understood is endemic to Jamaica. Trichomanes padronii clearly differs from T.
Pinnatifidum in its thicker, more robust rhizome, larger and more divided fronds
with much longer stipes, longer and more loosely-spreading hairs, and differently

*Dept. of Natural Resources, P.O. Box 5887, Puerta de Tierra, PR 00906.

8 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

fi
bp

=
co

FIG. 1. Portion of holotype of Trichomanes padronii (Proctor 39833, US).

located sori. A peculiar feature exists in the branching pattern of 7. padronii, which
is neither clearly anadromous (as in subg. Trichomanes and Pachychaetum) ies
catadromous (as is usually the case in subg. Achomanes). In fact, the basal divisions
of the pinnae are almost exactly opposite, as are many of the ultimate veins arising
from the costules of the segments. The species is assigned to subg. Achomanes
because of the branched and stellate marginal hairs.

LITERATURE CITED
CHRISTENSEN, C. 1936. The collection of Pteridophyta made in Hispaniola by E. L. Ekman 1917 and
1924-1930. Kungl. Svenska Vetens. Akad. Handl. III, 16(2):1-93, 1. 1-20. i.
LIOGIER, H. A. and MARTORELL, L. F. 1982. Flora of Puerto Rico and Adjacent Islands:
Systematic Synopsis. Universidad de Puerto Rico, Rio Piedras.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984) 4

Chromosome Numbers of Neotropical Isoétes
R. JAMES HICKEY*

Chromosome counts have been made for a number of /soétes found in eastern
North America, Europe, and India. Typically the haploid complement of n= 11, or
a multiple thereof, has been reported (Love et al., 1977; Kott & Britton, 1980).
Polyploid series range from 2n= 22 to 2n= 110. The aneuploid /. durieui is notable
in having the lowest chromosome number in the genus, with 2n=20. Several taxa
from India, including J. coromandelina Linn. f., 7. panchananii Pant & Srivastava,
I. indica L. Rao, and J. pantii Goswami & Arya, are reported to possess one or two
supernumerary chromosomes (Goswami, 1975; Pant & Srivastava, 1965). Only
Stylites gemmifera Rauh [=Isoétes andicola (Amstutz) L. D. Gémez] shows a
strong deviation from the presumed base number of x=11. According to Rauh and
Falk (1959), the diploid number for this species is ca. 58.

Mexico, Central America, the Antilles, and tropical South America, with over
fifty taxa, are exceptionally rich in /soétes. In addition, the morphological diversity
in these taxa is perhaps greater than in any other part of the world. Among the
neotropical species, only the count for J. andicola has been reported (Rauh & Falk,
1959). As part of an overall systematic revision of the Neotropical /soétes, the

of these counts have been previously mentioned in Tryon & Tryon (1982), at which
time I ascribed a count of 2n=44 in J. ticlioénsis H. P. Fuchs nom. nud. to I.
lechleri Mett. 7,

METHODS AND MATERIALS

The specimens of neotropical /soétes examined in this study were grown in the
alpine greenhouse of the Biological Sciences Group at The University of Connecti-
cut. All taxa, with the exception of J. andicola, were cultivated fully submerged in
glass aquaria in the Alpine Room under ambient day length. Specimens of I.
andicola were potted in rich humus constantly saturated from beneath and kept at
4°C in a cold room under a 16 hour day length. Voucher specimens are deposited in
the Gray Herbarium, Harvard University (GH), and duplicates are in the author's
herbarium.

To obtain roots for cytological study, corms were rinsed in water and all roots
removed; the plants were then grown as floating aquatics until new roots emerged
from the fossae. The roots were harvested when 1-2 cm long and were soaked for
4—6 hours in a saturated solution of paradichlorobenzene at room temperature. Roots
were harvested in the morning, as suggested by Kott and Britton (1980). Following
Pretreatment, the roots were blotted dry and fixed in Farmer’s Solution for a
minimum of 24 hours. The roots were hydrolyzed in hot IN HCl for 30 minutes,
deacidified in 95% ethanol, and stained with Whittman’s hematoxylin. The root tips
rp then destained in glacial acetic acid and squashed in Hoyer’s medium (Willey,

1).

*Box U-43, Biological Sciences Group, The University of Connecticut, Storrs, CT 06268.

10 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

ys Ys vad
¥ ; ™ .
‘= p : d

Diploid and dodecaploid species of Isoétes, x 1500. a. Isoétes storkii, 2n=22. b. Isoétes
Meslastils 2n= 22. c. Composite photo of I. novo-granadensis, 2n= 13

R. J. HICKEY: CHROMOSOME NUMBERS OF ISOETES ll

TABLE 1. NEW CHROMOSOME COUNTS OF NEOTROPICAL ISOETES
Isoétes alcalophila Halloy
2n=22: Argentina: Prov. Tucuman, Laguna Muerta, October 1980, Halloy s.n. (Fig. 1b).
Isoétes andicola (Amstutz) L. D. G6mez
2n=44: Peru; Dept. Junin, Lago Junin, April 1979, Karrfalt s.n. (Fig. 2c).
2n=44: Bolivia; Prov. Bautista Saavedra, Feuerer 59
Isoétes boliviensis U. Weber
2n=22: Bolivia; Prov. Omasuyos, Hickey 753 & Eshbaugh
Isoétes glacialis Asplund
2n=44: Bolivia; Prov. Murillo, Hickey 839 & Eshbaugh (Fig. 2b).
Isoétes herzogii U. Weber
2n=44: Bolivia; Dept. Cochabamba, Hickey 819 & Eshbaugh (Fig. 2a).
2n=44: Bolivia; Dept. Cochabamba, Hickey 821 & Eshbaugh
Isoétes novo-granadensis H. P. Fuchs
2n= 132: Ecuador; Prov. Napo, Sperling, Allgard & Roth 5189 (Fig. Ic).
Isoétes storkii T. C. Palmer
2n=22: Costa Rica; Prov. Alajuela, Volcan Pods, Churchill 3284 (Fig. 1a).
2n=22: Costa Rica; Cerro de La Muerte, Anderson 1439
Isoétes ticlioénsis H. P. Fuchs, nom. nud.
2n=44: Peru; Dept. Lima, Ticlio Pass, Hickey 840 & Eshbaugh
2n=44: Peru; Dept. Lima, Ticlio Pass, Hickey 842 & Eshbaugh (Fig. 2d).

Slides were examined with a Zeiss research microscope under bright field and
phase contrast. Photographs were taken using Kodak High-Contrast film.

RESULTS AND DISCUSSION
The recently described J. alcalophila Halloy (Halloy, 1979) from Argentina, /.

boliviensis U. Weber from Bolivia, and the Costa Rican /. storkii T. C. Palmer are
diploids, 2n=22 (Fig. la, b, Table 1). Isoétes glacialis Asplund, I. herzogii U.

Weber, and the undescribed /. ticlioénsis of H. P. Fuchs (Fuchs-Eckert, 1982), are

)
_ In 1980, L. D. Gomez transferred Stylites andicola Amstutz, the type of Stylites,
into Isoétes, and recognized the gemmiferous plants as /. andicola var. gemmifera
h G6

production is facultative, and so J. andicola should be considered a monotypic
species. Eight chromosome counts from both gemmiferous plantlets (Karrfalt mate-
tial) and young, non-gemmiferous plants (Feuerer material) yielded a diploid
number of 44. The discrepancy between these counts and the previous report of
2n=ca. 58 (Rauh & Falk, 1959) may well be the result of chromosome fragmenta-
tion, which I have found to be a common occurrence in many neotropical species.
However, the possibility of a cytologically distinct population cannot be ruled out.
My counts provide additional support for the inclusion of Stylites in Isoétes.

Two cells of J. novo-granadensis have yielded counts of 132 (Fig. Ic), and several
others have yielded counts of 126, 128, and 129. The dodecaploid J. novo-
Sranadensis has the highest chromosome count yet recorded for the genus. The

12 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

FIG. 2. Tetraploid species of Isoétes, x 1500. a. Isoétes herzogii, 2n=44. b. Isoétes glacialis,
2n= 44. c. Isoétes andicola, 2n=44. d. Isoétes ticlioensis, 2n= 44.

R J. HICKEY: CHROMOSOME NUMBERS OF ISOETES 3

The use of chromosome numbers and other cytological data promises to be of
considerable significance in understanding the evolution of Isoétes species. The
large size and variable morphology of the chromosomes (e.g., secondary constric-
tions, satellite DNA) in the neotropical species suggest that karyotype analysis might
lead to a better understanding of the evolutionary history of the genus. For example,
karyotype analysis has proven useful in the elucidation of triploid derived /soétes
aneuploids (2n = 33+ 1) in India (Kuriachan & Ninan, 1974). Such investigations
are particularly needed in /soétes, where morphological characters are notoriously
plastic.

I thank Gregory J. Anderson, The University of Connecticut, Hugh Churchill,
University of Vermont, Tassillo Feuerer of Munich, Stephan Halloy of Fundacion
Miguel Lillo, Argentina, and Calvin Sperling, Harvard University, for providing
plant material used in this study. I also thank Carl Taylor, Milwaukee Public
Museum, and Gregory J. Anderson for suggestions on cytological techniques.
Special thanks are due to W. Hardy Eshbaugh, Miami University, for assisting in the
collection of several taxa studied and for making this research possible. This study
was supported in part by NSF grants BSR 78-23389 to W. Hardy Eshbaugh and
BSR 82-07125 to G. J. Anderson.

LITERATURE CITED
FUCHS-ECKERT, H. P. 1982. Zur heutigen Kenntnis von Vorkommen und Verbreitung der
; sudamerikanischen Isoétes-Arten. Proc. Ned. Akad. Wet. C85:205-260
GOMEZ P., L. D. 1980. Vegetative reproduction in a Central American Isoétes (Isoétaceac). Its
morphological, systematic and taxonomical significance. Brenesia 18:1—14.
GOSWAMI, H. K. 1975. Chromosome studies in natural populations of Isoetes pantii, with heterosporous
sporangia. Cytologia 40:543-551.
ALLOY, S. 1979. Dos nuevos Isoétes (Lycopsida) de alta montafa, con datos ecolégicos de las
lagunas Muerta y Escondida (Cumbres Calchaquies, Tucuman, Argentina). Lilloa 35:65-95.
KOT. L. & and D. M. BRITTON, 1980, Chromosome numbers for Isoétes in northeastern North
America. Canad. J. Bot. 58:980-984. :
KURIACHAN, P. I. and C. A. NINAN. 1974. Karyomorphology of a triploid Isoetes coromendeline
A with a note on karyotype evolution in the species. New Botanist 1:160-167.
ave. A DLO R. E. G. PICHI SERMOLLI. 1977. Cytotaxonomical Atlas of the
Pteridophyta. J. Cramer, Vaduz.
BREED. D cua GK. SRIVASTAVA. 1965, Cytology ‘and reproduction oF Some Indian species of
Isoetes. Cytologia 30:239-251.

RAUH, W. and H. FALK. 1959. Stylites E. Amstutz, eine neue Isoétaceae aus den Hochanden Perus.

der Vegetationsorgane. Sitzungsber.

Heidelberger Akad. Wiss. Math.-Naturwiss. KI. 1959:3-83.
TRYON, R. M. and A. F. TRYON. 1982. Ferns and Allied Plants. Springer-Verlag, New York.

14 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984)

Two New Phenolic Glycosides in Asplenium septentrionale
FILIPPO IMPERATO*

On the basis of data from morphological and cytological analyses, Wagner (1954)
suggested the concept of reticulate evolution in the Appalachian Asplenium com-
plex, which was subsequently confirmed by Smith and Levin (1963) using chemical
analysis. It is interesting to note in this regard that the morphological characteristics
of the hybrids are generally intermediate between their parents, but that the chemical
constituents of the hybrids show a total addition of parental attributes; Appalachian
Asplenium plants clearly demonstrate additive inheritance of chemical characters.
Recently it has been shown that the chemistry of European Asplenium adiantum-
nigrum complex is analogous to that of Appalachian Asplenium complex (Richard-
son & Lorenz-Liburnau, 1982). In spite of the foregoing studies, the chemistry of
most species of Asplenium is not well known.

In the present investigation, a new kaempferol tetraglucoside (I) and a new
sulphate ester of 1-p-coumarylglucose (II) have been isolated from the fronds of A.
septentrionale; moreover two other sulphate esters of hydroxycinnamic acid-sugar
derivatives (III and IV) have been found in this plant material.

Previous work on the chemical constituents of A. septentrionale has led to the
identification of amino-acids (4-hydroxy-4-methylglutamic acid, 2-amino-4-hydroxy-
pimelic acid and the corresponding lactone, N-acetylornithine) by Berti and Bottari
(1968), keto acids (pyruvic, hydroxypyruvic, 2-oxoglutaric, 2-oxopimelic, and
4-hydroxy-2-oxo-pimelic acids) by Virtanen and Alfthan (1954), and cyanogenic
glucosides by Hegnauer (1961). Very recently Imperato (1983) reported isoquercitrin
and a new flavonoid (quercetin 3-0-(3’-sulphate) glucoside) from A. septentrionale.

MATERIALS AND METHODS

An A. septentrionale plant was collected on Mt. Etna, Italy. Fresh fronds were
homogenized and extracted with hot 95% ethanol. The extracts were concentrated to
a small volume in a vacuum and were filtered. Phenolic glycosides were isolated
by preparative paper chromatography on Whatman 3MM paper in n-butanol-acetic
acid-water (4:1:5, upper phase). The bands, revealed under ultraviolet light with
ammonia vapor, were cut off, eluted with 70% ethanol, concentrated, and rechro-
matographed in 15% acetic acid and in n-butanol-ethanol-water (4:1:2,2).

Total acid hydrolysis of isolated phenolic glycosides was carried out with 2N HCl
(1 hr at 100°C); controlled acid hydrolysis was carried out with 10% acetic acid (3.5
hr under reflux). Enzyme hydrolysis with B-glucosidase was carried out in citrate-
Phosphate buffer (pH 4.5 at 37°C for 20 hr). The aglycone:sugar ratio was
determined according to Nordstrém and Swain (1953). Aglycones were identified by
ultraviolet spectral analysis with the customary shift reagents (Mabry et al., 1970),
paper co-chromatography with authentic samples (four solvent mixtures) and poly-
amide TLC. Sugars were identified by paper co-chromatography (three solvent
mixtures) and SiO, TLC. Sulphate was identified using barium chloride. Flavonol

* Institute of Chemistry and Industrial Chemistry, University of Catania, V. le A. Doria 8,
Catania 1-95125, Italy.

F. IMPERATO: NEW GLYCOSIDES IN ASPLENIUM 15

SPECTRAL PROPERTIES AND R; VALUES OF KAEMPFEROL
3-SOPHOROTRIOSIDE-7-GLUCOSIDE

Spectral properties yoy Ry values
Shift reagent Nae Solvent mixture Ry (x 100)

265,352 BAW oF

NaOAc 263,375 BEW 48

NaOAc/H;BO; 260,3 H,0 90

AICI, 274,297 343,385 15% HOAc 74

AICI,/HCl 272 300,339,385 PhOH 51
273,385 (inc)

ZrOCl,/citric acid 263 332

BAW =n-butanol-acetic acid-water (4:1:5, upper phase), BEW =n-butanol-ethanol-water (4:1:2,2),
PhOH = phenol saturated with water, inc = shift accompanied by increase in intensity.

glycosides obtained by controlled acid hydrolysis and by enzyme hydrolysis were
identified by paper co-chromatography with authentic samples (four solvent mix-
tures), by ultraviolet spectral analysis with the customary shift reagents (Mabry et al,
1970) and by acid hydrolysis. Phenolic glycosides were methylated with methyl
iodide in dimethylformamide in the presence of silver oxide (18 hr in the dark at
room temperature under stirring), and the permethylated products were hydrolysed
by refluxing for 4 hr with 0.3 N HCl. Methylated sugars were identified by paper
co-chromatography with authentic samples (Petek, 1965) and SiO, TLC; the
partially methylated aglycones were identified by paper co-chromatography and UV
Spectroscopy.

RESULTS AND DISCUSSION

Color reactions (brown to yellow in ultraviolet + NH;), Ry values and ultraviolet
spectral characteristics (see Table 1) of the isolated flavonoid (I) were consistent
(Mabry et al., 1970) with those of a 3,7-disubstituted flavonol glycoside with free
Phenolic hydroxyl groups at positions 5 and A’. Total acid hydrolysis gave
kaempferol and D-glucose; quantitative examination gave an aglycone:sugar ratio of
1:4.1. Controlled acid hydrolysis gave kaempferol, kaempferol 7-O-glucoside,
D-glucose, sophorose (2-O-B-glucosylglucose) in small quantities, and a sugar which
On paper chromatography behaved as a trisaccharide. This sugar, isolated by
Preparative scale chromatograms, gave glucose by total acid hydrolysis, whereas by
controlled acid hydrolysis it gave glucose and sophorose; these data are consistent
with this trisaccharide being sophorotriose (O-B-glucosyl-(1 2)-O-B-glucosyl-
(1. 2)-glucose). Moreover, since this sugar was obtained from flavonoid (I) by H:O>
oxidation (Chandler & Harper, 1961), it must be linked to the 3-hydroxyl group of
kaempferol. On enzyme hydrolysis with B-glucosidase, (1) gave D-glucose,
kaempferol 3-O-sophorotrioside, and kaempferol in very small quantities. Methyla-
tion of (I) followed by acid hydrolysis gave 2,3,4,6-tetra-O-methyl-D-glucose,
3,4,6-tri-O-methyl-D-glucose, and kaempferol 5,4'-dimethyl ether. These results
show that the isolated flavonol tetraglucoside (1) must be kaempferol-3-O-sophorotri-
oside-7-O-glucoside, which is a new natural product (Fig. /).

Flavonol 3,7-diglucosides have already been found in two Asplenium species, A.
rhizophyllum (Harborne et al., 1973) and A. trichomanes (Imperato, 1979). From the

16 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

CH,OH

G. 1. Structural formula of compound (I), kaempferol-3-O-sophorotrioside-7-O-glucoside.
FIG. 2. Structural formula of compound (II), 1-p-coumarylglucose 3”-sulphate.

Systematic viewpoint, this is of interest because it has been suggested (Harborne,
1965) that flavonol 3,7-diglycosides are of restricted distribution.

Of the three hydroxycinnamic acid-sugar derivatives isolated from A. sep-
tentrionale, compound (II) has been proved to be a new natural product in the
following way. The compound (color reactions: colorless to blue in ultraviolet +
NH) was electrophoretically highly mobile (toward the anode) and strongly sugges-
tive of a sulphate derivative. On both total acid hydrolysis and controlled acid
hydrolysis, p-coumaric acid, D-glucose, and sulphate were obtained. Since the
ultraviolet spectrum (ANsC" 312 nm) showed a large bathochromic shift (50 nm) in
the presence of NaOMe, the phenolic hydroxyl group of p-coumaric acid must be
free (Harborne & Corner, 1961). Methylation followed by acid hydrolysis gave
p-methoxycinnamic acid and 2,4,6-tri-O-methyl-D-glucose. These results show that
compound (II) must be 1-p-coumarylglucose 3”-sulphate, which is a new natural
product (Fig. 2).

The two remaining phenolic glycosides (III and IV) isolated from A. septentrionale
have been characterized as sulphate esters of 1-p-coumarylglucose by the above
methods, but the position of sulphation on the glucose has not been determined
because these products could not be obtained in sufficient amount for their
characterization to be completed.

F. IMPERATO: NEW GLYCOSIDES IN ASPLENIUM 17

Sulphate esters of hydroxycinnamic acid-sugar derivatives were reported for the
first time by Cooper-Driver and Swain (1975), who found sulphate esters of
p-coumaryl- and caffeylglucose in Pteridium aquilinum and in ten Adiantum
species. Subsequently, the following compounds have been isolated from ferns:
2-O-p-coumarylglucose 6-sulphate from Asplenium fontanum var. obovatum
(Imperato, 1981), 2- and 3-sulphates of 1-caffeylglucose from Ceterach officinarum
(Imperato, 1981), and 1-p-coumarylglucose 2-sulphate and 1-caffeylgalactose
6-sulphate from Adiantum capillus-veneris (Imperato, 1982). Although sulphate
esters of hydroxycinnamic acid-sugar derivatives have been found mainly in ferns,
they have been isolated also from higher plants, €.g., from the grass Paspalum
convexum (Harborne, 1977).

The author thanks the Consiglio Nazionale delle Ricerche (Rome) for financial
support (grant CT 810164803) and Prof. H. Wagner (Institut fir Pharmazeutische
Biologie der Universitit Miinchen) for a sample of kaempferol 3,7-diglucoside.
Thanks are also due to Mr. A. D’Urso (Botanic Institute, University of Catania) for
identification of plant material.

LITERATURE CITED

BERTI, G. and F. BOTTARI. 1968. Constituents of Ferns. Jn L. Reinhold and Y. Liwschitz (eds).
Progress in Phytochemistry, vol. 1. Interscience Publishers, London, New York, Sydney.

CHANDLER, B. V. and K. A. HARPER. 1961. Identification of saccharides in anthocyanins and other
flavonoids, Aust. J. Chem. 14:586—595.

COOPER-DRIVER, G. C. and T. SWAIN. 1975. Sulphate esters of caffeyl- and p-coumarylglucose in
ferns. Phytochemistry 14:2506—2507.

HARBORNE, J. B. 1965. Characterization of flavonoid glycosides by acidic and enzymic hydrolyses.

Phytochemistry 4:107—120.

. 1977. Flavonoid sulphates. /n Reinhold, L., J. B. Harborne, a

Phytochemistry, vol. 4. Pergamon Press, Oxford.

,and J. J. CORNER. 1961. Hydroxycinnamic acid-sugar derivatives. Biochem. J. 81 242-249.

"'C. A. WILLIAMS. and D. M. SMITH. 1973. Species-specific kaempferol derivatives in
ferns of the Appalachian asplenium complex. Biochem. Syst. 1:51-54.

HEGNAUER, R. 1961. Distribution of hydrocyanic acid in cormophytes. IV. Distribution oo
sis. Pharm. Weekblad 96:577-596.

IMPERATO, F. 1979. Two new kaempferol 3,7-diglycosides and
trichomanes. Experientia 35:1134—1135

. 1981. New sulphate esters of hydroxycinnamic aci
(London) 19:691—692.

"~~: 1982. Sulphate esters of hydroxycinnamic acid-suga

veneris. Phytochemistry. 21:2717-2718.

. 1983. A new flavonol glucoside

(London) 9:390-391.

MABRY, T. J., K. R. MARKHAM, and M. B. THOMAS. 1970. The syste
flavonoids. Springer-Verlag, Berlin, Heidelberg, New York. ; see
NORDSTROM, CG. and T. SWAIN. 1953. The flavonoid glycosides of Dahlia variabih oh

General Introduction. Cyanidin, Apigenin and Luteolin glycosides from the variety “Dandy”.
bce J. Chem. Soc. 1953:2764-2773.
K, F. 1965. Chromatographies des sucres methyles. Bul
ICHARDSON, P. M. and E. LORENZ-LIBURNAU. 1982. C-glycosylxanthones
adiantum-nigrum _ Fern J. 72:103-106. a
SMITH, D. M. sary a ee A chromatographic study of reticulate evolution in the
Appalachian Asplenium complex. Amer. J. Bot. 50:952-958.

nd T. Swain (eds). Progress in

kaempferitrin in the fern Asplenium

d-sugar derivatives in ferns. Chem. Ind.

r derivatives from Adiantum capillus-

in the fern Asplenium septentrionale. Chem. Ind.

matic identification of

|. Soc. Chim. Fr. 1965:263-266.
in the Asplenium

18 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

VIRTANEN, A. I. and M. ALFTHAN. 1954. New a-keto acids in Green Plants. Acta Chem. Scand.
8:1720-1721

WAGNER, W.H.. Jr. 1954. Reticulate evolution in the Appalachian Aspleniums. Evolution 8:103-118.

REVIEW

L A MONOGRAPH OF THE FERN GENUS PLATYCERIUM (POLY-
PODIACEAE), by E. Hennipman and M. C. Roos. 1982. North Holland Publish-
ing Company, Amsterdam and Oxford, New York. $63.00.—This little 124-page
book on the staghorn ferns, Platycerium, is attractively presented on glossy paper
with numerous figures and plates, some of the latter in color. No cultivated ferns are
more spectacular than these showy plants, so familiar in conservatories the world
around. The book is divided into two sections: general and taxonomic. The former
contains the taxonomic history, habitats, morphology, phylogeny, generic relation-
ships, species concepts, and geography. Notable is the detailed character analysis for
primitive and divergent states, which emphasizes leaf structure and trichomes,
including scales, hairs, and paraphyses. The phylogeny is constructed according to
the sister group method, but the resulting graph, unfortunately, is so complicated,
the actual lines partially obscured by character bars that are too wide, that it
resembles a damaged Venetian blind. Character 18 (frond structure) is practically
chaotic, as are some of the other divergent, multistate characters in which the
Separate trends have not been sifted out. On the other hand, the geographical
analysis is especially interesting and well done.

The taxonomic section contains the generic description, keys, descriptions, and
range maps. The authors base their taxonomic conclusions on studies of dried
materials from 22 herbaria and living plants in seven botanical gardens. They reduce
the number of species from the 18 recognized by Barbara Joe Hoshizaki to 15 by
treating several of her species as infraspecific taxa. All characters considered,
Platycerium stemaria appears to be the most primitive of the present-day species,
being closest to the groundplan by having the least number of divergences. In the
taxonomic treatment, I was disappointed to see the taxa arranged alphabetically
rather than according to clusters of relationship worked out in the construction of
monophyletic groups. I agree with the authors that it is not necessary to name the
groups formally. The end of this fine book contains not only an index of collections,
but also valuable notes on collecting and preparing specimens of these ferns. All in
all this is a splendid contribution to pteridology, and I strongly recommend it to all
students of ferns, including fern horticulturists. The authors are to be congratulated.

aes H. Wagner, Jr., Department of Botany, University of Michigan, Ann Arbor, MI
8109.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984) 19

A Remarkable Cyathea Hybrid
R. E. HOLTTUM*

I recently discovered, in the Herbarium at Kew, specimens of a hybrid Cyathea
plant cultivated at the Birmingham, England Botanical Gardens in 1890 and of the
two plants stated to have been parents of the hybrid. The specimens were sent to
Kew by W. B. Latham, who was Curator of the Birmingham Gardens from 1868 to
1903. The only published reference to the hybrid known to me is in an account of
his career by Latham (1901), in which he stated that he raised the hybrid about 1870
and that the parent species were “the Mexican Cyathea insignis and the Norfolk
Island Alsophila excelsa”; he did not give a distinctive name to the hybrid. He also
referred to a Dicksonia hybrid which he raised in 1862 and which was named D.
lathamii by Thomas Moore (1885).! Thus Latham was a pioneer in fern hybridizing,
although he is not mentioned in the book by E. J. Lowe on the subject. Lowe (1895,
p. 11) recorded that when he began experiments in 1851, Thomas Moore and others
told him that hybridization of ferns was impossible; for comments on Lowe’s work
as recorded in his book, see Lovis (1967).

Although Latham accepted the nomenclature which placed in different genera the
parent plants of his Cyathea hybrid, he evidently recognized their close relationship.
They both belong to Cyathea subg. Sphaeropteris in my arrangement of Cyatheaceae
of the Old World (Holttum, 1963, 1964), where about 100 species of the subgenus
are recognized. —

In 1970, R. M. Tryon recognized Sphaeropteris as a distinct genus, including in it
not only the American species mentioned by me (1963, Pp. 124), but also several
others which in my judgement are quite unrelated. The six American species which
I regard as true Sphaeropteris were described by Tryon (1971) as “the group of
Sphaeropteris horrida.” In this group, two species are closely allied; they are named
by Tryon S. insignis (D. C. Eaton) Tryon (from the Greater Antilles) and S. horrida
(Liebm.) Tryon (from southern Mexico). Baker (1874, p. 17) united the two as
Cyathea insignis D. C. Eaton, but in cultivation the Mexican species became known
as C. princeps E. Mayer, and Latham so named the specimen of the Mexican parent
of his hybrid; see Tryon (1971) for a full synonymy. Latham’s specimen Is sterile,
but its scales indicate that it belongs either to S. insignis or to S. horrida, as named

‘Latham claimed that this was a hybrid between D. arborescens L
antarctica Labill. of Australia. The habit of the plant as described by ee
him to Kew, indicate that it is intermediate between the alleged parents piace : oie wad sae
plant, now 120 years old, is still alive (its height was lowered by air-layering #0

©xperimental work on it is now in progress; | hope a repo
Royal Botanic Gardens, Kew, Richmond, Surrey TW9 3AB, England.

20 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Latham’s specimen of the other parent shows clearly that it belongs to the species
from Norfolk Island originally named Alsophila excelsa R. Brown. When transfer-
red to Cyathea, the name must be changed to C. brownii Domin because there is a
different species named C. excelsa.

Thus there is no doubt that Latham’s two parent plants came from countries about
10,000 km apart. His hybrid is certainly intermediate in its characters of indusia and
scales; I therefore regard his claim as justified, and name his hybrid as follows:

Cyathea (Sphaeropteris) x lathamii Holttum, hybr. nov.

Int . princepem E. Mayer (Gartenflora 17:10. 1868) et C. brownii Domin
(Pteridophyta 262. 1929); a C. principe differt indusiis rudimentariis ambitu
irregulari, squamulis costarum pinnularum brevioribus; a C. brownii differt soris
indusiis rudimentariis, non squamulis discretis, cinctis, squamulis costarum
pinnularum majoribus, setis multis castaneis instructis.

TYPE: Cultivated in the Birmingham Botanical Gardens, England, 9 July 1890,
W. B. Latham (K).

In C. princeps the indusia are quite large, opaque, and of a firmer texture than
those of all other species of the subgenus except C. insignis. They completely cover
the young sori, but at maturity develop an apical aperture to form cups with
contracted rims; these cups soon develop two or three splits to the base (Tryon,
1971, figs. 7, 8). In C. brownii there are no indusia; each sorus is surrounded by
several narrow scales, as in most exindusiate species of the subgenus. In the hybrid
the base of the receptacle is surrounded by a continuous disc of irregular size and
very uneven outline.

The costal scales in C. princeps are commonly 5 mm long and nearly all have
firm, brown, marginal setae; the larger scales are wholly brown, whereas some
others are colorless but firm. In C. brownii the costal scales rarely exceed | mm
long and nearly all are thin and pallid with short, pallid, marginal hairs. The scales
of the hybrid are about 1.5 mm long and nearly all have firm, brown setae.

Schneider (1892, p. 354; 1893, p. 83) gave a contemporary report on the
cultivation of the parent species in Britain. His comment on C. brownii is that young
plants grow very quickly and are “every year sacrificed in enormous quantities for
indoor decoration.” His comment on C. insignis indicates that plants so named were
then well known in cultivation in English greenhouses; he did not distinguish
between C. insignis and C. princeps.

Among other specimens at Kew made from cultivated plants of Cyathea is one
labelled “Madeira (cult.), comm. C. L. Power, 25 April 1904.” All sori are old and
all show indusia comparable with those of Latham’s hybrid, although most are
somewhat larger. The costal scales are small, mostly pallid but with rather long,
marginal setae, colored or not. The pinnule lobes are very firm and rather deeply
lobulate, not evidently glaucous. I suggest that this represents another hybrid within
subg. Sphaeropteris, one parent being possibly C. cooperi (F. v. Muell.) Domin of
Queensland, which has become widely distributed in cultivation (some years ago it
was a weed in the fern houses at Kew).

I know of no other records of fern hybrids the parents of which are so widely
Separated geographically as those of C. x /athamii, but it is notable that such have

R. E. HOLTTUM: A REMARKABLE CYATHEA HYBRID 1

been produced in angiosperms (e.g., Eucryphia X intermedia Bausch, the parents of
which are E. glutinosa from Chile and E. lucida from Tasmania).

So far as I know, no other hybrids, either wild or cultivated, have been reported
between species of Cyathea subg. Sphaeropteris sect. Sphaeropteris Holttum (1963).
However, in sect. Schizocaena there is a probable wild hybrid between C. moluccana
R. Br. in Desv. (simply pinnate and indusiate) and C. squamulata (Blume) Copel.;
it is named C. alternans (Wall. ex Hook.) Pres] (Holttum, 1963, p. 145).

The existence of Latham’s hybrid is one more piece of evidence that in Cyathea
sensu Holttum (1963) species with indusia and species lacking indusia may be
related genetically. In his account of the genus Cyathea, as restricted by him, Tryon
(1976) described five putative hybrids between indusiate species which he includes
in Cyathea and exindusiate ones which he includes in Trichopteris. Tryon (1970, p.
15) separated these genera solely on the presence or absence of indusia; this
separation seems to me unnatural. In Holttum and Edwards (1983, pp. 163-164) I
suggested that Cyathea, Trichopteris, and part of Sphaeropteris sensu Tryon (1970)
need to be united and resubdivided in a new way; such a subdivision presents very
complex problems. Experimental hybridization might offer clues to their solution.
As Tryon remarked (1970, p. 42), hybrids between two indusiate, or two exindusiate,
species are difficult to detect from herbarium specimens and may be more abundant
than is at present apparent.

The problem of the origin of the American species of subg. Sphaeropteris
remains uncertain. As the subgenus is very much more diverse in the Old World (ca.
100 species, with rather distinct divisions named Schizocaena and Fourniera) than
in the New World, an origin in southeastern Asia is surely indicated. Tryon (1971,
Pp. 7) wrote “Sphaeropteris insignis and S. horrida [Cyathea princeps] may be
considered as the most primitive species in the [tropical American] group on the
basis of their open indusia and their crozier scales which bear wholly antrorse
teeth.” But as no Old World species has such indusia, I suggest that the condition 1s
more likely to be a local development, also the antrorse setae on the stipe scales of
the other species.

LITERATURE CITED

BAKER, J. G. 1874. Synopsis Filicum, ed. 2. Hardwicke, London.

HOLTTUM, R. E. 1963. Cyatheaceae. Flora Malesiana, II Pteridophyta, 1:65-176.
. 1964. The tree-ferns of the genus Cyathea in Australasia and the Pacific. Blumea
12:241-274. |

, and P. J. EDWARDS. 1983. The tree-ferns of Mt. Roraima and neighbouring areas of the
Guayana Highlands with comments on the family Cyatheaceae. Kew Bull. 38:155—188.

LATHAM, W. B. 1901. An account of his career. Gard. Chron. III, 29:296. :

LOVIS, J. D. 1967. Fern hybridists and fern hybridizing, I. The work of E. J. Lowe. British Fern Gaz.
9:301-307.

LOWE, E. J. 1895. Fern Growing. Nimmo, London.

MOORE, T. 1885. Dicksonia lathamii, n. hyb. Gard. Chron., 1s. 24: 584, 689, f. 154.

SCHNEIDER, G. 1892. The Book of Choice Ferns, vol. 1.Upeott Se London.

————. 1893. The Book of Choice Ferns, vol. 2. Upcott Gill, London.

TRYON, R. M., Jr. 1970. The classification of the Cyatheaceae. Contr. Gray Herb. 200:3-53.

——. 1971. The American tree ferns allied to Sphaeropteris horrida. Rhodora 73:1-19.

———.. 1976. A revision of the genus Cyathea. Contr. Gray Herb. 206:19-98.

22 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984)

The Ligule In Isoétes
B. D. SHARMA and R. SINGH*

The ligule was first illustrated in Lepidodendron by Solms-Laubach (1892).
Williamson (1894) described its structure under the name “adenoid organ.” Maslen
(1898) found the ligule to be a small triangular point in Lepidostrobus, while Seward
(1910) described the occurrence of “ligule pits” in the leaf bases of Lepidodendron
and Lepidophlois. Scott (1920) suggested size variation in the ligule, from a small,
pointed body as in Lepidostrobus to an elaborate structure, e.g. in Miadesmia.
Recently Grierson and Bonamo (1979) described the presence of a ligule in the
Middle Devonian species Leclercquia complexa. Among the living representatives
of lycopods, this structure exists in Selaginella, Isoétes, and Stylites; on this basis
these genera are sometimes grouped together as the “Ligulatae” (Sporne, 1975).

A number of functions have been assigned to the ligule. It is believed to be a
secretory or nutritive organ, or both, which by exuding water and mucilage serves to
keep the young leaves and sporangia moist (Foster & Gifford, 1959; Bierhorst, 1971;
Sporne, 1976). Michaux (1973) studied the ultrastructure of the cells of the ligule
and found that the mucilagenous secretion possessed polysaccharides. Horner et al.
(1975) believed the ligule to be a physiologically active organ which was involved in
the movement of selective substances. Goswami (1976) considered the basal portion
of the ligule (i.e., the glossopodium) to be a haustorial structure; Sporne (1976)
stated that no one knows its true function. Recently Kristen and Biedermann (1981)
and Kristen et al. (1982) studied the ultrastructure of the ligule of J. lacustris and
considered it to be a vestigial organ, which, in extinct genera, might have functioned
as a digestive organ.

The ligule is a comparatively well developed structure in Jsoétes which occurs
above the sporangium on the adaxial surface of the leaf. It has a large, embedded
portion called the glossopodium (Bierhorst, 1971; Goswami, 1976). The cells of the
glossopodium are isodiametric and distinct from the surrounding cells of the leaf
and the labium or velum. Sometimes the glossopodium is surrounded by transfusion
cells (Sharma et al., 1980). The free portion of the ligule is a thin, flat, triangular,
upward-facing structure parallel to the adaxial surface of the leaf.

MATERIALS AND METHODS

Three species of Isoétes, large I. coromandelina L. (JAC 2025), medium-sized I.
rajasthanensis Gena & Bhardwaja (JAC 2044), and small J. reticulata Gena &
Bhardwaja (JAC 2052) were studied (Gena & Bhardwaja, in press). The type
Specimens of the latter species are deposited at Government College, Ajmer, India.
Isoétes coromandelina was collected from Daosa, Jhalawar, and Atru, while /.
rajasthanensis came from Mt. Abu and J. reticulata from Atru in Rajasthan. The
material was fixed in FAA, and after passing through dehydration alcohol series was
embedded in wax. Microtome sections were cut at 8-12 jm (Johanson, 1940) and
stained with safranin and Fast green or safranin and haematoxylin. The latter
combination was better for studying the ligule.

* Department of Botany, University of Jodhpur, Jodhpur 342001, India.

SHARMA & SINGH: THE LIGULE IN ISOETES 23

FIG 2 FIG 3

GS. ~3. Reconstructions of Jsoétes ligules, X55. FIG. 1. I. coromandelina. FIG. 2. 1.
rajasthanensis. FIG. 3. I. reticulata. The abbreviations are: G = glossopodium, L=ligule.

DESCRIPTION

To study the ontogeny of the ligule, serial longitudinal and transverse microtome
sections were cut through the rhizomorphs of the three species of Jsoétes. In all
three species the ligule is fundamentally similar. In none of the species is there a
single apical cell, nor is there any plate of apical cells produced (Bhambie, 1957;
Paolillo, 1963). The superficial cells divide anticlinally, while the inner cells divide
at random. Leaf primordia appear on either side of the apex. The ligule develops
from a single initial cell and is differentiated adaxial to the leaf primordium. It
divides periclinally, and so a short filamentous structure is produced. In comparison
to the leaf, the ligule develops faster; a very young leaf contains an almost fully
developed ligule (Fig. 4). In I. coromandelina the glossopodium starts to form as
two anchor-shaped patches of parenchyma, one on either side of the leaf bundle
(Fig. 5). These are then connected by a transverse bar that is 5-7 cells thick (Fig.
6). The anchor-shaped patches also develop a proboscis-shaped structure from the
lower side of the bar (Figs. 7 and 8). The bar increases in thickness and forms a
large, pad-like structure towards the interior side (Fig. 9), while on the outer side a
large flap or whip-like portion of the ligule is produced. The pad remains embedded
in the tissue of the leaf. The glossopodium in /. coromandelina thus becomes a
Complicated structure with anchor shaped, curved side arms and a central pad-like

rtion

_ The free, flap-like portion of the ligule and the glossopodium are made up of
Similar, small, isodiametric, thin-walled cells (Fig. 10). However, the outer 3 or 4
Peripheral layers of cells of the glossopodium differ in being comparatively smaller

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

a
A

i oy a.

roy

"
mys.
s

- i
ee

Pane,
Ke

aie,

re es
a ®
* ad

1 Naa uP ha
oS <
11

FIGS. 4-12. Anatomical details of /. ake nagcseme FIG. 4. Longisection of apical portion of a
x 47. FIG. 5. Cross-section of a leaf showing origin of the
13 IG.

rhizomorph with young leaves and ligules,
glossopodium as anchor-shaped patches of parenchyma, x 13.5. Cross-section of a leaf
Showing anchor-shaped patches connected _ iG. . Cross- section of a sporophyll
showing glossopodium shape, x 13.5. FIG
Shape, x 13.5. FIG. 9. Longisection of a sporophyll showing the ligule w
upper labium is also seen, x 13.5. FIG. 10. Same, showing ligule ae different from those of the
labium, which have transfusion cells; *76,5. me, ri the non-vascularized ligule,

IG. 12. Basal portion of a labium with transfusion cells, x

SHARMA & SINGH: THE LIGULE IN ISOETES 5

(Fig. 10). The glossopodium does not receive any vascular supply (Fig. //).
However, in J. coromandelina it is surrounded by transfusion cells (Fig. 12) (Sharma
et al., 1980). In this species, the free portion of the ligule is 4.5-6 mm long. Its
epidermis is non-stomatiferous, but possesses unicellular and multicellular glandular
hairs (Fig. 13).

In J. coromandelina an upper labium covers the proximal portion of the ligule. In
the medium and small-sized species of /soétes, a distinct velum is present which
protects the sporangium (Fig. 74). In the latter species, the ligule is comparatively
smaller and the basal portion is less complicated (Figs. 14 and 1/5). The
glossopodium starts to differentiate either as a triangular structure, as in /. reticulata,
one on either side of the leaf bundle (Fig. /5) or as two circular patches of
parenchyma, as in J. rajasthenensis (Fig. 16). These structures are then connected
by a transverse bar (Figs. 17 and 18); as in I. coromandelina, an interior, pad-like
structure is produced (Fig. 19). However, the glossopodium is not surrounded by
transfusion cells.

On the basis of the present study, an attempt has been made to reconstruct the
structure of the ligule in the three species of /soétes studied (Figs. /-3). In J.
coromandelina the glossopodium is a complex, branched structure and the ligule is
quite large. With reduction in plant size, the complexity of the glossopodium and
the size of the ligule decrease and the transfusion cells found in association with the
glossopodium disappear.

DISCUSSION

Heterospory in pteridophytes originated in the lycopods sometime during the
Devonian Period (Pettitt, 1970). The ligule also appeared during the same geological
time (Grierson & Bonamo, 1979) and is known to occur in the extinct as well as
extant heterosporous lycopods. Whether its occurrence was just a coincidence or
Whether some relationship really exists between the ligule and heterospory needs
further investigation. Recent discovery of a ligule in the presumably homosporous
species Leclercquia complexa Banks (Grierson & Bonamo, 1972) increases further
the importance of this relationship. The structure of the ligule is not well preserved
in L. complexa, nor can its position on the leaf be related to any of the known
ligulate fossil or living genera. Could it be a reduced, sterile telome associated with
the fertile telome (sporangium)? In Lepidostrobus a number of megaspores were
Produced per sporangium and the ligule was a small structure, whereas Miadesmia
had a single megaspore in the sporangium but the ligule was quite elaborated (Scott,
1923; Eames, 1936; Bierhorst, 1971). In /soétes a large number of megaspores are
Produced per sporangium and the ligule is also a well developed structure. Thus
hardly any parallel can be derived between the evolution of heterospory and the
Sabon of the ligule.

_ In addition to its peculiar type of tracheary el
1S unique in having ; velum sae sporangium. In large plants the velum is absent,
but the upper labium is present (e.g., /. coromandelina, I. indica, and I. pantii),
Whereas in comparatively smaller species (e.g., /. rajasthanensis and I. reticulata) a

ements (Singh et al., 1982), [soétes

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

FIGS. 13— 19. Maa ian of Isoétes species. FIG. 13. Epidermis of J. i cnsatelieg ligule with
a glandular haris, x 180. FIG. 14. Longisection of an J. reticulata sporophyll showing ligule and
vel x27. FIG. 15. Same, _ Showing glossopodium, x54. FIGS. 16-18. Cross-sections of /.
siaschinan sporophylls Showing different stages of glossopodium development, 19.

ng 54.
“ohaonton ; n of an J. rajasthanensis sporophyll showing pad-like glossopodium and free portion of
igule, x 54.

SHARMA & SINGH: THE LIGULE IN ISOETES 7

distinct velum is present. The elaborated size of the ligule and the complexity of the
glossopodium of the larger species may be related to the presence of an upper
labium, while reduction in size of the ligule and simplification of the glossopodium
in smaller species are probably associated with the presence of a velum. However, this
conclusion needs further investigation.

The presence of glandular hairs on both the margins of the ligule favors its
secretory function, whereas the occurrence of transfusion cells surrounding the
glossopodium (Sharma et al., 1980) and mucilage with polysaccharides (Michaux,
1973) suggest its purpose is a water reservoir to protect the young leaves and
sporangia from desiccation. The present study does not favor the haustorial function
of the glossopodium, as suggested by Goswami (1976): because the glossopodium
cells are small, isodiametric, and uniform, and especially the 2 or 3 layers of cells
which occur in the interior portion of the glossopodium are smaller than the
surrounding leaf cells. Haustorium cells are generally large and irregular in shape
(Maheshwari, 1950).

e absence of a vascular supply and a cuticle in the ligule suggests that it can
neither be related to leaf nor to stem. Harvey-Gibson (1896) considered the ligule as
a specialized ramentum commonly found in pteridophytes. If this is true then the
phylogeny of such a specialized ramentum is yet to be discovered.

The authors are thankful to Prof. H. C. Arya, the Head, Department of Botany,
for providing laboratory facilities and to the U.G.C. and C.S.I.R. for financial
assistance.

LITERATURE CITED

BHAMBIE, S. 1957. The shoot apex of Isoetes coromandelina L. J. Indian Bot. Soc. 36:491-502.

BIERHORST, D. W. 1971. Morphology of Vascular Plants. Macmillan, New York.

EAMES, A. J. 1936. Morphology of Vascular Plants (Lower Groups). McGraw Hill, New York.

FOSTER, A. S. and E. M. GIFFORD, Jr, 1959. Comparative Morphology of Vascular Plants. Freeman,
San Francisco. Reprinted by Allied Pacific Private Ltd., Bomb

GENA, C. B. and T. N. BHARDWAJA. In press. Three new species
India. J. Bombay Nat. Hist. Soc.

GOSWAMI, H. K. 1976. A revision of ligule and labium in Isoétes. Acta. Soc. Bot.

ay.
of Isoetes L. from Rajasthan,
Poloniae

5:69-—76. wie ye
GRIERSON, J. D. and BONAMO, P. M. 1979. Leclerequia complexa: earliest ligulate lycopod

(Middle Devonian). Amer. J. Bot 66:474—476. :
HORNER, H. T. Jr., C. K. BELTZ, R. JAGELS and R. E. BOUDREAU, 1975. Ligule ge aga
and fine structure in two heterophyllous species of Selaginella Canad. J. Bot. 53:127-143.
JOHANSEN, D. A. 1940. Plant microtechnique. McGraw-Hill, New York. ie 4 os
KRISTEN, U. and M. BIEDERMANN. 1981. Ultrastructure, origin, and composition of the protein
bodies in the ligule of Isoetes lacustris L. Ann. Bot. 48:655-663. oe
KRISTEN, U., G. LIEBEZEIT, and M. BIEDERMANN. 1982. The ligule of Tsoctes aa
Ultrastructure, mucilage composition, and a possible pathway of secretion. Ann. Bot.
aang McGraw-Hill, New
MAHESHWARI, P. 1950. An Introduction to the Embryology of Angiosperms. S'c raw-Hill,

ork.

MASLEN, A. J. 1898. The ligule in Lepidostrobus. Ann. Bot. 12:256-259._
MICHAUX, N. 1973. La ligule chez rpostdt setacea Lam. C. R. Acad. Sci. Paris, D, engage
PAOLILLO, D. J ., Jr. 1963. The developmental anatomy of Isoetes. Illinois Biol. Monogr. 31:1-130.
PETTITT, J. 1970. Heterospory and origin of the seed habit. Biol. Rev. 45:401-415.

SCOTT, D. H. 1920. Studies in Fossil Botany, Part I, ed. 3. A. & C- Black, Ltd., London.

28 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

SEWARD, A. C. 1910. Fossil Plants, II. Cambridge Biological Series, London.

SHARMA, B. D., D. R. BOHRA, and R. SINGH. 1980. Transfusion cells in Isoetes coromandelina L.
Curr. Sci. 49:872-874.

SINGH, R., D. R. BOHRA, and B. D. SHARMA. 1982. Tracheary elements in Isoetes coromandelina
L. Bionature 2:23-25.

SOLMS-LAUBACH, H. 1892. Ueber die in den Kalksteinen des Kulm von Glatzisch-Falkenberg in
Schlesien erhaltenen structurbientenden Pflanzenreste. I. Bot. Zeit. 50:49-56, 105-113. pl. Il.

SPORNE, K. R. 1975. The Morphology of Pteridophytes, 4th ed. Hutchinson, London. Reprinted 1976
by B.I. Publications, Delhi.

WILLIAMSON, W. C. 1894. On the organization of the fossil plants of the Coal Measures, XIX. Phil.
Trans. R. Soc. London., B, 184:1-38.

REVIEW

PTERIDOPHYTIC FLORA OF GARHWAL HIMALAYA, by S. S. Bir et al.
Jugal Kishore & Co., viii + 83 pp. illustr. 1982. Rs. 95.—This is an account of the
pteridophytes in the Garhwal district of Uttar Pradesh, India (78-79° E Long,
30-31° N Lat), especially of the area within 50 km of the town of Mussoorie, an old
and ever popular collecting locality. The region mostly lies at 600-2400 m elevation
and has a tropical or subtropical monsoon climate. A total of 157 pteridophytes,
= of them widespread species, are enumerated, and others whose presence in the
region is problematical are mentioned. No new taxa are described. The generic
name Microsorum Link (1833), rather than Microsorium, is adopted following
Sledge (1960) and Pichi Sermolli (1977), although Link (1841) did correct the
original spelling to Microsorium. A map of the region, photographs of some of the
species, and an index conclude the volume, which is available from Jugal Kishore &

Co., 23-C Rajpur Road, Dehra Dun, India.—D.B.L

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984) 29

SHORTER NOTES

TWO SPECIES OF ADIANTUM NEWLY ESCAPED IN FLORIDA.—Two
previously unrecorded species of Adiantum have recently been discovered near
Homestead in southern Dade County, Florida. Both are commonly cultivated
tropical species known from single sites, so the plants almost certainly are escapes
from cultivation.

Adiantum trapeziforme L. was discovered by the late George N. Avery in February
1980. He found a single plant growing at the top of a low spoil bank in an area kept
humid by runoff from a nearby water-pumping station, and he collected a single
fertile frond (Avery 2207, FTG). The fern grows in an area shaded by the exotic
weed trees Schinus terebenthifolius and Albizzia lebbeck. Avery took us to see the
plant in late 1982; we visited the site again in January 1984. Both times we looked
for evidence of young Adiantum plants. No sporelings were ever found, although the
original plant appears to be growing well and produces spores regularly. The
Adiantum might represent a persistent plant from cuttings dumped at the site, but, as
no evidence for dumping was found, we concluded that the plant is a true escape.
The future of this species in South Florida is problematical. It grows in a relatively
protected area and is not likely to be destroyed by human activities in the near
future, but it apparently has not spread in the four years since its discovery. Whether
it is capable of becoming truly naturalized is not clear.

In March 1983 the senior author found approximately seven plants of Adiantum
anceps Maxon & Morton growing on the vertical walls of trenches cut into rocky
ground, along with Ctenitis sloanei (Poeppig ex Sprengel) Morton. The plants were
growing in the shade of Schinus terebenthifolius. Undoubtedly water collecting at
the bottom of the trenches helped maintain a humid environment. Two voucher
specimens have been collected (Herndon 702 and Avery 2501, FTG). This species
has spread within its rather specialized habitat, suggesting that the climate in South
Florida is conducive to its growth. The colony is in an area of active commercial
development and will likely be destroyed before the plants spread to more protected
areas. Still, the habitat in which the plants grow is quite similar to the “pinnacle
rock” areas found in some Dade County parks. Given the continuous supply of
Spores from cultivated plants and the availability of apparently suitable habitat in
Protected areas, it is likely that this species will sooner or later become an
established member of our naturalized flora.—Alan and Rhonda Herndon, 15301
S.W. 306th St., Homestead, FL 33033.

30 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984)

A NEW STATION FOR TRICHOMANES PETERSII IN ALABAMA.—The
filmy fern Trichomanes petersii A. Gray (Hymenophyllaceae) was first described
from Winston County, northern Alabama, and the fern occurs most abundantly in
the northern part of that state. Nevertheless, relatively few stations are known, and
the plant is considered rare and endangered by Dean (Ferns of Alabama and Fern
Allies, 1964, p. 31) and by Short (Distribution of Alabama Pteridophytes, M.S.
Thesis, Auburn University, 1978, p. 105). The entire range of 7. petersii in the
United States includes North and South Carolina, Georgia, Florida, Tennessee,
Illinois, Arkansas, Mississippi, and Louisiana. Only a single station is known in
many states.

While doing fieldwork in the vicinity of Tuscaloosa on 12 November 1983, I
discovered a dense colony of T. petersii covering a standstone boulder in a small,
north-facing ravine ca. 0.1 km south of Holt Lock and Dam on the Black Warrior
River at an elevation of ca. 65 m, in Tuscaloosa Co., west-central Alabama (Wyatt
1622, AUA, GA, NCU, UNA).

The new collection extends the range of 7. petersii approximately 100 km south
of the areas of its greatest abundance on the Sipsey Fork of the Black Warrior River.
Its occurrence in the Fall Line Hills of Tuscaloosa County on the southern edge of
the Cumberland Plateau parallels its restriction to ravines near the western (Franklin,
Lawrence, Winston, and Lamar Cos.) and eastern (DeKalb and Etowah Cos.) edges
of the Plateau. The only exception in Alabama is the collection by Correll (Amer.
Fern. J. 29:141. 1939) from the Valley and Ridge Province in Cleburne County. All
collections appear to be from sandstone of the Pennsylvanian Upper Pottsville
Formation.

The steep, protected ravines of the Holt Lake area north of Tuscaloosa harbor a
number of rare or disjunct angiosperms, including Neviusia alabamensis A. Gray,
Croton alabamensis E. A. Smith ex Chapman, Cladrastis kentuckea (Dum.-Cours.)
Rudd, and Aesculus parviflora Walter (Brooks, A Checklist of the Vascular Plants of
the Holt Lock and Dam Area, Tuscaloosa County, Alabama. M. S. Thesis, Univ. of
Alabama, 1969, p. 46). Fortunately, construction of the Holt Dam did not flood the
ravine in which T. petersii occurs, as happened at the type locality for this fern
(Dean, Ferns of Alabama and Fern Allies, 1964, p. 31) and for Leptogramma pilosa
var. alabamensis (Crawford) Wherry (Short & Freeman, Amer. Fern J. 68:1-2.
1978) on Sipsey Fork in Winston County. It is possible that 7. petersii occurs at
additional sites on the Black Warrior River north of Holt Lock and Dam and that this
very diminutive fern has simply been overlooked. In addition to the seven counties
for which herbarium records are known, attempts should be made to document
literature reports for Marion Co. by Mohr (Contr. U. S. Natl. Herb. 6:319. 1901),
Marshall Co. by Graves (Amer. Fern J. 10:78. 1920), and Walker Co. by Dean
(Ferns of Alabama and Fern Allies, 1964, p. 174).

I thank Ann Stoneburner and Ireneusz J. Odrzykoski for field assistance and
Robert R. Haynes for advice. A grant from the Whitehall Foundation helped make
this research possible.—Robert Wyatt, Department of Botany, University of Georgia,
Athens, GA 30602. ,

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 1 (1984) 31

TRICHOMANES GAMETOPHYTES AT BARTHOLOMEW’S COBBLE.
—Distribution of the little known asporophytic fern gametophytes of Vittaria and
Trichomanes in the northeastern United States was recently described by Farrar,
Parks, and McAlpin (Rhodora 85:83-91. 1983). Their study was prompted by the
discovery in 1976 by McAlpin and Farrar of Trichomanes gametophytes in Franklin
County, Massachusetts, more than 400 miles above the previously known northern-
most station (Amer. Fern J. 68:4. 1978). Both Trichomanes and Vittaria reproduce
asexually by gemmae, which develop on the gametophyte, and are not known to
develop sporophytes throughout their northeastern distribution. They are typically
found on heavily shaded, moist outcroppings of non-calcareous rock, often deep in
crevices or small caves. During the summer of 1982, I discovered a second
Massachusetts station for Trichomanes at Bartholomew’s Cobble in southern Berk-
shire County on rocks which are recognized as largely calcareous. This discovery is
of interest because of the historic nature of the site and because of the apparent
problem of the gametophytes growing on the “wrong substrate.”

Bartholomew’s Cobble was aptly described by Weatherby (Amer. Fern 12 STA.
1947) as a picturesque natural preserve located in the southwestern corner of
Massachusetts in the Housatonic River valley, several miles from Ashley Falls. Like
other cobbles in the valley, it is formed as a rocky hill projecting from the
underlying limestone. It is a National Natural Landmark protected and administered
by The Trustees of Reservations of Massachusetts and is recognized by field
naturalists for its impressive natural concentration of plants. Weatherby (Amer. Fern
J. 37:1. 1947) cited a total of 276 species of flowering plants and ferns within the 25
acre preserve; however, literature distributed at the Cobble boasts almost 500
wildflowers, 100 kinds of trees, and 49 species of ferns and fern-allies, including
somewhat elusive specimens of Scott’s Spleenwort.

During the summer of 1982, while hiking the picturesque Ledges Trail overlook-
ing the Housatonic River, I was attracted to investigate the little pockets and caves
scattered around the outcrops, which are covered with various ferns and other plants.
Having recently accompanied Dr. Donald Farrar in the field in Lancaster County,
Pennsylvania, where he was studying the distribution of Trichomanes and Vittaria, |
developed some skill in their field identification. To my surprise, I discovered the
typical little fuzzy green tufts of Trichomanes growing on the sidewalls of several of
these little “limestone” caves. These tiny plants are superficially very moss-like and
may be easily overlooked; however, with a good hand lens and a little experience,
they can be readily recognized. Farrar (pers. comm.) has indicated that neither
Trichomanes nor Vittaria have ever been reported on a calcareous substrate.

_ [was pleasantly surprised to discover that Wherry reported a somewhat similar
incongruity when he discovered acidophilic Spike-moss ( Selaginella rupestris)
growing on the same ledges of Bartholomew’s Cobble (Amer. Fern J. 41:1. 1951).
The explanation, he found, was that acid soils accumulate in the pockets formed in
the veins of milky-white quartz mingled with the limestone. It was In these very
pockets that I discovered the Trichomanes. Dr. Wherry warned against erroneous
inferences as to the soil preference of individual species on the basis of superficial

32 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

observations. I would like to think he would be pleased that another acid-loving
plant was discovered growing on this predominantly limestone cobble.

Bartholomew’s Cobble is surely an atypical site for Trichomanes. And yet, as
field naturalists become more familiar with these interesting little ferns and involved
in the search for new sites in the interest of extending the distributions recently
mapped by Farrar, et al. (Rhodora 85:83-91. 1983), we do well to note Wherry’s

warning and not overlook habitats on the assumption that they are calcareous and
therefore unfavorable to Trichomanes.—Kenneth G. Miller, Roddy Science Center,
Millersville University, Millersville, PA 17551.

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QUARTERLY JOURNAL OF THE AMERICAN FERN SOCIETY

A Western Holly Fern, Polystichum X scopulinum,

in Newfoundland WARREN H. WAGNER, JR. and ERNEST ROULEAU
Drymoglossum Under Water Stress C. S. HEW
Problems in Asplenium, with Some New Species :

from Ecuador ROBERT G. STOLZE
Frequency of Cyanogenesis in Bracken

in Relation to Shading and Winter Severity

. I. SCHREINER, D. NAFUS and D. PIMENTEL

New Combinations and Some New Names in Ferns DAVID B. LELLINGER
Gaulterio Looser (1898-1982)
Shorter Notes: Equisetum ramosissimum in Louisiana;

Three New Combinations in Loxogramme; Notes on

North American Ferns, Il; Graves’ Spleenwort

in Ohio
Reviews 36, 50,

Suggestions to Contributors

wrssoUF BOTANICAR
JUL 27 1984

GARDEN LIBRARY

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The American Fern Society

Council for 1984
TERRY R. WEBSTER, Biological Sciences Group, University of Connecticut, Storrs, CT 06268.
President
FLORENCE S. WAGNER, Dept. of Botany, University of Michigan, Ann Arbor, MI 48109.
Vice-President
MICHAEL I. COUSENS, Faculty of Biology, University of West Florida, Pensacola, FL 32504.
Secretary
JAMES D. CAPONETTI, Dept. of Botany, University of Tennessee, Knoxville, TN 37916.
Treasurer
LESLIE G. HICKOK, Dept. of Botany, University of Tennessee, Knoxville, TN 37916.
Records Treasurer
DAVID B. LELLINGER, Smithsonian Institution, Washington, DC 20560. Journal Editor
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DENNIS Wm. STEVENSON, Barnard College, Columbia University, New York, NY 10027.
Fiddlehead Forum Editor
American Fern Journal
EDITOR

DAVID B. LELLINGER U.S. Nat’] Herbarium NHB-166, Smithsonian Institution,
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AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984) 33

A Western Holly Fern, Polystichum x scopulinum,
in Newfoundland
WARREN H. WAGNER, JR.* and ERNEST ROULEAU**

The holly ferns of North America have recently received considerable study (W.
Wagner, 1973; D. Wagner, 1979), which has resulted in a number of changes in their
taxonomic interpretation. In the west, 11 sexual taxa are recognized, in the east only
four. Three of the latter are northern, occurring mainly in Canada. The fourth, the
familiar Christmas Fern, P. acrostichoides (L.) Schott, with distinctive semi-
dimorphic leaves, is southern in distribution and is by far the most abundant and
widespread member of the genus. The rarest Holly Fern in the east is the mainly
western Hybrid Holly Fern P. Xscopulinum D. C. Eaton (fertile tetraploid form),
known previously from a single mountain on the Gaspé Peninsula of Quebec. This
disjunct has attracted much attention because it is separated by over 2000 miles
(3200 km) from its range in the western United States and southwestern Canada.
Fernald (1924) considered it a relict which had survived the Ice Age on an
unglaciated mountain or “nunatak.”

Fernald (1924, p. 89) considered P. Xscopulinum a northern variety of the
southern hemisphere P. mohrioides (Bory) Presl, stating that “this anomalous plant
[var. scopulinum (D. C. Eaton) Fern.] ... has been treated as a variety of
Polystichum aculeatum or of P. lonchitis, as P. mohrioides and as a distinct species
standing midway between P. aculeatum and P. mohrioides.” W. Wagner (1973)
found that P. scopulinum is actually a hybrid, with one parent P. mohrioides s. 1.
and the other P. munitum (Kaulf.) Presl s. /.' The hybrid origin of P. Xscopulinum
was deduced from a number of data which show that it Is an allopolyploid
intermediate between the parents. In expeditions with A. R. Kruckeberg in serpen-
tine areas of Washington state backcrosses of P. Xscopulinum with its parents and
also primary sterile diploid hybrids were found. Thus, P. Xscopulinum occurs both
in a sterile diploid form and a fertile tetraploid form, the latter having the ability to
disperse and reproduce sexually.

The occurrence of P. Xscopulinum is of special interest because the polyploid
hybrid occurs so far from its parents. Similarly, Wright's Cliff-brake, Pellaea
Xwrightiana Hook., the allopolyploid hybrid of P. longimucronata Hook. and P.
ternifolia (Cav.) Link, is known from two stations in North Carolina some 1000
miles (1600 km) east of its metropolis in the southwestern United States (Wagner,
1965, 1972). i,

Polystichum Xx scopulinum occurs on Mt. Albert in the Shickshock Mountains of
the Gaspé Peninsula, where it has been collected many times. The colonies grow on
serpentine cliffs and rock slides in ravines at altitudes of 2000-3000 ft. At a

‘David Wagner (1979) interpreted the North American P. mohrioides as distinct > Sen

America and treated it as P. lemmonii Underw. The variety of P. munitum known es — Est

imbricans D. C. Eaton he considered to be a distinct species. P. imbricans (D. C. Eaton) D. Wagner.

He thus give the hybrid formula as P. imbricans x lemmonit.

Herbarium and Department of Botany, University of Michigan, Ann Arbor, MI ge H1X 2B2

eect Marie-Victorin, Institut Botanique, 4101 Est, Rue Sherbrooke, Montreal, Que. 2B2,
ada,

including new record from Newfoundla

ons
wetwarin
: >

\ hr

a 7

Polystichum Xscopulinum, southerly slopes of dry serpentine ridge, near ee head of North Arm, Newfoundland, Rouleau
own range of Polystichum & scopulinum,

S pen “wl:
oe te

884 (MT). FIG.

(p861) 2 SWMOA ‘TWNYNOL NY34 NVOINSWY

WAGNER & ROULEAU: WESTERN HOLLY FERN IN NEWFOUNDLAND 35

distance, the plant may be confused with P. lonchitis (L.) Roth, a common northern
species, and thus may be overlooked in routine collecting. It may also superficially
resemble exposed or dwarfed forms of P. braunii (Spenner) Fée, a fern with
bipinnate, thinner-textured, and scalier fronds. These three species are contrasted in
Fernald’s key (1950, p. 37).

Rouleau found P. Xscopulinum for the first time in Newfoundland on 18 July
1950 while exploring the southerly slopes of a dry serpentine ridge near the head of
North Arm in the Humber District (Rouleau 884, MT). The plant resembled a small,
fertile P. braunii (Fig. 1), and its true identity was not recognized until March 1978.
Rouleau has found both P. braunii and P. lonchitis to be common and widespread in
Newfoundland, but thus far he has collected P. x scopulinum only at this single site.
This discovery extends the known range of P. X scopulinum nearly 400 miles (560
km) northeast of the colony on Mt. Albert in Quebec. Although fully fertile, the
herbarium specimen is smaller than average for this species. The stipes are unusually
long, nearly equal in length to the blades, presumably as a result of growing in a
deep pocket. The pinnae of the lower half of the blade are strongly auricled at the
anterior base, and some of the auricles are contracted basally to a stalk.

A distribution map (Fig. 2) of P. x scopulinum based on records from W. Wagner
(1973), D. Wagner (1979), Taylor (1970), and the Newfoundland specimen shows
not only the vast disjunction between west and east and the far greater number of
localities in the west, but also the extension of the taxon in the west to much lower
latitudes.

Damman (1965), in his review of distribution patterns in the flora of Newfound-
land, recognized a category of species that are restricted to serpentine areas, to
which P. x scopulinum must now be added. He described the habitats as bleak and
exposed with sparse vegetation. He believed that serpentine species occur on these
sites because of a “lack of competition, the basic soils, and their ability to tolerate
frost churning.” Damman discussed Fernald’s “Nunatak Theory” and concluded
from reports by a number of workers that Fernald’s nunatak areas had been covered
with ice at some time or other during the glacial period. :

Rather than assume that P. Xscopulinum persists as a relict ona Newfoundland
nunatak, it may be more reasonable to attribute the occurrence of P. x scopulinum
to long distance dispersal by spores. Bouchard et al. (1977) recently gave another
striking example of probable long distance dispersal in Newfoundland: Thelypterts
limbosperma (Allioni) H. P. Fuchs [syn. Oreopteris limbosperma (Allion!) Holub]
was unknown in eastern North America until 1973, when a large colony was found
in Gros Morne National Park. The species is known to occur otherwise in North
America only from Alaska to central Washington.

In view of the number of examples now known 0
Canada and the United States and the evidence that single spores have the
potentiality for establishing new colonies (Wagner, 1972), the presence of P.
*scopulinum in the east seems no longer as exceptional as It was once thought he

. The existence of this fern in Newfoundland several hundred miles from wes
nearest other colony could have resulted from a wind-blown spore from as close as
the Gaspé Peninsula or as far as the west coast of North America.

f west to east disjunctions in

36 AMERICAN FERN JOURNAL: VOLUME 74 (1984)
We thank André Bouchard, A. W. H. Damman, David H. Wagner, and Florence
S. Wagner for help in various ways.

LITERATURE CITED

BOUCHARD, A., D. BARABE, and S. HAY. 1977. An isolated colony of Oreopteris limbosperma
(All.) Holub in Gros Morne National Park, Newfoundland, Canada. Naturaliste Can.

104:239-244.

DAMMAN, A. W. H. 1965. The distribution patterns of northern and southern elements in the flora of
Newfoundland. Rhodora 67:363-392.

FERNALD, M. L. 1924. Polystichum mohrioides and some other subantarctic or Andean plants in the
northern hemisphere. Rhodora 26:89-95.

————.. 1950. Gray’s Manual of Botany, 8th ed. American Book, New York.

TAYLOR, T. M. C. 1970. Pacific Northwest Ferns and their Allies. Univ. of Toronto Press, Toronto
and Buffalo.

WAGNER, D. H. 1979. Systematics of Polystichum in western North America north of Mexico.
Pteridologia 1:1—64.

WAGNER, W. H., Jr. 1965. Pellaea wrightiana in North Carolina and the question of its origin. J.
Elisha Mitchell Sci. Soc. 81:95-103.
. 1972. Disjunctions in homosporous vascular plants. Ann. Missouri Bot. Gard. 59:203-217.
. 1973. Reticulation of holly ferns (Polystichum) in the western United States and adjacent
Canada. Amer. Fern J. 63:99-115

REVIEW

FERNS AND FERN ALLIES OF GUATEMALA, PART III. MARSILEACEAE,
SALVINIACEAE, AND THE FERN ALLIES (INCLUDING A COMPRE-
HENSIVE INDEX TO PARTS I, II, AND IID, by Robert G. Stolze. Fieldiana:
Botany, NS, 12. 91 pp. 1983.—This is the final part of the Ferns and Fern Allies of
Guatemala, and includes the heterosporous ferns and fern allies—eight genera in
all—plus an index to all parts. Lycopodium is treated by B. @llgaard and Isoétes
with the help of R. J. Hickey. The third part follows the first two in format (Part I
reviewed in Amer. Fern J. 67:94, Part II in 72:14), and the clear, concise style is
maintained here also. I found no errors of consequence. The keys seem well
constructed with contrasting statements in each pair of leads. I tried several species
of Lycopodium and found no problems in use.
A nice feature is the discussion within the generic treatments of excluded species
In Marsilea, Azolla, and Salvinia which have not been found but could be expected
in Guatemala, and how these could be distinguished from included taxa. Stolze also
points out the need for monographic work in these genera and the difficulties in
taxonomy and identification when much (most in Azolla) material is vegetative only.
The author is to be commended on the completion of this important work on
tropical American pteridophytes. Stolze’s Ferns and Fern Allies of Guatemala and
Man R. Smith’s Flora of Chiapas—Pteridophytes have made an excellent start on
filling in our knowledge of the ferns on this region.—James D. Montgomery,
Ichthyological Associates, Inc., Berwick, PA 18603.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984) 37

Drymoglossum Under Water Stress
C. §, Hew?

Drymoglossum is an epiphytic fern found commonly on the trunks of some local
trees. It exhibits typical CAM features (Hew & Wong, 1974; Wong & Hew, 1976).
It has fleshy fronds for storing water, but it is not uncommon to observe shrivelling
of its fronds during dry spells.

Different CAM plants seem to respond to drought differently. Under water stress,
CAM plants such as Tillandsia increase their CAM activities, but plants such as
Opuntia reduce their CAM activity instead (Kluge & Ting, 1978). We have reported
recently that tropical succulent orchids behave just like Opuntia when placed under
water stress (Fu & Hew, 1982).

240Fr
= 200
D
a,
D
g
= 160 -
se, e
3)
Pink A Control
© 120 A * -10bars
3 % ® -i5bars
S
2 =e Oo -18bars
e
40
x

0
0800 3200 1600 2000 « e..0400 0200 1200

Time (hours )
FIG. 1. Effect of water stress on diurnal fluctuation of titratable acidity in Drymoglossum fronds after
imposing two days of water stress.
s in amino acid metabolism are aspects

Inhibition of protein synthesis and change olist
been extensively studied in relation to

of metabolic changes in plants which have :
water stress (Hsiao, 1973). Accumulation of proline concentration was the main
reason for the increase of free amino acids (Singh et a 1973a, b). Proline
accumulation is a result of a stimulation of its synthesis and an inhibition of its
utilization (Stewart, 1980). Works on proline accumulation under water stress have

* as 5 OADOES. > ic of Singapore.
Department of Botany, National University of Singapore, Singapore 0511, Republic of Singapore

38 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

> 80
<
= 160} 2
ad x
2 560 f
a A ret
3 120+ Cg Control g
so -10 bars c
3 - nee
< 80F . -15bars o x
£ £
3 me S
w Q20F-
= 40+ ~
—18 bars
i i J f- 1 Ll i.
2 0 -10 -12 6 =18
No.of Days under Stress PEG concentration (bars )

FIG. 2. Effect of length of time of water stress on titratable acidity in Drymoglossum fronds. FIG.
3. Proline accumulation in Drymoglossum fronds under various PEG concentrations 56 hrs after
imposition of water stress.

been carried out with Chlorella (Greenway & Setter, 1979), barley (Hanson et al.,
1977; Singh et al., 1973a, b; Tully et al., 1979), Bermuda grass (Barnett & Naylor,
1966), halophytes (Cavalieri & Huang, 1979), spinach (Huang & Cavalieri, 1979),
and tobacco (Boggess & Stewart, 1980). This paper reports the effect of water stress
on CAM activity and proline accumulation in Drymoglossum.

Sterile, mature fronds of Drymoglossum piloselloides (L.) Presl were collected
from Acacia tree trunks. Eight uniform-sized fronds were placed in each petri dish
with filter paper serving as substratum. These were kept in a temperature-controlled
room (26° C) with a 12 hr photoperiod and at a light intensity of 1200 ft-c. The
light source was a 700-watt HPLR mercury lamp.

Water stress was imposed by placing the fronds in various concentrations of
polyethylene glycol (PEG) 1000, giving a range of osmotic potential from about
~ 10 bars to — 18 bars. The control was kept in distilled water. All treatments were
done in triplicate.

Titratable acidity was measured as described by Wong and Hew (1976). The
extraction and determination of proline were done according to the method de-
scribed by Singh et al. (1973a) and by Troll and Lindsley (1955). Unless stated
otherwise, measurement of titratable acidity was done at 0800 hr.

Both controls and stressed plants displayed pronounced diurnal fluctuation of
titratable acidity (Fig. /). Pronounced decrease in titratable acidity was observed 2
days after water stress. Titratable acidity decreased with a corresponding increase un
PEG concentration and with days of water stress (Figs. | & 2). The decrease In
titratable acidity in Drymoglossum under stress, therefore, resembled that of tropical
orchids (Fu & Hew, 1982). The drop in titratable acidity in water-stressed

C. §. HEW: DRYMOGLOSSUM UNDER WATER STRESS 39

Drymoglossum could also be attributed to a decrease in CO) fixation, as has been
observed in orchids (Fu & Hew, 1982) and in other plants (Osmond, 1978

Proline accumulation increased in fronds of Drymoglossum following water stress
(Fig. 3). This is in agreement with previous reports obtained for barley, grass,
wheat, spinach, and tobacco. Our findings support the generalization that most if
not all plants have the ability to accumulate proline, provided that the imposed water
stress is severe enough (Hsiao, 1973).

Proline accumulation in succulent halophytes under stress has been reported
(Cavalieri & Huang, 1979), but it is yet to be shown that the succulent halophytes
exhibit CAM features (Webbs & Burley, 1965). We now have evidence to indicate
that under PEG stress, CAM plants such as Drymoglossum start to accumulate
proline around — 10 to —15 bars. The same has also been observed for succulent
tropical orchids, also CAM plants (Hew, unpublished data).

LITERATURE CITED

BARNETT, N. M. and A. W. NAYLOR. 1966. Amino acid and protein metabolism in Bermuda grass
during water stress. Pl. Physiol. 41:1222-1230.

BOGGESS, S. F. and C. R. STEWART. 1980. The relationship between water stress induced proline
accumulation and inhibition of protein synthesis in tobacco leaves. Pl. Sci. Letter. 17:245-252.

CAVALIERI, A. H. and A. H. C. HUANG. 1979. Evaluation of proline accumulation in the adaptation
of diverse species of marsh halophytes to the saline environment. Amer. J. Bot. 66:307-312.

FU, C. F. and C. S. HEW. 1982. Crassulacean acid metabolism in orchids under water stress. Bot. Gaz.
143:294—297.
GREENWAY, H. and T. L. SETTER. 1979. Accumulation of proline and sucrose during the first hour
after transfer of Chlorella emersonii to high NaCl. Austral. J. Pl. Physiol. 6:69-79.
HANSON, A. D., C. E. NELSON and E. H. EVERSON. 1977. Evaluation of free proline accumula-
tion as an index of drought resistance using two contrasting barley cultivars. Crop ac
17720-7236,

HEW, C. S. and Y. S. WONG. 1974. Photosynthesis and respiration of ferns in re
Amer. Fern J. 64:40—48.

HSIAO, T. C. 1973. Plant responses to water stress. Ann. Rev. Pl. Physiol. 24:519-570.

HUANG, H. C. A. and A. J. CAVALIERI. 1979. Proline oxidase and water stress-induced proline
accumulation in Spinach leaves. PI. Physiol. 63:531-535. ae oe

KLUGE, M. and L. P. TING. 1978. Crassulacean Acid Metabolism. Analysis of an Ecological
Adaptation, Springer. Berlin.

OSMOND, C. B. 1978. Crassulacean acid metabolis
29:379-414.

SINGH, T. N., L. G. PALEG, and D. ASPINALL. 1973a. Stress metabolism I.
and growth in the barley plant during water stress. Austral. J. Biol. Sci. 26a =e

SINGH, T. N., D. ASPINALL. L. G. PALEG and S. F. BOGGESS. 1973b. Stress = ;
Changes in proline concentration in excised plant tissues. Austral. J. Biol. Sct. 26:57-63. s

STEWART, C. R. 1980. Proline accumulation: Biochemical aspects. /” Paleg. site ms “as 7
(eds). Physiology and Biochemistry of Drought Resistance 10 Plants. Harcourt brace
Jovanovich, New York.

TROLL, W. and J. LINDSLEY. 1955. A photometric metho
Chem. 215:655— i

ies e a ene and C. E, NELSON. 1979. Proline accumulation In sere oot
barley leaves in relation to translocation and the nitrogen budget. PI. one =) Sas It

WEBBS, K. L. and J. W. A. BURLEY. 1965. Dark fixation of CO, by obligate and facultative sa
marsh halophytes. Canad. J. Bot. 43:281-285.

WONG, S. C. and C. S. HEW. 1976. Diffusive resistance, tit
tropical epiphytic ferns. Amer. Fern. J. 66:121-124.

—

lation to their habitat.

m: a curiosity in context. Ann. Rev. Pl. Physiol.

Nitrogen metabolism

d for the determination of proline. J. Biol.

ratable acidity. and CO, fixation in two

40 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984)

Problems in Asplenium, with Some New Species
from Ecuador
ROBERT G. STOLZE*

While studying the genus Asplenium for the “Flora of Ecuador,” I encountered
great difficulties with circumscription of A. myriophyllum (Swartz) Presl. This is a
highly dissected species of wet forests, reported from Florida, the Greater Antilles,
Mexico, much of Central America, Colombia, Venezuela, Ecuador, and Peru. It
occurs rather frequently throughout much of its range, has several very close
relatives, and is itself highly variable. Among its most diagnostic characters are:
tripinnate or subtripinnate (often only bipinnate in Florida) laminae rather abruptly
reduced at the base; dull grayish or reddish brown petioles up to 15 cm long;
castaneous to blackish, rather shiny, scarcely clathrate rhizome scales; and pinnae
about 20-25 pairs, mostly ascending, sessile, with the basal pinnules overlapping
wae rachis.

splenium myriophyllum has been found in herbaria, often identified as A.
Poise Fée, A. cristatum Lam. (syn. A. cicutarium Swartz), A. divaricatum
Kunze, and A. rutaceum (Willd.) Mett. (syn. A. conquisitum Underw. & Maxon ex
Christ). It is easy to understand the confusion, for these species share many similar
features, and whereas each seems to be readily identifiable in the typical habit, there
can always be found several collections in which characters tend to merge with those
of other species in the group. Asplenium rutaceum (common in the neotropics) is
the most distinctive, as it is the only species which has highly lustrous, castaneous
to blackish petioles and lower rachises. Furthermore the lamina is strongly and
gradually reduced to a naked and flagelliform apex, this often with a proliferous tip.
In the other species of the group, the petiole is dull and lighter in color, and the
lamina tapers to a pinnatifid apex—this sometimes attenuate, but never flagelliform
and radicant.

Asplenium cristatum (also with wide distribution in the neotropics) is the next
most distinctive of the species listed above, as the lamina is scarcely, if ever. reduced
at the base; i.e., the basal pair of pinnae are nearly or quite as long as the central
ones. The lamina in all the other species is strongly (either gradually or abruptly)
reduced at base, with two to many proximal pairs of pinnae conspicuously shorter
than the central ones. Nevertheless, I have found several specimens of A. cristatum,
from Guatemala to Ecuador, in which one or two proximal pairs of pinnae are
significantly reduced.

Asplenium cladolepton and A. divaricatum are species which more properly might
be considered variants of A. myriophyllum. The former, known from Colombia,
Venezuela, and rarely in Ecuador, differs from A. myriophyllum primarily in its
smaller size and narrower lamina. The epipetric Peruvian species, A. divaricatum.
appears to be a depauperate A. myriophyllum with leaves only 3-10 cm long and
tiny ultimate segments. It may be merely an ecological variant.

*Department of Botany, Field Museum of Natural History, Chicago, IL 60605.

R. G. STOLZE: PROBLEMS IN ASPLENIUM 4]

Asplenium myriophyllum varies strongly throughout its range. Many collections
from Florida and Guatemala are much smaller and often only bipinnate, whereas
leaves in South American plants are typically luxuriant, up to 40 cm long and fully
tripinnate. Petioles are commonly 8-15 cm long, yet those of some plants from
Florida and the Greater Antilles measure only 2-3 cm. It is no wonder then that
ferns in this group are universally misunderstood. Obviously detailed research is
needed, in which comparisons are made of specimens throughout the neotropics.
Furthermore, it is probable that much hybridization is occurring here—a situation
not uncommon in the genus. Mickel (“How to Know the Ferns and Fern Allies,” p.
56. 1979; Wm. C. Brown, Dubuque, Iowa) delineates several known hybrids in
North America which involve A. myriophyllum. But no such studies have been
undertaken yet throughout the broad range of its distribution.

One should be cautious in naming a new species within a group already so
confused. However, in studying the genus for “Flora of Ecuador,” I came upon a
large number of Ecuadorean specimens which appeared to be clearly distinct in
lamina architecture from the other components of the species complex. These were
variously determined as A. cicutarium, A. conquisitum, A. cristatum, A.
myriophyllum, A. rhizophyllum, and A. rutaceum. As herbarium specimens were
separated on the basis of obvious morphological features, other, less obvious,
characters came to light, which convinced me that this was a species much more
distinct than most others already recognized in the group. Oddly enough, although it
has been collected frequently in Ecuador, I can find no record of its occurring IM
neighboring countries.

Asplenium ecuadorense Stolze, sp. nov. Figs. 96
Rhizoma erectum, paleaceum; paleae 2-4 mm _ longae, deltato-lanceolatae ve
-ovatae, brunneae vel fuscae, conspicue clathratae: folium 20—45 cm longum, 0—
ipl , fascic

plerumque brunneus, numquam_ nitidus, lamina gradatim:
pinnatifidam vel flagelliformem versus, saepe apice prolificanti, ‘ Ee arater
angustata; pinnae 34—60-jugae, lanceolato-oblongae, confertae vel poten i
patentes, vel pinnae proximales deflexae, sessiles, pinnulis basalibus a i.
Superpositis supra rachin; venae segmentorum ultimorum simplices; sori in quog
Segmento solitarii; indusium delicatum, subintegrum

Chimborazzo [Prov. Bolivar, Ecuador], 500 m,
ornans,” Rimbach 16 [Rosenstock exsicc. no.
isotypes GH, S, US). ae

In wet forests, commonly on trunks of trees, very rarely on the forest floor,
100-1400 m, thus far found only in Ecuador.

Plants epiphytic strial; rhi stout, erec
_ rarely terrestrial; rhizome * Pole: -
lanceolate aa. : n or gray-brown scales, these 2-4 mm long, conspicu

10] (US, 2 sheets: 1197005-6:

tripinnate or subtri asciculate; petiole 1-3 ¢ , terete, ¢
reddish brown oe “ mang glabrous OF with ee tek
Nute, filiform scales; lamina glabrous, thin-herbaceous to membranaceous,

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

FIGS.
adaxial side, x 6. FIG. 3. Rhizome seale,: <1

1-3. Asplenium spipgorneb as FIG. 1. Habit, x 0.5
12.

_ FIG. 2. Portions of rachis and two pinnae.

R. G. STOLZE: PROBLEMS IN ASPLENIUM 43

green, elliptic-attenuate, strongly and gradually reduced to a pinnatifid or flagelli-
ex and often with a proliferous tip, strongly and gradually narrowed at base,

with 8-18 slightly to greatly reduced pinnae; rachis dark or reddish brown,
marginate or narrow-alate; pinnae (30)34—-60 pairs, crowded to imbricate, mostly
ome imal ones commonly deflexed, oblong-lanceolate, their

of their length, sessile, the basal pinnules strongly

overlapping the rachis; pinnules 10-20 pairs, cut nearly or quite to the costa into

of segments, these elliptic or ovate-elliptic, entire (or often the basal

SELECTED SPECIMENS EXAMINED:

ECUADOR: Bolivar: Western Cordillera, western slope of the river Suquibi, 500 m., Rimbach 29
(topotypes GH, US). Cotopaxi: Epiphyte, produces new crowns from a terminal bud. On road from
Quevedo to Quito, ca. 500 m, Haught 2917 (GH, S. UC, US). Guayas: Crescit in silv. trop. prop.
Puente de Chimbo, 8/1891, Sodiro 20/47 (P). Los Rios: Epiphytic fern, lightly disturbed forest on hills:
Centinela Ridge area, 12.5 km E of Patricia Pilar, 1400 ft, Hansen et al. 7746 (AAU, MO). Manabi:
La Morena, ca. 15 km NNE of Flavio Alfaro, remnants of
seasonal rain forest, ca. 100 m, Harling & Andersson 18909 (GB). Pichincha: Seasonal rain forest, 20
km W of Santo Domingo de los Colorados, 1000 ft, Cazalet & Pennington 5264 (NY, UC, US).
Tungurahua: Western Cordillera, W of town of Ambato, 500 m, moist forest region, Rimbach 640 (F).
Zamora-Chinchipe: Fallen trees on hilside, on tree trunk, road from Loja to Zamora, km 42, 1400 m,
Dodson & Thien 843 (MO).

The general aspect of this new species 1s that of A. rutaceum in that both have the
lamina strongly reduced at the apex and base, with the tip of the rachis often
flagelliform and radicant. The pinnae of both are commonly borne at right angles to
the rachis and are sessile, with the basal pinnules strongly overlapping the rachis,
and the pinna margins parallel for most of their length. In addition, the rhizome
scales are gray-brown and conspicuously clathrate. However, A. ecuadorense ditters
obviously from A. rutaceum in that the pinnae are much more crowded (often
strongly imbricate) and numerous (ca. 3550 pairs vs. 15-30 pairs), and the petioles
and lower rachises are dull brown or reddish brown, VS- castaneous to atropurpureous
and highly lustrous in A. ruftaceum. Sem

Asplenium myriophyllum is the other species with which A. ecuadorense mig A e
easily confused. Both are similar in gross morphological characteristics. anal’
or subtripinnate lamina, strongly reduced to both apex and base; dull scsi ae
usually reddish brown petiole; pinnae sessile, with basal pinnules oan a
rachis. But the new species has gray-brown rhizome scales that are fe ks
clathrate, whereas in A. myriophyllum these are castancous to blackish and only
obscurely clathrate (i.e., the lumina are greatly constricted). Also, <egnee
pinnae (ca. 35-50 pairs) mostly spread at right angles from the rachis. oe ach ie
to imbricate, and are predominantly oblong. i.e. the margins are ‘alee! eae ;
their length. In comparison, A. myriophyllum has fewer pinnae (ca. ; —2 a Me :
which are mostly ascending and much less crowded, and are broadest at or ne
base, tapering abruptly or gradually to apex.

Three more species came to light during
they are described here.

studies on the “Flora of Ecuador” and

44 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

pu
&

OS
oe (SS

Ok
NG

Qe
Cf

FIGS. 4-6. Asplenium oellgaardii. FIG. 4. Habit, x 0.5. FIG. 5. Apex of leaf, with proliferous tp.
x3. FIG. 6. Central portion of rachis, with several pinnae, x6.

R. G. STOLZE: PROBLEMS IN ASPLENIUM 45

Asplenium oellgaardii Stolze, sp. nov. Figs. 4-6.
Asplenium trichomanes var. herbaceum Sod. Anal. Univ. Quito 8(58):269. 1893 (seors. 145). 1897.
TYPE: “In sylv. vulc. Pasochoa, 1X/90.” Sodiro s. n. (not seen; probable isotype US1196744).
Rhizoma parvum, erectum, paleaceum; paleae 1.5—2.5 mm_longae, lineari-
lanceolatae, attenuatae, nigricantes, obscure clathratae, folium 7—18(22) cm long-
um, 1.2-2.5 cm latum, pinnatum; petiolus 1.5—4.5 cm longus, brunneus, numquam
nitidus, paleis sparsis, tortuosis, filiformibus, castaneis vel nigricantibus; lamina
; apic

e S,
nese obtusae, ad basin valde inaequilaterales; venae 2-4-jugae, simplices,
manifestae vel indistinctae.

TYPE: San Miguel, on margin of paramo between Atantaqui and Hacienda Pinan,
Prov. Imbabura, Ecuador, 3420 m, Wiggins 10333 (UC; isotypes F, US).

In andine forests, commonly on shaded slopes and banks of quebradas, 2150-3400
m, thus far found only in Ecuador and in one location in Colombia.

Plants terrestrial; rhizome small, compact, erect, provided with linear-lanceolate,
attenuate scales, these 1.5—2.5 mm long, blackish, or occasionally with very narrow
brownish margins, obscurely clathrate, the lumina small an the walls relatively
thick: leaves 7—18(22) cm long, 1.2-2.5 cm broad, pinnate: petiole 1.5-4.5 cm

(9) on a pinna, commonly borne nearer to the
Hlowish to greenish, 2-4 mm long, linear to

72)

i)
|

acroscopic one |-
pinna apex than to the base; indusium ye
narrow oblong, the margin entire.
SELECTED SPECIMENS EXAMINED:
COLOMBIA: Caldas: Clearing, “Pinares.” above Salento. ; Stee tes
ened El Oro: Along Quebrada, on S-, SW-, and W-facing ee
achi eee ‘ 3 d Re oe e 1 rti “i
icaran, tributary to Rio Minas Nuevas, ON ee Colambo, SE of Gozanama. 2750 m,

Steyermark 54117 (F). Loja: On shaded bank on uppeT slopes of m

ere : 12 km W of Loja. 8000-9000 m. Wiggins 10987 (MO. UC,

sta Solis 14061 (F). Ad Cotocollao, 2750 m. Mille s.
pro

waa
=
—~
iF)
—
4

: . Nono, 9/1899, Sodiro s. n. (NY). Cotocallao, ne
unknown: “In silv. regionis interand.” Sodiro s. 1. (US | : :
As is ic aphne wim most of Sodiro’s new taxa, it is difficult to be ae
authentic type material, but probably US1196744 is an isotype of ¥ ie a
manes var. herbaceum. This and the other Sodiro specimens cited above ap
parently were intended for this new variety, but on the species level, the opis
herbaceum previously had been used by Fée (1857) and by Baker Lal iil .
plants, thus requiring a different epithet. Furthermore, A. oellgaardi is - : gfe
related to A. trichomanes, which is a fern of temperate regions having a dark an

46 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

FIGS. 7-9. Asplenium subdimidiatum. FIG. 7. Habit, x 0.5. FIG. 8. Central portion of rachis, with
pinna, abaxial side, x 3. FIG. . Portion of ate with scales, x 25.

R. G. STOLZE: PROBLEMS IN ASPLENIUM 47

lustrous petiole and rachis, a non-proliferous leaf apex, and subequilateral pinna
bases. A. oellgaardii has dull reddish to grayish brown petioles, strongly inequilateral
pinna bases, and the leaf tip is generally proliferous.

This new species has been found in various herbaria also determined as A. fragile
Pres] and as A. debile Sod. Leaves of the closely related A. fragile may often bear
proliferous buds in pinna axils, but rarely, if ever, at the tip of the rachis. Also, the
pinnae are irregularly obovate to rhombqid, whereas the pinnae in A. oellgaardii are
oblong. Less closely related is A. ruizianum (syn. A. congestum C. Chr., a new
name for A. debile Sod., non Fée, 1869). This differs from A. oellgaardii by its
fewer pinnae (4-10 pairs). Also, the lamina is equal to or shorter than the petiole
and bears no proliferous buds; whereas in A. oellgaardii the lamina greatly exceeds
the short petiole.

This new species is named in honor of my good friend Benjamin Ollgaard, who
not only is coordinating the contributions of pteridophyte treatments for the “Flora
of Ecuador,” but is actively engaged in field work for the Flora.

Asplenium subdimidiatum Stolze, sp. nov. Figs. 7-9.
Rhizoma erectum, paleaceum; paleae |.5-2.5 mm longae, deltato-lanceolatae vel

rmes, subdimidiatae; venae
subdichotomae vel subflabelliformes; indusium crassum, brunneolum, nigrescens
ubi affixum.

TYPE: Epiphytic. Lagunas de Cyabeno, second lake, situated some 3 km W of
the first lake, Prov. Napo, Ecuador, primary forest with some periodically flooded
parts, 300 m, 23 Aug. 1981, Brandbyge, Asanza, Werling & Leth-Nissen 36016,

ate, dark brown scales, these 1.5—2.5 mm long, clat rate. th

i to filiform scales to 2 mm long,

gradually reduced to a pinnatifid or attenuate apex. not pro
care] narrowed at base, with 4-6 P “a
provided sparsely (at least on costae and veins ade =
regularly stellate. gland-tipped trichomes; rachis dark brown, caer a,
scaly as on the petiole; pinnae 15-28 pairs, approximate, patent to s ightly cet ~
i bparallel at least toward the pinna base, sessile,
’ is: pinnules 3-6 pairs, Deere to
i subdimidiate,
short-stalked, cuneate and flabelliform or more commonly, trapeziform, su a
truncate, and strongly produced acroscopically, narrowly cuneate OF strongly excised
pasiscopically, the margins sha :
asiscopically; veins forking subdichotomous!y @ —
terminating é teeth of the pinnules; sori |-3 in each pinnule elliptical, entire,
rather thick-textured, brownish, but bec

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

>
OD

AQT?

Text
P8

=~
a"

—_
as a

Beceag Om
ae ek cn NS
SS

= Sy

EES
<o
ee
tH

——S

—

exe
sell

FIGS. 10, 11. Asplenium pseudoangustum. FIG. 10. Habit, x 0.5. FIG. 11. Rhizome scale, * 12.5.

R. G. STOLZE: PROBLEMS IN ASPLENIUM 49

Normally I would choose not to describe a novelty on the basis of a single
collection, but this is a very distinctive fern and is not likely to be confused with any
other species in the genus.

Two particularly distinctive features of this new species help separate it from most
others of the genus: the conspicuous scales of the petiole and rachis and the peculiar,
branched trichomes on the veins and costae. The scales are of a type common to
species included in sect. Sphenopteris by Morton and Lellinger (Mem. New York
Bot. Gard. 15: 5. 1966), i.e., dark brown to castaneous, very narrow and usually
tortuous, often appearing more like trichomes than scales, clathrate, some of them
3-7 cells broad near the base, yet often attenuate to a single-ribbed apex, and more
commonly single-ribbed throughout, the lumina reduced to just a few short, lateral
cross walls, but not enclosed. An even more distinctive feature of A. subdimidiatum
is the inconspicuous indument which sparsely dots the pinnules, particularly on the
abaxial surface. Here and there along the veins and occasionally on the costae are
scattered, appressed, brownish trichomes which are branched from the point of
attachment, the two to several arms often terminating in a slightly enlarged,
colorless gland. Admittedly they are tiny (0.1-0.2 mm) and perhaps will go
unnoticed except under higher magnification, but to my knowledge they have not
been found in other species of Asplenium.

The scaly axes and the shape of some of the pinnules are suggestive of A.
cuneatum Lam., but there the resemblance ends. In the latter species, the lamina is
broadest at or near the base, so also are the short-stalked pinnae, and the sor! are
commonly linear (many times longer than broad). In A. subdimidiatum, the lamina
is conspicuously narrowed at base, the pinnae have their margins parallel at least
toward the base and are quite sessile, with the basal pinnules strongly overlapping
the rachis, and the sori are elliptical, usually only about twice as long as broad.

Superficially the new species resembles A. cristatum Lam. in the crowded, sessile
pinnae and the general aspect of the lamina. But A. cristatum has far more pinnules
(8-12 vs. 3-6 pairs on larger pinnae), the laminae are not narrowed at base, and the
axes are not or scarcely scaly.

Figs. 10-11.
e 1.5—3 mm longae,

ongum, 1.3—2.5(3) cm latum, sessile vel subsessile, simplex; lamina glabra. soho
herbacea, elliptica, acuta vel subacuta, ad basin attenuata,
lineares, recti vel aliquantum arcuati, distantes. ed at
YPE: Tingo Maria, Prov. Hudnuco Peru, 700 m, hillside forest, epiphytic on
trunks and branches of small trees, R. M. & A. F. Tryon 52357 (GH; isotypes F, eet
In deep shade of wet forests, on trunks or branches of trees, 350-1400 m:

Ecuador; Peru. ee
Plants epiphytic; rhizome small, erect to ascending. provided —— a oti
iia clathrate scales, these lanceolate io Oe acu ie a) nike 4 630 Ga
ong; leaves spi ssile or subsessile, SI ang uU .
g es aay: oye itose, sessile 0 lacking; lamina firm-herbaceous to

50 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

twice-forked, obscure, spreading at a 30—45° angle from the midrib; sori straight or
somewhat arcuate, well-spaced, elongated, medial between midrib and margin;
indusium narrow, elongated, pale green or yellow-green.

Caceres, 500-600 m, Ferreyra 17038 (GH). Palo Blanco al oeste del Puente, Mariscal Caceres,
Tocache Nuevo. 600-700 m, Schunke 5671 (F, US). Puerto Pizana, Mariscal Caceres, Tocache Nuevo,
350 m, Schunke 6951 (F).

A single Ecuadorean collection came to my attention which at first I assumed was
a depauperate specimen of A. serratum L. or which perhaps represented a range
extension for A. angustum Swartz. Closer examination of this Asplenium with small,
undivided laminae proved it was quite distinct from either, and a further search of
the Field Museum collections and other herbaria turned up a number of specimens
from Peru. Most of these had been determined “Asplenium angustum vel aft.”

This superficially does resemble A. angustum of Colombia, Venezuela, the
Guianas, and Brazil (which is perhaps itself merely a variant of A. serratum). But
there are differences in texture and indument of the lamina, rhizome scales, and
soral arrangement. In A. angustum the rhizome scales are linear to filiform,
castaneous to black, and commonly terminate in a thin, unicostate tip, there are
minute, dark, clathrate scales scattered on the midrib of the rather thick lamina, and
the sori are crowded and borne at a very acute angle. In A. pseudoangustum, the
rhizome scales are ovate to lanceolate, lighter in color, with the apex merely acute:
the lamina is thinner in texture and lacks scales; and the sori are widely spaced and
borne at a less acute angle from the midrib.

REVIEW

= _USOS DE LOS HELECHOS EN SURAMERICA CON ESPECIAL REFER-
ENCIA A COLOMBIA, by Maria Teresa Murillo P. 156 pp. Instituto de Ciencias
Naturales, Universidad Nacional de Colombia, Bogota, 1983.—The introductory
third of this well printed paperback book includes a brief history of Colombian
botany and a key to the families of pteridophytes found in South America. Numerous
beautiful line drawings by Eugenia de Brieva illustrate the families. The main text is
an alphabetical list by Latin name of ferns and fern-allies with their uses, as
recorded in botanical literature. A brief description of each plant, its vernacu ar
name(s), its distribution in Colombia, and its uses (often including quotations from
the literature) are given. Fourteen color habit photographs taken in the field are
included. Indices to vernacular and Latin names and a bibliography conclude the
volume. This book will be of interest to many botanists, but is especially important
to ethnobotanists and pharmacologists.—D.B.L.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984) 5]

Frequency of Cyanogenesis in Bracken in Relation
to Shading and Winter Severity
I. SCHREINER,* D. NAFUS,* and D. PIMENTEL**

A number of studies have shown that cyanogenic phenotypes of plants are less
palatable to, and less damaged by, many herbivores than are acyanogenic phenotypes
of the same species (Jones, 1962; Angseesing & Angseesing, 1973; Cooper-Driver
& Swain, 1976; Dritschilo et al., 1979; Schreiner et al., 1984). Yet many plant
species are polymorphic for cyanogenesis, and the acyanogenic morphs are common
(Fikenscher & Hegenauer, 1977). Studies on this polymorphism in Trifolium repens
and Lotus corniculatus have shown that a variety of environmental stresses are more
deleterious to cyanogenic plants than they are to acyanogenic ones. In Trifolium, low
winter temperatures are correlated with low frequencies of plants containing cyano-
genic glucoside (Daday, 1954; De Araujo, 1976). However, the correlation between
low winter temperatures and a high incidence of acyanogenic plants is poorer for L.
corniculatus (Jones, 1977). Cyanogenic Lotus and Trifolium plants were less tolerant
of drought (Abbott, 1977; Foulds & Grimes, 1972). In some situations, salt spray
may also select more strongly against cyanogenic Lotus plants (Keymer & Ellis,
1978). Cooper-Driver (1976) found that Bracken, Preridium aquilinum (L.) Kuhn,
was polymorphic for cyanogenesis and that cyanogenic Bracken fronds growing In
the shade had higher cyanide levels than those growing in open, sunny habitats
(Cooper-Driver et al., 1977). The incidence of cyanogenic and acyanogenic fronds in
the two habitats was not measured. In an attempt to follow up on this work, the
current study examined the incidence of cyanogenic and acyanogenic fronds in open
and shady habitats. A survey was also made to determine if there was any correlation
between winter severity and the incidence of cyanogenesis in Bracken stands.

MATERIALS AND METHODS

The relationship of habitat to the incidence of cyanogenesis in Bracken was
examined by surveying fronds within a 20 km radius of Ithaca, New York, primarily
in the Connecticut Hill Wildlife Management Area. Bracken stands were classified
as shady if a closed tree canopy shaded all the fronds. Stands growing In open fields
Were classified as sunny, and stands growing at the edge of woodlands were
classified as partially shaded. Fronds were collected by walking across the longest
dimension of the stand and collecting a frond every meter. Ten fronds were sampled
from stands of less than 100 m2, and 20 fronds were taken from larger stands. The
ronds were collected in late May and early June. : : ae

To test for cyanide levels in the Bracken fronds, 6 mm from the end of ane OF the
lowest pinnae of the fronds was removed (about 0.2 g plant material). The sample
was placed in a glass vial and crushed with a glass rod to disrupt the tissue. If bots
Cyanogenic glucoside and one or more cyanide-releasing enzymes were present In :
frond, this disruption would cause them to come into contact and release cyanide ions.

oe eae .
College of Agriculture and Life Sciences, University of Guam, Mangilao, Guam 9691.
Department of Entomology, Cornell University, Ithaca. NY 14853.

5 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

TABLE |. DISTRIBUTION OF CYANOGENIC AND ACYANOGENIC BRACKEN FRONDS UNDER

VARIOUS LIGHT CONDITIONS.
A. Percentage of cyanogenic fronds in various habitats

Type of Open-sunny Partial shade Shade
frond (170 fronds) (150 fronds) (140 fronds)
HCN = 66%
HCN + 18 al 34
B. Levels of cyanide in the cyanogenic fronds above

Amount of Open-sunny Partial shade Shade

HCN (31 fronds) (31 fronds) (48 fronds)
Low 8%
Medium 48 61 58
High 16 26 33

After the frond was crushed, a strip of filter paper (3.5 cm x 0.5 cm) impregnated
with 0.5 ml picric acid solution was inserted and the vial stoppered (Dawson, 1941).
After one hour, the paper was examined. If it had turned purplish brown, the frond
was scored as highly cyanogenic. If the paper had turned orange within the hour, the
frond was scored as medium cyanogenic. The vials containing papers which showed
no color change were kept overnight. If the paper had turned orange within 24 hours,
the cyanide level of the frond was scored as low. If there was no change in the color
of the filter paper within a 24 hour period, the frond was scored as acyanogenic,
lacking either the glucoside, the enzyme(s), or both, so that cyanide ions were not
released.

The relationship between the incidence of cyanogenesis in Bracken and winter
severity was investigated by sampling Bracken stands about ever 150 km along a
1000 km transect from Ithaca, New York southwards towards Fayetteville, North
Carolina. Every 150 km, 20 Bracken stands were found along rural roads. Five
fronds were taken from each stand. Cyanogenesis in the fronds was measured as
described previously, except that only the release of cyanide ions was recorded in
these samples, with no attempt to quantify the amounts. Locations south of
Washington, DC were sampled in the first week of May, 1978, and those to the
north, in mid-June.

RESULTS

Two-thirds of the large Bracken stands and half of the small ones sampled

contained both cyanogenic and acyanogenic fronds, indicating that at least two
ecotypes of Bracken were present. Differences in the appearance of the fronds
Suggested that in many cases, more than two ecotypes were present.
Fronds from shaded stands were nearly twice as likely to be cyanogenic as were
fronds from sunny stands (34% vs. 18%, Table 1A). The raw data were analyzed
using a Chi-square test, and the difference was significant at the 0.025 level. There
was also a 90% probability (Chi-square) that fronds growing in the shade released
more cyanide than did those growing in the sun (Table 1B).

Differences in winter severity did not seem to be related to the incidence of
cyanogenesis in Bracken populations (Fig. /). About 25% of the fronds were
cyanogenic in both the northernmost and the southernmost samples, whereas the
intermediate sites had lower frequencies.

|. SCHREINER ET AL.: CYANOGENESIS IN BRACKEN 53

Sons 1. Percentage of cyanogenic Bracken fronds (large numbers) at sites in the eastern United States in
relation to winter severity expressed as the average minimum temperature for February (small numbers)
at the nearest weather station (U.S. Dept. of Commerce, 1979, pp. 1-2).

DISCUSSION

_ Cyanogenesis in Bracken was significantly associated with habitat. Fronds grow-
Ing in shaded stands were twice as likely to be cyanogenic as were fronds growing In
open fields. Cyanogenic fronds growing in the shade may also have released more
cyanide ions than cyanogenic fronds growing in the sun (90% probability), even
though fronds in the woods came up one to two weeks earlier than those growing In
open areas. Note, however, that older fronds would be expected to have —
cyanide levels, as the quantity of cyanide is highest in the fiddleheads and decreases
rapidly as the fronds age (Greshoff, 1908; Moon & Raafat, 1951; Cooper-Driver et
al., 1977; Schreiner, et al., 1984). Our results agree with those of Cooper-Driver .
: - (1977), who also reported higher levels of cyanide in fronds growing in shade
Sites than in those growing in the open.

The earlier appearance of Bracken fronds in shaded habitats | 9 boom
Was unexpected since open areas generally warm Up earlier in the spring (Geiger,
1966). Hellum and Zahner (1966) suggested that in Michigan, early at
Bracken fronds were likely to be damaged by late frosts while still in the weal layer, »
that plants in Open areas were set back compared to those 1n woodlands. It seems
Probable that the same is true in rural New York.

than in sunny habitats

54 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

One possible cause for the different frequency of cyanogenic fronds in wooded
and open areas is a difference in fertility between the sites. Cyanogenesis in Bracken
has been found to be markedly enhanced by application of urea fertilizer (J.H.
Lawton, pers. comm.). The Connecticut Hill area appears to have infertile soils, but
perhaps the Bracken growing in wooded areas get some additional nutrients from the
litter of the trees shading them.

Lawton (pers. comm.) has found that in Britain moths mining the young croziers
appear to induce cyanogenesis in the vicinity of their mines. In rural New York,
crozier-mining lepidopterans were most common in shaded sites (Schreiner, 1980).

owever, no correlation between the presence of mines and cyanogenesis was
observed. (Schreiner et al., 1984

Cyanogenesis in Bracken did not appear to be related to overall winter severity, as
the incidence of cyanogenic plants was lower at the intermediate sites than at the
most southern and northern ones. This contrasts with the results found for 7. repens
(Daday, 1954; De Araujo, 1976), but not with those observed for L. corniculatus
(Jones, 1977).

Partial funding of this research was provided by NSF Grant DEB-8010912. We
wish to thank Dr. Clive G. Jones for valuable comments.

LITERATURE CITED

ABBOTT, R. J. 1977. A renege association between soil moisture content and the frequency of the
cyanogenic form of Lotus corniculatus L. at Birsay, Orkney. Heredity 38:397-400.

pata bP A ae a J. ANGSEESING. 1973. Field observation on the cyanogenesis
polymorphism in Trifolium repens. Heredity 31:276-282.

COOPER- DRIVER G. 1976. Chemotaxonomy and phytochemical ecology of bracken. Bot. J. Linnean
Soc. 73:35
: re db SWAIN. 1976. Cyanogenic polymorphism in bracken in relation to herbivores. Nature

; SD. FINCH, T. SWAIN and E. BERNAYS. 1977. Seasonal variation in secondary plant
compounds in relation to palatability of Pteridium aquilinum. Biochem. Syst. Ecol. 5:177-183.
DADAY, H. 1954. Gene seta sacra in wild populations of Trifolium repens L. IV. Mechanism of
natural selection. Heredity 8 8.
DAWSON, C. D. R. 1941. Tetrasomic inheritance in Lotus corniculatus L. J. Genet. 42:49-72.
DE ARAUJO, A. M. 1976. The relationship eee altitude and cyanogenesis in white clover
(Trifolium repens L. ). Heredity 37:291-2
DRITSCHILO, W. J., KRUMMEL, D. ae and D. PIMENTEL. 1979. Herbivorous insects
colonizing cyanogenic and acyanogenic Trifolium repens. Heredity 42:49-5
FIKENSCHER, L. H., and R. HEGENAUER. 1977. Der Verbreitung der Blausiiure bei den
Cormophyten. II. Ueber die cyanogenen — ngen bei einigen a bei dem
Oliniaceae und in der Rutaceen-Gattung Ziera. Pharm. Weekblad. Nedl. 112:1
FOULDS, W. and J. GRIMES. 1972. The eae of grannee and acyanogenic faite of
rifolium repens to soil moisture supply. Heredity 28:181-187.
GEIGER, R 1966. The Climate near the Ground. Harvard Raves Press, Cambridge, MA.
GRESHOFF, M. 1908. Transistorisch Blauzwir in Varens. Pharm. Weekbl. Nedl. 45:770-773.
— A. K. and R. ZAHNER. 1966. The frond size of bracken fern on a forested outwash sand in
orthern lower Michigan. Proc. Soil Sci. Soc. Amer. 30:520-524.
JONES, D. A. 1962. Selective eating of the acyanogenic form of the plant Lotus corniculatus L. by
various animals. Nature 193:1109-1110.
- 1977. On the polymorphism of cyanogenesis in Lotus corniculatus L. VII. The distribution
of the cyanogenic form in Western Europe. Heredity 39:27—44.

|. SCHREINER ET AL.: CYANOGENESIS IN BRACKEN 55

KEYMER, R., and W. M. ELLIS. 1978. Experimental studies on plants of Lotus corniculatus L. from
Anglesey polymorphic for cyanogenesis. Heredity 40:189-206.

MOON, F. E., and M. A. RAAFAT. 1951. Some biochemical aspects of bracken poisoning in the
ruminant animal. II. The significance of the cyanogenetic principle of bracken. J. Sci. Food
Agric. 2:327—336.

SCHREINER, I. H. 1980. Cyanogenesis and the herbivorous insects of bracken fern (Pteridium
aquilinum). Ph.D. Thesis, Cornell University, Ithaca, NY.
., D. NAFUS and D. PIMENTEL. 1984. Effects of cyanogenesis in bracken fern (Pteridium
aquilinum (L.) Kuhn) on associated insects. Ecol. Entomol. 9:69-79.

U.S. DEPT. COMMERCE. 1979. Climatological Data. National Summary, vol. 30.

FLORA OF ECUADOR 14(4). POLYPODIACEAE—THELYPTERID-
OIDEAE, by Alan R. Smith. Flora of Ecuador 18:1—148. 1983.—Thelypteris is

alphabetically within each subgenus. Each has a synonymy, de
specimens, ane notes. eh ese species are described and illustrated, and .
new combinations are made. For students of the genus. it would have been helpful 1

the sections of subg. Amauropelta had been included wit
Constraints of format probably did not permit this.
the notes. The paper includes an index and a map of th
Welcome features. The paper may be purchased from th
Publishing House. Box 6710, S-11385 Stockholm, Sweden. —D-B.-

56 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984)

New Combinations and Some New Names in Ferns
DAVID B. LELLINGER*

The forthcoming publication of microfiches and an index of the pteridophyte
types in the U.S. National Herbarium (US) requires that the following combinations
and new names be made.

Asplenium radicans var. alloeopteron (Kunze ex Klotzsch) Lellinger, comb.

nov.

Asplenium alloeopteron Kunze ex Klotzsch, Linnaea 20:353. 1847. SYNTYPES: Guyana, Schomburgk
1150 (LZ destroyed: isotype B not seen photo 9509'), and Guyana, Schomburgk 1206 (LZ destroyed;
isotype B not seen photo 9508 fragm US).

Campyloneurum angustipaleatum (Alston) M. Meyer ex Lellinger, comb. nov.
Polypodium angustipaleatum Alston, J. Bot. Brit. & For. 77:346. 1939. TYPE: Vicinity of
Cochabamba, Depto. Cochabamba, Bolivia, Bang 1/288 (BM not seen; isotype US).

¥Ctenitis aripensis (C. Chr. & Maxon in Maxon) Lellinger, comb.
Dryopteris aripensis C. Chr. & Maxon in Maxon, J. Washington Acad. Sci. 14: vee "T924, “TYPE:
Heights of Aripo, Trinidad, Britton & Freeman 2349 (US).

Ctenitis haitiensis (Brause) Lellinger, comb.
Dryopteris subincisa var. haitiensis Brause, Ark. for oe 17(7):67. 1922. TYPE: Ma Blanche,
Morne de la Hotte, Haiti, ca. 1400 m, Ekman 556 (B not seen fragm & photo US).

Ctenitis paranaensis (C. Chr.) Lellinger, comb.

Dryopteris falciculata var. paranaensis C. Chr. Danske cae Selsk. Skr. VII, 10(2):92. 1913.
TYPE: Villa Nova, near the Rio Negro, Est. Parand. Brazil, Annies [Ros. Fil. Austrobras. Exs. 79| (S
not seen; isotype US).

Ctenitis protensa var. dicksonioides — oe comb.
Aspidium dicksonioides Fée, Crypt. Vasc. Brés. 1:143. SL YPE: Neat = Gael de Cachoeria,
Rio Vaupes, Est. Amazonas, Brazil, Spruce 2129 (P or a. not seen; isotypes BM not seen, US)

Ctenopteris cryptosora (C. Chr.) wa neer, comb.
Polypodium decipiens Mett. ex Kuhn, Linnaea 36:129. 1869, non Fae (1862), nom. illeg. TYPE:
mbang, Borneo, Korthals 171 (L not seen ee 844)
Polpodiam cryptosorum C, Chr. Ind. Fil. 520. 1906. TYPE: A renaming of Polypodium decipiens
Mett. ex Kuhn, and so based on the type of that name.

Culcita novae-guineae (Rosenst.) Lellinger, comb. nov.
Davallia novae-guineae Rosenst. Repert. Sp. Nov. Fedde 5:36. 1908. TYPE: Mt. Gelu, New Guinea,
700 m, Werner [Ros. Fil. Novoguin. Exs, 45| 73 (B not seen; isotype US).

Cyathea atahuallpa (Tryon) Lellinger, ined
Sphaeropteris atahuallpa Tryon, Rhodora 74:442, f.5— ae TYPE: E side of Cerros Calla Calla

above Balsas on road to Leimebamba, Depto. nee dates Peru, 3000-3100 m, Wright 6922 (GH not
seen; isotype US).

|
Here and elsewhere the word * ‘photo” followed by a number refers to the numbered series of photographs
taken by C. V. Morton and distributed from the U.S. National Museum

*Dept. of Botany, Smithsonian Institution. Washington, DC 20560.

D. B. LELLINGER: NEW COMBINATIONS AND NAMES IN FERNS 57

Cyathea bradei (Windisch) Lellinger, comb. n
Sphaeropteris bradei Windisch, Bradea 1:372, f. J, ae eee TYPE: Base of Cerro Mitu, Comis.
Vaupés, Colombia, st Raffauf & Soejarto 24229 (GH not seen).

Cyathea intramarginalis (Windisch) Lellinger, comb. nov.

Sphaeropteris intramarginalis Windisch, Bradea 1:373, t. 1, f. 5; t. HI, f. 3, 4, 10. 1973. TYPE:
Sarvén-tepui, Edo. Bolivar, Venezuela, 1750 m, Wurdack 34154 (GH not seen; isotypes NY not seen,
US).

Cyathea lockwoodiana (Windisch) Lellinger, Sige ats
Sphaeropteris lockwoodiana Windisch, Bradea 2:57. 197 TYPE: go rial Rio Macaya,
Rio Apaporis, Comis. Vaupés, Colombia, Schultes 5635 i not seen; isotype

Cyathea parianensis (Windisch) Lellinger, see nov

Sphaeropteris parianensis Windisch, Bradea 2:58, f. 35,8. 1916, TYPE: Cerro Patao, N of Puerto de
Hierro, Peninsula de Paria, Edo. Sucre, Venezuela, 800— ss m, Steyermark & Agostini 9/129 (GH not
seen; isotypes US, VEN not seen).

Cyathea sipapoensis (Tryon) Lellinger, comb. nov
Sphaeropteris sipapoensis Tryon, Rhodora 74:441, f. 1-4. 1972. TYPE: Intermediate Camp, Cerro
Sipapo (Pardque), Terr. Amazonas, Venezuela, Maguire & Politi 28765 (NY; isotype GH).

aig subquadripinnatum (Copel.) Lellinger, primes no
Athyrium subquadripinnatum Copel. Occ. Pap. Bishop Mus . 14:61, es 1938. TYPE: Moerai,
Rurutu, ae Islands, 10 m, H. St. John 16640 (BISH not seen, aioe US).

Diplazium tenuifolium (Copel.) gre aeeens
rine tenuifolium Copel. Phil. J. Sci. 40:3 929. LECTOTYPE — here): San Ramon,
mboanga, Mindanao, Philippines, sbi set Phil. Exs. 198 (MICH, 2 sheets not seen,
mane US). Copeland cited three syntypes instead of designating a aes in describing A.
tenuifolium. According to M. G. Price (pers. comm.), Copeland marked as type the specimen chosen
here as lectotype.

Gleichenia angusta (Klotzsch ex Sturm in Mart.) Maxon ex Lellinger, comb.
nov.

Mertensia angusta Klotzsch ex Sturm in Mart. Fl. Bras. 1(2):225. 1859. TYPE: Brazil, Sellow (B not
seen).

Gleichenia boliviensis (Maxon & Morton) Lellinger, comb. n a ae
Dicranopteris boliviensis Maxon & Morton, Bull. Torrey Bot. Club 66:44. 1939. COP
Bolivia, 3000 m, Tate 35/ (NY; isotype US).
Gleichenia Lellinger, comb. n
peruviana (Maxon) pine ex :
hig peruviana Maxon, Amer. Fern J. 33: 133. 1943. TYPE: poised Depto. Huanuco,
= 2700 m, Macbride 4510 (F: ne US)

Glyphotaenium insidiosum (Slosson) Lellinger, ee si 4-8. 1912. TYPE: Near
Polypodium insidiosum Slosson, Bull. Torrey Bot. Club 39:287, ¢. pa “ey
Camp La Gloria, $ of Sierra Moa, Pcia. Oriente, Cuba, Se 8043 (NY; isotype

Glyphotaenium integrum Laces. & goin ee Tye in c fle SE pen
Enterosora integra Lellinger & Morton, Act _ Venez. 2:123 5 puis
of the NW arm of Auyan-tepui, Edo. Bolivar. aed 1850 m, Stevermar

Glyphotaenium nesioticum (Maxon) sy comb. : pet Vicinity of Vinegar Hill,
Polypodium nesioticum Maxon, Smiths. Misc. Coll. 47:410. 1903 5
ortland Parish, Jamaica, 1200 m, Maxon 2773 fe

58 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Grammitis allosuroides (Rosenst. ) Lellinger, comb. n
he allosuroides Rosenst. Meded. Rijks Herb. Leiden 19:16. ei TYPE: Lagodos Valley, Bolivia,
4000 m, Herzog 2373 (S not seen; isotype US).

Grammitis angustipes (Copel. , emer wicnae
Ctenopteris — Copel. Philip. J. eo El Silencio, Yanaconas, Depto. El
Valle, Colombia, 1900-2200 m, Killip & sh ia co. (U .

Grammitis basalis (Maxon) Lellinger, comb. n
Polypodium basale Maxon, Amer. Fern J. 52:110. 1962. see: us Puyo, Pcia. Pastaza, Ecuador, 400 m,
Mexia 6930 (US)

Grammitis buesii (Maxon) Lellinger, comb. n
Sinan buesii Maxon, Contr. Gray Herb. 165:72. 1947. re Cerro Chuyapi, Depto. Cuzco, Peru,
00 m, Bués A45 (U

Grammitis chaseae eo Lellinger, comb. nov.
Ctenopteris chaseae Copel. Philip. J. Sci. 84:416, 1. 5. 1956. TYPE: Serra do Caparao, Est. Espiritu
Santo, Brazil, ca. 2100 m, 27 Nov 1929, Chase (US).

Grammitis ciliolepis (C. Chr.) Lellinger, comb.
Polvpodium ciliolepis C. Chr. Ark. for Bot. 20A(7):21. 1926. noes eg Chaco Village, Pcia. Sur Yungas,
Depto. La Paz. Bolivia, Axpliind 1501 (presumably C not seen fragm U

Grammitis congesta (Copel.) Lellinger, comb. nov.
Ctenopteris congesta Copel. Philip. J. Sci. 84:397, t. 3. 1956. TYPE: Loma Grande, La Convencion, Peru,
13000-14000 ft, Bués 2/72 (US).

Grammitis fendleri (Copel.) ey me comb. n

c — eg Copel. Phil. J. Sci. 84:463. 1956. TYPE: ve renaming of Polypodium radicale Moritz
ex Hie based on the type of oe

Phesdian ale Moritz ex Hieron. Bot. Tat ae Engler 34:511. 1904, non Moritz ex Baker, 1867. TYPE:
Near Colonia Tovar, Edo. Aragua, Venezuela, Fendler 2/5 (B not seen; isotype US).

Grammitis humilis nen in Tr. As Planch.) socraaay comb. nov.
Polypodium humile Mett. in Tr. & Planch. Ann. Sci. Nat., Paris, V, 2:251. 1863. TYPE: Chapinerro,
Depto. Cundinamarca, a 2700 m, Fane 137 (K not seen es US: isotype US).

Grammitis kegeliana (Kunze) Lellinger, comb.

—— kegelianum Kunze, Linnaea 21:210. 1848. TYPE: Sie Suriname, Kegel 1072 (GOET
not see

Grammitis killipii (Copel.) Lellinest, comb.
Xiphopteris killippi Copel. Amer. Fern J. 42:105, 1. 10. ibis eyes San Antonio, W of Cali, near summit
of Cordillera Occidental, Depto. El ie sshd 1900-2350 m, Killip & Garcia 33887 (US).

Grammitis longipinnata dig ) Lellinger, comb. nov.
Ctenopteris longipinnata Copel. Philip. J. Sci. 84:459, +. 12. 1956. TYPE: Cabecera del Koribeni. Alta
Huallaga, Depto. Hudnuco?, Peru, 9300 ft, Bués 1952 (US).

rammitis maxoniana Lellinger, non. nov.
era eerie Maxon, Contr. U. S. es 17:597. t. 42. 1916. non Grammitis flexuosa
Humb. & Bon 6) [=Eriosorus flexuosus i 4 K.) Copel.].
INN eg (Maxon) Copel. Phil. J. Sci. 84:434. 1955.

D. B. LELLINGER: NEW COMBINATIONS AND NAMES IN FERNS 59

Grammitis mayoris ieee ) Lellinger, comb. _
Polypodium mayoris Rosenst. Mém. Soc. Sci. Nat. Neuchatel 5:53, 1. 4, f. 6. 1912. TYPE: Paramo del
Ruiz, Depto. Caldas, coficbee ca. 3700 m, Mater 69 (S not seen: eee US).

Grammitis melanotricha (Baker in im Thurn) Lellinger, comb. n
Polypodium melanotricha Baker in im Thurn, Timehri 5:216. 1886. TYPE: Guyana, im i 125 (K not
seen; isotype fragment US).

Grammitis mortonii co ) Lellinger, comb. nov.
Xiphopteris mortonii Copel. Amer. Fern J, 42:97, t. 8. 1952. TYPE: Crest of the Sierra Maestra between
Pico Turquino and La Bayamesa, Peis. Oriente, Cuba, 1350 m, Morton & Acuna 3547 (US)

Grammitis perpusilla (Maxon) eiiorge comb. n
Polypodium perpusillum Maxon, Contr. U. S i! Herbs 173 ene “1914. TYPE: Serra de Caraga, Est.
Minas Gerais, Brazil, March 1892, Ule (US).

Grammitis shaferi —_ ee comb. no
Polypodium shaferi Ma atl. Herb. 17:410. aie TYPE: Camp La Gloria, S of Sierra
Moa, Pcia. Oriente, ie. ae Ao cite

Grammitis williamsii ao deseo! comb. n
Polypodium williamsii Maxon, Contr. U. S. Natl. Herb. 17:547, t. re 1916. TYPE: Near Apolo, Depto. La
Paz, Bolivia, 5000 ft. eink 1167 (US).

Microgramma ce Ube ge eg comb. nov.

Drynaria acuminata Fée, Cry Brés. bay Eo 1869. TYPE: Vicinity of Barra [Manaus],
Prov. Rio Negro [Est. Amaz abe il, Gee. in ee 51 (RB not seen). [Mickel and de la Sota (pers.
comm.) indicate that Fée’s epithet is nee oldest one for this species, which had been known as Polypodium
loretense Maxon].

Microgramma latevagans perce & C. Chr. in Maxon) Lellinger, comb. nov.
Polypodium latevagans Maxon & C. Chr. in Maxon, Proc. Biol. Soc. Washington 52:120. 1939. TYPE:
Unduavi, Depto. La Paz, Bolivia, 2400 m, hiss 36] (US; isotype NY not seen).

Microgramma recreense (Hieron.) Lellinger, — no
Polypodium recreense Hieron. Bot. Jahrb. Engler 34:537. 190 yer (chosen here): Near El
Recreo, Pcia. Manabi, Ecuador, Eggers 15838 (US: isolectotype Bn me SEER}

Microgramma tuberosa (Maxon) Lellinger, comb. n oe
Polypodium tuberosum Maxon, Amer. Fern J. 33:135. 1943. ie aa of La Chonta, Pcia, Oro,
Ecuador, Rose, Pachano & Rose 23468 (US).

Pecluma camptophyllaria var. lachnifera (Hieron. ) ee comb. n

Polypodium lachniferum Hieron. Bot. Jahrb. Engler 34:515. 1904. : Mt. igen Lbews
Tungurahua, Ec ador, Lehmann 458 (B not seen: iohecrotgties LE not seen, OS) mete by A. vans
(Ann. Missouri Bot. Gard. 55:254. 1968)

Pecluma eurybasis var. glabrescens (Rosenst.) rng ae ee bo ae seri
Polypodium lac Pia var. glabrescens Rosenst. Repert. Sp. . Fedde 11:57. ea n

North Yun ngas, Depto. La Paz, Bolivia, Buchtien 2770 (S not seen: pean US).

Pecluma eurybasis var. villosa (Evans) Lellinger, Sang OS 3 1968. TYPE: Above

Polypodium eurybasis var. villosum Evans, Ann. Missouri Bot. Gard. 55: 25, f sé F ie 19688
— just N of the mouth of Quebrada El Obispo. Depto. A eek Colombia, 2700 m, Fosberg .
(US)

60 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

paecsouge ptilodon var. robusta (Fée) geri comb. nov.
Polypodium robustum Fée, Crypt. Vasc. Bres. . TYPE: Rio de Janeiro, Est. Rio de Janeiro,
Braz zil, "Glee 2407 (presumably P not seen ol og

Pleopeltis pleolepis (Maxon & _—- in Copel.) compan oor
Polypodium pleolepis Maxon & Copel. in Copel. Univ. Calif. Publ. Bot : pit ‘oat. Hr Bo
Barranca del Rubelcruz, Depto. Alta ee ens. 3000 ft, von = ie a ns

Polypodium mckeei (Tindale) methnger, comb. n
Dictymia mckeei Tindale, Amer. Fern J. 50:122. 1960. TYPE: ace! to |’'Hermitage, Nouméa District, New
Caledonia, 400 m, McKee 2126 (NSW; isotype US).

Polystichopsis cubensis (Kuhn) Lellinger, comb.
Aspidium cubense Kuhn, Linnaea 36:108. 1869. TYPE: Cuba, hia 1099 (B not seen; isotype US).

Thelypteris a, ae bis Smith ex Lellinger, nom. nov.
Bi Segetes reducta C. _ Svenska Vetens.-Akad. Handl. III, 16(2):18, rt. 2, f. 1-3. 1937, non
elypteris reducta Small rine TYPE: Valle Nuevo, Pcia. La Vega, Dominican facet ca. 24
bites H13839 (C not seen; isotype US).

Thelypteris ireneae oehned Aeon cu. nov.
Dryopteris ireneae Brade, Sellowia 17:5 1965, as “irenae.” TYPE: Estrada do Contérno, Serra das
Araras, Est. Rio de Janeiro, eat 800 m, Sea 5609 (HB: isotype US).

Thelypteris scabra (Presl) en come nov

Polyvpodium scabrum Presl, Del. Prag. 1:169. IPE: Brazil, without collector (PR not seen).

I wish to thank Alan R. Smith and Michael G. Price for their helpful comments on
this paper.

GAULTERIO LOOSER (1898-1982)

Gaulterio Looser was born in Santiago, Chile of Swiss parentage on 4 September
18 He was an industrialist and an engineer and owned a factory that made
agricultural implements. Some of the manufacturing machinery was of his own
esign. He joined the American Fern Society in 1928 and started to publish papers
on the pteridophytes of Chile shortly thereafter. He was made an honorary member
of the Society in 1949. His herbarium, containing about 8,000 specimens, was given
for the most part to the Conservatoire et Jardin Botaniques, Geneva, Switzerland. He
was a Doctor Honoris Causa of the Universidad Catélica de Chile and of a Swiss
university. His interests and scientific output were broad; he published over 200
papers on ferns, archeology, anthropology, flowering plants, biography. phytogeog-
raphy, ecology, and geology. He died in Santiago on 22 July 1982.—From material
provided through the kindness of Dr. Clodomiro Marticorena.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984) 61

SHORTER NOTES

EQUISETUM RAMOSISSIMUM IN LOUISIANA.—In a recent article
(Hauke, R. L. 1979. Amer. Fern J. 69:1-5) I reported the occurrence of Equisetum
ramosissimum Desf. in North America, from one locality in North Carolina and
another in Florida. Dr. A. Murray Evans, of the University of Tennessee, who had
sent me the North Carolina specimen of E. ramosissimum wrote me in February
1982 about a collection of Eguisetum from Louisiana that he had noted “looks just
like the NC specimen.” Since the North Carolina specimen turned out to be E.
ramosissimum, he suggested I look at this one also.

The specimen in question (Point Coupee Parish, locally common in loose sand
near river, Morganza Spillway, St. Hwy 1, west of New Roads, La., 30 Sept 1962,
C. A. Brown & B. B. Exner 17610, LSU 000056) looked to me like E. x ferrissii,
but a scanning electron micrograph of the surface was needed to be certain. The
SEM shows clearly that the silica micromorphology is not that of E. X ferrissii, but
is consistent with that of E. ramosissimum or E. laevigatum. Anatomically, the stem
has both the carinal and the vallecular collenchyma prominent, a feature of E.
ramosissimum, but not of E. laevigatum. Also, the rows of stomata are occasionally
doubled, a character of E. ramosissimum. Hence, this specimen appears to docu-
ment the presence of Eguisetum ramosissimum in Louisiana, an addition to the
previous records of that species in North America.—Richard L. Hauke, Dept. of
Botany, University of Rhode Island, Kingston, RI 02881.

THREE NEW COMBINATIONS IN LOXOGRAMME.—As a result of my
annotating many specimens of Loxogramme in different herbaria with the following
names, I consider it expedient to publish them in advance of a treatment of the
genus as a whole:

Loxogram ini i b
me Price, comb. nov.
: sess capa ogra l { TYPE: Gerra, Abyssinica, Schimper

Gymnogramma abyssinica Baker, Syn. Fil., ed. 2: 517. 1874.
1445 (K).

Loxogram i i b V.
me cuspidata (Zenker) Price, comb. Nov. Se
Grammitis cuspidata Zenker, Plantae Indicae 1: t. 2. 1835. TYPE: Near Utacamund, Nilgiris, South

India, Rev. B. Schmid (not seen; isotypes BR, E, L. P).
Loxogra zal: i b Vv.
mme wallichiana (Hooker) Price, com). Nov. BS aie
Selliguea wallichiana Hooker, Icon. Pl. 3: t. 204. 1840. LECTOTYPE (chosen here): Penang,
Malaya, Wallich 10 (K). The other syntype is Penang, Malaya, 7
M. G. Price, Herbarium, North Univ. Bldg., Univ
MI 48109,

ady Dalhousie (8).
ersity of Michigan, Ann Arbor,

62 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984)
NOTES ON NORTH AMERICAN FERNS, II.—Since the publication of the

first paper in this series (Amer. Fern. J. 71:90—94), I have found a nomenclatural

tangle in one Dryopteris hybrid and two more combinations that need to be made:

Adiantum pedatum subsp. subpumilum (Wagner in Wagner & Boydston)
Lellinger, comb. & stat. nov.

Adiantum pedatum var. subpumilum Wagner in Wagner & Boydston, Canad. J. Bot. 56:1727, f. J.
1978. TYPE: South of Orchard Point, Brooks Peninsula, NW Vancouver Island, British Columbia,
Canada, Pojar & Boas 770191 (MICH; isotypes UBC, V

This subspecies, known in nature only from Vancouver Island, surely is at least as
distinct from subsp. pedatum as are subsp. aleuticum and the newly described
subsp. calderi Cody (Rhodora 85:93, f. /. 1983) from Quebec, Newfoundland, and
Vermont, with a few outliers along the coast of the northwestern United States.

Dryopteris Xslossoniae Wherry ex Lellinger, sp. nov.

Aspidium cristatum x marginale Davenp. Bot. Gaz. 19:494. 1894. AUTHENTIC SPECIMENS:
Boxford, Newbury, and Merrimac, Essex County, Massachusetts, Raynal Dodge in 1892 (GH ex Herb.
Davenport none seen; duplicates US and presumably elsewhere).

Dryopteris cristata X marginalis Davenp. Bot. Gaz. 19:497. 1894, nom. provis. et illeg.

Nephrodium cristatum =< marginale Davenp. Bot. Gaz. 19:497. 1894, nom. provis. et illeg.

Lastrea cristata < marginalis Davenp. Bot. Gaz. 19:497. 1894, nom. provis. et illeg.

Nephrodium slossonae Hahne, Allg. Bot. Zeitschr. 10:103. 1904, nom. nud.

Dryopteris < slossonae (Hahne) Wherry, Bartonia 21:15. 1942, nom. nud.

Plant hybrida inter D. cristatam et D. marginalem posita: ex D. cristata a

differt, ex D. marginali in laminis pallidioribus soris submarginalibus et ie
abortivis differt
re Rosie, Essex County, Massachusetts, Aug 1893, G. E. Davenport 4
)
The original publication of the epithet honoring the morphologist Margaret

Slosson was by Hahne in a paper on fern hybrids; Hahne listed the hybrid formulae
nown to him, mostly from Europe, along with their corresponding binomials. In
those cases where no binomials had been published, he proposed them, all as
nomina nuda. Because there is no indication that Hahne knew of Davenport's paper
describing Aspidium cristatum Xx marginale, or of Slosson’s (Pl. World 2:4-7.

1878) describing the same hybrid less technically, and because Davenport’s combi-
nation in Nephrodium is a provisional and illegitimate name, the works cannot serve
as the basis (protologue) for Hahne’s epithet. Wherry (Bartonia 21:15. 1942) did not
cite Davenport’s paper, and probably he did not see Hahne’s paper in accepting his
epithet, for he did not publish any description. Nor did his later citation (Fern
Guide. 128: 1961) of the hybrid formula Aspidium cristatum * marginale Davenp.
in synonymy under D. x slossonae validate the name because a hybrid formula is not
equivalent to a species epithet, and so a description accompanying a hybrid formula
is not a valid protologue. Therefore, the name Dryopteris X< slossoniae has until now
not been validly published, for it has lacked a description. Lastly, the correct
spelling of the epithet is slossoniae; the —ii following the consonant n must be
altered to —iae because of Slosson’s gender, in conformity with the International
Code of Botanical Nomenclature.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984) 63

Lycopodium X brucei (Cranfill) Lellinger, comb. nov.

Lycopodiella X brucei Cranfill, Amer. Fern J. 71:97. 1981. TYPE: Borrow pit ca. 200 m ENE of
road KY-280 0.6 mi from its junction with road KY-121, Calloway County, Kentucky, Bruce 76006
(MICH not seen).

This is the hybrid between L. appressum (Chapm.) Lloyd & Underw. and L.
prostratum Harper.

I wish to thank Drs. J. D. Montgomery, D. H. Nicolson, and A. R. Smith for
their helpful comments.—David B. Lellinger, Dept. of Botany, National Museum of
Natural History NHB-166, Smithsonian Institution, Washington, DC 20560.

GRAVES’ SPLEENWORT IN OHIO.—Asplenium * gravesii Maxon is the
apparent hybrid between A. bradleyi D. C. Eaton and A. pinnatifidum Nutt. Graves’
Spleenwort has been reported only from Alabama, Arkansas, Georgia, Illinois,
Missouri, Tennessee, and West Virginia (Cranfill, “Ferns and Fern Allies of
Kentucky,” p. 164-166, 1980; Wagner, Evolution 8:103-118. 1954: Wagner &
Darling, Brittania 9:57—63. 1957; Werth & Taylor, Amer. Fern J. 70:28. 1980).
Among these sources, Ohio also is included in the distribution of Graves’ Spleen-
wort by Cranfill and Wagner. A search of herbaria housing large collections of Ohio
pteridophytes has failed to locate a specimen confirming these reports. Moreover, the
important’ description of this hybrid by Wagner & Darling (1957) does not list this
taxon from Ohio. This is now a moot point, however, since a population of Graves
Spleenwort is extant in the state.

Three plants of Asplenium X gravesii were discovered on 22 February 1983 in
Hocking County, Ohio (Cusick 22355, MICH, OS). The site was revisited on 15
February 1984 in the hope of locating additional specimens, but no others were
found. The hybrid grows on a sunny, southeast-facing exposure of the Black Hand
Sandstone, a massive subacid sandstone of Mississippian age. The xeric, nearly
Vertical cliff also supports an extensive population of Asplenium x trudellii Wherry,
With scattered individuals of A. pinnatifidum. No plants of A. bradleyi_ were
discovered, despite a careful search of this exposure and of other rock faces in the
immediate vicinity. Indeed, Bradley’s Spleenwort has been collected only once in
this part of Ohio. A single population of A. bradleyi grew on a similar ve in
adjacent Fairfield County until at least 1971. It is no longer known from that site,
however. Three other spleenworts grow on rock exposures within 2 km of the <
* 8ravesii population. These are: A. montanum Willd., A. platyneuron (L.) B.S.P.,
and A. trichomanes L. The first two species are widely scattered through the
Whole valley system, while the Maidenhair Spleenwort is decidedly uncommon.

I thank Dr. Charles Werth, University of Virginia, for sharing with me his
thoughts on spleenwort hybrids and Daniel Rice, ODNR, for field assistance.
—Allison W. Cusick, Division of Natural Areas & Preserves, Ohio Department of
Natural Resources, Fountain Square, Columbus, OH 43224.

64 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 2 (1984)

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LITERATURE CITED

AMERICAN Edis eh OF BIOLOGICAL SCIENCES, Committee on Form and Style of the
Conference of Biological Editors. 1964. Style Manual for Biological Journals, ed. 2.
Washington, D.C.

Committee on Form and Style of the Conference of Biological Editors. 1972. CBE
e Manual, ed. 3. Washington, D.C.
HOLMGREN, PATRICIA K., W. KEUKEN, and EILEEN K. SCHOFIELD. 1981. Index Herbariorum.
The Herbaria of the World, ed. 7: Reg. Veg. 106:1-4

LAWRENCE, 6. M. et al. 1968. Botanico-Periodicum- Huntianum. Hunt Botanical Library,

ittsburgh.

SCHWARTEN, LAZELLA, and H. W. RICKETT. 1958. Abbreviations of titles of serials cited by
botanists. Bull. re Bot. Club 85:277-300. See also the supplement in Bull. Torrey Bot.
Club 88:1-10. 1961.

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The Habitat Characteristics and Abundance
of Equisetum x ferrissii and its Parent Species,
Equisetum hyemale and Equisetum laevigatum, in lowa
LORENZ M. RUTZ and DONALD R. FARRAR 65

The Organic Nutrition of Botrychium Gametophytes
DEAN P. WHITTIER 77

Anti-microbial Activity of Phenolic Acids
in Pteridium aquilinum
MICHAEL SAN FRANCISCO and GILLIAN COOPER-DRIVER 87

Reviews 86, 96

IAN fe

The American Fern Society

Council for 1984
TERRY R. WEBSTER, Biological Sciences Group, University of Connecticut, Storrs, CT 06268.
President
FLORENCE S. WAGNER, Dept. of Botany, University of Michigan, Ann Arbor, MI 48109.
Vice-President
MICHAEL I. COUSENS, Faculty of Biology, University of West Florida, Pensacola, FL 32504.
Secretary
JAMES D. CAPONETTI, Dept. of Botany, University of Tennessee, Knoxville, TN 37916.
Treasurer
DAVID S. BARRINGTON, Department of Botany, University of Vermont, Burlington, VT 05401.
Records Treasurer
DAVID B. LELLINGER, Smithsonian Institution, Washington, DC 20560. Journal Editor
ALAN R. SMITH, Dept. of Botany, University of California, Berkeley, CA 94720. Memoir Editor
DENNIS Wm. STEVENSON, Barnard College, Columbia University, New York, NY 10027.
Fiddlehead Forum Editor

American Fern Journal
EDITOR

DAVID B. LELLINGER U.S. Nat’] Herbarium NHB-166, Smithsonian Institution,
Washington, DC 20560.

TERRY R. WEBSTER ....... Biological Sciences Group, University of Connecticut, Storrs, CT 06268.

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AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 3 (1984) 65

The Habitat Characteristics and Abundance of
Equisetum ferrissii and its Parent Species,
Equisetum hyemale and Equisetum laevigatum, in lowa
LORENZ M. RUTZ and DONALD R. FARRAR*

Equisetum Xferrissii Clute is the naturally occurring sterile hybrid of E.
laevigatum A. Br. and E. hyemale var. affine (Engelm.) A.A. Eaton (Hauke, 1958).
Although it is morphologically distinct and relatively easy to identify, it does not
consistently appear on floristic lists of Iowa’s prairies, nor in broader discussions of
lowa or Midwest floras. Only recently has the commonness of E. X ferrissii become
recognized. Peck (1976) documented the presence of E. x ferrissii in 74 of lowa’s
99 counties. He felt (pers. comm.) that a thorough search would turn up E.
x ferrissii in the remaining counties as well, making it as widespread in Iowa as its
parent species. E. hyemale has been recorded in all counties, and E. laevigatum in
91 of 99 counties. The current study was designed to quantify the presence of E.
x ferrissii in one county considered representative of much of Iowa and to address
ecological questions regarding the dispersal and maintenance of the sterile hybrid
and its parental species.

Ecological information on these three Equisetum taxa is sparse. Hartman (1958)
recognized what are now known as E. x ferrissii and E. laevigatum as plants of
ditches and embankments, noting also that E. x ferrissii grew in moister sites than
E. laevigatum and that E. hyemale preferred a moister habitat than either E.
Xferrissii or E. laevigatum.

Because they are agricultural weeds and toxic to livestock, E. palustre and E.
arvense have received considerable attention. Borg (1971) did an extensive study on
E. palustre as a weed in Finland. He recorded rhizome growth of 30 cm to several
meters per year, reaching depths of 180 cm below the soil surface. In regard to
vegetative dispersal, he found that rhizome fragments of E. palustre were not good
propagules and did not support the idea that E. palustre was spread by road building
activity. a

As field weeds, E. palustre and E. arvense have been tested for perbientc
responses. E. arvense topgrowth was found to be sensitive to most chemicals, and
thizome growth could be retarded to various degrees (Hoyt & Corder, 1962).
Equisetum palustre, on the other hand, was found to be quite resistant to the
chemicals tested. Topgrowth could be damaged, but rhizomes remained virtually
unaffected (Holly, 1953). -

The ease of vegetative propagation in Equisetum has been noted severa ue
(Hauke, 1963: Praeger, 1934; Schaffner, 1931; Wagner and Hammitt, 1970). "he
reports concentrate on the proliferation of aerial culm fragments floating on water or
placed in or on saturated soil. Hauke (1963) suggested that E. X ferrissit culms =
transported outside the range of its parent species by human activity. He note ;
however, that preventing desiccation during translocation was critical for pub ge
Dosdall (1919) also found that culms rapidly lost viability upon eye ts
“Department of Botany, Iowa State University, Ames. IA 50011.

Volume 74, number 2, of the JOURNAL was issued July 18, 1984

66 AMERICAN FERN JOURNAL: VOLUME 74 (1984)
contrast to Borg’s findings (1971), that rhizome fragments were the most suitable
propagules for starting Equisetum in drained soil.

Sexual reproduction in Equisetum has been studied mostly in laboratory cultures,
but Duckett and Duckett (1980) described the ecology of in situ gametophytes of E.
arvense, E. fluviatile, and E. palustre in Great Britain. The sites described were
previously flooded muds, bare of vegetation. They found that spores and gameto-
phytes had a very low range of moisture tolerance and proposed this as an
explanation for the frequent absence of gametophytes in apparently suitable sites.

Gametophytes in the American Midwest were discovered by Walker (1921, 1931)
on stream banks in Iowa, Nebraska, and Kansas. All gametophytes were found on
the narrow mudbelt along the water, typically associated with Riccia. Unfortunately,
Walker’s identifications must be treated with caution for two reasons. She reported
gametyophytes of “E. kansanum” and E. “laevigatum,” which are now known
respectively as E. laevigatum and the sterile (!) E. Xferrissii, and, according to
Walker (1931), the identifications depended “entirely upon the species growing in
the vicinity.”

fey 4
oe @
92° ae £24 ;
‘ = Field
o> 6
S <
S a ie
9° o
Fes ay oy
=
Depth @

Width
Fence-
row
Lisanti pels ar oe er eis

FIG. 1. Schematic cross section of a typical lowa road showing ditch features; not drawn to scale.

METHODS

Natural areas in central and northwestern Iowa were visited to gain familiarity
with the three taxa in their native habitats and in roadside ditches. Hauke’s (1963)
monograph was used to identify the plants.

To quantify the relative frequencies of the three taxa, and to obtain ecological
information on ditch populations, a survey of 64 miles (103 km) of roadside in Story
County, central lowa, was made on foot in the summer of 1981. Story County 1s
composed of 16 townships of 36 sections each. Sections are one mile (1.6 km)
square and usually are delimited on four sides by roads. The roads are bordered on
each side by ditches 4-10 m wide and 1-1.5 m deep, depending on the road type
(Fig. 1). Information regarding the design, construction, and maintenance of Iowa’s
roads and ditches was obtained from the Iowa Department of Transportation (IDOT),

RUTZ & FARRAR: HABITAT CHARACTERISTICS AND ABUNDANCE OF EQUISETUM 67

TABLE 1. PRESENCE OF Equisetum SUBGENUS Hippochaete IN IOWA PRESERVES OF
NATIVE VEGETATION.

Vegetation Equisetum Equisetum Equisetum
Preserve type laevigatum x ferrissii hyemale
Bixby State Park Forest - = 4
layton Co.
Black’s Prairie Prairie + + -
tory Co.
Cayler Prairie Prairie + + _
Dickinson Co.
Crossman Prairie Prairie + i
oward Co.
Dolliver State Park Forest = = x
ster Co
Doolittle Prairie Prairie aie 5 mE
ory Co.
Freda Haffner Kettlehole Prairie rk aE a
kinson Co.
Gitchie Manitou State Preserve Prairie & + + +
Lyon Co. forest
Hayden Prairie Prairie ae + .
Howard Co.
ee gee *
Kalsow Prairie Prairie - =
Pocahontas Co
Ledges State Park Forest oe ng. =
Boone Co.
Woodman Hollow State Forest as ne i
eserve, Webster Co.
+ = present, — = not observed, * = present only as invasion from bordering roadside ditch

the Story County Engineer, and the Story County Weed Commissioner.

The survey unit was the four ditches bordering a section. If any of the ~— were
missing, the equivalent ditch of the adjacent section was surveyed. One primary
section in each of eight alternating townships was selected randomly. Along primary
sections, the following ditch and Equisetum stand characeristics were measured:
drainage, topographical position, ditch depth, apparent soil drift, species aang
(number of species present), road type, ditch position relative to field (N, E, 2 :
estimate of snow drift, *type and width of fencerow, *adjacent field use, . .
material, *slope of ditch bottom, kind of Equisetum, overall density of the -~- :
maximum density found in the stand, height of Equisetum, stand length, stan
Width, *percent of culms with cones, and *health of plants. Asterisked characteris-
tics were found to be closely linked to others or essentially non-varying. For clarity
of presentation, these are not included in the results. : ee

To obtain a larger statistical base for abundance data, two “secondary —
Were likewise selected from each of the eight townships. Along these sections, only
the taxon present, stand length, and overall density were recorded. ae ar
Secondary sections is included only in Table 2. Equisetum stands typically eae
the entire ditch from fence row to road bed. Thus stand length, eed aa e
the road, was directly proportional to the total physical extent of the stand.

68 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

TABLE 2. RELATIVE ABUNDANCE OF Equisetum laevigatum, E. * ferrissii, and E. hyemale IN
ROADSIDE DITCHES IN STORY COUNTY, IOWA.

Sum of Average No. of stands!
stand stan
No. of length length Low Medium High
Taxon Sections stands (m) (m) density ensity density
E. laevigatum imary 166 22,370 135 0 5 By
Secondary” 198 22,445 113 93 74 34
Total 364 44,723 123 173 133 87
E. X ferrissii i 3,868 64 a3 7
Secondary” 2,923 59 40 10 1
Total 110 6,791 62 93 17 I
E. hyemale im 8 611 l 3 4
Secondary” 9 1,363 152 2 1 6
Total 17 1,974 116 3 4 10
All Grand total 49] 53,291 109 268 154 98

'Low = 0-50 culms per square meter, med = 50-200, high = 200-400. Some extensive stands were
measured at several points along their length.

*Time constraints allowed the survey of only half of the secondary sections initially selected, con-
sequently their distribution was biased towards the north portion of the county.

OBSERVATIONS AND RESULTS IN NATURAL HABITATS

Prior to European settlement, Iowa lands formed a network of upland prairies,
prairie potholes, and prairie and woodland streams, providing an abundance of
habitats suitable to E. x ferrissii and its parent species. The distribution of the three
taxa, as observed in natural areas of a number of Iowa’s parks and preserves, reflects
this earlier distribution (Table 1).

Equisetum hyemale is common on the banks and terraces of Iowa’s creeks and
rivers. It may become a dominant part of the understory of the gallery forest,
attaining densities, at Ledges State Park for example, of 500 culms per square meter
over large areas. Stands continue along tributary branches to various degrees,
sometimes well into steep gullies with only intermittently flowing water. Low
density stands occasionally appear on old terraces well above the present floodplain.
An example of this occurs at Woodman Hollow State Preserve. Equisetum hyemale
grows on or very near the water table. Subsurface soils of this habitat are unlikely to
ever become dry.

Stands of E. laevigatum and E. x ferrissii in native Iowa habitats always occur in
full sunlight on prairie uplands. In undisturbed prairie, both may occur conspicu-
ously or in very low densities. The presence of E. laevigatum at Crossman Prairie
was substantiated by only two individual culms. In disturbed areas of native prairies,
densities become substantially higher. Along an old drainage ditch cut through the
northwest corner of Kalsow Prairie, for example, E. laevigatum attains densities of
60 culms per square meter. E. ferrissii is less frequent than E. laevigatum, but it
also 18 not uncommon in native prairie habitats.

Soils in Iowa’s prairies range from dry to very wet, reflecting the poor drainage
conditions of much of northern and central Iowa prior to artificial drainage by
European settlers. As with soils of the native riparian habitat of E. hyemale, it is
unlikely that the more poorly drained prairie soils ever become dry. Equisetum
laevigatum, however, grows on higher, well-drained sites, along the rim of the Freda

RUTZ & FARRAR: HABITAT CHARACTERISTICS AND ABUNDANCE OF EQUISETUM 69

Haffner Kettlehole in northwestern Iowa, for example. The substrate there is a
coarse glacial till, which becomes very dry in summers of sparse rainfall. E.
x ferrissii grows in the same area, but is confined to the lower slopes of the kettle
im

Two sites with Equisetum gametophytes were located. Sporophytes from these
gametophytes were transplanted, allowed to mature in the greenhouse, and identified
as E. hyemale in both cases. In the summers of 1980 and 1981, gametophytes of E.
hyemale were common on the vertical cut bank of the Des Moines River at Ledges
State Park. A few gametophytes were also found on the silts and muds of the level
floodplain a few meters back from the bank. The second site was along the cut
banks of Kegley’s Creek in Story County (section 36, Lafayette Township). The cut
banks at both sites were 1 —2 m high, kept moist by constant seepage, faced north
to west, and received only late afternoon sun, if any at all. New sporophytes were
common in late summer, but their fate was uncertain, as frost wedging cleaves most
of the bank faces, burying the sporelings in mud. Associated plants included
Ricciocarpus natans and Riccia frostii, several unidentified mosses, and a variety of
angiosperm seedlings and annuals.

OBSERVATIONS AND RESULTS IN ROADSIDE DITCHES

As the prairies were converted to farmland, roads were cut into the landscape in
the middle 1800's. Equisetum x ferrissii and E. laevigatum were displaced from
their native prairie but found a suitable habitat along the roadsides where they are
now found in abundance. Today, with the virtual elimination of prairies, roadside
ditches have become the primary habitat for E. x ferrissii and E. laevigatum, and
often provides suitable habitats for E. hyemale as well. ae

Characteristics of ditch stands of Equisetum.—The number, size, and distribu-
tion of stands among three density classes of each taxon was surveyed (Table 2).
Equisetum laevigatum was by far the most common, having the largest number of
stands and the greatest sum of stand length. Because of the difficulty in delimiting
individual stands, the latter figure probably expresses the dominance of E. laevigatum
more meaningfully than does the number of stands. The total length of ditch
occupied by E. laevigatum was over six times that of E. x ferrisstl, which in turn
was over three times that of E. hyemale. The total stand length of E. laevigatum
alone represents 43% of the 103 kilometers surveyed. Discounting overlaps In
stands, 52% of the extent of central lowa’s roadside ditches contains one of these
three Eguisetum taxa.

The Sianbation of stands in three density categories is different for each of the
taxa (Table 2). Equisetum laevigatum stands regularly occur In all categories, re a
more frequently of low or medium density. Equisetum x ferrissil occurs oop
exclusively as low density stands. Equisetum hyemale shows the opposite, ces
mostly as high density stands. This relationship 1S repeated in the mean be ‘ ao
shown in Table 3. The maximum density also indicated that E . X ferrissit ( : = .
m*) had by far the lowest density of the three taxa. Equisteum hyemale = s
highest mean maximum density (156 culms m2), but did not greatly exceed ©.
laevigatum (105 culms m7”).

70 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

BS E. laevigatum
60--
E. X ferrissii

=
50 E.hyemale

w
a
z
<q
% 40h
Be a ie
°o
x 30h

lof

an = ee ee 2
v
UPLAND STEP SWALE SWELL SLOPE TERRACE FLOODPLAIN

a
FIG. 2: The distribution of roadside stands of E. laevigatum, E. X ferrissii, and E. hyemale by
topographical position. Values are the percent of the total number of stands for each taxon. A—A’ =
topographical cross section illustrating the categories listed.

The mean species richness (Table 3) in the ditches fell near nine for stands of all
three taxa, a low, but expected value for such an impacted environment. Species
richness ranged from | to 24 species, in addition to the Equisetum present. At
several locations the single species growing with E. laevigatum was Bromus inermis.

‘he mean index values for road type, ditch depth, drainage, and soil drift (Table
3) indicate that E. hyemale occurred more frequently in deep ditches along paved
roads than did E. laevigatum and E. x ferrissii; the latter tended to occur in
shallower ditches along gravel roads. Deeper ditches correspond with greater soil
and snow accumulation and poorer drainage.

The topography of Story County was classified into the seven categories dia-
grammed in Figure 2, and the distribution of stands among these categories
recorded. Equisetum laevigatum had a higher proportion of its stands in well drained
areas (slopes and swells) than did E. x ferrissii. In less well drained areas (steps,
Swales, and terraces), E. x ferrissii had a higher proportion than E. laevigatum.
Both were infrequent on the floodplains where E. hyemale had the greatest
proportion of its stands. All three taxa occurred on the level upland.

The topographical preferences of Equisetum are repeated in the position of the
maximum stand densities within ditches (Table 4), Equisetum hyemale grows most
densely in the bottom of the ditch more often than does E. laevigatum or E.

RUIZ & FARRAR: HABITAT CHARACTERISTICS AND ABUNDANCE OF EQUISETUM 71

TABLE 3. SELECTED MEANS FOR VARIABLES ON STANDS OF Equisetum IN PRIMARY
SECTIONS, WITH STANDARD ERROR FOR CONTINUOUS VARIABLES IN PARENTHESES.

Variable E. laevigatum E. X ferrissii E. hyemale

No. of stands 166 60 8

Max. density 105(7.8) 21(3.6) 156(52)

culms per m7)

Overall density! 8 1.1 73

Height (cm.) 70(1.5) 66(2.7) 92(9.1)

Diversity” 8.6(0.37) 8.5(0.60) 9.1(1.01)
oad type* 21 a2 2.4

Ditch depth (ft.) 2.7(0.09) 2.5(0.13) 4.1(0.65)

Drainage Pees 1.9 12

Soil drift° 9 2.0 2.9

Snow drift® 5 1.6 1.9

; or, 2= fair, 3 = g00
‘ =none, 2=light, 3= medium, 4= heavy
1=light, 2= medium, 3 = heavy

TABLE 4. POSITION OF MAXIMUM DENSITY IN THE DITCH PROFILE AS DIAGRAMMED
IN FIGURE 1!
Aspect of ditch slope

E WwW

N Shoulder Bottom
E. laevigatum 1% 31% 19% 18% 16% 9%
- X ferrissii 15% 26% 22% 1% 21% ok
E. hyemale 9% 9% 18% 18% 18% 27%

'Since maximum densities sometimes occurred in more than one position in a single stand, the total
number of scored positions is somewhat greater than the total number of stands.
Xferrissii, the latter preferring the slopes and shoulders. Equisetum laevigatum
shows this effect most strikingly, often growing hedgelike, tallest and densest, on the
ditch shoulders (Fig. 5). Of the slopes available, south-facing seems to be most
preferred by E. laevigatum and E. ferrissit. 3
Characteristics of the ditch habitat.—To ecologically evaluate the roadside
habitat of Equisetum in Iowa, it is necessary to consider the activities and conditions
imposed upon plants in this habitat. The factors influencing the distribution and
abundance of Eguisetum in the ditch can be considered in two groups: human
activities of construction, maintenance, and vegetation management, and inherent
itch characteristics, including the incidental effects of human activity.
he roads in Iowa can be classified into four categories: (1) interstate and other
limited access highways, (2) paved state an
im and (4) dirt road ie access highways contain many Equisetum stands
ut these were not studied for legal and safety reasons.
minimally graded, lack the aes ditches characteristic of the other sa — oy
be bordered directly by row crops or vegetation similar to that in the itches.
roads were included in the survey as they appeared in the selected sections. ian
far the most numerous roads are the paved state and county roads an =
gravel section roads. The most important feature of these two groups 1S :
conspicuous ditch on either side of the roadbed (Figs. /, 3-5). On new constructio

2 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Figs 3. Winter ditch in say County, stg showing snow drift deposited by northerly winds. FIG.
. Newly excavated ditch in Story County, Iowa. Vegetation in distance is still intact. FIG. 5. June
pe vegetation, showing strong growth eo E. laevigatum along roa

and reconstruction projects, these are cut to an average depth of five feet (1 -5m) for
paved roads and three feet (0.9 m) for gravel roads. The ditches have two functions:
water drainage and capture of drifting snow in winter to maintain a clear road with
minimal plowing (Fig. 3). With time, the ditches become partially filled with soil
and gravel, necessitating reconstruction.

Since there are few abrupt topographic changes in central Iowa, relatively little
earth is moved in preparing new roadbeds. Most earth moving involves onsite
relocation of ditch soil to the roadbed. However, in reconstruction, soil is often
relocated considerable distances from shallow ditches to low areas on the roadbed.
In one case which we observed, soil was transported over three kilometers. The
excavated ditch, in this case, showed broken ends of Equisetum rhizomes, indicating
that rhizome material was being relocated. These activities virtually eliminate other
existing ditch vegetation (Fig. 4), but seem to have no long-term negative effects on
Equisetum because the rhizomes generally extended deeper than the excavation. The
remaining Equisetum rhizomes quickly resume growth in an environment with little
competition from other species.

Another maintenance activity involves gravel roads only. About once per week
during the summer, gravel roads are graded to smooth their surface. Two passes are
made, moving gravel and chance roadside vegetation first to the road center, then
across to the opposite shoulder. Inspection of debris moved in this operation
frequently revealed Equisetum rhizomes.

RUTZ & FARRAR: HABITAT CHARACTERISTICS AND ABUNDANCE OF EQUISETUM 73

Story County began a blanket 2,4-D spraying program shortly after World War II.
The program lasted 25 years, ending in 1973. As a result, populations of dicotyle-
donous plants were reduced in size, if not eliminated. Only spot spraying is now
practiced by the county; consequently, native and introduced dicotyledonous flower-
ing plants are again becoming more common. Private spraying occurs along the field
side of some ditches where farmers attempt to control weeds. The Equisetum taxa in
this study occasionally showed herbicide damage, but were not eliminated by any of
the commonly used chemicals. This herbicide resistance was emphatically attested
to by farmers, extension personnel, and by the County Weed Commissioner.

The IDOT reseeds new or reworked roadsides with Bromus inermis (Smooth
Brome), Festuca arundinacea (Kentucky 31 Fescue), Coronilla varia (Crown-vetch),
and Lotus corniculata (Birdsfoot trefoil). County seeding is the same, with the
addition of some prairie grasses, i.e. Andropogon gerardii (Big Bluestem) Panicum
virgatum (Switch Grass), and Sorghastrum nutans (Indian Grass). Additional prairie
species have been used in experimental plantings (Landers, 1968, 1970), but these
have not been incorporated into the general management program for roadside
vegetation.

The ditches are mowed by state and county governments and property owners.
Official mowing is limited to one sickle bar’s width along the road edge. Private
mowing is not prohibited, however, and these practices range from manicured lawn
to short, isolated cuts with sickle bar once to several times in mid- to late summer.
Equisetum does not regenerate with sufficient rapidity to survive repeated close
mowing. The effects of sickle bar cutting depend on the timing, frequency, and
thoroughness of the cuts. Equisetum laevigatum and E. x ferrissii put up a single
flush of culms in late spring. Early cuts can be quite damaging, but the later the cut,
the less damage to the stand. After a first flush, E. hyemale adds new culms all
summer. It can recover from individual cuts, but a high frequency of cuts is more
damaging to that species. Most ditches escape mowing entirely, either by choice of
the landowner, or because they are too deep and narrow for mowing machinery. ;

A few landowners burn their ditches regularly. Equisetum laevigatum, in a ditch
burned annually, was exceptionally healthy, uniform, and quite attractive.

Where roads cut through the rolling topography of Iowa, their ditches become
areas of less extreme moisture conditions. In low, wet areas, the ditch locally
depresses the water table and is artificially drained, making it a drying factor. On
high, dry areas, the ditch is more moist than surrounding land, being closer to the
Water table and serving as a sink for seepage and runoff. In both situations, the ditch
generally provides a drained but consistently moist habitat.

The ditches are bordered on both sides by extensive areas 0 ;
Soil resources. The roadbed may be built of rich topsoil removed from the .
Adjoining fields (in 90% of the surveyed Equisetum stands) are fertilized an
cultivated to within a meter or less of the ditch shoulder. On the other hand, soil in
the ditch bottom is usually poor, being only slightly altered glacial till. It generally
1S dense and calcareous, and has little organic materia

The ditch serves as a sink for various materials. Chemic
the form of nutrients as well as toxic herbicides and pesticid

f unused or underused

hemicals leach from fields in
es. Automotive wastes

74 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

and trash drift in from the roadside. Large quantities of road gravel and topsoil
accumulate, the latter delivered by winter winds and spring and summer runoff.
hen adjacent to fields without mulch or standing crops, dormant vegetation may
be covered by several centimeters of powdered topsoil. Drifting snows accumulating
in the ditches (Fig. 3) provide winter protection for ditch plants and augment spring
moisture.
DISCUSSION

Equisetum x ferrissii and E. laevigatum were inhabitants of the once extensive
Iowa prairies. The principal habitat of these taxa in Iowa at present is the continuous
system of roadside ditches. Equisetum laevigatum is by far the most frequent,
occurring to some extent in each mile of road surveyed. FE. Xferrissii stands are
usually much smaller, but their number is almost one third as great as that of E.
laevigatum. Equisetum hyemale, a natural component of the gallery forests along

owa’s streams, has also successfully colonized these roadsides. It is the least
frequent of the three taxa in this habitat, but thrives where it is established.

The current high frequency of E. laevigatum and E. x ferrissii probably reflects
their broad distribution before European settlement. The occurrence of F. laevigatum
in all eight prairie preserves explored by the authors suggests that it was a ubiquitous
component of the prairies that once covered most of the state. Equisetum ~ ferrissii,
though not so frequent in the prairies as E. laevigatum, is present in over half of the
prairies surveyed and may also be assumed to have been quite common. As the land
was surveyed and roads were cut in the middle 1800’s, E. laevigatum and E.
ferrissii probably lingered to some extent, more or less in place, and adopted the
roa rders as suitable habitats. Their current abundance cannot, however, be
entirely explained through their historical occurrences. Few new roads are being
built in Iowa today, but old ones are continually being maintained with attendant
disturbance to ditch vegetation. Both taxa are present along newly constructed
interstate and state highways. Furthermore, E. hyemale is not a natural component of
the prairies; its presence in the ditches is due to colonization from some distance.
Therefore, an explanation of the relative abundance of the three Equisetum taxa in
the ditches must also include current patterns of reproduction and dispersal.

If one accepts Walker’s (1921, 1931) identification of wild E. /aevigatum
gametophytes, they occur in the same habitat as those of E. hyemale described
herein. Assuming temporal concurrence, this habitat concurrence makes the sexual
formation of E. Xferrissii, although undocumented in nature, regularly possible,
and its abundance in pre-settlement vegetation understandable.

Although possible, it is unlikely that existing ditches can provide the combination
of conditions necessary for the sexual formation of E. x ferrissii, considering the
narrow habitat tolerance of gametophytes (Duckett & Duckett, 1981). Although
excellent habitat for mature sporophytes, modern ditches are too exposed, too well
drained, and vegetatively too dense to provide suitable gametophyte habitats. It 1S
conceivable that in early days of road construction in Iowa, the ditches were subject
to greater erosion and greater moisture seepage, since the land was not yet
artificially drained. Such ditches would have been more similar to stream banks, and
perhaps suitable for gametophytes of E. laevigatum and E. hyemale.

RUTZ & FARRAR: HABITAT CHARACTERISTICS AND ABUNDANCE OF EQUISETUM 75

Although Duckett and Duckett (1981) feel that sexual reproduction must occur for
Equisetum dispersal, we feel that for the ditch habitat and the three taxa in this study
it is not essential. Their presence and abundance in the ditches can be attributed to
vegetative dispersal and persistence alone. Growth of rhizomes is rapid and can
account for clonal expansion over large areas of continuously suitable habitat. If
Borg’s (1971) conclusions regarding the clonal spread of E. palustre are applied to
these three taxa, it is also possible that their rhizomes could traverse the 30 feet (9.1
km) of roadbed between ditches. A more likely mechanism for crossing gravel roads
is transport by grading.

Expansion of clones cannot account for the many disjunct populations found in
the ditch habitat; some form of dispersal must occur. Aerial stem fragments of
Equisetum have excellent propagative properties (Hauke, 1963; Praeger, 1934;
Schaffner, 1931; Wagner & Hammitt, 1970), but their lack of resistance to
desiccation and the general lack of continuous standing or moving water in the
ditches make it improbable that culm proliferation accomplishes dispersal in
ditches. Rhizome fragments transported in soil are far more likely propagules.
Although apparently not the case for E. palustre (Borg, 1971), rhizome fragments of
E. hyemale, E. laevigatum, and E. ~ ferrissii maintain their viability if kept moist,
and under such conditions even small fragments sprout readily (Rutz, 1982).

Earth-moving activities involved in road construction and maintenance also move
Equisetum rhizomes and provide a very effective dispersal mechanism for these three
taxa. Rhizomes are transported to adequately moist sites bare of competing vegeta-
tion. Once established, plants of Equisetum rapidly produce extensive rhizome
systems (Borg, 1971; Rutz, 1982; Weaver & Albertson, 1943). Standard earth-
moving practices are unlikely to eradicate deep rhizomes from a ditch site once they
have become established.

Ditch habitats provide conditions both beneficial and detrimental to plant growth.
Ditch moisture conditions are ideal for Equisetum, being drained but continually
moistened by seepage and runoff. In the adjacent fields and on roadbeds, nutrients
and moisture are available to plants able to exploit them. That the extensive rhizome
system enables Equisetum to do this is evidenced by the appearance of culms up to
10 meters into cultivated fields, from the observation that Equisetum Is a nuisance in
plugging drainage tiles, and from the frequently observed increase in stand height
and density along the road edge (Fig. 5).

Detrimental characteristics of the ditches inc
other chemicals and the constant additions of soil and gravel. However, E. hyemale,
E. laevigatum, and E. x ferrissii, like E. palustre (Holly, 1953), appear t me ag:
immune to herbicide damage and possibly benefit through reduced competition 0
dicotyledonous plants. Likewise, soil sedimentation presents no problem to the

thizomatous Equisetum. :
Perhaps ie onty aspect of the ditch habitat truly detrimental to nucle
mowing. None of the three Equisetum taxa can SUTVIVe repeated close mowing, :
this condition is rare. The degree to which they are affected by maintenance atte .
depends on frequency and timing of the cuts. Generally mowing aims to € imin
dicotyledonous weeds and is not of sufficient frequency t0 damage Equisetum.

lude the high level of herbicides and

76 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Although artificial, the continuous network of Iowa’s roadside ditches is a
significant habitat, serving as a refuge for many native species. The common
occurrence of all three Equisetum taxa in the roadside habitat and the suitability of
this habitat to quantitative survey techniques and experimentation provide an
excellent opportunity for the ecological comparison of the three taxa. We feel that
results based on roadside ditch observations and experiments provide relevant insight
into the ecology of Equisetum in native habitats, as well as providing an explanation
for the remarkable success of the sterile hybrid E. x ferrissii and its parent species
in Iowa’s roadside vegetation habitats.

LITERATURE CITED

BORG, P. J. V. 1971. Ecology of Equisetum palustre in Finland, with special reference to its role as a
noxious weed. Ann. Bot. Fennici 8:93—144.
DOSDALL, L. 1919. Water requirement and adaptation in Equisetum. Plant World 22:1-13, 29-44.
DUCKETT, J. G., and A. R. DUCKETT. 1980. ascealapialaag Soi and population dynamics of wild
etophytes of Equisetum. Bot. J. Linn. Soc. 80:1
jopuca E. L. 1958. The taxonomy and ecology of the, het laevigatum complex. Trans.
sas Acad. Sci. 61:125—148.

HAUKE, R. "1988. Is Equisetum laevigatum a hybrid? Amer. Fern. J. 48:68—72.

Tkaceoreeteane . A taxonomic monograph of the genus Equisetum, subgenus Hippochaete. Beih. Nova
Hedwigia 8:1-123.

HOLLY, K. 1953. The effect of synthetic growth regulator herbicides on Equisetum palustre. Proc. First
Brit. Weed Control Conf., pp. 227-233.

HOYT, P. B., and A. C. CARDER. 1962. Chemical control of field horsetail. Weeds 10:11 1-115.

LANDERS, R. Q. 1968. Using Iowa’s prairie species to fight roadside weeds. Iowa Farm Sci.
22:13-14.

. 1970. The use of prairie grasses and forbs in lowa ees and park landscapes. Second

Midwest Prairie Conference, University of Wisconsin, Madiso

PECK, J. H. 1976. The pteridophyte flora of Iowa. Proc. Iowa Acad. Sci. 83:143-160.

PRAEGER, R. L. 1934. Propagation from aerial shoots in Equisetum. J. Bot. Brit. For. 72:175—-176.

RUTZ, L. M. 1982. The biology of the genus Equisetum, subgenus Hippochaete in central Iowa. M. S.
Thesis. Iowa State University, Ames, Iowa.

SCHAFFNER, J. H. 1931. Propagation of Equisetum from sterile aerial shoots. Bull. Torr. Bot. Club
58:531-535.

WAGNER, W. H., Jr. and W. E. HAMMITT. 1970. Natural proliferation of floating stems of scouring

rush, Equisetum hyemale. Mich. Bot. 9:166—174.

WALKER. R. 1921. The gametophytes of Equisetum noah ites Bot. Gaz. 71:378-391.

seers, S958; Ee irae of three species of Equisetum. Bot. Gaz. 92:1—22.

WEAVER, J. E., and F. W. ALBERTSON. 1943. sient of sub forbs and underground plant
parts at the end of pee drought. Ecol. Monogr. 13:63-

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 3 (1984) 77

The Organic Nutrition of Botrychium Gametophytes
DEAN P. WHITTIER*

In studies on the green, thalloid gametophytes of leptosporangiate ferns, sugars
have been added to the nutrient media to promote gametophyte growth. The
commonly used sugar has been sucrose (Dyer, 1979). In comparative studies with
several sugars, sucrose and glucose have given the best growth (Hurel-Py, 1955;
Whittier, 1964). Other sugars which have supported the growth of some gameto-
phytes are fructose, maltose, ribose, and xylose (Hurel-Py, 1955; Courbet, 1957;
Whittier, 1964). Other studies have shown how exogenous sugars can alter gameto-
phyte development and morphology (Whittier & Steeves, 1960).

Very little is known about the organic nutrition of the white, fleshy gametophytes
of the Ophioglossaceae, compared with the green, thalloid gametophytes of the
leptosporangiate ferns. It is generally accepted that the mycorrhizal fungus supplies
the organic nutrients required because young gametophytes from germinating spores
fail to grow beyond a few cells unless infected by a mycorrhizal fungus (Campbell,
1911) and mature gametophytes from nature always contain an endophytic fungus.
There have been a few reports of gametophytes of the Ophioglossaceae in nature
turning green at the soil surface, but it is doubtful if these gametophytes were
self-sufficient (Boullard, 1963). The few studies on Ophioglossum and Botrychium
gametophytes in axenic culture have shown that the gametophytes will grow to
maturity if the nutrient medium contains sucrose (Whittier, 1972; Gifford &
Brandon, 1978: Whittier, 1983). Thus, the endophytic fungus is not required for
gametophyte growth or normal morphology in axenic culture.

The seeds of orchids are similar to the gametophytes of the Ophioglossaceae
because they require a mycorrhizal fungus in nature and grow on nutrient media
supplemented with soluble sugars in axenic culture. Orchid seeds will not germinate
on a medium containing insoluble carbohydrates in axenic culture. Arditti (1967)
has noted that if orchid embryos did exhibit extracellular digestion, the role of the
mycorrhizal fungus would have to be reexamined. It has never been determined if
Botrychium gametophytes can metabolize insoluble organic materials in —
culture or from the soil. Any possibility that these gametophytes can obtain all or
some of their carbon from the soil should be investigated. Preliminary tests can be
performed to determine if these gametophytes exhibit extracellular digestion with the
techniques of axenic culture.

Since only sucrose has been use
Ophioglossaceae, additional studies are nece
nous carbon sources, soluble or insoluble, wi

d as a carbon source for the gametophytes of the
ssary to determine what other exoge-
1] support gametophyte growth. More

gametophytes.

ne
*Department of General Biology, Vanderbilt University, Nashville, TN 37235.

78 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

MATERIALS AND METHODS

This study was initiated with spores of Botrychium dissectum f. obliquum (Muhl.)
Fern. collected in Nashville, Tennessee. Voucher specimens of plants from which
the spores were collected are on deposit at the Vanderbilt University Herbarium
(VDB). The experimental conditions were tested on both spores and gametophytes
of B. dissectum. The spores were surface sterilized and sown on nutrient media by
the methods of Whittier (1973). For the experiments which were initiated with
gametophytes, the gametophytes were supplied from stock cultures maintained on
the basic mineral medium with 0.5% sucrose in the dark.

The basic mineral medium was composed of Knudson’s solution of mineral salts,
FeEDTA, minor elements, 0.6% agar, and had a pH of 6.1 (Whittier, 1973). The
cultures were grown on 15 ml of medium in 125 x 20 mm culture tubes with screw
caps. The cultures were maintained in the dark at 24+1°C. The duration of the
experiments averaged 4 months for those initiated with spores and 3 months for
those initiated with gametophytes.

The majority of the experiments involved supplementing the basic mineral
medium with various potential carbon sources. The organic supplements and their
concentrations are given with the results. There were several replicates for each
treatment, and many experiments were repeated. The percentage of germination was
obtained by observing 1000 spores from each experimental treatment. Thirty young
gametophytes growing from spores and an average of 20 older gametophytes were
measured to determine the gametophyte growth for each treatment. It should be
noted that spores of B. dissectum will germinate and the young gametophytes will
undergo a limited amount of growth on a nutrient medium without a carbon source.

Growth of the young gametophytes from spores is expressed as the percent
increase in gametophyte length over spore diameter. In experiments with older
gametophytes, growth is expressed as the percent increase in length over the initial
gametophyte size. In the experiments which were initiated with measured older
gametophytes, efforts were made to start with gametophytes of similar size. All
measurements were made with a calibrated ocular micrometer.

In many experiments, especially those on the effect of substances other than
sugars, a medium with 0.5% sucrose was included as a control to demonstrate that
the spores were viable and that the older gametophytes, obtained from stock
cultures, were alive and capable of growth. The growth of the gametophytes on the
medium with 0.5% sucrose should only be used for comparative purposes within
individual experiments. The percent increase in the lengths of both young and older
gametophytes on 0.5% sucrose varied considerably among the experiments for two
reasons. The durations of the experiments varied to some extent. Secondly, the
initial size for the older gametophytes was not constant for all experiments. Thus,
any increase in length would give a higher percent increase in length if smaller
gametophytes were used instead of larger gametophytes to initiate an experiment.

The data on gametophyte growth are presented with the standard error of the
mean. The analysis of variance and Duncan’s multiple range test (Li, 1964) were
used to determine if the differences among the means were statistically significant at
the 1% confidence level. Within individual experiments, the means for the growth of

D. P. WHITTIER: ORGANIC NUTRITION OF BOTRYCHIUM GAMETOPHYTES 79

TABLE 1. THE EFFECT OF SURFACE STERILIZATION OF SPORES ON YOUNG
GAMETOPHYTE GROWTH.

% increase in
Surface sterilized gametophyte length % germination
Spore 880.7+50.8a 6.1
Sporangium 905.7 + 59.2a 4.8

TABLE 2. THE EFFECT OF LIGHT ON THE GROWTH OF OLDER GAMETOPHYTES.

(4) A
Medium gametophyte length
Light Dark
Minerals only 4.4+1.5a 2.1+2.3a
Sucrose 0.25% 283.4+22.4b 300.8 + 38.6b

the young or old gametophytes followed by the same letter are not significantly
different at the 1% confidence level.

These are the basic procedures and conditions used to determine effective carbon
sources for spore germination and gametophyte growth with Botrychium dissectum.
Any exceptions or modifications to these procedures, which had to be made in some
experiments, are discussed with their related results.

RESULTS

In the past studies on Borrychium gametophytes in axenic culture, the spores were
surface sterilized with 20% Clorox for 2 minutes. Because the spores germinated
and the gametophytes grew, it was assumed that the Clorox treatment had no adverse
effect. Because young gametophyte growth is used as a measure of the effectiveness
of a carbon source, the effect of the Clorox treatment on germination and on young
gametophyte development was examined. Sporangia were surface sterilized for 20
minutes with 25% Clorox. The spores from these sporangia were collected under
axenic conditions and divided into two batches. Spores from the first batch were
dusted on the nutrient medium. Spores from the second batch were surface sterilized
with the usual 20% Clorox treatment, washed with sterile water, suspended in sterile
water, and pipetted onto the nutrient medium. Between the two treatments, there :.
little difference in the percentage of germination and the difference in the
young gametophytes was not statistically significant (Table 1). In ae ot .
experiments have shown that spores of Botrychium dissectum can be treated up to
minutes with 20% Clorox before germination is inhibited. ns

Since light inhibits the germination of Botrychium spores — oi .
preliminary experiment was carried out to determine if light ohare t a ie .
older gametophytes. Gametophytes were cultured in the light (12 _ ? ag L08
day; intensity 1400 lux) on media with or without a carbon source (Ta : = -
did not promote gametophyte growth on the basic mineral inedium- Lig : a orem
no effect on gametophyte growth with sucrose present. in eae oe
gametophytes to light for transferring and measurine will not al - = si
growth. Since growing the gametophytes in the dark better sim

; ini iments.
conditions, all gametophytes were grown In the dark for the remaining experim

80 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

TABLE 3. THE EFFECT OF SOIL ON GAMETOPHYTE GROWTH.

% increase in
Treatment gametophyte length
Old Young
Soil A Lo) -6a 79.3+4.6a
Soil B =+0:2+1.3a
Minerals only 1.2+1.4a 82.5+3.6a
Sucrose 0.5% 69.8 + 23.0b 1051.6+57.8b

TABLE 4. THE EFFECT OF VARIOUS SUCROSE CONCENTRATIONS
ON GAMETOPHYTE GROWTH.
% increase in

Percent sucrose gametophyte length % germination
Old Young

0.25 56.1+9.7d 180.7+18.1b 9.7

0.5 101.9+11.8cd 262.0 + 30.8ab 9.2

1.0 176.2 418.3c 276.0+23.4a 9.8

2.0 470.4+54.9b 283.6+ 18.6a 93

4.0 587.9+35.9a 206.0 + 16.5ab 8.7

Campbell (1911) observed that soil would not support the growth of young
Ophioglossum gametophytes. He found that without the interaction of a mycorrhizal
fungus, the germinating spores produced gametophytes of only 3 or 4 cells. To test
these observations, spores were sown on autoclaved soil from a site in Tennessee
where Botrychium gametophytes had been collected (Table 3). The percentage of
germination is not reported because it was impossible to recover large numbers of
spores from the soil cultures. The growth of young gametophytes on the soil was
similar to the minimal growth of gametophytes which occurred on the basic mineral
medium. The young gametophytes on the soil and on the mineral medium stopped
growing at the 4- or 5- celled stage. The older gametophytes were taken from the
stock cultures and placed on the basic mineral medium or on autoclaved soil from
two sites in Tennessee where gametophytes had been found. No gametophyte growth
occurred on the soil cultures or on the mineral medium after 2.5 months. The
gametophytes from the stock cultures were healthy and capable of growth because
those placed on the medium with 0.5% sucrose did grow (Table 2).

The carbon source for the previous studies on gametophytes of the Ophioglossaceae
in axenic culture has been sucrose (Whittier, 1972, 1983; Gifford & Brandon,
1978). In these studies, the sucrose had been autoclaved in the basic mineral
medium. Minor changes do occur to sucrose during autoclaving (Ball, 1953). Spores
were sown on a medium with filter-sterilized sucrose (0.5%) to determine if
gametophyte growth was improved over that on a medium with autoclaved sucrose
(0.5%). There was no significant difference in the percent increase in length of
young gametophytes on filtered sucrose (304+ 18.6) and autoclaved sucrose
(322+ 19.8). For the remaining experiments, the sucrose and other organic
components of the media were not filter sterilized.

Xperiments to determine the optimal concentration of sucrose for gametophyte
growth were carried out with sucrose concentrations of 0,.25-4.0% (Table 4). There
was little variation in the percentage of germination on the various sucrose concen-

D. P. WHITTIER: ORGANIC NUTRITION OF BOTRYCHIUM GAMETOPHYTES

TABLE 5. THE EFFECT OF VARIOUS SUGARS AT 0.5% CONCENTRATION ON
GAMETOPHYTE GROWTH.

% increase in
gametophyte length

Treatment % germination
Old Young
Sucrose 365212327340 372.0 + 34.4b 4.5
Glucose 431.5+31.9a 416.6+44.8ab 4.8
Mannose 438.7+42.9a 425.4+50.2a 4.7
Trehalose 223,3225-1ed 408.4 + 48.7ab 4.8
Fructose 306.9 + 19.3be 250.7+8.7c 4.6
Cellobiose 111.2+14.2de 209.0 + 15.2cd 4.7
Maltose 73.1 + 16.2e 180.2 + 12.7d 4.4
Minerals only 0.7+ 1.8e 16,322.36 4.2

trations. The older gametophytes grew best on 4% sucrose. However, the difference
in growth on 4% and 2% sucrose was less than the difference between the 2% and
1% sucrose concentrations. It would appear that the growth of these gametophytes
was beginning to level off at 4% sucrose. The younger gametophytes grew best at
2% sucrose. However, the differences in the amount of growth on the 0.5-4.0%
concentrations of sucrose were not statistically significant. Less growth occurred on
the medium with 4% sucrose than at 2% sucrose, which may have been due to the
Osmotic concentrations of the medium with 4% sucrose.

Because the higher concentrations of sucrose inhibited the growth of young
gametophytes, all other sugars were tested at a 0.5% concentration. Preliminary tests
showed that the pentoses, xylose and ribose; the hexoses, galactose, sorbose, and
thamnose; the disaccharides, lactose and melibiose; and the trisaccharide, raffinose,
did not support the growth of young gametophytes. Further testing was completed on
SIX sugars, in addition to sucrose, which had supported the growth of young
gametophytes in the preliminary tests (Table 5). The best growth for both the young
and older gametophytes was obtained with the hexoses, glucose and mannose.
However, the difference in the growth by the older gametophytes on glucose,
mannose, and sucrose was not statistically significant. With young gametophytes,
trehalose gave almost the same growth as glucose and mannose. In both young aia
older gametophytes, the poorest sugars for supporting growth were the disaccharides,
maltose and cellobiose. Spore germination was essentially the same for the various
sugars.

In symbiotic associations between fungi and other plants, sugar alcohols may be
produced by either of the symbionts and may move between the symbionts ( Lewis &
Harley, 1965: Smith, 1967: Hill & Smith, 1972). Also, some fungi are known to
accumulate mannitol in their mycelium (Turner, 1971). Consequently, three ee
mon sugar alcohols were tested on Botrychium gametophytes.. Mannitol, inositol,
and sorbitol at concentrations of 1% had no effect on germination and did not
Support the growth of young or older gametophytes.

Although the gametophytes appear to require a myc
the soil, the possibility that Botrychium gametophytes
insoluble organic materials in the soil for growth has not
Soils used in the earlier experiments (Table 3) may not h

orrhizal fungus for growth in
might be able to break down
been adequately tested. The
ave contained enough

82 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

TABLE 6. THE EFFECT OF INSOLUBLE CARBOHYDRATES ON GAMETOPHYTE GROWTH.

se in

Treatment gametophyte length % germination
Ol Young

Starch 2.0% 0.4+2.6a 67.43. 1a 4.4

Cellulose 2.0% 0.6+2.2a 61.1+2.8a 4.2

Sucrose 0.5% 60.4+6.5b 372.0 + 34.4b 4.5

Minerals only 0.8+1.5a 68.1+2.5a 4.2

TABLE 7. THE EFFECT OF GLYCERINE ON YOUNG GAMETOPHYTE GROWTH.

% increase in

Treatment gametophyte length % germination
Glycerine 0.5% 266.0 + 20.8a 9.6
Sucrose 0.5% B99 22 30.24 9.0
Minerals only 67.4+3.1 9.5

humus or organic matter to support gametophyte growth. The use of insoluble
organic materials in the soil by the gametophytes depends on their ability to carry out
extracellular digestion. Therefore, it was of interest to determine if Botrychium
gametophytes could metabolize insoluble carbon sources in the external medium.

Several insoluble carbon sources were incorporated into the nutrient medium to
determine if gametophytes could utilize exogenous, insoluble organic materials. In
preliminary experiments, dextrin did not support the growth of older gametophytes.
Starch (2%) and cellulose (2%) had no effect on germination and did not support the
growth of young or old gametophytes (Table 6). Treating the nutrient medium with
I,KI demonstrated that there was no starch breakdown in the medium adjacent to the
older gametophytes. Spores also were sown in petri plates on emulsified nutrient
media containing triglycerides. Young gametophytes on media with emulsified
trihexonin, tristearin, and tripalmatin did not exceed gametophyte growth on the
basic mineral medium. Germination was unaffected by the triglycerides.

Of the carbon sources tested on these gametophytes to this point, only the soluble
Sugars promoted gametophyte growth. However, sugars are not the only organic
materials which will support the growth of Botrychium gametophytes. Spores sown
on a medium supplemented with glycerine (Table 7) germinated and the gameto-
phytes grew. Although the growth with glycerine was somewhat less than that on the
same concentration of sucrose, the difference was not statistically significant. The
effect of glycerine on the growth of older gametophytes was not examined.

As controls for many experiments, older gametophytes were placed on the basic
mineral medium to show how much growth occurred after the — had
been removed from the stock cultures which contained sucrose. The maximum
change in length in the dark, the usual condition for these experiments, was a 2%
increase after 4 months (Table 2). These gametophytes taken from stock cultures
contained starch, yet essentially no growth occurred. In addition to the gametophytes
placed on the basic mineral medium, gametophytes placed on media containing
Starch and cellulose also exhibited no growth (Table 6). The possibility that these
older gametophytes cultured on the basic mineral medium or on a medium without a
usable carbon source had died was tested. Gametophytes on a mineral medium

D. P. WHITTIER: ORGANIC NUTRITION OF BOTRYCHIUM GAMETOPHYTES 83

which increased in length 1% in two months, when transferred to a medium with
0.5% sucrose had an increase in length of 34% in one month. Gametophytes which
had a minimal increase in length on media with 2% starch or 2% cellulose after 4
months (Table 6) increased in length 57% and 64% respectively in four months on
0.5% sucrose. Thus, they remained alive for as long as four months without a
usable, exogenous carbon source and resumed growth when sucrose was available.

DISCUSSION AND CONCLUSIONS

Although the primary objective of this study has been to investigate the organic
nutrition of Botrychium gametophytes, information about other questions on the
biology of these gametophytes has been obtained.

Early observations (Campbell, 1911) that wet areas in the soil were the best places
for the germination of Ophioglossum spores may not be true for all spores of the
Ophioglossaceae. Botrychium spores obtained sterilely from sporangia and dusted
directly on the solid nutrient medium germinated without difficulty. Liquid water is
not necessary for the germination of Botrychium dissectum spores.

In most of the experiments, the percentage of spore germination was determined.
There was some variation in the percentage of germination among the experiments,
which would be expected since the duration of the experiments was not exactly the
same. However, it should be noted that none of the treatments in this study
stimulated (allowed) more than 10% of the spores to germinate. All the spores
appear to have normal cytoplasmic components and are fully formed, thus it would
seem that all the spores should germinate. If the spores are normal and viable, as
they appear to be, then conditions for maximum germination have not been found
with this study.

The carbohydrate nutrition of plant tissue and plants in axenic culture has been
investigated over the years by determining which carbohydrates will support growth.
Sugars are the usual carbon source for these cultures, although insoluble carbohy-
drates have been used occasionally (Street, 1969). The sugars that commonly
support excellent growth are sucrose, fructose, and glucose for plant tissues ( ee
1969) and orchid seedlings (Withner, 1959). Similar results have been obtained Ms
gametophytes of leptosporangiate ferns, although for some species fructose ase
give the growth of glucose and sucrose (Hurel-Py, 1955). Since sucrose ones y
supports excellent growth or growth as good as other sugars, It 1s usually prov! a si
the carbon source in attempts to grow new plants or plant tissues In axenic cu ture.

The few gametophytes of the Seance sagt which a enagt Taps.
cu ith s carbon source ; + ;

lture were supplied with sucrose as ita te te in can have

Gifford ncentrati
retin, ee m gametophytes. Increasing the concen-

, ignifi ount.
tration above 0.5% did not promote early gametophyte growth a significant amo
r concentrations produced significant

times the growth as 0.5% sucrose. Thus, increasing the
. ophytes
accelerate the growth of older gametophytes and can produce mature gametophy

in short periods of time.

84 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Of the other two sugars, glucose and fructose, which have been excellent carbon
sources for many plant tissues, 0.5% glucose supported better growth of Botrychium
gametophytes than 0.5% sucrose. Gametophytes grew on fructose, but to a lesser
extent than on glucose or sucrose.

Trehalose was the only disaccharide, other than sucrose, to support good growth
of these gametophytes. It was superior to sucrose as a carbon source for older
gametophytes, but not for young gametophytes. Trehalose has not been commonly
employed in other axenic cultures, but is known to support the growth of female
gametophytes of Selaginella (Koller & Scheckler, 1982) and some orchid seedlings
(Ernst, 1967).

An unusual aspect of the carbohydrate nutrition of Botrychium gametophytes was
their use of mannose for growth. This was the only other sugar which supported
growth of the gametophytes at a higher rate than sucrose. This is somewhat
surprising because in other studies employing axenic culture techniques mannose is
usually a poor carbon source (Street, 1969). With root and callus cultures, it ranges
from being toxic, like galactose, to inert. Some gametophytes of leptosporangiate
ferns can use mannose, but it supports a lower level of growth than do glucose or
sucrose (Courbet, 1957: Whittier, 1964). However, for some orchid seedlings in
axenic culture mannose is a better carbon source than sucrose (Wynd, 1933; Ernst,
1967).

With one or two minor exceptions, the use of soluble sugars by Botrychium
gametophytes is comparable to that found in other plants or plant tissues in axenic
culture (Street, 1969). Glucose and mannose supported better growth than sucrose
for both older and younger gametophytes. It is possible that high concentrations of
glucose and mannose will support more rapid growth and gametophyte maturation
than higher concentrations of sucrose. Since at the lower concentrations the increase
in growth on glucose or mannose over that on sucrose is relatively small, it is
possible that higher concentrations of glucose or mannose may not give the hoped
for increases in growth.

The only other substance which was found to support the growth of these
gametophytes was glycerol. Thus, something besides a sugar will support gameto-
phyte growth, although somewhat less than for sucrose at the same concentration.
Although glycerol is not a common carbon source for plant tissues, it has been
shown to support the growth of some callus tissues (Gautheret, 1948; Hildebrandt &
Riker, 1955) and Osmunda gametophytes (Stephan, 1929).

The endophytic fungus has been considered the source of the organic materials for
these gametophytes since Campbell (1911) reported that young Ophioglossum
gametophytes did not grow beyond the three- or four-celled stage unless infected
with a mycorrhizal fungus. This idea has been supported by the observations on
older gametophytes from nature, which always contain a mycorrhizal fungus.
Although it appeared doubtful, the possibility that the gametophytes obtain some OF
Qs ie ae organic materials from the soil was investigated. The gametophytes of B.

ectum did not grow on sterilized soil which had been obtained from areas having
gametophytes. Thus, it would appear that the gametophytes cannot obtain enough
organic nutrients from the soil to support growth.

D. P. WHITTIER: ORGANIC NUTRITION OF BOTRYCHIUM GAMETOPHYTES 85

Extracellular digestion would certainly be a requirement for these gametophytes to

grow in the soil without a mycorrhizal fungus. Some plant tissues have demonstrated
the ability to digest and use exogenous starch as a carbon source (Hildebrandt &
Riker, 1953; Straus & LaRue, 1954). Botrychium gametophytes did not grow on
media containing starch, cellulose, dextrin, or emulsified neutral lipids. In addition
no starch digestion occurred in the nutrient medium in contact with the gameto-
phytes. There was no indication that Botrychium gametophytes have the ability to
carry out extracellular digestion.
The organic nutrition of gametophytes of Botrychium dissectum in axenic culture
is, with the exception of glycerol, dependent on the availability of soluble sugars.
Although gametophyte growth remains relatively slow, increasing the sugar concen-
tration, at least with sucrose, accelerates growth and maturation. With the appropri-
ate concentrations of sucrose and possibly glucose or mannose, it would appear that
the maturation time for gametophytes of B. dissectum can probably be reduced to
about a year.

The theory that the mycorrhizal fungus supplies Borrychium gametophytes with
organic materials necessary for growth has not been brought into question by any
observations in this study. The gametophytes failed to grow on sterilized soil and on
insoluble organic nutrients in axenic culture. These gametophytes did not carry out
extracellular digestion. Thus, no evidence has been presented to alter the view that a
symbiotic association with a mycorrhizal fungus is necessary for the organic
nutrition of gametophytes of the Ophioglossaceae.

LITERATURE CITED

ARDITTI, J. 1967. Factors affecting the germination of orchid seeds. Bot. Rev. 33:1-97.

BALL, E. 1953. Hydrolysis of sucrose by autoclaving media, a neglected aspect of culture of plant tissues.
Bull. Torrey Bot. Club 80:409-411.

BOULLARD, B. 1963. Le gametophyte des Ophioglossacées. Considérations biologiques. Bull. Soc. Linn.
Normandie 4:81—97.

CAMPBELL, D. H. 1911. The Eusporangiatae. Carnegie Inst. Washington Publ. 140.

COURBET, H. 1957. Action de quelques sucres sur la germination des spores et la croissance des prothalles
d’Athyrium Filix-femina Roth en culture aseptique. Compt. Rend. Acad. Sci. (Paris) 244:107-110.

rimental investigation. Pp. 253-305 in A. F.

Dyer, ed.
ERNST, R. 1967. Effect of carbohydrate selection on the growth rate of freshly germinated Phalaenopsis and

Dendrobium seed. Amer. Orchid Soc. Bull. 36:1068-1073.

GAUTHERET, R. J, 1948. Sur l'utilisation du glycérol par les cultures de tissus végétaux. Compt. Rend. Soc.
Biol. 142:808-810.

GIFFORD, E. M., Jr., and D. D. BRANDON. 1978. Gametop
axenic culture. Amer. Fern J. 68:71-75.

HILDEBRANDT, A. C., and A. J. RIKER. 1953. Influence of cone
on callus tissue growth in vitro. Amer. J. Bot. 40:6

a... 1955. Inhibition by alcohols of diseased plant gr

, D. J., and D. C. SMITH. 1972. Lichen physiology.

hytes of Botrychium multifidum as grown in
entrations of sugars and polysaccharides

ths in tissue culture. Cancer Res. 15:517-522.
XII. The inhibition technique. New Phytol.

71:15-30.
HUREL-PY, G. 1955. Action de quelques sucres sur la croisance des prothalles de fougeres. Compt. Rend.

: Acad. Sci. 240:1119—-1121.
OLLER, A. L. and S. E. SCHECKLER. 1982. Culture
Amer. Misc. Publ. No. 162:16.

of megagametophytes of Selaginella. Bot. Soc.

1
|

86 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

LEWIS, D. H. and J. L. HARLEY. 1965. Carbohydrate physiology of mycorrhizal roots of beech. I. Identity
of endogenous sugars and utilization of exogenous sugars. New Phytol. 64:224-237.

LI, J. C. R. 1964. Statistical inference I. Edwards Brothers, Ann Arbor, Michigan.

SMITH, S. E. 1967. Carbohydrate translocation in orchid mycorrhizas. New Phytol. 66:371-378.

STEPHAN, J. 1929. Entwicklungsphysiologische Untersuchungen an einigen Farnen. I. Jahrb. Wissen.
Botanik 70:707-742.

STRAUS, J. and C. D. LARUE. 1954. Maize endosperm grown in vitro. I. Cultural requirements. Amer. J.

Bot. 41:687-694.

STREET, H. E. 1969. Growth in organized and unorganized systems. Pp. 3-224 in F. C. Steward, ed. Plant
Physiology, vol. VB. Academic Press, New York.

TURNER, W. B. 1971. Fungal Metabolites. Academic Press, New York.

WHITTIER, D. P. 1964. The influence of cultural conditions on the induction of apogamy in Pteridium
gametophytes. Amer. J. Bot. 51:730-736.
. 1972. Gametophytes of Botrychium dissectum as grown in sterile culture. Bot. Gaz. 133:336-339.
. 1973. The effect of light and other factors on spore germination in Botrychium dissectum. Can. J.
Bot. 51:1791-1794.

————.. 1983. Gametophytes of Ophioglossum engelmannii. Can. J. Bot. 61:2369-2373.

and T. A. STEEVES. 1960. The induction of apogamy in the bracken fern. Can. J. Bot.

38:925-930.

WITHNER, C. L. 1959. Orchid Physiology. Pp. 315-360 in C. L. Withner, ed. The orchids: A scientific
survey. Ronald Press, New York.

WYND, F. L. 1933. Sources of carbohydrates for germination and growth of orchid seedlings. Ann. Missouri
Bot. Garden 20:569-581.

REVIEW

A Taxonomic Revision of the Genus Grammitis ... in New
Guinea, by B. S. Parris. Blumea 29:13-222. 1983.—Of all the regions of the
Old World tropics, New Guinea is the least known botanically. Copeland (Phil. J.
Sci. 81:81-119, t. 1-9. 1953) published the only other account of the genus in
New Guinea, and this contained only keys and a list of species with synonyms and
distributions. Copeland included 48 species in Grammitis sensu stricto, whereas

arris has 64; the difference is due no doubt to the botanical explorations which
have taken place since Copeland wrote. Parris’ monograph is unusually complete. It
contains both a dichotomous key and a multiple-entry key, as well as a conspectus of
the species groups. A great many Grammitis species are endemic to New Guinea.
Those which range more widely often go no farther than Java, Sumatra, or Borneo,
fewer extend to the Philippines or the small islands of the Pacific. The accepted
species names are given with their synonyms, a detailed description, a statement of
range and detailed distribution within New Guinea, ecological and other notes, and
a map and illustration. There is an extensive section on the phytogeography of
Grammitis in New Guinea, as well as an index to scientific names and an index to
collections. —D.B.L.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 3 (1984) 87

Anti-microbial Activity of Phenolic Acids
in Pteridium aquilinum
MICHAEL SAN FRANCISCO and GILLIAN COOPER-DRIVER*

All land plants are continually exposed to a wide variety of microorganisms
ranging from those which are beneficial to those which are pathogenic. Most plants
remain remarkably healthy, and this seems to be especially true of ferns, which
reportedly are rarely subject to diseases (Braid, 1940; Hutchinson, 1976; Bandoni,
pers. comm.). It is now generally recognized that the resistance of plants to
pathogens (and other predators) is mainly due to their containing fungicidal and
bactericidal secondary compounds. The few surveys which have been carried out on
ferns have confirmed that they do contain such antibiotic substances (Maruzzella,
1961; Khrisagar & Mehta, 1972; Lynch-Brathwaite et al., 1975; Banerjee & Sen,
1980; Cooper-Driver et al., in prep.).

One major class of secondary compounds with known antimicrobial activity is the
phenolics, which range from simple phenols to complex polymeric tannins
(Harborne, 1982, pp. 230-264). Their main role in plants appears to be as
protective compounds against fungi, bacteria, and viruses (Friend, 1979; Swain,
1979). For example, the flavonoids present in the farinose exudate on the leaf
surfaces of some ferns have antimicrobial properties (Chowdhury et al., 1974;
Wollenweber, 1978). fe

Simple phenolics, such as p-hydroxybenzoic, protocatechuic, and vanillic
(hydroxybenzoic) acids, and p-coumaric, caffeic, and ferulic (cinnamic) acids (Fig.
1) have been shown to be present in the fronds of ferns (Bohm & Tryon, 1967;
Bohm, 1968; Cooper-Driver, 1976) and to play an important role in the ecology of
Bracken (Preridium aquilinum (L.) Kuhn) by inhibiting the germination and growth
of neighboring plants (Stewart, 1975; Glass, 1976; Gliessman & Muller, 178):
Such acids are also known to have inhibitory effects on feeding by insects (Jones &
Fir, 1979). The concentration of these acids in t
quantitatively during growth and development (Glass & Bohm, 1969; aairtngee
et al., 1977: Jones, 1983), and thus must affect the antimicrobial, allelopathic. an
anti-insecticidal properties of the plants differentially during the apie a -

Simple phenolic and cinnamic acids must also play a major role int : ET ch
fern gametophytes against pathogens, particularly since more complex pheno , “Be
as flavonoids, have been shown to be absent during the pre-antheridial stag
(Petersen & Fairbrothers, 1980).

However, it has been reported that fern prothalli are readily a in eT
species of “damping off” fungi, like Pyshium specis® ee aos cinerea,
and gametophytes grown in pure culture can be readily invaded by Bol on Ga

ythium debaryanum, and Rhizoctonia solani (Hutchinson & cal ee ae aul
metophytes grown in pure culture appear [0 be more susceptible

ifi i Fusarium
ene re specific pathogens of angiosperms,
cule iti et sail 1976). In angiosperms, many of these

Solani and Verticillium dahliae (Hutc
gton St., Boston, MA 02215.

“Department Biological Sciences, Boston University, 2 Cummin,

88 AMERICAN FERN JOURNAL: VOLUME 74 (1984)
CINNAMIC ACIDS HYDROXYBENZOIC ACIDS

R R
HO CH=CH-COOH HO COOH
R -H p-coumaric acid ROH p- hydroxybenzoic acid
R - OH caffeic acid R - OH Pprotocatechuic acid
R - OCH, ferulic acid R- OCH, vanillic acid

FIG. | Commonly occurring phenolic acids in Pteridium aquilinum.

damping off fungi attack only the seedling stage, and plants become progressively
more resistant to attack as they mature. Infections are most severe at high moisture
levels. These pathogens survive for relatively long periods in the soil (Wheeler,
1976, pp. 9-27), posing a potential threat to fern gametophytes.

It was of interest, therefore, to investigate the effect of the concentration of
phenolic acids isolated and identified from the gametophytes and sporophytes of
Bracken on the growth of fungal and bacterial plant pathogens. All nine organisms
tested, with the exception of Erwinia, have been reported to cause infections of
Bracken (Gregor, 1938; Hutchinson & Fahim, 1958; Hutchinson, 1976). Changes in
the concentrations of the acids were measured in Bracken over the May to October
growing season, and observations were made of visible signs of fungal infections.
We also isolated fungi and bacteria at frequent intervals throughout the year.

MATERIALS AND METHODS

Isolation of phenolic acids from gametophytes.—Spores of Pteridium aquilinum
were collected in August and following sterilization (Haufler, pers. comm.) were
sown on an agar slant of Moore’s medium (Klekowski, 1969) as a slurry of spores
and water. The tubes were incubated at 25° C, 12 hours day length. Germination
was observed after approximately 1.5 weeks, and the gametophytes were grown until
they were eight weeks old. The 80% methanolic extract of the gametophytes was
base hydrolysed, and the acids were separated by thin layer chromatography (TLC)
on cellulose using benzene-acetic acid-water (6:7:3, upper phase) in one direction
and IN sodium hydroxide—formic acid—water (150:8:42) in the second direction. The
acids were detecied by spraying with diazotized sulfanillic acid. Gametophytes were
subcultured 7 to 8 weeks following inoculation, which proved to be a good method
for increasing the amount of gametophyte tissue and also for initiating antheridium
formation and subsequent sporophyte growth.

SAN FRANCISCO & COOPER-DRIVER: ANTI-MICROBIAL ACIDS IN PTERIDIUM 89

The quantification of phenolic acids present in the gametophyte tissue was carried
out by direct extraction with 80% methanol and by base hydrolysis with 2N NaOH
for 2 hours under nitrogen followed by acidification, extraction of the released acids
with ether, purification via re-extraction into sodium bicarbonate, acidification, and
final extraction into ether. The solution was then evaporated to dryness and the
residue taken up in a few drops of methanol. Absorbance was read at 310nm and
related to known concentrations of p-coumaric acid as a standard. The methanol-
insoluble residue was also base hydrolysed and examined for the presence of
phenolic acids.

Isolation of phenolic acids from sporophytes.—Dried material was extracted
with 80% methanol. The extract was base hydrolysed as described above and the free
acids separated by thin layer chromatography using the solvent systems isopropanol-
t-butanol—n-butanol-ammonia—water (4:2:2:1:2) in the first direction, followed by
benzene—acetic acid—water (6:7:3, upper phase) in the second direction. Both Rp
values of the acids and their ultra-violet spectral properties were compared with
standards (Cooper-Driver et al., 1972).

Quantitative estimates were made on dried samples of Bracken collected from
three sites of the Massachusetts Audubon Bird Sanctuary at Natick at bi-weekly
intervals. The procedures used for the quantification of the phenolic acids were
identical to those described above for the gametophytes.

Micro-organisms.—Test organisms included the fungi Botrytis cinerea, Rhizocton-
ia solani (Deuteromycetes), Pythium debaryanum, P. middletonii, and P. ultimum
(Oomycetes) and the bacteria Corynebacterium poinsettiae, C. fasciens, Erwinia
amylovora, and E. carotovora. The Erwinia species were chosen as examples of
economically important pathogens as causal agents of fireblight disease (Quamme et
al., 1976). Cultures of B. cinerea and R. solani were obtained from the American
Type Culture Collection and the Pythium cultures from Dr. C. Bracker, Purdue
University. The Corynebacterium species were obtained from Dr. M. Starr, Univer-
sity of California, Davis, and the Erwinia from Dr. A. Chatterjee, University of
Kansas.

Fungal cultures were maintained on potato dextrose agar (PDA) at 27° C, and
Phenolic acids were added to the plates either singly or in combination to give final
concentrations of 0.005%(SOppm), 0.01%(100ppm), 0.05%(500ppm) and 0.1%
(1000ppm) w/v. The plates were inoculated with cultures of fungi, by inversion, and
Were incubated for 5 days at 27° C. Fungal growth was measured and expressed as
the extent of hyphal growth as a percentage of the control (without added acid). All
tests were run in triplicate. are

Bacteria were grown in nutrient broth. Phenolic acids were added either individu-
ally or in combination in order to give final concentrations of 0.005%(50ppm),
0.01%(100ppm), 0.05%(500ppm) and 0. 1%(1000ppm) w/v. The medium was first
autoclaved and then inoculated with 2% 10° bacteria in exponential growth (10
cells per ml); bacterial growth was measured by determining the absorption at

80nm and was expressed as a percentage of the control (Khrisagar & Mehta, 1972).

Microbial infections.—Field observations on microbial infections were made on

20 labelled Bracken plants at bi-weekly intervals. Typical signs of infection were

90 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

TABLE 1. MINIMUM CONCENTRATION OF PHENOLIC ACIDS (in ppm) CAUSING 50%
INHIBITION OF FUNGAL OR BACTERIAL GROWTH.

~ 3

z 5

3 :

2 3. 5 &

a 6 = - =

5 s 3 3 fe 3 i

S 2 = £ re 3 E

7 5) ! 3s a °

Fungi aX a 0 x > Q. o

Rhizoctonia solani 50 50 100 100 > 1000 »1000! 100

Botrytis cinerea 5 500 >1000 >1000 1000! >1000! 500

Pythium debaryanum 100 5 5 500 500 1000 50

P. middletonii 50 50 50 50 50 50

P. ultimum 50 50 100 100 500 >1000! 50
Bacteria

Corynebacterium 100 500 1000 1000 500 100 50

poinsettiae

C. fasciens 500 500 100 500 500 1000 50

Erwinia amylovora 50 50 100 50 50 50 100

E. carotovora 100 50 100 50 100 100 50

‘stimulatory at low concentrations

local necroses, indicated as leaf spots or brown necrotic tissue especially around
damaged areas, and ‘Curl-tip’, which was shown by curling and darkening at the
apex of the pinnae. The amount of infected tissue was estimated.

In order to isolate micro-organisms from infected plant tissue, the tissue was
surface sterilized with 1 ppm mercuric chloride and washed in 70% ethanol followed
by sterilized distilled water. The tissue was dried and placed on suitable growth
medium, PDA for isolation of fungi and nutrient agar (NA) for bacteria. Plates were
incubated at 30° C. Seventeen different bacteria and eight fungi were isolated from
infected areas and obtained in pure culture. Attempts were made to demonstrate the
pathogenicity of these fungi, but it was difficult to demonstrate re-infection of
Bracken plants in the field (cf. Hutchinson, 1976), either by inoculating the leaf
surface by scraping or by injecting the stem at the junction of the petiole and the
stem.

RESULTS
Bracken gametophyte tissue was surprisingly completely devoid of flavonoids.
The major phenolic compounds present were p-coumaric, caffeic, ferulic acids, and
p-hydroxybenzoic acid; vanillic, protocatechuic, and o-coumaric acids, which were
present in the sporophyte, could not be detected. There were few qualitative or
quantitative changes in these acids in the _—_* from one week up to ten
weeks. The acids were present mainly as gluc or quinic acid esters, and
electrophoresis at pH 2.2 and treatment with Sa confirmed the presence of

SAN FRANCISCO & COOPER-DRIVER: ANTI-MICROBIAL ACIDS IN PTERIDIUM 9]

sulfated derivatives of caffeic and p-coumaric acids (Cooper-Driver & Swain, 1975).
Phenolic acids in the gametophytes were present at concentrations of 0.15—0.20%/gm
dry weight. Base hydrolysis of the alcohol-insoluble fraction gave no indication that
hydroxy-cinnamic acids were bound to the cell walls (Hartley & Jones, 1977),
whereas p-coumaric and ferulic acids were isolated from the base hydrolysate of the
methanol insoluble residue of the sporophyte.

No flavonoids were detected in sporophytes growing from the gametophytes,
whereas vegetatively produced sporophyte plants on emergence synthesized flavonoid
glycosides.

The effect of these acids on the growth of Botrytis cinerea, Rhizoctonia solani,
and Pythium species, together with Corynebacterium poinsettiae, C. fasciens, and
Erwinia species is shown in Table | and is expressed as minimum concentrations of
acids causing 50% inhibition of fungal or bacterial growth. Overall, cinnamic acids,
especially ferulic and p-coumaric acids, were the most effective inhibitors of fungal
growth (the growth of Rhizoctonia solani was inhibited at concentrations as low as
50ppm). On the other hand, protocatechuic acid was stimulatory to growth of R.
solani at all concentrations tested, and vanillic acid was only slightly inhibitory
(18%) at concentrations as high as 1000ppm. In the case of Botrytis cinerea, the
hydroxybenzoic acids together with caffeic acid had little effect on growth, and
p-coumaric and ferulic only inhibited growth at concentrations of 500ppm or above,
and then only by 55%. In combination, the acids were inhibitory to growth of R.
solani at 100ppm (50%) and were completely inhibitory to growth at higher
concentrations. For B. cinerea, 500ppm was inhibitory (74%), and no growth
occurred at higher concentrations. Overall, however, R. solani was much more
affected by phenolics than was B. cinerea. ae

All Pythium species also showed much greater susceptibility to phenolic acids
than did Botrytis. Growth of P. debaryanum was uniformly inhibited by phenolic
acids at concentrations above 500ppm, and 100ppm ferulic acid and p-coumaric acid
inhibited growth 13% and 50% respectively. Protocatechuic acid was the least
inhibitory. The growth of P. middletonii was inhibited at all concentrations of the
acids tested, whereas the growth of P. ultimum was stimulated by protocatechuic
acid. In all Pythium species, no growth occurred when a combination of acids was
added to the medium.

The bacterial species tested responded to the phenolic acids in a more uniform
manner. At 50ppm, the acids were mildly stimulatory or caused only slight
inhibition of growth (<25%). As the acid concentration increased, there was a
corresponding increase in inhibition. There were no differences in the inhibitory
action of hydroxybenzoic acids or cinnamic acids. Combinations of the ants
inhibited bacterial growth in all cases. No growth was observed at concentrations
above 100ppm and, with the exception of E. amylovora, 98% inhibition was
Observed at concentrations as low as 5Oppm.

Seasonal changes in the concentration of ph
Fig. 2. There was a slight increase during '
followed by a slight drop and a gradual increase reachin
of 0.75% phenolic acids/gm dry weight of leaf material

enolic acids in Bracken are shown in
May, the period of leat expansion,
g a maximum concentration
(0.25%/gm fresh weight) in

92 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

0.8

May | June | July | August |September !

FIG. 2 Seasonal changes in the concentration of phenolic acids (in % per gram of dry weight,
@, and fresh weight, ©) in Pteridium aquilinum leaves.

late July. There was then a drop in phenolic acid concentrations during August
following a period of heavy precipitation (1.2 in rain) that caused leaching of the
acids from the leaves. The concentration of eke acids in the leaf tissue dropped
markedly as senescence set in during September and October.

The number of fungal and bacterial species isolated from sterile leaf tissue showed
a general increase as the fronds matured, with an especially sharp increase in the
amount of infected tissue in early August.

DISCUSSION AND CONCLUSION

Phenolic acids, when added to a microbial growth medium, inhibit the growth of
both fungi and bacteria. They have been shown to react with fungal enzymes and
other proteins. Phenolic compounds in infected tissues are often oxidized and
polymerize to form reactive oligomeric products that may have increased anti-
microbial activity often associated with increased inhibition of the cell wall degrada-
tion caused by extracellular enzymes produced by pathogens (Friend, 1979). This
may account for the high inhibition of Rhizoctonia solani by phenolic acids, as

SAN FRANCISCO & COOPER-DRIVER: ANTI-MICROBIAL ACIDS IN PTERIDIUM 93

oxidation products were seen as brown rings around the edges of the fungal
mycelium. Hunter (1974) has demonstrated that the resistance of cotton plants to R.
solani is due to oxidized catechin, which inhibits the polygalacturonase activity of
the pathogen. Phenolic acids also act on membrane permeability and electric
potentials, causing an enhanced ratio of oxygen consumption by uncoupling electron
transfer (Glass & Dunlop, 1974).

The variation in response of the fungi, Botrytis and Pythium to phenolic acids
may be a reflection of the chemical composition of their cell walls. Chitin is known
to be far more resistant to microbial decomposition than is cellulose (LeJohn, 1971).
This may explain the relative resistance of Botrytis over Pythium species to the
phenolic acids, as Rhizoctonia and Botrytis have cell walls of chitin, whereas those
of Pythium are composed primarily of cellulose (Bartnicki-Garcia, 1968). Cell
membranes of Pythium also lack sterols, which are present in both species of
Deuteromycetes (Hall, 1979). Differences in cell wall composition between the two
genera of bacteria (Stanier et al., 1976, p. 871) are not reflected in their response to
the individual acids. The high resistance of B. cinerea to phenolic acids is
interesting in light of the fact that this fungus has often been found associated with
Bracken. Godfrey (1974) reported B. cinerea on the leaf surface (phylloplane); we
also found this species to be present in leaf isolates. In addition, B. cinerea
has been recorded on decomposing Bracken petioles (Frankland, 1976), and prothalli
are also very susceptible to attack by this organism (Hutchinson, 1976).

It is also interesting that the cinnamic acids, ferulic and p-coumaric, are generally
the most inhibitory acids to the growth of fungi and bacteria. The cinnamic acids are
of prime importance in lignin formation, and polysaccharide esters of p-coumaric
and ferulic acids have been shown to be released from the cell walls of grasses by
cellulase action (Hartley & Jones, 1977). It has been suggested that such acetylation
of the cell wall polysaccharides by hydroxycinnamic acids may be a primitive

; i (Swain, 1979).
Although gametophytes do not have bound cinnamic acids i
shown by the lack of detectable acids in the hydrolysed residue, nevertheless these
compounds were present in the soluble fraction and were the most effective
anti-microbial compounds against the pathogenic organisms tested.

Mixtures of simple phenolic acids have a synergistic effect and are better
inhibitors of microbial growth when present in combination. These same acids have
also been shown by Adams and Bernays (1978) to be more effective deterrents to
feeding by Locusta migratoria in combination. Increased synergistic inhibitory
effects of acids on radish and sorghum germination have also been shown ( Rasmus-
sen & Einhellig, 1977; Einhellig & Rasmussen, 1978).

From the field and laboratory studies, it was evident t
Present in high enough concentrations in Bracken sporophytes
4500-7500ppm, phenolic acids/gm dry weight) and gametophytes
1500-2000ppm, phenolic acids/gm dry weight) 7
antimicrobial compounds. Damage to the tissues Dy :
releases the acid vi as free ids (Woodhead & Cooper-Driver, 1979). mmnee
Phenolic acids are effective inhibitors of microbial growth at concentrations of the

hat phenolic acids are
(0.25-0.75%,
(0.15-0.20%,

94 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

order of 100ppm w/v, they obviously play an important anti-microbial role in
Bracken. The concentration of phenolic acids present in gametophyte tissue was
much lower, but was still high enough to inhibit microbial growth.

Concentrations of phenolic acids in the leaves of Pteridium aquilinum changed
over the season depending not only on the age of the plant, but also on abiotic and
biotic factors. The sharp increase in infected tissue and the numbers of isolated
microorganisms detectable in early August may be due to a preceding period of
heavy precipitation followed by warm, humid weather around 40° C. The heavy
rainfall would have resulted in leaching of the phenolic acids from the leaves, and
thus a decrease in chemical protection. In July there was a heavy infestation of
aphids on the Bracken fronds, and the presence of these insects could well have
increased the points of entry for the fungal hyphae, as well as promoting greater
cree of plant defense chemicals from the leaves.

comparison of phenolic compounds in sporophytes and gametophytes of
Polypodium aureum, Thelypteris patens, T. normalis, Asplenium aethiopicum,
Cyrtomium falcatum, Gymnopteris rufa, and Pteris straminea (Cooper-Driver,
unpubl.) has shown that there are two distinct patterns: (a) ferns in which the
phenolic complement in gametophytes and sporophytes is identical with often a wide
range of flavonoids being present in both, and (b) those in which the phenolics in the
gametophyte are restricted to simple phenolic acids, as in the present case of
Bracken. It is interesting to speculate whether this difference in gametophyte
chemistry reflects the dependence of the species on the eae cs for reproduc-
tion. As is well known, Bracken reproduces mainly vegetatively.

Thus it has been shown that phenolic acids differentially enseon the growth of fern
pathogens and other bacteria in vitro and are present for most of the growing season
in high enough concentrations in vivo in the plant tissues to inhibit the growth of
pathogenic microbes.

We would like to thank Bruce Lund, Director of the Massachusetts Audubon
Bird Sanctuary, Natick, MA, for his help and cooperation during the field work,
Matthew Zavitovsky for his expert field advice, and Tony Swain for his evaluation of
the manuscript.

LITERATURE CITED

ADAMS, C. M. and E. A. BERNAYS. 1978. The effects e combinations of deterrents on the feeding
behaviour of Locusta migratoria. Entomol. Ex I. 23:101—109.

BANERJEE, R. D. and S. P. SEN. 1980. Antibiotic =e of pteridophytes. Econ. Bot. 34:284-298.

BARTNICKI-GARCIA, S. 1968. Cell wall chemistry, morphogenesis and taxonomy of fungi. Ann. Rev.
Microbiol. 22:87-105.

BOHM, B. A. 1968. Phenolic sep ers in ferns III. An examination of some ferns for caffeic acid
derivatives. Soyer 7:1825-
, and R. M. TRYON te aes ona in ferns I. A survey of some ferns for caffeic
and benzoic acid peared Canad. J. Bot. 45:585-593.

BRAID, K. W. 1940. The eradication of bracken. Scott. J. Agr. 33:31-36.

CHOWDHURY, A., N. MUKHERJEE and N. ADITYACHAUDHURY. 1974. Sensitivity of some plant
pathogenic fungi towards plant metabolites: antifungal activity of some chalcones,
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COOPER-DRIVER, G. A. 1976. Chemotaxonomy and phytochemical ecology of bracken. Bot. J. Linn.

Soc. 73:35—46.

——, and T. SWAIN. 1975. Sulphate esters of caffeyl- and p-coumaryl glucose in ferns.
Phytochem. 14:2506—

———., J. J. CORN ER-ZAMODITS, and T. SWAIN. 1972. The metabolic fate of hydroxybenzoic
acids in plants. Z. Naturforschung. 27B: te 946.

, S. FINCH, T. SWAIN, and E. A. BERNAYS. 1977. Seasonal variation in secondary
anpounds in relation to the ens of Pteridium aquilinum. Biochem. Syst. Eco
5:177-183.

EINHELLIG, F. A. and J. A. RASMUSSEN. 1978. Synergistic inhibitory — of vanillic and

p-hydroxybenzoic acids on radish and grain sorghum. J. Chem. Ecol. 4:425-436.

FRANKLAND, J. C. 1976. Decomposition of bracken litter. Bot. J. Linn. Soc. 5 145-150.

FRIEND, J. 1979. Phenolic substances and plant disease. Jn T. Swain, J. Harborne and C. F. Van

mere, eds. Biochemistry of Plant Phenolics. Rec. Adv. ana 12:557-588.

GLASS, a. D. M. 1976. The allelopathic potential of phenolic acids associated with the rhizosphere of
Pteridium aquilinum. Canad. J. Bot. 54:2440—2444
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Pteridium aquilinum and Athyrium filix-femina. Phytochem. 8:371-377.
, and J. DUNLOP. 1974. Influence of phenolic acids on ion uptake IV. Depolarization of

membrane potentials. Plant Physiol. 54:855-858.

GLEISMANN, S. R., and C. H. MULLER. 1978. Allelopathic mechanisms of oe in bracken
(Pteridium aquilinum) in Southern California. J. Chem. Ecol. 4:289-

GODFREY, B. E. S. 1974. Phylloplane mycoflora of bracken, Pteridium ie Trans. Brit.
Mycol. Soc. 62:305-311.

GREGOR, M. J. F. 1938. Associations with fungi and lower plants. /n F. Verdoon, ed. Manual of
Pteridology. M. Nijhoff, The H

HALL, R. 1979. PS iaiswane and ser oe Bot. Rev. 47:1—

HARBORNE, J. B. 1982. Introduction to Ecological Biochemistry, bi ed. Academic Press, New

York.
ad R. D. and E. C. JONES. 1977. Phenolic — and degradability of cell walls of
s and legume species. Phytochem. 16:1531
HUNTER, “ E. 1974. Inactivation of pectic enzymes be polyphenolics in cotton seedlings of different

HUTCHINSON. S. A. 1976. The effects of fungi on bracken. Bot. J. Linn. Soc. 73:145-150.
—, and M. M. FAHIM. 1958. The effects of fungi on the gametophytes of Pteridium aquilinum

L. Kuhn. Ann. Bot., , 22:117-1 ae
JONES, C. G. 1983. iaisciburtcad variation, colonization and insect communities in the case of
bracken fern, Pteridium aquilinum (L.) Kuhn. /n R. F. Denno and M. S. McClure, eds.

Variable Plants and Herbivores in Natural aa Aone Systems. Academic Press, New York.
ilinum and their involvement

, and R. D. FIRN. 1979. Some allelochemicals of Pteridium aquil
in resistance to Pieris brassicae. Biochem. Syst. Ecol. 7:187-192. s
KHRISAGAR, M. K. and A. R. MEHTA. 1972. Antibacterial substances in ferns. Pl. Med.

22:386-390.

KLEKOWSKI, E. J. 1969. Reproductive biology of the Pteridophyta III. A study of the Blechnaceze.
Bot. J. Linn. Soc. 62:361-377.

LeJOHN, H. B. 1971. Enzyme regulation, lysine

r lines of evolution in fungi. Nature 231:164-168. icin

LYNCH- pele WAITE, B. A., E. J. DUNCAN and C. E. SEAFORTH. 1975. A survey of ferns To
antibacterial activity. Pl. Med. 27:173—200.

MARUZZELLA, J. C. 1961. Antimicrobial substances from ferns. Nature 191: 518.

PETERSEN, R. L. and D. E. FAIRBROTHERS. 1980 Flavonoid synthesis and antheridium initiation in
Dryopteris gametophytes. Amer. Fern J. 70: 93-95.

pathways and cell wall structures as indicators of

96 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

QUAMME, H. A., T. VAN DER ZWET and V. DIRKS. 1976. Relationship of fireblight resistance to
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RASMUSSEN, J. A. and F. A. EINHELLIG. 1977. Synergistic inhibitory effects of p-coumaric and
ferulic acids on germination and growth of grain sorghum. J. Chem. Ecol. 3:197-205.

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New York.

STEWART, R. E. 1975. Allelopathic potential of western bracken. J. Chem. Ecol. 1:161-169.

SWAIN, T. 1979. Phenolics in the environment. /n T. Swain, J. B. Harborne, and C. F. Van Sumere,
eds. Biochemistry of Plant Phenolics. Rec. Adv. Phytochem. 12:617—640.

WHEELER, B. E. J. 1976. An Introduction to Plant Diseases. Wiley, New York.

WOLLENWEBER, E. 1978. The distribution and chemical constituents of the farinose exudates in
gymnogrammoid ferns. Amer. Fern J. 68:13-28.

WOODHEAD, S. and G. COOPER-DRIVER. 1979. Phenolic acids and resistance to insect attack in
Sorghum bicolor. Biochem. Syst. Ecol. 7:309-310.

REVIEW

The Phylogeny and Classification of the Ferns, A. C. Jermy, J. A. Crabbe, and
B. A. Thomas, eds. Koeltz Scientific Books. 284 pp. Reprint 1984. DM150 (ca.
$60.00).—This authorized reprint makes available again the results of the Sympo-
sium on the Phylogeny and Classification of the Filicopsida, which was held in
London in April 1972. The original volume was reviewed by C. Page (Fern Gaz.
11:47-48. 1974). The reprint is on slightly thinner and less opaque paper than is the
original and there is a slight but inevitable loss in definition of the photographic
illustrations, but the volume is sturdily bound and fully serviceable, and will make
an excellent addition to any fern student’s library.—D.B.L.

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G. Coke, Robin Hill, Stinchcombe, Dursley, Gloucestershire, England.

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QUARTERLY JOURNAL OF THE AMERICAN FERN SOCIETY

The Identification of Hawaiian Tree Ferns
of the Genus Cibotium RICH BECKER 97

Two New Tree Ferns from Panama _ ROBERT G. STOLZE 101

Trunk Length and Frond Size in a Population

of Nephelea tryoniana from El Salvador RALPH L. SEILER 105

An Unusual New Elaphoglossum from Peru ROLLA TRYON = 108
New Tropical American Ferns JOHN T. MICKEL 111

Shorter Note: An Urban Locality for
Asplenium platyneuron

1985 A.I.B.S. Meeting—Call for Papers
American Fern Journal
Index to Volume 74

Errata

MISSOURI BOTANICAL

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DENNIS Wm. STEVENSON, Barnard College, Columbia University, New York, NY 10027.

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AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 4 (1984) 97

The Identification of Hawaiian Tree Ferns
of the Genus Cibotium
RICH BECKER™

The identification of Hawaiian tree ferns is difficult for most laymen and
taxonomists because of past nomenclatural confusion, misapplication of the specific
names, and the absence of a complete key to all of the species. Although Sadleria, a
Hawaiian endemic genus of the Blechnaceae, has some arborescent species which
are referred to as tree ferns, these are not treated in this paper. The correct
nomenclature of the six Cibotium species found in Hawaii is presented. A simple key
is given for the species. This is supplemented with descriptions of their characteris-
tics as seen in their native habitats. I have deposited in the herbarium of the
University of Hawaii my reference collection of entire fronds for each of the
Hawaiian species described.

The range of Cibotium extends from Assam through China, Malaya, Indonesia,
the Philippine Islands, and Hawaii to southern Mexico and Guatemala. There are
three Asiatic species (Holttum, 1963, pp. 164-166).: C. barometz (L.) J. Smith, C.
cumingii Kunze, and C. arachnoideum (C. Chr.) Holttum. The Central American
species are: C. schiedei Schlecht. & Cham. (Mexico), C. regale Versch. & Lem.
(Mexico to Honduras), and C. wendlandii Mett. ex Kuhn (Mexico and Guatemala),
according to Smith (1981, pp. 75-76) and Stolze (1976, pp. 94-97).

The six endemic Hawaiian Cibotium species are: C. glaucum (J.E. Smith) Hook.
& Arn., C. chamissoi Kaulf., C. splendens (Gaud.) Krajina ex Skottsb., C. nealiae
Degener in Degener & Hatheway, C. hawaiense Nakai & Ogura in Ogura, and C. st.-
johnii Krajina. Cibotium glaucum and C. chamissoi are found in the rain forest areas
of Kauai, Oahu, Molokai, Lanai, Maui, and Hawaii. Cibotium splendens is common
on Oahu; forms of it may also be on Molokai and Maui, but this has not been
confirmed. Cibotium nealiae and C. st.-johnii are apparently restricted to Kauai.
Cibotium hawaiense is endemic to the island of Hawaii; it is locally abundant on that
island. I have seen it close to Wright Road in the Olaa Forest, at Puu Makaala near
the Stainback Highway, at the end of the Olaa Back Road, above Laupahoehoe, and
in the Kohala Mountains. The most common species on the island of Hawaii is C.
glaucum. Cibotium chamissoi is the next most abundant. -

The key to resolving the nomenclatural confusion associated with the Hawaiian
tree ferns lies in understanding Hooker's treatment of the genus in the ny
Filicum (1844). He separated C. chamissoi Kaulf. and C. glaucum (J.E. Smith)
Hook. & Arn. and described a new species, c. menziesil Hook. Krajina in
Skottsberg (1942, pp. 30-41) showed the latter to be identical to C. .
Kaulf. and the name C. chamissoi sensu Hooker to be C. splendens (Gaud.) Krajina
ex Skottsb.

*P.O. Box 1358, Station A, Vancouver, B.C. V6CT2T, Canada.

Volume 74, number 3, of the JOURNAL, was issued December 6, 1984.

98 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Hillebrand may have used the name C. chamissoi Kaulf. in its original sense on
some of his first collection labels (Skottsberg, 1942, p. 40). However, in the Flora of
the Hawaiian Islands, Hillebrand (1888, pp. 545-548) used the incorrect nomencla-
ture from Hooker’s Species Filicum, as did Rock (1913), in his The Indigenous
Trees of the Hawaiian Islands. Both are standard works on the Hawaiian flora that were
for a long time assumed to be correct, causing many later authors to unwittingly use
an incorrect nomenclature. The credit for uncovering the synonymy belongs to
Krajina, who has carefully studied the genus.

The key given below is based on relatively constant characteristics that are easily
observable in the field on mature plants. Other characteristics, such as trunk height,
frond length, pinnule width, and number of sori per pinnule, vary greatly with the
age and vigor of the plants. They are not reliable diagnostic features and have not
been used in the key.

KEY TO THE HAWAIIAN SPECIES OF CIBOTIUM
1. Stipes with reddish-brown or stiff purplish-black hairs along most of their length.

2. Fronds distinctly glaucous beneath; costae villosulous

2. Fronds not glaucous beneath; costae glabrous 2. C. chamissoi
1. Stipes naked, or with dull brown or golden brown hairs mainly at their base.

2. One or both of the basal pinnule segments auriculate, fronds glabrous and distinctly glaucous.

3. C. glaucum

1. C. st.-johnii

2. None of the pinnule segments auriculate; fronds not or only slightly glaucous beneath.

3. Stipes very dark brown to blackish with a conspicuous, white line along each side when fresh; pin-

nule segments covered with tufts of hairs (as seen under 10 X lens) beneath ...... 4. C. nealiae

3. Stipes brown or greenish when fresh; pinnule segments covered with cobweb-like hairs (as seen
under 10 x lens) beneath.

4. Trunk slender and smooth, stipes covered with inconspicuous, soft, matted, dull, brown hairs.

5. C. hawaiense

4. Trunks rough with stubs of dead fronds; stipe bases with golden brown hairs 6. C. splendens

1. Cibotium st.-johnii Krajina, Stud. Bot. Cechosl. 1(2):94. 1938.

The most characteristic feature of C. st.-johnii is the copious amount of matted
hairs on the full length of the stipes, and on the costae and costules. The hairs at the
base of the stipes are, when fresh, reddish-brown. Further up the stipes and on the
costae they are grayish-brown to whitish. Cibotium st.-johnii closely resembles C.
glaucum, but has much smaller fronds. In 1966, Dr. Krajina annotated Heller 2818
(BISH 01703, 01703a 01703b) as “Cibotium glaucum (Sm.) H. & A. subsp.
St.-Johnii (Krajina) c.n. var. fallax Krajina.” The description of Hillebrand’s (1888,
Pp. 547). C. chamissoi var. B can be used to confirm the identity of C. st.-johnii.
Distribution: Alakai swamp, Kauai.

2. Cibotium chamissoi Kaulf. Berl. Jahrb. Pharm. 21:53. 1820.

C. menziesii Hook. Sp. Fil. 1:84, 1. 29C. 1844.

This is the most robust species. The stipe bases are covered with long, reddish-
brown hairs; further up the stipe, the hairs are still and blackish. The rachis and the
costae are often straw-colored. The fronds are not at all glaucous beneath and
usually are glabrous, but sometimes have minute, scattered tufts of hairs visible
under 40x magnification. Distribution: Kauai, Oahu, Molokai, Lanai, Maui, an
Hawaii. Hawaiian name: hapu’u’i’i.

R. BECKER: IDENTIFICATION OF HAWAIIAN CIBOTIUM 99

On the islands of Molokai, Lanai, and Hawaii, I have seen several tree ferns that
may be hybrids between C. glaucum and C. chamissoi. Their fronds were distinctly
glaucous beneath and their basal pinnule segments auriculate. These are characteris-
tics of C. glaucum; however, their stipe hairs were bristly and either dark purplish-
brown or dark orangy-brown.

3. Cibotium glaucum (J. E. Smith) Hook & Arn. Bot. Beech. Voy. 3:108. 1832.
Dicksonia glauca J.E. Smith, Rees’ Cyclop. 11(22). 1808.

Cibotium glaucum is the most abundant species on the Island of Hawaii. The stipe
bases are covered with a mass of glossy, golden-yellow hairs and are lustrous when
dry. The distal part of the stipes and the rachis are essentially naked. The fronds are
glabrous and bright glaucous beneath (slightly less so in the wetter parts of the range
of the species). The pinnule segments closest to the costae are auriculate and overlap
the costae. The auricles are largest on plants from the island of Hawaii; they are
present but much smaller on plant forms from the other islands. One plant I
examined on Molokai had one frond exauriculate. In this species, the stipe bases
remain attached to the trunks for some time after dead fronds drop off. Distribution:
Kauai, Oahu, Molokai, Lanai, Maui, and Hawaii. Hawaiian name: hapu’u pulu.

4. Cibotium nealiae Degener in Degener & Hatheway, Fl. Haw. 5: Family 12.
1951.

The stipe bases of this species are very dark brown, almost black, and are covered
with a thick mass of matted, yellowish hairs. Distally, the stipes are greener and
naked. A conspicuous, long, white line (pneumatothode) extends along each side of
the fresh stipes. I found the pneumatothodes to be very obvious when I saw the
species in the field. Although the lines are a useful field identification, they did not
remain visible on my herbarium specimens. The costules and veins are yellow and
prominent. The frond underside is green, but not glaucous beneath. The costules are
covered with very small tufts of hairs beneath. Distribution: Hanalei, Kauai.

5. Cibotium hawaiense Nakai & Ogura in Ogura, Bot. Mag. Tokyo 44:468.

1930 e
The spelling of the epithet hawaiense follows Nakai and Ogura’s bala
publication. The stipes of C. hawaiense may appear naked, but actually are aes
with thick, flaky mats of dull brown hairs. When a stipe is pulled back eae the
trunk, the basal part is seen to be covered with golden, glossy, and slightly sae
hairs. The laminae are thin and bright green, the underside is slightly paler an
somewhat glaucescent. Fine, cobwebby hairs can be seen, with a 10 x a see
the undersides of the pinnules. The trunk is slender, and the fronds, alter arc ri
upwards, are held almost horizontally. The dead fronds hang down close to the
trunk, similar to those of the Date Palm. Eventually they break away cleanly, leaving
a smooth trunk. Distribution: Olaa, Laupahoehoe, Kohala Mountains, Hawaii.
Hawaiian name: meu.
6. Cibotium splendens (Gaud.) Krajina ex Skottsb. Acta Horti Gothob. 15:40.
1942.
Uranie 96, 369, f. 21. 1827.

Pinonia splendens Gaud. in Freyc. Voy.
1:83. 1844.

Cibotium chamissoi sensu Hook. Sp. Fil.

100 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

This is the most delicate appearing species. It has been confused with C.
hawaiense, although their ranges do not overlap. A glaucescent form from Molokai
(Fosberg 9600, BISH 01741) was annotated with the unpublished name C. splendens
var. glaucescens by Krajina. This specimen has pinnule segments that are somewhat
glaucous and with arachnoid hairs on their undersides. Distribution: Oahu, and
perhaps also on Molokai, and Maui, but not seen by me on these islands, or on
Lanai or Hawaii.

I wish to thank Sandra Becker for her assistance in mounting my reference
collection of herbarium specimens. The review of the preliminary draft and discus-
sion on the nomenclature of C. nealiae by Dr. V. J. Krajina were most helpful. The
continued encouragement and support of Drs. D. Mueller-Dombois, K. W. Bridges,
and C. Lamoureux, are much appreciated.

LITERATURE CITED

CHRISTENSEN, C. 1905-1906. Index Filicum. H. Hagerup, Copenhagen.

HILLEBRAND, W. F. 1888. Flora of the Hawaiian Islands. Reprinted 1965, Hafner, New York.
HOLTTUM, R. E. 1963. Cyatheaceae. Flora eee. I, 1:66-176.

HOOKER, W. J. 1844. Species Filicum, vol. 1. ndon

soe ce % F. 1913. The Indigenous Trees of ee Islands. Published under patronage, Honolulu,

Haw:
SKOTTSBERG, C. 1942. Vascular plants from the Hawaiian Islands. Part 3. Pteridophytes collected
during the Hawaiian bog survey 1938. Acta Horti Gothob. 15:35-14
SMITH, A. R. 1981. Flora of Chiapas, part 2. Pteridophytes. Calif. Ac ad. Sci., San Francisco.
STOLZE, R. G. 1976. Ferns and Fern Allies of Guatemala, part I. Fieldiana, Bot. 39: 1-130.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 4 (1984) 101

Two New Tree Ferns from Panama
ROBERT G. STOLZE*

Recently I was sent two specimens of tree ferns to identify, which were thought to
be Cnemidaria: one by Henk van der Werff at the Missouri Botanical Garden, and
another by David Lellinger at the U.S. National Herbarium, which was a duplicate
he had received from Missouri. One proved to be new species of Cnemidaria, the
other a new, simply pinnate Trichipteris. 1 am always hesitant to describe new
species based only upon single collections, as is the case with these, but each is
highly distinctive from others in their respective genera. It is curious that both of
these new Panamanian species collected between 1975 and 1982 are very small (for
tree ferns) and are simply pinnate.

Cnemidaria glandulosa Stolze, sp. nov. Fig. 1.
Folium pinnatum, tenuiter herbaceum, ca. 130 cm longum, | 25 cm latum,
segmento apicali non conformi; petiolus haud spinosus, petioli paleis abundantibus,
bicoloribus: lamina in pagina abaxiali dense glandulosa; glandes minutae,
bacilliformes; rhachis plus minusve nuda; pinnae subse
ine integro et revoluto; venae 2- vel 3-furcatae, liberae, vel raro venae basales
anastomosantes et areolas costales efformantes, indusium atrobrunneum, plus
minusve semicirculare; receptaculum ovideum, sine paraphysibus.

VTYPE: North of San Felix at Chiriqui-Bocas del Toro border, on Cerro Colorado
copper mine road along continental divide. Lower montane rain forest, 5,000—5,500 ft,
Province of Chiriqui, Panama, Mori & Kallunki 5908 (US; isotype MO).

Thus far known only from the type collection.
Caudex not seen; leaf pinnate, thin-herbaceous, to 130 cm long and 25 cm broad,
abruptly terminating in a nonconform apical section (7.e. broader at base than the

i ntially naked, or with a few scattered scales or
i ; pinnae about 12

bas we
only abaxially, the glands dark brown, minute, appressed, bacilliform; costae naked
adaxially, sparsely provided with broad, flaccid, amorphous, whitis
abaxially; veins 2—3-forked, essentially = hg spaced He ge ee
Be thon aie ae oe ay pe pitts indusia dark brown, more Or less
id. lacking paraphyses, spores pale yellow to srl
white, globose-tetrahedral, trilete, with one large aperture near the center of each face
near the equator

The he: species bears closest resemblance to C. decurrens of peas om
northern Central America, especially in that some forms of the latter have cou
entire pinnae. However, in C. decurrens the lamina is eglandular and coe - sen
reduced to the apex, veins commonly (not rarely) merge to form costa pe eae
the indusia are pale or yellowish (not dark) brown. The occurrence 0

*Department of Botany, Field Museum of Natural History, Chicago, IL 60605.

102 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

FIG. 1. Holotype of Cnemidaria glandulosa, Mori & Kallunki 5908 (US).

glands on the abaxial surface of pinnae is unique in the genus. In Cc glandulosa the
glands may be observed even at low magnification, and by using higher power
(25 x) one can easily discern their rod-like shape.

R. G. STOLZE: TREE FERNS FROM PANAMA

FIG. 2. Holotype of Trichipteris
oi S3

pinnata, Knapp 5881 (MO).

Spanitade pinnata Stolze, sp. nov.

m pinnatum, tenuite
incor anicnll conformi,

r vel firme herbaceum,

la rca cecal
es x :e i a
sia eee:
ere
| 58822 > bhi
PANAMA
Fa
ae 180 N
terrestrial. Sori browne
MISSOURI BOTANICAL GARDEN HERBARIUM (
Fig. 2.

circa 50 cm longum, 20

cm |
segmentum apicale ad basim articulatum; petiolus haud

103

104 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

spinosus vel interdum sparse muricatus, squamis ovatis vel lanceolatis, nitidis, brunneis
vel interdum bicoloribus; pinnae nonnullae, breviter petiolulatae, ad rhachim articulatae,

/TYPE: Santa Rita Ridge road, 21-26 km from Transisthmian Highway. Tropical wet
forest, 500-550 m, Province of Colén, Panama, Knapp 588] (MO; isotype F).

Thus far known only from the type collection.

Caudex erect, ca. 6 cm long, | cm broad; leaf pinnate, thin- to firm-herbaceous, to
50 cm long and 20 cm broad, abruptly terminating in a conform apical segment which is
articulate at the base; petiole 18-22 cm long, yellowish or grayish brown, lacking spines,
but sometimes slightly muricate, amply scaly at least near the base, the scales 5-7 cm
long, ovate to lanceolate, lustrous brown or some of them bicolorous (with narrow,

short-stalked, articulate to the rachis, the margins crenate, the apex acuminate, the base
subtruncate to broadly rounded, the tissue between the veins rather densely gland-dotted
on the abaxial side; costae glabrous abaxially, densely provided with appressed tri-
chomes as on the rachis adaxially; veins 4—6 pairs along the costule, only the distal ones
reaching the pinna margin; sori borne on the proximal 1/3 or 1/4 of each vein branch;
indusia lacking; receptacle globose, with short paraphyses among the sporangia; spores
pale yellow to whitish, trilete, rather sharply tetrahedral.

Compared to most tree ferns, this is a very diminutive one—the fronds are only half a
meter long and the caudex is small, not trunk-like. Of course, it must be Pp 1 that
more collections are made of this species, much larger plants also may be found.
Trichipteris pinnata is most likely to be confused with T. williamsii Maxon of Panama,
Colombia, and Venezuela, one of two species in the genus with entire or subentire
pinnae (Trichipteris typically has huge leaves, 2—3-pinnate or more). Both species also
have the lamina abruptly terminating in a conform apical segment which is articulate to
the rachis, and the lateral pinnae are nodose-articulate as well. In 7. williamsii, however,
the pinnae are subcoriaceous, with their margins entire, or at best slightly undulate, and
the basal veins often or occasionally merge to form costal areoles. Pinnae of T. pinnata
are thin- to firm-herbaceous, with their margins rather deeply crenate and veins are all
free. Leaf tissue in T. williamsii is eglandular, whereas the abaxial surface of the pinnae
in T. pinnata is copiously dotted with minute, castaneous glands.

There is some disagreement about the valid spelling of the name Trichipteris, some
authors preferring “Trichopteris.” Tryon (1970, p. 41) adhered to Presl’s original
spelling, as did Barrington (1978, p. 19) in his revision of the genus. Holttum and
Edwards (1983, p. 161) and others prefer to adopt Schott’s alteration to “Trichopteris”
for orthographic reasons. I do not intend here to belabor either argument, as the rules
according to the I. C. B. N. are rather ambiguous and/or flexible in cases of this kind. I
prefer to follow Tryon, who separated the many species of Trichipteris from Alsophila,
and Barrington, who did the revision.

LITERATURE CITED

TRYON, R. M., 1970. The classification of the Cyatheaceae. Contr. Gray Herb. 200:1-53.
BARRINGTON, D. S. 1978. A revision of the genus Trichipteris, Contr. Gray Herb. 208:1-93.
HOLTTUM, R. E., and P. J. EDWARDS. 1983. The tree-ferns of Mt. Roraima and neighbouring areas

of the Guayana Highlands with comments on the family Cyatheaceae. Kew Bull.
38(2):155-188.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 4 (1984) 105

Trunk Length and Frond Size in a Population
of Nephelea tryoniana from El Salvador
Ralph L. Seiler*

Field observations of tree fern fronds suggested that small tree ferns have longer
stipes relative to lamina length than tall tree ferns of the same species. This study
was done to determine how total frond length, lamina length and stipe length vary
within a population.

In April 1979, measurements were made of 146 fronds on 45 individuals of
Nephelea tryoniana Gastony from a dense cloud forest at 2250 m in Bosque
Montecristo, a wildlife preserve in northwest El Salvador. The forest is described
more completely in Seiler (1981). The trunk length of each tree was measured, as
well as the stipe and lamina length of all undamaged fronds. Usually three fronds
per tree were measured, but occasionally four or five or as few as two per tree were
measured. The majority of the fronds were sterile, but the presence of fertile fronds
should have no effect on the results, for fertile and sterile leaves are practically
monomorphic in Nephelea (Gastony, 1973).

RESULTS

The length of the fronds of a tree is fairly constant. The standard deviation within
a tree averaged 4.55% (s=2.96, n=45). Analysis of 139 fronds on 43 trees
indicates frond length is weakly related to trunk length (r=0.39, P<0.01). Two of
the 45 trees were not included in the statistical analysis for reasons that will be
discussed later. In Figure 1, frond length is plotted against trunk length. Each point
represents the mean frond length from one tree.

The ratio of lamina length to stipe length within a tree is also fairly constant, the
standard deviation within a tree averaged 5.72% (s=3.01, n=45). Analysis of 139
fronds on 43 trees indicates that shorter tree ferns do have longer stipes relative to
lamina length and that the lamina/stipe ratio is strongly correlated to trunk length
(r=0.90, P<0.01). In Figure 2 the ratio of the lamina length divided by the stipe
length is plotted against the trunk length; each point represents the mean lamina/stipe
ratio for one tree.

Two trees, numbers 17 and 22 (Figs. 1 and 2), were found that had been knocked
over and, although the main trunks were covered with humus and litter, the —
had subsequently turned and grown upright. Usually the trunk length is more or =
equal to the height of the crown above the ground, but in both of 8 a =
actual height of the crown was more than a meter less than the length 0 e wea ;
The data gathered from these two trees were not included in the statistical ana yses
because these two trees were different from the rest in that the effective height en
much less than the trunk length and being forced to change the direction of growt

may have had an unknown effect on the apical meristem. cacaanel
The effective height of the crown is better predicted by the measured a ies
ratio than is the total trunk length on the two fallen trees. In tree 22 the lamina/stipe

*3977 S. 775 W., Bountiful, UT 84010.

106 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Mean Frond Length, in meters

Mean Lamind/Stipe Ratio

' T r T “Tr Gi T
2

3
Trunk Length, in meters

ms. I, Relationship between mean frond length and trunk length in Nephelea tryoniana. See text for
explanation of numbered points. FIG. 2. Relationship between lamina length divided by stipe length
(lamina/stipe) and trunk length in Nephelea tryoniana. See text for explanation of numbered points.

RL. SEILER: TRUNK LENGTH AND FROND SIZE IN NEPHELEA 107

ratio is 1.23, the trunk length is 160 cm, and the effective height is 16 cm. The
predicted trunk length is 10 cm for this tree. Similarly, tree 17 has a lamina/stipe
ratio of 1.42, a trunk length of 137 cm, and an effective height of 27 cm. The
predicted trunk length is 41 cm.

DISCUSSION

More measurements of trees taller than two meters would have been desirable, but
would have required cutting the trees down. Since all tree ferns are very rare in El
Salvador and are protected in Bosque Montecristo, only the 5 m tall tree was cut
down. The sampling bias towards small trees could cause. undue weight to be put on
the values for the two tallest trees, suggesting a correlation between frond and trunk
length that may not really exist. Omission of the data from the two tallest trees
reduces the correlation coefficient; however, the correlation is still significant
=0.22, P<0.05).

The lamina/stipe ratio is strongly correlated with the trunk length. The effective
height of the crown seems to be the determining factor for the lamina/stipe ratio.
The relative length of the stipe in the two trees that had been knocked over is much
better predicted by the effective height of the crown, rather than by the total length
of the trunk.

Although leaf angle was not measured, it was observed that the fronds of the
smaller tree ferns are held more erect than those of larger ferns. Functionally, it may
be that the more erect leaf angle and the relatively longer stipes on small tree ferns
combine to lift the lamina further toward the light. For mechanical reasons, the stipe
may have to be shorter to support the more massive, more horizontal lamina of taller
tree ferns.

Further investigation is needed to determine what combination of environmental
and internal developmental factors determine the relative stipe length and its func-
tional significance. The developmental age of the apical meristem of the fern may be
important in determining the lamina/stipe ratio, while the most probable environ-
mental factors would be light intensity and quality.

LITERATURE CITED
GASTONY, G.. 1973. A revision of the fern genus Nephelea. Contr. Gray Herb. 203:81-148.
SEILER, RL. 1981. Leaf turnover rates and natural history of the central American tree fern Alsophila
salvinii. Amer. Fern J. 71:75-81.

108 AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 4 (1984)

An Unusual New Elaphoglossum from Peru
ROLLA TRYON*

A distinctive species of Elaphoglossum has been recognized among specimens
received for identification from the Missouri Botanical Garden.This new species has
a combination of characters that adds to the diversity of its large genus. It also
emphasizes the need for more intensive collecting in the species-rich eastern Andes
of Peru.

Elaphoglossum pascoense R. Tryon, sp. nov. Figs. 1, 2.

Caulis terrestris dorsiventralis valde repens, petioli adaxialiter sulcati fasciculis
vascularis sex, phyllopodiis nullis, lamina sterilis oblongo-ovata ca. 12—24 cm longa

.5—12 cm lata profunde cordata subcuspidata venis liberis longis, lamina fertilis
oblonga ca. 12 cm longa 2 cm lata, sporae reticulato-echinatae.

Say ¢ = 4

FIG. 1. Spore of Elaphoglossum pascoense, X 5555.

= res vith dark, appressed scales, these few except at the apex, stele dorsiventral;
eaves 28-54 cm long; petioles gradually merging into (long-decurrent on) the stem
spaced, as long as to usually longer than the lamina, more or less covered by

scales; fertile lamina densely covered beneath by sporangia, these intermixed with
es

*Harvard University Herbaria, Harvard University, Cambridge. MA 02138.

R. TRYON: A NEW ELAPHOGLOSSUM FROM PERU
109

x 0.56.

FIG. 2. Holotype of Elaphoglossum pascoense, with fertile leaf in center,

110 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

TYPE: Road between Oxapampa and Villa Rica, Dept. Pasco, Peru, 75°20”W,
10°73”S, on flatter parts of steep ridge, 11 October 1982, Robin B. Foster 9127
(GH; isotypes MO, others to be distributed).

The most unusual features of Elaphoglossum pascoense are the relatively
broad, deeply cordate sterile lamina with wholly free venation. The sterile lamina
can only be compared to that of E. crinitum, which has fully areolate venation.

This new species agrees with Elaphoglossum sect. Setosa (Mickel & Atehortua,
1980) in the lack of phyllopodia, the long-creeping stem, the presence of hyda-
thodes, and the reticulate-echinate spores. It has the long-creeping stem and dark
stem scales of subsect. Alpestria. However, it is quite different from all of the
enumerated species of that subsection, which have the sterile lamina rather narrow
and more or less cuneate at the base.

I am indebted to Dr. Alice F. Tryon for SEM preparations of the spores. This
research was partially supported by National Science Foundation Grant DEB
81-05726 to Rolla Tryon and Alice Tryon.

LITERATURE CITED
MICKEL, J. T. and L. ATEHORTUA, G. 1980. Subdivision of the genus Elaphoglossum. Amer. Fern
. 10:47-68.

AMERICAN FERN JOURNAL: VOLUME 74 NUMBER 4 (1984) 111

New Tropical American Ferns
JOHN T. MICKEL*

In the course of identification work for various Latin American field programs at
the New York Botanical Garden, several new species of ferns have come to light.
Three of the new taxa are in Anemia, a genus of which there is yet much to be
learned.

trols
Anemia elaphoglossoides Mickel, sp. nov. Fig. 3B

Frondes dimorphae, lamina sterilis indivisa integra glabra coriacea, lamina fertilis
bi- ad tripinnata.

Rhizome horizontal, compact, ca. 4 mm diam., clothed with red-orange hairs,
5-8 mm long; fronds clustered, 10-20 cm long, dimorphic, the stipe /2-¥s of the
frond length, stramineous, glabrous, slender, 0.5 mm diam., 7-13 cm long; sterile
blade simple, elliptic, glabrous, coriaceous, 3.5—4.5 cm long, 1.0-1.2 cm wide,

broadly cuneate at the base, the apex obtuse; veins free, dichotomously forking,
leaving the midvein at an acute angle; margin thickened; fertile frond about equalling
the longest sterile frond in length, bi- to tripinnate; spores unknown.

UTYPE: Brazil: Est. Goids: ca. 15 km S of Niquelandia; gravelly hillside and sandy
cerrado, rooted at base of overhanging rock; elev. 1000 m; 21 Jan 1972, H. S. Irwin
et al. 34664 (UB; fragment NY).

Known only from the holotype, two plants, one sterile and one fertile. This is the
only known species of Anemia with totally undivided sterile blades. Its relationship
within the genus probably in subg. Anemia, but the precise relationship is not known.
The spores are too young to see the ornamentation; the stomates are floating.

Anemia marginata Mickel, sp. nov. Fig. ~
Frondes dimorphae, lamina sterilis ternata pinnata, segmenta lanceolata coriacea
glabra viridi-grisea ad marginem incrassata flava, frondes fertiles bi- ad tripinnatae
quam steriles multo longiores. ;
Rhizome horizontal, compact, short-creeping, Ca. 3 mm diam., hairs on
colored near the apex, maturing to orange-brown; fronds dimorphic, sterile see
erect, 3.5-7 cm tall; stipe /2—%4 of the frond length, stramineous, glabrous; - e
ca. 1 cm broad, ternate; lateral pinnae small, ca. 7 X 3 mm, entire, ssi ation ps
ascending, with an acute apex, the base broadly attached; terminal sere arger, ¢ “a
14x4 mm, lanceolate, entire to slightly , Oe atc glabrous, Veins
free; ish- with distinct thickened yellow margin, ©. |
a a 53, 15 cm tall, much surpassing the sterile fronds in length, 2-3
pinnate.
TYPE: Brazil: Est. Goids: Alto Paraiso, Chapada dos Veadeiros, 5 km E of od
Paraiso, elev. 1600 m; locally common in patches of humus and sand os ees as
spots near boulders in high sandy flat exposed place; 15 Feb pr, eres

Estabrook 206 (UB; isotype NY).

*New York Botanical Garden, Bronx, NY 10458.

112 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

FIG. 1. SEM photomicrographs of _— spores. A, B. Proximal and distal faces of A. salvadorensis

ale 947), 510. C. Proximal face of A. sanctae-martae (Steyermark & Rabe 96462), x510.

Spr face of A. hirsuta (Mickel 62 41), X510. E. Distal face of A. oblongifolia (Irwin 34329),
F. Proximal face of A. clinata (Williams 2584), x 510

J. T. MICKEL: NEW TROPICAL AMERICAN FERNS 113
ois Tt
Anemia salvadorensis Mickel & Seiler,' sp. nov. Fig. 2C.

Rhizomata adscendentia; frondes usque 17 cm longae, usque 3.5 cm latae; stipes
quam lamina sterilis quintuplo brevior;, lamina sterilis anguste obovato-oblonga vel
oblonga pinnata, sparse pilosa; pinnae steriles 3-9 jugae oblique oblongae obtusae
dimidiatae margine denticulatae; nervi liberi; frondium fertilium stipites laminas
subaequantes; pinnae fertiles patentes vel oblique adscendentes, a pinnis sterilibus
remotae.

Rhizome ascending, clothed with reddish-brown hairs; fronds to 17 cm long, 3.5
cm wide; stipe of sterile frond ca. 1/6 of the frond length, deeply grooved adaxially
and laterally, stramineous, wiry, moderately hirsute with reddish, tortuous hairs,
glabrate; blade narrowly obovate-oblong or oblong, terminating in a flabelliform or
obcordate apical segment, pinnate, thin, dull, sparsely pilose on both surfaces; sterile
pinnae 3-9 pairs, subsessile, slightly ascending, to 1.9 cm long, .8 cm broad,
obliquely oblong, obtuse, strongly inaequilateral at the base (dimidiate), truncate
above and cuneate below, erose denticulate, the margin slightly cartilaginous, a
distinct midrib lacking; veins free, evident, slightly raised on the adaxial surface;
stomates floating; fertile fronds similar to the sterile but longer-stiped, stipes up to
half the total frond length, the fertile pinnae 0.7—1.7 cm below the sterile blade;
fertile pinnae to 3.2 cm long, spreading to slightly ascending, short-petiolulate,
tripinnate, the ultimate divisions slender and virtually lacking lamina; spores
tetrahedral-globose, striate, the ridges slender, few, with noticeable space between
6-7 «um, the space being about the same width as the ridges), 76-94 xm diam.
TYPE: El Salvador: Depto. Ahuachapan: Montana de Agua Prieta, near San
Benito, among grass on steep, disturbed, sunny slope, elev. 700 m, 20 Feb 1979,
Seiler 947 (NY; isotypes F, MHNES). Known only from the type collection.

This species is distinct in the habit of the fertile pinnae and pinnate blade. In the
habit of the fertile pinnae, the ascending rhizome, and floating stomates it resembles
members of subg. Coptophyllum sect. Adetostoma (A. brandegeea, A. intermedia,
and A. clinata), which are found in western Mexico (the first two) and Panama to
Bolivia (A. clinata), according to Mickel (1962, 1966). However, the pinna form is
more like that of A. sanctae-martae of Colombia, and in form of the pinnae except
for their undivided nature like that of the more ubiquitous A. hirsuta, which 1s
closely related to A. sanctae-martae (subg. Anemia sect. Hirsutae). The very narrow
form of the blade and the very short stipe suggest a relationship to A. oblongifolia
(subg. Anemia sect. Oblongifoliae). The spores of A. salvadorensis most closely
resemble those of the A. hirsuta complex (incl. A. sanctae-martae) (Figs. 1A-E).
t clear, it is our conclusion that the
the evolution of the genus, lying

The section Adetostoma was placed in subg.
its spores with many closely set ridges, as In “. ;
The floating stomates were thought to be an anomaly or ete pone ane
subgenus. On the other hand, they are known in some species of SUD. = a
and in all species of subg. Anemia. On the basis of the ascending rhizome,

(Mr. Seiler served for two years in the Peace Corps in Fl Salvador. ye a uae
much of the country collecting plants and preparing specimens ee

Historia Natural de El Salvador (MHNES).

114 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

. 2. A. Habit of Saccoloma membranaceum. B. Pinna of S. membranaceum. C. Habit of Anemia
salvadorensis. D. Habit of Anemia marginata.

J. T. MICKEL: NEW TROPICAL AMERICAN FERNS 115

floating stomates, and the distinctive, obliquely held fertile pinnae, we would place
the new species in sect. Adetostoma, but with the similarities to sect. Hirsutae and
sect. Oblongifoliae of subg. Anemia, we would also remove sect. Adetostoma from
subg. Coptophyllum and place it in subg. Anemia.

Patent fertile pinnae are known from several groups within the genus Anemia: A.
aspera and A. perrieriana (subg. Coptophyllum sect. Anemiaebotrys), A. elegans
and A. eximia (subg. Coptophyllum sect. Trochopteris), A. colimensis (subg.
Anemiorrhiza) and all of sect. Adetostoma (subg. Anemia). These four groups are
probably not closely related, and whether any of them actually represents the
ancestral type in the genus is not known. It is clear that the patent fertile pinnae
represent the primitive condition in the genus (Mickel, 1967), being derived from
fronds with sporangia distributed over the abaxial surface and then limited to the
basal pair-of pinnae, but it is not known whether the four groups within the genus
today that have patent fertile pinnae are remnants of this condition or whether they
are reversions to this condition. There are very likely other species of Anemia to be
found in Latin America that hopefully will shed light on the phylogenetic relation-
ships within the genus.

fol
en acutiserratum (Hieron.) Mickel, comb. nov. me
10'S 2 Asplenium cirrhatum L. C. Rich. var. acutiserratum Hieron., Hedwigia 60:259. 1919, as
“acutiserrata.’~ LECTOTYPE (selected here): Trinidad, Fendler 140 (presumably B; isolectotype NY).

|
i ticaiam longicaudatum Mickel & Stolze, sp. nov. Fig. 3A.

Ab. A. cirrhato Rich. ex Willd. frondibus grandibus et apice pinnarum acuto
differt.

Plants terrestrial; rhizome stout, erect, provided with lanceolate to ovate, acute
scales, these 2-4 mm long, 0.6-1.0 mm wid k brown or (more commonly)
bicolorous, with narrow, lighter brown margins, subclathrate, the lumina isodiamet-
ric or slightly longer than broad; fronds 70-100 cm long, 12-18 cm pir cet

iole 10-30 cm long, terete, dart brown to atropurpureous, lustrous or sublustrous,

ith e

iv crenulate; veins indistinct or obscure, commonly
eto: ot se ee pees inéar to narrowly oblong, slightly arcuate, indu-
sium linear, yellowish, firm-textured, subentire. oe Ri
“TYPE: Ecuador: Pcia. Morona-Santiago: Cordillera Cutucu, ridge between K10s
Itzintza and Chupiasa, 1200-1350 m, Camp E-1283 (NY: esate oe.

This species belongs to the A. radicans complex. Bis. somewnat fe bide is
widespread A. cirrhatum in that they are beg, ones pee vie She more
considerably smaller with obtuse to subacute pinna apices, It resem a. ‘ata
closely A. acutiserratum (Hieron.) Mickel of Trinidad, aan ee pig ee nee
apices but with serrate pinna margins, auriculate pinna bases, an

normally reach 35—45 cm in length.

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

116

Sal
+ se

1
| Mi,
M 4
= WA
| Wy,
WIR tA
CPI V4

gicaudatum. B. Habit of Anemia elaphoglossoides. C. Habit of

FIG. 3. A. Habit of Asplenium lon

Grammitis kirkbridei.

J. T. MICKEL: NEW TROPICAL AMERICAN FERNS 117

Grammitis kirkbridei Mickel, sp. nov. Fig. 3C.
. knightii (Copel.) Seymour frondibus parvioribus, lobis approximatioribus,
stipitibus brevioribus differt. Haec species Joseph H. Kirkbride, Jr. botanico dicatur.
Plants epiphytic; rhizome erect, ca. 0.6 mm diam, clothed with yellow-brown,
lanceolate scales ca. 1 mm long; fronds fasciculate, 3-5 cm long, 5—9 mm wide;
stipes 3-9 mm long, clothed with stiff, spreading, reddish-brown hairs 1-—1.5
mm long; blade pinnatifid, cut /2-% to the midvein, the lobes obtuse, slightly
ascending, 10-13 pairs, with abundant, stiff, erect, reddish-brown hairs ca. 0.4 mm
long; veins obscure; sori mostly 3 or 4 pairs per lobe, ca. | mm iam., medial
between the midvein and the margin, which is often slightly recurved.

TYPE: Panama: Pcia. Darién: Tres Bocas on the Rio Coasi, 1 May 1968, J. H.
Kirkbride Jr. & J. A. Duke 1381 (NY).

Grammitis kirkbridei belongs to the group of G. basiattenuata (Jenm.) Proctor
of the West Indies and G. truncicola (Klotzsch) Morton of South America, but has
fewer lobes and shorter laminar hairs. It most nearly resembles G. knightii (Copel.)
Seymour, from which it differs in its smaller, less deeply cut fronds, pinnae closer
together, and shorter stipe.

Polystichum hottense C. Chr. Figs. 4A, B.

For many years this species was known only from the type collection from the
Massif de la Hotte in southwesternmost Haiti (Ekman H-10130) and part of a mixed
collection from Ma Blanche, Haiti (Ekman H-5/3). It has recently been collected at
the mining site of Las Abejas, Prov. Pedernales, in southwestern Dominican
Republic (Liogier 14173, 26784; Mickel 8168; Zanoni 16718). This striking plant
with bipinnate fronds and abundant, spreading, often black-centered rachis scales
has never been well illustrated and is thus presented here.

/ O l ¢ L
Pteris striphnophylla Mickel, sp. nov. Figs. 4C, D.

b P. longifolia L. frondibus parvis, lamina sclerotica glabra, apice pinnarum

obtuso differt. ‘ ;
compact, the scales re guage
light brown, 1.5-2.5 mm long, in a tangle of wiry roots and old stipe bases; sti
2-4 cm son with hairs eine to narrow scales at the base; fronds approximate,
7-11 cm long, 1.5-2.4 cm wide, pinnate, with a conform terminal Sag pinnae
6-9 pairs, mostly 10-12 mm long and 4-5 mm wide, entire, pe 0 sag ie
obtuse, the base rounded to cordate, extremely coriaceous, pinna er es tops
rachis swollen, the pinnae tending to be deciduous; veins obscure, = = =P fo
l-forked; blade glabrous except for a few short hairs at pinna pores meals
continuous along the margin, interrupted only at the pinna apex; 1n us
coriaceous. :
TYPE: Dominican Republic: Pcia. Pedernales: Sierra de Baoruco, a i
Las Mercedes to Aceitillar, 1000 m, growing in a ravine, the Sieg a
rock cliff, 11 Feb 1969, Bro. Alain Liogier Spe (NY; — JBSD).

This is clearly related to P. longifolia, judging trom Its
spores, vine bases, swollen pinna attachments, and arent te conceal
easily distinguished by its small size, glabrous blade, and heavily

texture.

118

“esa

AA
Sx
4
=>

"ag

“thi

x SS

LAY

LAG

SOD) aoe HL

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

N
4 .

UC / eZ
WS,
eZ LUTE?
Saad (WISI

‘AW Wag

(Pkéic,

Wiki.

kes

WAOe az,
Stam NI

arg...
a
: ANNA és
i wa
prnttitce.

Liz
SSK.

DUBBO: .
-) NQ) ThKiz,
Wii

ES
AM

Sane

A. Habit of Polystichum —
pin

FIG. 4.
Pteris striphnophylla. D. Same,

5mm

(Liogier 14173). B. Same, rachis and pinna base. C. Habit of

J. T. MICKEL: NEW TROPICAL AMERICAN FERNS 119

Saccoloma membranaceum Mickel, sp. nov. Figs. 2A, B.
Ab aliis speciebus generis frondibus parvibus et ne membranacea differt.
Rhizome horizontal, short-creeping, 2-3 mm dia with coarse, black,

roots 0.40.6 mm diam.: rhizome clothed with Sadie reddish-brown, hea

indurated, linear-deltate scales 1-3 mm long; fronds many, approximate, 10-1

ong, 2.0-3.4 mm wide; stipe 2-3 cm long, castaneous to light brown, bina

0.3-0.4 mm diam aa ade n arrowly lanceolate, 1-pinnate to a pinnatifid apex, the

t pinnae cut less than y e midvein, the basal acroscopic lobe
slightly hip accept veins free, Me simple or dichotomously orking, more
divided in larger lobes; lamina membranous, al on veins, ca. |

labrou
mm from the margin, indusia cup-shaped; pried se with low, dichoto-
mously branching ridges.

VTYPE: Brazil: Est. Acre: Cruzeiro do Sul, Rio Jurua and Rio Moa, Serra do
Moa, Rio Moa 6 km above school; forest on mountain; 25 Apr 1971, Prance et al.
12432 (INPA; isotype NY).

The label states that the type specimen is epiphytic, but the roots hold consider-
able sand and all the relatives are strictly terrestrial.

This species is strikingly different from other members of the genus in size and
texture. The spores are similar to those illustrated by Tryon and Tryon (1982, p. 386)
in their low, dichotomously forking ridges. The rhizome scales and sorus structure
and location also support its placement in Saccoloma.

ACKNOWLEDGMENTS

These studies were supported in part by a grant from the National Science
Foundation (DEB 77-25582). Thanks also to Dr. Rupert Barneby and Joann Ward for
their help with the Latin diagnoses, Mr. Joel Huang for the SEM photomicrographs,
and Bobbi Angell and Charles Clare for the drawings. Furthermore, Mr. Seiler
expresses his appreciation to the Museo de Historia Natural de El Salvador and to
the Unidad de Parques Nacionales y Vida Silvestre for their support during his stay
in El Salvador.

LITERATURE CITED
MICKEL, I T. 1962. A monographic study of the fern genus A
22. ce 36:349-482.
"1966. A new species of Anemia from South
—_———. 1967. The phylogenetic position of Anemia co
TRYON, R. M. and A. F. TRYON. 1982. Ferns and Allie

nemia, subgenus Coptophyllum. lowa

America. Amer. Fern J. 56: 58-60.
limen sis. Amer. J. Bot. 54: 432-437.
d Plants. Springer-Verlag, New York.

120 AMERICAN FERN JOURNAL: VOLUME 74 (1984)

SHORTER NOTE

An Urban Locality For Asplenium platyneuron—The ebony spleenwort,
Asplenium platyneuron (L.) B.S.P., was recently discovered at a site in Brooklyn
(Kings County), New York. Vouchers consisting of only sterile fronds are deposited
at BKL (W. Sax-Cedar & J. Goldbaum, 15 Feb 1984; T.J. Delendick, 22 Feb 1984).
The population consists of fourteen individuals growing in a vertical stone wall at a
well-developed and heavily trafficked intersection in the Park Slope district, approx-
‘imately midway between Prospect Park and the Gowanus Canal. The wall is part of
an access well to a below-ground basement entrance to a commercial building. The
well is approximately 12 feet deep, 5 feet wide and 40 feet long, with an east-west
orientation. The ferns occur in the uppermost seven feet of the wall, among clumps
of moss in the mortar between the granite stones; they are restricted to the east end
of the well, thus having a western exposure. None of the plants examined had fertile
fronds.
This Brooklyn station represents the second verified locality for A. platyneuron
known to exist on western Long Island at the present time, and the only one which
is current for Kings County. The other site for the species in the western portion of
the island is at Fort Totten on the north shore of Queens County, reported by Greller
and Locke (Amer. Fern J. 73:6-8. 1983). This situation contrasts strongly with the
distribution of the species on Long Island in the 19th and early 20th Century, when
it was noted as “frequent throughout the island” by Jelliffe (The Flora of Long
Island, 1899), including at least three locales in Kings County: East New York,
G. B. Brainerd in 1866; Rockaway Beach, G.D. Hulst in 1889; and Central Park
[now designated as Prospect Park], J. McCallum in 1902 (all BKL). It is, or at any
rate was, fairly common in the eastern counties of Nassau and Suffolk. Stalter,
(Bull. So. Carolina Acad. Sci. 41:72. 1979) cited it as occurring on Long Island,
and the late Stanley Smith of the New York State Museum, Albany, noted it in both
Nassau and Suffolk counties (manuscript checklist).

This find also represents the only current site for A. platyneuron on Long Island
south of the terminal moraine that forms the backbone of the island. There is a
specimen of the species at BKL from Rockaway Beach (Kings County, 1889) and at
NY from Valley Stream, Nassau County (Bicknell, 8 Aug 1903). The latter includes
the collector’s note, “Bank of Aqueduct causeway. The only plants I have found
south of the terminal moraine.” The outwash plains from the glacial moraine which
form the southern portion of Long Island provide few habitats suitable for Asplenium
species. The occurrence of A. platyneuron there seems, on the basis of available
data, to be restricted entirely to man-made sites. It is remarkable that A. platyneuron
has been able to extend its range into an area which has been so long and so heavily
used as to lack virtually any trace of the original flora, and that its extension was
made possible through commercial development.

I am grateful for the assistance of Wynne Sax-Cedar, who made the first
collections from the newly discovered locale, and to Andrew Greller and Richard
Stalter for their comments and review of the text.—Thomas J. Delendick, Brooklyn
Botanic Garden, 1000 Washington Ave.., Brooklyn, NY 11225.

AMERICAN FERN JOURNAL: VOLUME 74 (1984) 121

AMERICAN FERN JOURNAL

Manuscripts submitted to the JOURNAL are reviewed for scientific content by
one or more of the editors and, often, by one or more outside reviewers as well.
During the past year we have received the kind assistance of J .D. Caponetti, W. D.
Hitz, H. T. Horner, Jr., J. D. Montgomery, M. G. Price, H. E. Robinson, A. R.
Smith, R. G. Stolze, W. C. Taylor, D. P. Whittier, E. Wollenweber, and J. J.
Wurdack, to whom we are deeply indebted. We welcome suggestions of other
reviewers.

Dr. Alan R. Smith, Department of Botany, University of California, Berkeley, CA
94720, will become editor of the American Fern Journal effective 1 Jan 1985. Dr.
David B. Lellinger, U. S. Nat’l. Herbarium NHB-166, Smithsonian Institution,
Washington, DC 20560, will become editor of Pteridologia on the same date.

—D.B.L

1985 A.1.B.S. MEETING-CALL FOR PAPERS
The American Fern Society and the Botanical Society of America will meet with
the A.I.B.S. Annual Meeting at the University of Florida, Gainesville, on 11—I5
August 1985. Those members of the American Fern Society wishing to present a
paper or poster and who have not received abstract forms may obtain forms from the
program chairman of the Pteridological Section of the Botanical Society: Dr.
Christopher Haufler, Dept. of Botany, University of Kansas, Lawrence, KS 66045.

INDEX FOR 1984

divaricatum, 40: ecuadorense, 41-43: fontanum var. obovatum,
jongicaudatum, 115, 116:

Adiantum, 29; anceps, 29; capillus-veneris, 17; pedatum at
: 40, 41, 43: oellgaardii, 44. 45.

aleuticum, 62. subsp. calderi, 62, subsp. pedatum, 62,
ilum, 62, ee subpumilum, 62: trapeziforme. 9
, 19,

an
0

Ampelopteris prolifera, 5 ; ;

A . TL, 112, 115; sect. Adetostoma, 113, | subg tum, 50: sect. Sphenopteris, 49 . : pees =
Anemia, 111, 113, 115; sect. Anemiaebotrys, 115: subg 47, 49, trichomanes 4 var. herbace
Anemiorrhiza, 113, 115: aspera, 115: brandegeea, 113: clinata x trudellii. 63 plier

12, 113; colimensis, 115: subg ee 113. 115: Athyrium subquadripinnatum. 57; tenuifolium,
elaphoglossoid 11, 116; elegans, 115; exima, 115: hirsuta, Azolla,
112, 113: sect rsutae, 113, ae Ses ae i Becker. R. es identification of Hawaiian tree ferns of the genus

111; 114: oblongifolia, 112, 113: s
ig salvadorensis, | 12-114:

: oF OP
12, 113; s t. Trochopteris, 115

Anti- saleechlal activity of phenolic acids in Pteridium aqilinum,
8

as gaa cristatum X marginale, 62: cubense. 60: dicksonioides.

Coke 14. 15. 40, 49, 50, 120; acutiserratum.

adiantum-nigrum, 14: aethiop
S|

a
conquisivem, 40, 41: cristatum,

ect. Oblongifoliae. 113.
sanctae-martae.

icum, 94: cegriscee oe
cirrhatum,

Cibotium, 9
ah S.S. etal. era flora of Garhwal Himalaya (rev.).
Botrychium, 79_85: dissectum, 78. 79. 83-85, f: ‘ikem.

na Sctgeetaieel ——e oe
of cma Isoétes. 9
Cibotium, 97. 98: sist 97: barometz, 97: ae
97-99: cumingii, 97: glaucum 97-99, var. fallax. 98:
- menziesii, 97. ‘98: nealiae, 97-100: regale
_ var. glaucescens, 100:

97, d
st.-johnii, 97, 98: we
Cnemidaria. 101: glandulosa. oa so decurrens. 101

122

Cooper-Driver, G. (see M. San Francisco)
Crabbe, J.A. (see A.C. Je
— ae 56: haitiensis, 56; protensa var. dicksonioides,
56; sloanei, 29
Cees Biggs 58; chaseae, 58; congesta, 58; cryptosora,
8

56; fen . 58; flexuosa, 58: longipinnata, 5
Pie roman eae, 56
usick, A.W. Graves’ > ort in Ohio, 63
pris 19-21: alternans, 21; atahuallpa, 56; bradei, 57:
nii, 2 staal 0: excelsa, 20: insignis, 19, 20:
intramarginalis, 57; lathamii, 20: oe ots
luccana, 21; parianensis, rs princeps, 19-21; sect
Oe ar 21: sipapoensis. 57: sect. Spinco oI;
g. Sphaeropteris, 19-21: squamulata, 2
Pr eae,
Cyrtomium falcatum, 94
Cyclosorus supine 6
Davallia novae-guineae, 56
Death notice: Gu alts Looser (1898-1982), 60
Delendick, T.J. An urban locality for Ae pa platyneuron,
120
Dicksonia, 19; antarctica, 19: arborescens, 19; glauca, 99,
lathamii, 19
Dicranopteris rent 57; peruviana, 57
Dictymia mc
ah Bor ot ‘sce, 6; ee 6; petri, 6; subquadri-
7: tenuifolium
Pescporceend 37-39; ites 38
Drymoglossum under $s, 37
Drynaria acuminat. “_
Dryopteris aripensis, 56; cristata, 62: cristata x marginalis, 62:
——: var. Paranensi 56: ireneae, 60: marginalis, 62:
, x slossonae, 62; x slossoniae, 62; subincisa var.
ensis, 56
———— 108: subsect. Alpestria. 110: crinitum, 110:
ee 110; sect. Setosa, 110

opine inte;
Equiset 61, GL: 69-73, 75, 76; arvense, 65: 66; x ferrissii,
61 67 . 73-76; fluviatile, 66: subg. dopasanee
. affine, 65, 67-71,

6l, 65-75; palustre, 65, 66, 75; ramosissimum, 61
ramosissimum in Louisiana, 61
Exonotins flexuosus,
Farrar, D.R. (see L.M. Rut
Ferns and fern allies of Sau temala, part [II (rev.), 36
Flora of Ecuador 14(4): eR Se TARE (rev.),

Frequency of cyanogensis in Bracken in relation to shading and
winter severity, 51

Gleichenia angusta, 57:
ponderous insidiosum, 57; integrum

boliviensis, 57; peruviana. 57
m, 57: nesioticum, 57

Gra saiaagrcnie in Ohio, 63

sont roides, 58; angus' _ 58: basalis, 58:
besiattenuata, 7 uesii, 58: chase , 58: ciliolepis. 58:
congesta, 58: oe 61: fendle: eure feta 58; humilis,
58: Kegelian SDs, killipii, 58: cick idei. 116, 117; knightii
117; Seana . 58; maxoniana ayoris, 59: mela-
notricha, 59: ene 5: perp la. 59: shaferi. 59:
truncicola, 117: williamsii,

Gualterio Looser (1898-1982) (death notice), 60
Gymnogramma a . 61
Gymaoptens rufa,

pee

ITissii
S parent species, Equisetum hyemale and Preah
rae in Iowa, 65
aki Hag isetum ramosissimum in Louisia
Hensipean. E. .C. Roos. A monograph Py sy genus

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

Platycerium Sync (rev.),

Herndon, A. & R mdon. Two sah of Adiantum newly

m under water stress. 37

R.J. Chro of neotropical Isoétes, 9
The ‘ion of Faveiies tree apie of the genus Cibotium,

fiscal of pteridophytes of Japan, volume 3 ( ,6
new phenolic shail i in aia

11; andicola, 9,

Isoétes Ho, 22 , 25, 36; alcalophila, 10,
11, 12; boliviensis, 11; coromandelina 26; durieui
glacialis, 11, 12; herzogii, 11, 12; indic 9 25; lacustris, 11.
2; lechleri, 9; macrospora, 11; anadensis, 10, 11;
anchananii, 9; pantii, 9, 25; rajasthanensis, 22, 23 3120;
retic slat 22, 23, 25, 26; storkii, | 1; ticlioensis, 9, 11, 12

Jermy, A. C., J. A. Crabbe, and B. A. Thomas (eds.). The
6

tag and — tio a)
Kur. . & T. Nakaike. hataliéod of saison of Japan,
pay

ata x are 62

aa; 22)

ew Sena. and some new names in
$,.96; rns on North American ferns,

ee rev.
Lastrea cristat:
Leclercquia aes
Lellin _ D.

Leptogram ar ee var. alabamensis, 30
iol e ligule of —

, 61; cuspidata, 61; wallichiana, 61

‘ scopodiolin x brue ei, 63

Lycopodium, 36; appressum, 63; X brucei, 63; prostratum, 63
Marsilea, 36

Matte euccia, 1; struthiopteris, 1-5; orientalis, 5

Mickel, J. vee w tropical American ferns, 111

Microgramma acuminata, 59; latevagans, 59; recreensis, 59:
tuberosa, 5

Microsorium, 28

Microsorum, 28

Miller, K. G. Trichomanes gametophytes at Bartholomew's cob-
bie, 31

A monograph of the fern genus Platycerium (Polypodiaceae)

. T. Usos de los helechos en Suramérica con
especial relists a Colombia (rev.),
Nakaike, T. (see S. Ki
slossonae, 62
names in ferns, 56
A new station for ‘aerate eer in Alabama, 30
New — American ferns, 111
Not rican ferns, II, 62

rma, 35

organic nutrition of Botrychium gametophytes, 77

Osmunda, 84: cinnamomea, 5

Parris, B. S. PY taxonomic revision of the genus Grammitis . . .
in New Guinea (rev.), 86

Pecluma camptophyllaria var. lachnifera, 59; eurybasis var.

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

glabrescens, 59. var. villosa, 59; ptildon var. robusta, 60
Pellaea < wrightiana, 33: longimucronata. 33: ternifolia, 33
Phegopteris conne ee 6
The ee and classification of the ferns (rev.), 96
Pinonia splen ship
Platycerium, iar semana. 18

“Shai pleolepis, 60

lypodium alike 58: angestipalcani, 56: aureum. 94:
asale, 58; buesii, 58: ciliol 8. cryptosorum, 56:
decipiens urybasis var. villosum, 59; flexuosum, 58
umile, 58: insidiosum, 57: keg um, 58: lachniferum, 59
var. glabrescens, 59: latevagans, 59; loretense, 59; mayoris, 5
mckeei, 60; melanotricha, 59: ticum, 57: perpusillum
59; cape 60: sans 58: recreense, 59: robustum

; : sha feri

: . 59; tubero:
Po fae cubensis, 60
Polystichum 6; "none 33; aculeatum, 33: braunii, 6, 35:
hottense, 117, 118: imbricans, 33, mbricans. 33:
imbricans X lemmonii, 33: lonchitis, = eh aa BA

osum, 59: williamsii, 59

e, 61
es from Ecuador. 40
m Pue rto pe p
Promotion ok apogamy in Matteuccia struthiopteris, the Ostrich
=k

. 5; aquilinum, 1, 5. 17, 51. 87,
a softs flora of Garhwal Himalaya (r
Pteris longifolia, 117; peas 94. pence 117, 118
A remarkable Cyathea hybrid,
Revi Ferns and lies i. roe part III, 36; Flora of

— = 94

: ge
sine Guinea, 86; Usos de los helechos en
| referencia a Reins ia
~ MC. (see EB. ‘Heonipmad

nape E. (see W. H. Wagner, Jr.

utz, L. M. & D. R. Farr ee habitat characteristics and
bundance of Equise ‘cial and its parent species
uisetum hyemale ne Equisetum laevigatum, in Lowa, 65

Saccoloma, 119: membranaceum, 114, 119

Sadleria

Salvinia, 36

: . Cooper-Driver. Anti- microbial activity
of phenol acid in Pteridium aquilinum,
Schre _ Nafus & D. Pimentel. Frequency of cyanogensts

in Baas in relation to diading and winter severity, 51
Seiler, R. L. Trunk length and frond size in a population of

123

Nephelea mares from El oo 105
Selaginella, 22, 8 a
Selliguea sem sitg
harma, B. D. & ee The ligule of Isoétes, 22
e B. D. Sharma)
. R. Flora of Ecuador 14(4): Polypodiaceae—Thelyp-
ih otis Wi} 30
sia Ne oe atahuallpa, 56: bradei, 57; horrida. 19. =
intramarginalis, 57:
nsis,

ri ® (se

lock woodiana.

36: Problems in Asplenium gon some new y tpecied Sonn
; a, 101

gemmifera, 9

A taxonomic revision of the genus Grammitis . . . in New Guinea
. 86

Breer 55: subg. Amauropelta, 55:

ekmanii, 60; subg. Glaphyropteris. 55:

ireneae ; limbo: ;

subg. Cyclosorus, 55:

Tmy)
Three new cabiioieicie in » bicsecaise, 6l,

odea ara,

cee Fess; ies
chaetum, 7: padronii
Trichomanes, 7

subg. Achomanes, 7: . Pachy-
nii, 7, 8; petersii, 30: sien 7: subg.

tophytes at scoieisirenmial s cobble. 31
. 1 i
wo new p lic g r ov icgieeiinne. 4
Ti rs Dp. ma 1 ]
rwo new

0
Two species of Adiantum ig escaped in Florida, 29
Lane Se 104: pinnata, 103. 104; williamsii, 104
richopteri aes

ae os i + was ee ae |

— El — =

usu.

Elap! gone um a thon Peru
An urban ie . de platyneu
Usos de los helechos en Suramérica con cde referencia a

Colombia (rev.) ee
Vittaria, 31

on Aderkas, P. Promotion of apogamy in Matteuccia struthiop-

teris. the Ostrich Fern,
H.. Jr. & E. Rouleau. A sara Holly Fern.
“aE x scopulinum, in N riccinconil n
Whittier, P. The organic nutrition of ae gameto-
ytes
Wyatt, R. A new Station for Teichomanes petersii in Alabama, 30
Xiphopteris killipii, 58: morton

ERRATA eae ih

Page 59, line 27: For “recreense’” read “recree

Page 60, lines 23 and 24: For “Gaulterio” ae mGualterio.”

124

AMERICAN FERN JOURNAL: VOLUME 74 (1984)

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PS Form 3526, July 1984 (See instruction on reverse,

A GUIDE TO HARDY FERNS
by Richard Rush

A Special Publication of the British Pteridological Society

Contains over 600 species and varieties believed to have potential
horticultural value for the cool temperate zone, especially western
Europe and North America. Each entry is annotated with world distribu-
tion, bibliographical references to descriptions and illustrations, ecolog-
ical and horticultural requirements, growth habit, and variations. A
comprehensive introduction and discussion on categories of hardiness
facilitates maximum use of this book. 70 pages, softbound.

Distributed in the U.S.A., Canada, and Mexico by Trainpower, Inc., Publications
Division, P.O. Box 1255, Vienna, VA 22180 ($8.00 postpaid). Elsewhere: The
British Pteridological Society Booksales, 46 Sedley Rise, Loughton, Essex
1G10 1LT, England (£4.50 postpaid).

WANTED DEAD OR ALIVE!

High quality, heart-shaped fern prothallia, live or
FAA-preserved, for slide making.

We need monoecious and dioecious prothallia
as well as stages showing sporophyte develop-
ment. Please send samples for viewing. We pay

for high quality specimens only!

We also supply high quality prepared microscope
slides to those who need the very best for their
teaching or research.

TRIARCH, INC. - P.O. Box 98 - RIPON, WI 54971
(414)-748-5125

BRITISH PTERIDOLOGICAL SOCIETY Oe
Open to all who are interested in growing and studying fens ane “OF THE
Full members receive THE FERN GAZETTE and the BULLETIN ake
BRITISH PTERIDOLOGICAL SOCIETY. Membership OF eerie and
for full members, £ 5 for ordinary members (not receiving the san : :
£5 for student members (under Fe years of age). For sguoteelt pene
residents should apply to Dr. J. Skog. Biology Songhai iy Se
University, Fairfax, VA 22030. Non-U. S. residents ca ag uF nd
5 Coke Robin Hill, Sanchcomie, OMe Oe

Ferns of Jamaica
A guide to Pteridophytes
G. R. Proctor

amaica has one of the richest fern floras in the world, yet hitherto there has been no
publication dealing authoritatively with the Jamaican ferns i in their entirety. Ferns of
amaica will rectify this situation, by p of all the
knowz ferns and fern allies of the island. The aim has been to present a scheme of
classification with broad validity, reconciling as far as possible older views which empha-
sized resemblances with more recent attitudes that emphasize differenc
This work names and describes the 579 ‘Species and 30 varieties of ferns recorded ir in
Jamaica. Keys to the subfamilies of P
eae ie ails relevance and potential for wide use in the whole ‘American tropics.
the min distinguishing features of closely related plants.
ni stily mp the eighty- -three genera are illustrated, many with splendid nineteenth
century drawings taken from rare and often inaccessible sources. Where suitable
illustrations could not be found, new drawings have been specially commissioned for this
work. Each taxon is succinctly but fully described, and data on ga local and general
geographical distribution are given. Habitat and altitudinal range are stated for most of
the species. The author has _ — waioaiae all previous literature dealing with
Jamaican ferns in the course of his research, a An
extensive bibliography is seid ae care has been taken to present all
information in a standardized format for ease of consultation

Ferns of Jamaica is an immensely practical flora, Sapctially in the information it
provides on habitat and ecology. The work’s definitive and authoritative nature will
ensure its recognition as the standard work on the subject for many years to come.
sabe bleh oo Acknowledgements, — ion Pteridophyta, Conspectus of
Major Taxa, Systematics and Descriptions of Species: Class I. Psilopsida, Class II
pe sats Class IIL Lycopsida, Class IV. Nera ag Bibliography, Glossary, Index.
The Author

One time Senior Botanist in agence of the ephyge of the Science Museum, Kingston,
Jamaica, Dr. G. R. Proctor is botanist with thirty years of experience
in the West Indian islands, including Tanciaize. and the Central American countries.
610pp, 135 line illustrations, 22 maps. Hardback. 0 565 00895 1

Publication due 3rd January 1985.

ORDERS TO: Rudolph Wm. Sabbot, Natural History Books, 5239
Tendilla Avenue, Woodland Hills, Calif, 91364. Tel: (818) 346 7164.
[Authorised agent of the British Museum (Natural History)].

Price $75.00 plus 10% postage and packing.

British Museum (Natural History)

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