4V

tssued February 7, 1910.

U. S. DKPARTMKNT OF AGRICULTURE

BUREAU OF ANIMAL INDUSTRY.— Bulletin 118.

A. D. MELVIN . Bureau.

CULTURAL STUDIES OF SPECIES
OF PENICILLIUM.

BY

BOTANICAL
GARD

CHARLES THOM, Ph. D.,
Mycologist in Cheese Investigations \ Hairy Division.

WASHINGTON:

GOVERNMENT PRINTING OFFICE.

1910.

• .?4
T5I5

THE BUREAU OF ANIMAL INDUSTRY.

Chief: A. D. Melvin.
Assistant Chief: A. M. Farrington.
Chief Clerk: Charles C. Carroll.
Biochemic Division: M. Dorset, chief.
Dairy Division: B. H. Rawl, chief.

Inspection Division: Rice P. Steddom, chief; Morris Wooden, R. A. Ramsay,
and Albert E. Behnke, associate chiefs.
Pathological Division: John R. Mohler, chief.
Quarantine Division: Richard W. Hickman, chief.
Zoological Division: B. H. Ransom, chief.
Experiment Station: E. C. Schroeder, superintendent.
Animal Husbandry Division: George M. Rommel, chief.
Editor: James M. Pickens.

II ITER OP TRANSMITTAL.

U.S. Department of Agriculture,

Bi im.ai of Animal Industry,
Washington, D. C, July 12, 1909.
Sir: I have the honor to transmit herewith and to recommend for
publication as a bulletin of this Bureau a manuscript entitled "Cul-
tural Studies of Species of Pt n ic'iUiu m ," by Dr. Charles Thorn, my-
cologist in the cooperative soft-cheese investigations carried on at
Storrs, Conn., by tlie Storrs Agricultural Experiment Station and
the Dairy Division of this Bureau. A previous paper of this author
dealing with certain species of PeniciUium which are characteristic
factors in the ripening of Camemhert and Roquefort cheese has been
published as Bulletin 82 of this Bureau. The investigations con-
nected with the study of the organisms referred to necessitated the
culture and com pari -mi of many other species and an examination of
the nomenclature for the whole genus P< nicillium. Considerable con-
fusion was found to exist regarding the identification of the various
specie-, hence it seemed important that a comprehensive study from

cultural data of all obtainable species should be undertaken.

Acknowledgment is made of suggestions and advice by Dr. Erwin
F. Smith, of the Bureau of Plant Industry of this Department, be-
sides the persons whose names are given in the text.

The illustrations, with one exception, are from original drawings
1>\ t he nut hor.

Respect fully, A. D. Mi:i.\ i\.

( Tiief of Bureau.
I lou. James Wilson,

Seen tary of Agriculture.

CONTENTS.

] 'age

Introduction 9

The basis for specific characterization LO

Necessity for describing culture media and temperal lire conditions 11

Habit, -i run ure, and appearance of colonies 15

Physiological effects upon media 19

Culture media 22

Nomenclature 23

The generic name 23

Nomenclature of Bpecies 23

The type species 25

Explanation of drawings 27

/', niiill in in i i pn ii sin, i Link 27

I', nicillium italicum Wehmer 29

/', nicillium digitatum Saccardo 31

/', nirill in m roqueforti Thom 34

P< a i rill in in purpurogt num 0. Si nil 36

/', nicillium pinophilum Hedgcock (nomen oovum) 37

/', nicillium rubrum 0. Stoll 39

/', nirill in in I hi, a a, Zukal 39

/', nicillium duclau ri I >elacroix 42

I', nicillium clavifornu Bainier 13

I', nicillium granulatum Bainier 1 1

/', // :,-illiiini In, ri, -mi I, Saccardo 15

/', nicillium br< vicauL Saccardo, var. album Thom, n. var 17

Penicillium brevicauh Saccardo, var. glabrum Thom, n. var is

/', nicillium roat tun Link (?) in

I't nicillium runt, mberti Thom 50

/', nirill in in nun, m In rli var. rogt ri Thom, n. var 52

I', n icillium bifornu d. sp -''1

/', nicillium commune a. sp 56

Penicillium No. 22 :

I', nil ill in in chrysogenum n. sp 58

Penicillium rugulosum d. sp no

I', nir, Ilium rilrin u m n. ~|> lil

Pen icillium X... :{? 63

/'. nicillium X" 12 64

/', nicillium alranu nto&um a. sp 65

Penicillium No. 24 66

I', nirill in in slnlniiiji r a in n. sp 68

/', nirilHii tit Jtintrtilnstiiii n. sp 69

/', II iri 1 1 in m il, ninth, lis II. .- ] > 71

I', nicillium divaricatum a. sp 72

I', nicillium lilacinum n. >p 7:1

6 CONTENTS.

Tajce.

/'< nicillium intricatum a. Bp 75

Penidllium spinulosum n. sp 7(>

Penicillium No. 28 77

Species forming pink sclerol ia 78

Penicillium No. 29 78

Penicillium No. 30 79

Penicillium No. 31 80

Penicillium No. 32 80

Comparal ive cultural data 81

Cultures in distilled water 82

Agar-agar as a source of food 83

Agar-agar as a source of carbon 83

Various sources of carbon 83

Cultures in Raulin's fluid and in Cohn's solution 86

Color in conidial areas 87

Effect of concentrated media 88

The grouping of species 89

Odors 90

Anaerobic cultures (with carbon dioxid) 90

Incubation tests 91

Summary of data from comparative culture 94

Keys to cultural identification of species 94

Key 1. — Analysis of species in cultures upon gelatin and agar 95

Key 2. — Species determinable from substrata 97

Tabular statements 98

Table 1. — Tabulation of salient characters of species 98

Table 2. — Gelatin and color reactions 100

Table 3. — Comparative cultures 101

Table 4. — Comparative cultures in synthetic fluid (Dox's) 102

Table 5. — Decimal summary of comparative cultures in synthetic fluid

(Dox's) 104

Table 6. — Incubation experiments 105

References to literature 106

Index to species 109

ILLUSTRATIONS

Page.

1'iL. I . /'. nidllium t tpansum Link 28

2. Penidllium italicum Wehmer 30

:;. /', nidllium digitatum Saccardo '•'<■'•

I. /'. nidllium roqueforti Tin mi '■'•■)

5. /'. nil-ill in in jiiir/iunii/i ii a in ( ). Si (.11 37

6. I'i nidllium pinophilum Bedgcock :;s

7. I'i iiirill in in r ii hr ii in 0. Si oil 10

8. I', nidllium luU um Zukal 41

«t. I', nidllium duclauxi Delacroix • -'

10. I'i nidllium davifonru Bainier 13

11. /'< nidllium granulatum Bainier 45

12. I'i nii-illiiim hniiriiiili Saccardo l(i

1:;. /'< nidllium bn iricaul* . var. album 17

11. l't nidllium br< wicauh . var. gl brum Is

L5. I'i nil i Hi a in roseum Link (?) 19

L6. /'- nidllium cam* mb< rti Thom 51

17. I'rnii-illiiini camemberti, \ ar. rogt ri 53

18. I'i nirill in in bifornu

I'i. Penidllium commum 57

l'ii. I'i nidllium chrysogt num 59

i!l . /'< nidllium rugulosum 61

•j_'. /'. nirilliii in dtrinum 62

23. I'i nirill in m .\n. :;7 64

24. /'- nidllium atrarru ntosum 66

25. Penidllium No. 24 67

26. I'i nidllium stoloniferum 68

L'7. /'- nidllium funiculomm 'u

/', nirilliii ill ill ill lliln ns '1

l'Ii. /'« nidllium dii aricatum '• • :>

30. I'' n'n ill iii iii liladnum - '

31. I'i nidllium intracatum <•>

32. /'< nidllium spinulosum 76

;;:;. I', nidllium No. 28 • '

34. I', nil-ill in in No. 29 '■'

35. I'i nn ill in in No. :il 80

36. /'< nidllium No. 32 81

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

INTRODUCTION.

In a previous paper ' two species of PeniciUium were shown to
secrete the proteolytic enzyms which ripen certain varieties of
cheese. The effort to identify these organisms necessitated the
culture and comparison of numerous other species and a study of
the literature of nomenclature for the whole of thegenus Penidllium.
The difficulties encountered in deciding whether to discuss these
forms under old names or to describe them anew from cultural data
led to the extension of this study beyond the forms occurring in a
dairy investigation so as to include any obtainable species, and
especially all species whose identification under published names
could 1 stablished.

Many recent studies have linked particular chemical and physio-
logical activities with the presence of particular species of fungi.
When such data relate to known species -identifiable species — our
knowledge of the metabolism of these forms has been greatly in-
creased. When, as has often happened, the generic name alone is
given in a group so diverse in its activities as PeniciUium, such data
only add to the confusion. Whether the observations apply to all
of the species of the genus or to a single unnamed species is left un-
settled. To give real utility to such work, some one must test the
applicability of the data to each species under consideration. On
account of their ease of cultivation and the notion that there was a
single common green species legitimately named P. glaucum, species
of PeniciUium have formed the subjects of many such investigations.
In very few of these cases are sufficient data regarding morphology
given to w arrant even a guess as to t he species used. An e.xaiuinat ion
of the present stat us of specific uomenclat lire in t he genus will t here-
fore furnish a sounder basis for further studies of their activities.

This paper represents cultural work which has continued more
than four years and includes those species lor which the dat a obt ained
seem abundantly t<> justify the characterization offered. Some of
these forms have been cultivated for the whole period, others for a
less time. \o claim to monographic completeness can be made. In

"The figures refer to the list of literature at end of bulletin.

10 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

collecting these forms, many of them have been isolated in this
laboratory from dairy products and from fruits, or collected in the
field by the writer. One series of forms was purchased from Krai, of
Prague. Several species have been contributed by or verified by
those who described them — two by Dr. G. Bainier, two by Dr. C.
Welimer, one by Dr. G. Delacroix, and one by Dr. G. G. Hedgcock.
Many correspondents have sent cultures for examination or study.

THE BASIS FOR SPECIFIC CHARACTERIZATION.

The available sources of identification of these species will be
discussed first. By the kindness of Professor Thaxter the exsiccati
of the genus in the herbarium of Harvard University were examined.
Cultures of certain species were tried but no living spores were
found. No specimen was found in such condition as to be used to
identify material by comparison. Similar courtesy was extended
for the examination of specimens in the herbarium of Kew Gardens
and at the University of Berlin. In no case was material found by
which cultural material could be identified or identification verified
by comparison. The plant bodies are too evanescent to retain the
criteria necessary for identification for any great length of time, as
ordinarily preserved. In most cases after a few years of handling
the specimens were found reduced to powder. Welimer 33 has noted
for certain species of this genus that the conidia do not retain their
power to germinate beyond a very few years. Specimens as formerly
prepared therefore become useless for the identification of species
by the method of types. The method described by Hedgcock9 for
preserving specimens has thus far been applied only to one species
of Penidllium and entirely too recently to test its permanent value.
Aside, therefore, from certain species which will be discussed later,
it has been impossible to determine material belonging to this genus
from herbarium specimens.

. There remain two methods of identification: (1) The identification
of material or cultures by the authors of descriptions; (2) identifica-
tion by critical study of the descriptions and illustrations published.

Regarding the first method, fortunately some of the more recent
authors have either preserved their organisms in culture, as is done
at the Ecole de Pharmacie in Paris with the cultures of Bainier, or
placed them in one other of the distributing laboratories where
such organisms are maintained in continuous culture. But the earlier
authors are dead and have left only their published descriptions and
figures as a means of determining what organisms they studied.

Turning to the critical comparison of material with published
descriptions, we find many difficulties. These descriptions give in
meager outline observations of mold masses showing penicillate coni-
dial fructifications, as found in nature upon substrata more or less

BASIS FOB SPECIFIC CHARACTERIZATION. 11

accurately specified. No cultures were made. The original masses
are assumed to be comprised each of a single species. Parasitism or
selective saprophytism is assumed, bul the substratum is rarely
designated with sufficient care to make a duplication of the original
culture possible. Hence the characters would include whole series
of forms whose differences are marked. No account is taken of their
omnivorous nature nor of the marked variations introduced in ap-
pearance by changed conditions. The method of types and the met hod
of substrata as represented by the herbarium material and the pub-
lished descriptions we have are. therefore, not sufficient for the
identification of species of P< niciUium.

Alt hough we a hand on the "method of substrata" as the sole has is
of description, the force of natural selection as shown by the distri-
bution of certain of these species in nature is a most valuable acces-
sory. Sonic species occur so constantly upon particular substrata
and under particular conditions as to simplify their identification
greatly. Examples of this are P. italicum Wehmer and P. digitatum
Saccardo, as they are found upon citrus fruits. Unfortunately very
few species restrict themselves to particular substrata, so that except
for some few species and a very few media, the occurrence of a
Penicillium in any given situation is hut slight evidence for its
identification. Tin' constant occurrence of species of PenidUium
in the laboratory, in connection with foods, and in factory processes,
such as cheese-ripening, all point to controlled culture as the proper
source of diagnostic characters.

If we look to cultural study" for our conception of species we have
t wo methods of procedure : (1) The exhaustive study of the limits of
variability for each species; (2) the comparative study of numerous
species under arbitrarily chosen conditions uniformly maintained.

The first is the best method known for gaining complete knowledge
of single species, hut it is too cumbrous for taxonomic purposes.
Physiological and chemical studies have commonly been restricted
t<> part icula r classes of reactions for single species or groups of species.
These have contributed much to our knowledge of fungous varia-
bility, but t Hen give no hint, except the vague nomenclature

used, upon which to judge which species were actually studied. In
experimental cultures changes in the chemical nature of the medium
or in the conditions, or both, have been found to produce great
changes in the morphology of the fungi studied. With the exception
of a few fundamental group or generic characters, nearly every

aWeidemann, ' in a recenl paper, has described several new species of this genus
in their relation to culture media. En this he lias followed the bacteriologists fun her
than the writer has thought necessary by giving formulas for a considerable number
of media and detailed notes a- to reactions upon such media, instead of the more
formal descriptions attempted here.

12 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

attribute used in specific description has been shown to be a reaction
to environment, hence changeable with such environment (for some
species at least). Exhaustive study of all the species would be
endless.

The alternative is to select certain media and a particular set of
conditions, then to cultivate all the organisms under investigation
in a uniform manner and to base distinctions of species upon differ-
ences in the reactions obtained, and upon the differing characters of
the several species, in this common environment.

To test the reliability of such data, species obtained in the dairy
laboratories were first studied carefully upon the peptone-milk-sugar
gelatin described by Conn4 and upon potato agar as described in
our previous bulletin (Thorn,25 p. 7). From these cultures trans-
fers have been made to media of very different composition — Cohn's
solution, Raulin's fluid, milk in various forms, synthetic fluids pre-
senting different sources of carbon — alwa}'s bringing cultures back
to the original media. Many species differ so materially in gross
characters when grown upon these different media that successive
cultures, if not known to be pure transfers, might be supposed to
be different species; but when returned to the original media and
conditions these forms have immediately produced the characters
and reactions first found, with a large degree of uniformity. Abso-
lutely uniform reactions are not to be expected from living organ-
isms, at any rate under our imperfect control of working conditions;
but when such reactions are definitely recognizable as essentially
the same, the result may be judged as satisfactory. It is even more
confusing to find that two or more species may react very similarly
upon a particular substratum. A transfer of these organisms to a
medium of markedly different composition brings out the contrasting
characters, however. The desirability of recording the widest pos-
sible distinctions in such descriptive work makes necessary the use
of media differing in composition as much as practicable, in such
comparative cultures.

It is worthy of note that the species P. roqueforti and P. camemherti,
essential to the cheese industry, have been isolated repeatedly from
cheeses of widely different origin. The Roquefort species has been
obtained from laboratories not concerned with Roquefort cheese
studies, from ensilage, and from other substances. The same char-
acters have been found in cultures of these two species from these
variable sources under conditions in which the possibility of close
genetic connection between cultures is thus remote. There seems to
be little possibility of question that these species at least are well-
fixed conidia-bearing forms, not cultural varieties of other species.
The question at issue was not whether or how variations could be
produced, but whether a particular variation is constantly produced

BASIS FOB SPECIFIC CHARACTERIZATION. 13

by a species in a particular environment. Th«v correlative question
whether the characters of a given species of mold can be perma-
nently changed by passing through a series of cultures upon different
media is involved in the same investigation. It is asserted by some

workers that the physiological react ions of molds (if not also the
morphology) can be changed, and that such changes persisl after
the return of the species to the original environment. So far as this
investigation has gone such a view is certainly not supported by the
conduct of the species of P< niciUium which have hen studied. In
those species most thoroughly studied both the physiological and
morphological reactions have appeared to be very reliable. A sum-
mary of these observations follows:

1. The same species may differ greatly in morphology and physi-
ological reactions when grown upon different media.

2. Two species closely similar, when grown parallel in one en-
vironment, may differ characteristically when transferred to a differ-
ent medium or a different set of conditions.

3. With these species the repetition of culture under particular
conditions produces fairly constant morphology and reactions.

Forms arising in this way have been designated as ecads by Clem-
ents^ in a recent discussion of the "Aspects of the species question."
This name is used preferably for forms whose origin is known either
because produced by cultivation or under such carefully determined
natural conditions as admit of full description. With species of
this genus in which part at least of the morphological characters of
every culture are definitely attributable to the conditions and to the
chemical character of the substratum, practically all known forms
would therefore be properly designated as such ecads. If described
as ecads, however, each description must be limited strictly to the
data obtainable upon a single medium, whereas by writing into the
description the reference to media it has been found possible to in-
clude a much more complete set of characters than would readily be
worked out from a single set of conditions.

These observations lead to the conclusion that the cultural descrip-
tion of species of molds demands the recognition of the points noted
below:

1. The culture media and conditions must he described SO fully as
to make the repetition of the culture upon t he same medium and under
approximately the same conditions easily possible anvw here.

2. The habit, .structure, and appearance of the colony must he
given as it develops upon at least two standard media of decidedly

different composition.

3. The physiological effects of the colony upon these media should
be noted.

14 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

4. Full drawings or photographs should show hahit as well as micro-
scopic details of cells and cell relations.

5. Other morphological or physiological data obtainable should
be given as accessory information. Very striking characters are
often found under accidental or unique conditions which immedi-
ately differentiate particular species. Some of these characters come
up in the ordinary course of cultural study, others are found under
accidental conditions but could rarely occur in laboratory routine.

It is clear that such descriptions can only result from repeated cul-
ture of a species under constant observation. If the range of such
culture has been wide, it will bring out the most striking characters
and thus reduce the number of minute distinctions necessary.

NECESSITY FOR DESCRIBING CULTURE MEDIA AND TEMPERATURE

CONDITIONS.

In a previous paper 27 the characters available from cultural studies
of species of Penicillium were discussed. The series of observations
have been greatly extended in the three years intervening. These
characters may therefore be profitably reviewed, following the sum-
mary just given.

Culture medium.- — The composition of the substratum is shown in
this paper to affect the character of the colonies grown upon it to
such a degree as to make the exact description of the medium essen-
tial. Examples of these effects may be cited to show how conspicu-
ous these differences may be. In a medium free from certain sugars
P. duclauxi produces upon the surface of the medium very short coni-
diophores with conidial fructifications, whereas when such a sugar
is added numerous coremia are formed, which in well-nourished colo-
nies often become 10 mm. in height. This species also produces a
rich purple color in certain media, but not in others. Hedgcock has
noted that P. aureum Corda (as distributed by him) produces colo-
nies orange-red upon alkaline media, but lemon-yellow in acid media.
P. digitatum Sacc. grows sparingly, if at all, in media offering
nitrogen only as nitrates, whereas many other species grow equally
well from nitrates and organic nitrogen. One species is included
which produces feeble gray or brownish cultures when carbon is pre-
sented from gelatin, starch, or lactose, but becomes a clear green
when cane sugar is added.

A culture medium must offer not only the proper chemical ele-
ments for fungous growth, but must offer these in assimilable form.
Different species of Penicillium make quite different demands as to
the form of food substances required, hence a description must specify
the substratum accurately enough to enable the duplication of cul-
ture conditions. If, however, the proper substances in the proper
form are offered, changed percentages in the concentrations of these

HABIT, STRUCTURE, AND APPEARANCE. 15

substances have been found to affect the growth of cultures more
slowly. Most species grow normally in extremely dilute solution-.
hut continue to irrow w ell or even richly until the solutions reach con-
cent rat ions of considerable osmotic pressure.

Temperature. The species studied are mostlj saprophytes of wide
distribution. They are therefore able to thrive within a consider-
able range of temperature. Comparatively few of the species tested
grew normally at blood heat (37° C). At this maximum many of
them either failed to grow or were actually killed. Within the range
of IJ to :!o ('. the rapidity of development in mosl species, as
indicated by the production of colored fruit, increased with the rise
in temperature. The lower temperatures affect the rate of fruit for-
mation without, as a rule, preventing such development. Cultures
held at 10° to 20° (\ reached a development fully equal to those held
at higher temperature, only in longer periods.

HABIT, STRUCTURE, AND APPEARANCE OP COLONIES.

Many series of comparative cultures indicate that any colony of a
particular species will reproduce the same characters whenever
grown upon a particular medium under particular conditions. The
differences between many species of Paticilliuiu are so striking to the
eye as to enable one familiar with t hem to identify the several species
immediately. These same differences are, however, so difficult to
define, and in many cases so transitory, as to render their expression
in form to insure recognition very difficult. Recognition of specie-
depends at best upon a series of observations of these characters
throughout one, and usually more than one, generation upon two or
nioie substrata. A discussion of these characters follows:

Fruiting period. — In comparing cultures of related forms it is
found that the time necessary for development from the spore to the
production of ripe conidia differs for the different species, but that
periods in the different species bear a comparatively constant rela-
tion to each other when all are grown under the same conditions.
Similarly the length of the growing and fruiting period varies in the
several species. In some species the mycelium produces new coni-
diophores and masses of conidia among or overgrowing the old for a
considerable length of time. In some a secondary growth of white
or even colored hyphae often overgrows the fruiting area. In others
the chains of conidia once started continue to increase in length for
several weeks, until the conidia! fructification becomes a mass 1 mm.
or even more in length. In still others but a single crop of conidia
is developed and t he mycelium apparently dies or is totally exhausted

ill ;i few days. In some species the center of the colony unit II res and

dies while the margin continues to grow and produce new conidio-

phores for some time. The habit of the species in the production of

16 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

conidial fruit is usually a well-marked character and is often found
very useful in the separation of forms found growing together.

Color. — Color and color changes are difficult to describe on account
of the deficiencies in the standards of color, but they form the first
character noticed. The variety of greens, blue greens, gray greens,
yellow greens, and shades of brown in the genus Penicillium baffles
one seeking descriptive terms. The shades of color peculiar to each
species under oft-repeated conditions are easily recognized and are
quite reliable. The alterations in the color of spores due to changing
the composition of the medium, as shown in the recent work of Stoll24
and in this paper for species of this genus, and by others (Milburn,15
Bessey1) for other genera, emphasize again the necessity of uniformity
in and careful description of the culture medium. But in spite of
the admitted difficulties in color description, the careful observation
and record of the color of the colony at every stage of its development
is very necessary to identification. This is complicated by the
changes which occur at different ages of the colonies, so that it would
be easily possible to place certain species in at least two of the color
groups as designated by the older authors if we simply make our
observations a few days apart.

The color of the mass of mycelium as observed from below (" re-
verse" as designated by Dierckx5) in cultures gives striking con-
trasts. This must not be confused with discoloration of the sub-
stratum which may or may not be produced by the same species in a
given medium. The mycelium itself viewed in reverse has character-
istic colors in certain species which are useful in diagnosis and are
entirely independent of discolorations of the substratum. A colony
colorless itself may color the medium brightly, while a colony bright
colored itself may make no change in the color of the medium.

Surface. — As a convenient term " surface" may be used to desig-
nate the general appearance or the texture of the aerial portion of
the colony. Perhaps the word "habit" would be in some measure
more accurate, but that would apply % also to the submerged myce-
lium. Comparison of the surface appearances of many cultures
shows that this is one of the most stable characters when the same
medium and conditions are used. For species of Penicillium and
allied genera two types of surface will include most of the species
met with.

In the one type all or a large majority of the conidiophores in the
rapidly growing colony arise directly from strictly vegetative hyphse
which may be submerged in the substratum or lie upon its surface
or be alternately prostrate and submerged. Each conidiophore
stands separately, therefore, and usually all are found to be so nearly
of the same length that the surface appears to be velvety. Such a
surface may be called either velvety or "strict." A strict surface
may be called "closely strict" when the conidiophores are so short

SURFACE A.xn MARGIN 01 COLONIES. IT

that the conidial fructifications are barely raised above the surface,
or it may be loose or lax If longer conidiophores give a deeper
velvety appearance.

In the other type all or most of the conidiophores air lateral
branches of definitely aerial hyphae. These hyphae and conidiophores
form felted masses or loose networks of aerial mycelium for which
the term " floccose" is descript ive.

Margin. In seeking the origin of all structures direct observation
of the margin of the young and growing colony is essential. Such
observation determines how the fungus spreads in the substratum,
theseptatioE and measurement of hyphae, ami t he origin, order of de-
velopment, and relative positions of aerial structures. As the colony
matures growth ceases, ripe conidial areas extend to the very margin,
the masses of conidia often change color and fall apart, and the
conidia-bearing branches may curl up or drop off. Some species seem
to inhibit their own further growt h after a, short period, while in other
cases they dry up the culture medium by transpiring water. Many
things thus contribute to render the old colony an unintelligible mass
of spores and hyphae.

In colonies with surface strict or velvety (consisting of conidiophores
only I t here i- a succession of structures from the center to the periph-
ery. In the center are conidiophores with ripe conidia, marked by the
colored area. This shades into a white margin of developing conidio-
phores and conidial fructifications, while the extreme margin consists
of submerged vegetative hyphae. The relative width of these areas
and rate of the development of conidiophores and colored conidia give
characterist ic appearances to colonies of part icular species. In some
there is a broad submerged vegetative border, then a similar white
band of developing conidiophores. In others the area of colored
conidia extends so closely to the margin that the white border is
barely discernible. In the floccose species aerial mycelium often ex-
tends as rapidly at the margin as does the submerged part. In such

Cases the area of coloration follows the expansion of the colonj more

slowly.

Although close resemblances in culture are not uncommon, the
relative development of these areas i- quite typical and often sharply
distinctive of species. Colonies showing a broad submerged and white
margin usually spread over wide areas of the substratum, whereas
those bearing ripe fruit to the verj edge of the growing colony rarely
develop beyond restricted areas.

The gross characters already discussed have purely specific value,
or ma\ e\cn he more closely restricted as characters distingui hin
particular ecads of specie.-. The generic characters and \c\\ impor-
tant specific characters are microscopic, and include cell relations and
details of spore formation.
8108 — Bull. 1 I ^ in -2

18 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

Conidiophore. — The essential data as to conidiophores are their
length, septation, the diameter of their cells, and especially their origin
and relation to the substratum and to each other. Although ex-
tremes of variation in length of conidiophore may be very marked in
any culture, the majority of conidiophores in any such culture ap-
proximate an average length. This length to be most reliable must
be taken from the origin in another hypha to the lowest branch of the
fructification. If the conidial fructification were counted into the
length, the length of conidiophore would in many case's be doubled
with the maturing of the spores. The actual length, however, is little
changed with such maturity. Valid data on these points can be
secured in many species only by direct observation of the undis-
turbed colony in the air under the microscope, instead of by the study
of fluid mounts. This is equivalent to saying that Petri dishes, or
other vessels which can be uncovered for study, must be used. The
student must expect, therefore, to make many cultures and jeopardize
the purity of one such culture every time he undertakes its proper
examination.

Conidial fructification. — For lack of a better term conidial fructifica-
tion maybe defined as including the chains of conidia, the conidiiferous
cells, and the branches bearing them back to their junction with the
conidiophore proper. In a species of Penicillium fructifications vary
greatly in detail, so that satisfactory illustration becomes difficult.
Comparison of large numbers of fruiting branches in many cultures of
the several species establishes specific types of appearance which can
be shown in a conventionalized series of sketches for each species.
The data found of value have been the mode of branching, the meas-
urements of branches and conidiiferous cells, the relation of these to
each other, the arrangement of the chains of conidia with reference to
each other, and the measurement and appearance of the fructification
as a whole. These penicillate fructifications vary from close columns
of spores arising from single verticils of cells borne directly upon the
apices of the conidiophores to widely spreading "brooms" whose
numerous divergent chains arise from verticillately or complexly
arranged branches from the original conidiophores. In the study of
these conidial fructifications mounting in fluid commonly greatly dis-
arranges these complex masses of branches and spore chains. Direct
observation of the undisturbed colony is therefore essential to a cor-
rect conception of the habits of the species, though the details of
branching and spore bearing must be sought in fluid mounts.

The term " conidiiferous cell " is used for the cell at the base of every
chain of conidia in preference to "sterigma, " as often used, or to
"basidium" as used by Stoll.24 The term was proposed by Professor
Atkinson as meeting the objection that basidium implies a relation-
ship net justified in fact, while sterigma usually designates not a spore-

PHYSIOLOGICAL Kill.' I> UPON CULTURE MEDIA. 19

producing cell l>ut a spore-bearing process of a cell, hence is properly
applicable to the narrowed apices of these verj cells. In the Latin
descriptions the term "basidium" has been retained, however.

( 'onidia. — The usual data with reference to the conidia are of equal
service in our cultural studio, viz, shape, size, color, arrangement,
markings, mode of germination, and conditions of growth. A record
of the changes in color at different ages of the colony is essential.
Completeness in observation is as necessary in spore characters as
elsewhere. The variations in the size of conidia are notable in some
species : in others conidia appear to be either globose or elliptical, even
in the same chain. Where the outer cell wall is marked or spiny these
markings often do not appear until the conidia are fully mature.

PHYSIOLOGICAL EFFECTS UPON MEDIA.

Certain physiological effects of fungous growth which are incident
to the use of ordinary culture media have boon found to be reliably
characterisl IC of species. Some of these react ions are so conspicuous
a- to aid greatly in separating nearly related organisms. Such re-
actions a- have been found uniform and definite or unique are intro-
duced into the technical diagnoses of spceies in this paper. Other
physiological data are appended as accessory cultural information.

Among t he data observed in repeated series of eomparat ive culture
are the following: Odor, litmus reaction of medium at different
stages of growth, liquefaction of gelatin media, the production of
coloring substances in the media, the changes produced in milk, and
the production of drops of transpired fluid upon surface of colony.
Among the accessory data, observations upon carbon assimilation,
upon proteolytic reactions, and upon the production of enzymes have
been made for certain species.

Odor. — The production of definite odors by colonies is confined to a
small number of species, and even among these often to particular
media. When definitely present it is a character immediately recog-
nizable and in certain species diagnostic; in others it associates the
organism at once wit h a part icular group of species.

Litmus. The use of an indicator in the medium gives in very
many species a sharp reaction. The value of this react ion is more nar-
rowly restricted, however, than is indicated in previous papers
(Thorn25 :>. The in t roil net ion of sterilized litmus or azolitmin into
complex media brings contradictory results when the composition of
the medium is slightlj altered, [f, for example, a solution of puregela-
tin in distilled water be used as a nutrient .the reactions are definitely
alkaline, with very few exceptions, which are just as definitely acid.
The gelatin solution itself is acid. The alkaline reaction indicates
that t he products of t he digest ion of gelatin in such cases are of alka-
line nature. If. however, the gelatin solution be neutralized and

20 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

5 per cent cane sugar be added, an equally large majority of the same
species cause an acid reaction. Some few remain alkaline. These
species growing upon cane sugar produce acids in quantity both to
neutralize the alkaline products of the decomposition of gelatin and
to change the reaction of the mass. The litmus reaction has been
found generally reliable in a medium composed of 15 per cent gelatin
in distilled water, in lactose-peptone gelatin after Conn's4 formula,
and in potato agar during four years of cultural work where many
successive lots of media have involved the use of materials from
different sources. In complex media the decomposition products of
organic nitrogenous constituents and those of carbohydrates tend
either to neutralize each other or to intensify the reactions, but add
greatlj' to the difficulties of analyzing the results. The litmus
reaction therefore may be used with the simplest organic media, but
is best applicable to cultures in synthetic media where analysis of the
results is feasible.

Gelatin. — The liquefaction of gelatin media by an organism in cul-
ture shows its ability to produce a particular form of proteolysis.
This reaction is so conspicuous and so adaptable to cultural use that
it has been recorded in all cultural studies. Various investigators
have pointed out the limitations of this reaction. In cultivation
nearly all of the species of Penicillium have been found to grow well
upon a 15 per cent solution of gelatin alone in distilled water. Com-
parison of the results in this medium with liquefaction of several forms
of gelatin media experimented with showed essential agreement.
Since the advantages all lie in the simplification of formulae, the data
as to liquefaction by these species have been compiled from series of
parallel cultures of all the species upon 15 per cent gelatin in dis-
tilled water.

The value of this reaction is measurably vitiated by the fact that
any species which can subsist upon gelatin alone must be capable of
more or less proteolytic action upon it. The results of observation
confirm this statement. The value of these observations therefore
depends upon the indication of the comparative rate of activity of
different species in inducing proteolysis, not upon the presence or
entire absence of this action. Numerous tests, involving a range from
15° to 25° C, show that slight changes of temperature affect this
reaction only to the extent to which they affect the rate of growth,
but do not disturb the comparative value of the data obtained.
Within this range of temperature the most active species produce
liquefaction in 5 to 8 days, other vigorous liquefiers require 8 to 15
days, while many species producing no liquid or but traces of soften-
ing in 15 days produce a gradual liquefaction in the succeeding 2 to 4
weeks. In studying species of Penicillium, liquefaction of gelatin has
been found to separate such species as produce this proteolysis within

PHYSIOLOGICAL EFFECTS UPON CULTURE MEDIA. 21

the time needed for other observat ions upon these cultures about 15
days. Where liquefaction occurs during the active growth of the
colony it definitely indicates the secretion of ectoenzyms capable of
tins digestion. Long-deferred liquefaction may result from such
secretion or from the liberation of endoenzyms by the disorganizat ion
of mycelium.

Color production. Certain species cause marked changes in color
in some of the substrata in which they grow. Such reactions are
usually produced upon certain media and qoI on others. They are
therefore selective reactions. For example, one species produces a
bright yellow color in media containing milk sugar, such as milk or
peptone-milk sugar-gelatin; but no color in plain potato agar. Some
<>\' these colors are soluble in alcohol to form brightly colored solu-
tions. The chemical nature of these substances has not been studied.
but the reactions themselves have been thoroughly tested for a few
species. In these species color production uniformly follows the
proper culture condit ions.

Production of fluid upon surfaci of colony. — The presence upon the
surface of the colony of Large drops of transpiration water which may
be colorless or brightly colored by excreted products is common.
Although a transient character dependent to some measure upon
the humidity of the air in the culture vessel for its prominence, there
i^ much difference between species in this particular.

Technical descriptions of these fungi have always been based.
theoretically, upon morphology only. In such descriptions the
host or substratum has been more or less definitely indicated. Where
the organisms are parasites or saprophytes closely restricted to par-
ticular substrata and conditions, this practice can perhaps be justly
supported. With omnivorous cosmopolitan saprophytes cultural
study quickly shows morphology to be affected greatly by changed
conditions or altered composition of media. Further, very many
of these orga.ni.sms have no known special habitat. A description
based upon a specimen of one of these species found in the held,
without comparative culture, might possibly contain some peculiar
character which would identify a specimen of the same species when
next found, but this is only a possibility. In practice the descrip-
tions we have are nearly useless.

The introduction of cultural study carries with it the necessity of
recognizing at least the most striking physiological data. With
these species of PeniciUium such data have been found as reliable as
morphology. Further, the careful definition of morphology and
reactions upon specified substrata under known conditions is only
amplifying and rendering definite the two items habitat and local-
ity- which have always been included in plant descriptions. In
recent years the students of ecology and of experimental evolution

22 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

have .shown these to be much more important in flowering plants
than was formerly supposed. The introduction of these characters
is thus in full conformity with the best systematic practices of
to-day, where the life history of the species is investigated as fully as
possible.

CULTURE MEDIA.

In developing the descriptions presented the following media have
been used:

1. Potato agar. This was made as described in another paper25
(p. 7). Essentially this is simply a potato infusion to which agar-
agar is added in any proportion desired. Since this medium is free
from sugar and very dilute in nutrient value, sugar may be added to
cultivate certain species.

2. Bean agar. The directions for making bean decoction were
obtained from Maze at the Pasteur Institute in Paris. Common
white beans are heated in five volumes of water. Boiling is stopped
just before the swelling of the cotyledons would rupture the seed
coats. This gives a clear, slightly yellowish liquid which filters
readily yet contains sufficient nutrients to grow many species nor-
mally. Agar may be added as desired. Since this decoction is
poor in available carbon, the addition of sugar is often desirable for
many species.

3. Peptone-milk sugar-gelatin, as described by Conn.4 This stand-
ard bacteriological formula, with and without litmus, wras used in
developing the original draft of these descriptions.

4. Fifteen per cent gelatin in distilled water. As a stock medium,
this wTas used without neutralization. Comparative cultures neu-
tralized with NaOH seemed less adapted for most species than un-
changed gelatin. With few exceptions the common species of
Penicillium grow readily in plain gelatin and give the same reaction
as in Conn's more complex formula.

5. Synthetic media. A. W. Dox, chemist to this investigation,
has modified Czapek's formula for a synthetic medium, intended to
present in a nearly neutral solution unaffected by sterilization the
elements necessary for fungous growth. In stock form neither nitro-
gen nor carbon is presented in this fluid. It has consequently been
found an excellent means of testing the availability of these elements
in various forms. For convenience this will be referred to as Dox's
solution, the formula of which is —

Distilled water centimeters. . 3, 000. 0

Magnesium sulphate grams. . 1. 5

Dipotassium phosphate (K2HP04) do 3.0

Potassium chlorid do 1.5

Ferrous sulphate do .03

Nn.MI N< I.ATCKI.. 23

In the work here reported nitrogen \\ ras added as sodium nitrate, 6
grams. Parallel experiments in which monopotassium phosphate
(KH.I'Oy. which gives a perfectly clear solution of a strongly acid
reaction, was substituted for dipotassium phosphate, K.I I P( ),, gave
no advantages in cult n re to offset the advantages of a neutral medium
except the disappearance of the traces of precipitated magnesium
phosphate. The availability of carbon in any organic form can be
tested readily in this solution. Solidified media are obtained by the
addition of agar.

NOMENCLATURE.
THE GENERIC NAME.

The generic name, PenidUium Link, is held -in this paper in its
hyphomycete sense to designate all species which continue to propa-
gate themselves for an indefinite number of generations by penicil-
late asexual fructifications. Such grouping does not imply the
author's belief in the phylogenetic relationship of all such forms.
The penicillate type of fructification is a definite character which
binds together in this way into a "form-genus" a large number of cos-
mopolitan and omnivorous saprophytes, very few of which are
known to produce sexual fructifications. Within this heterogeneous
group, several series of forms possessing particular groups of char-
acters have been separated and generic names have been based by
some workers upon such segregation. Wehmer28 founded the ^enus
Citromyces upon two such species causing citric acid fermentation in
sugar media, with its morphological basis in the presence of a single
whorl of conidia-bearing cells at the apex of the conidiophore. Fur-
ther .study slows that the presence of this character alone would
group together forms not so closely related to each other as to other

species of PeniciUium lacking this character. It seems best, there-
fore, to use the name l\ iiirillium to designate the entire group and
leave further investigation to establish permanent genera when
really genetic relationships shall have been discovered.

NOMENCLATURE OF SPECIES.

In considering the actual problem of nomenclature of species
several positions may he taken. It is not difficult to find published
descriptions of single species in this genus which are sufficiently
indefinite to include a large percentage of all the know n species. A- ;i
rule, the morphological character-, of the various forms under culti-
vation would not exclude them from half a dozen of the older
species so far as current descriptions go. hi many cases there would
not, however, he the leas! reason for the adoption of one name in
preference to another. Shall the investigator adopt for his material

24 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

a name previously used when he has little or no reason to believe that
he has the organism originally described under that name ? Granting
the apparent impossibility of contradiction, he might find it a safe
practice. If, however, contradiction should arise, the position
becomes entirely indefensible. In case there- is a fair probability that
the forms are identical, the use of the old name may be justified per-
haps without direct proofs. The alternative position calls for the
complete description of the form studied and its distribution in cul-
ture to different centers of cultural work under a new specific name
associated with this definite material and description. It can not be
contended that these organisms are new to science, for it is entirely
possible that certain of them have formed the basis for already pub-
lished descriptions. This course can be justified by the contention
that such publication applies to definite material, available for
examination, culture, and comparison by others, and that there is less
probability of confusion from such publication than would ensue
from the use of names long published, without any real evidence of
the identity of the organisms with those originally studied. After
careful examination of all material available and consultation with
many workers in closely similar fields, new names are attached to
such forms as by continued cultural study appear to be sharply
marked species but not identifiable by older descriptions.

The name P. glaucum is not used. Careful examination of literature
and of cultural material from several sources, together with confer-
ences and correspondence with investigators in numerous laboratories,
does not afford evidence as to what form was originally used by Link
or even secondarily by Brefeld2 under this name. The name as used
at present seems to be applied collectively to the common green
forms which under examination are quickly found to be not one but
several species. Further study may give some indication as to
where the name really belongs, but until that time there is little
profit in applying it to any particular form. It might, perhaps, be
withheld until some worker succeeds in repeating Brefeld's classic
studies in ascus production, and then applied to the form so found.

The present paper is not intended to be a monograph of the genus.
It is presented as a report covering several thousands of cultures
of a group of common forms, in the hope that the descriptions and
key offered may be useful to others. The author has included
studies of as many authenticated cultures of the more recently
described forms as it was possible to secure. The verified cultures
secured have already been listed. Much assistance and advice were
freely given by Dr. C. Wehmer. On the other hand, it was impossi-
ble to secure cultures of the species recently described by Oude-
mans,17 or those listed by Dierckx;5 and one recently described by
Peck18 also appears to be lost. No claim to completeness can be

THE TYPE SPECIES. 25

made, though the material may possibly form the oucleus of a real
monograph later.

In the examination of the early literature of this genus the author
is greatly indebted to l>r. C. I.. Shear, of the Bureau of Plan! Indus-
try of this Department, for the use of hooks and for much careful
cooperation in examining and discussing the subject of types and
descriptions.

THE TYPE SPECIES.

Link'-1 in his "Observationes," published in 1809, established the
genus Penicittium to include the common green molds having a
penicillate type of conidial fructification- a conidiophore branching
more or [ess complexly at its apex, such branches becoming or being
tipped by cells, each of which produces a chain of conidia. The w hole
produces a brush-like appearance, the chains of conidia serving as the
hairs or bristles of the brush. This generic name has been universally
accepted t<> include in a form genus all species reproducing themselves
indefinitely by such conidial fructifications. However doubtful we
may be as to the forms originally examined, there is no question that
we know the general type of structure which Link intended in his
description of the genus.

Under the genus PancUlbuii, Link placed three species. The first
specie- listed was P. glaucum, but the description given is equally
applicable to many different forms. This was noted as frequent in
decaying bodies and said to be most closely related to P. expansum
(the third species listed), of which he suggests his material may have
been but undeveloped specimens.

The second species, P. cand'uhuit, is described as producing round
colonies, with mycelium and spores white, upon decaying fungi and
herbs. Although many authors have used this name for material
from differenl sources, and Morini16 has described an ascigerous form
under this name, there has been no means of determining what form
I ink had in mind u hen writing his description. Many forms will pro-
duce white mycelium and spores under special conditions, while but
• me of those examined has been shown to do this under all conditions,
and this one is so specialized in its habit as to be excluded by Link's
statement of habitat from the application of this name. Numerous
authors have suggested that the /'. "candidum" forms are probably
colonies of species, colorless under special condil ions, but green under
other cultural conditions.

The third species, P. expansum, is slightly better described, while
it- habitat is primarily given as rotten fruit, although the author has
manifestly extended his use of the name to forms growing upon other
substrata which he believed to be identical with the fungi grown upon
decaying fruit.

26 CULTURAL STUDIES OF SPECIES OF PEXICILLTUM.

On page J!) <>f his " Observationes " Link describes the genus
Coremium with a single species, C glaucum, which he specifies as found

upon decaying fruits. This fungus can be traced through a con-
nected series of publications giving descriptions and figures which
show the original conception to include (if not entirely to be drawn
from) the large green coremia which develop upon apples and related
fruits decaying in storage.

This organism is figured by Greville8 as Floccaria glauca; it is cited
cited by Fries7 as a variety of Penicillium crustaceum, from which he
has "seen it originate upon apples in the autumn." It is twice
referred to by Corda in Icones Fungorum (Vol. II, p. 17); he cites
Coremium glaucum, C. cUrimum, and 0. candidum as synonyms;
again in Prachtflora (p. 54, taf. XXV, figs. 3, 4, 17, 18, 19, 20, and 21)
under the name of Coremium vulgare he manifestly had this same spe-
cies, although he groups it with figures which appear to be different
organisms.

These figures and descriptions cited are definite enough to show
that workers contemporary with Link applied the name Coremium,
glaucum Link to the coremiform rot of the apple. As a result of
observations of living material Fries considered this only a form
of Penicillium crustaceum, which he made to include P. glaucum and
P. expansum Link.

The present writer has collected this fungus upon decaying apples
and related fruits repeatedly in America; also upon pears and
mespilus in Hanover/ Germany.

Repeated cultures have shown that the ability to produce coremia
is a definite character of this species, recognizable under many con-
ditions of culture, but not shown under other conditions. The same
culture will commonly show both simple penicillate fructifications
and coremium production. The species must therefore be regarded
as one of the several species of Penicillium which always produce
coremia under proper cultural conditions.

Examining Link's species of Penicillium , we find that he specifies
P. expansum as primarily found upon rotten fruit. P. expansum
Link clearly included Coremium glaucum Link, therefore, probably
with others; but from its known abundance in Germany there can
be little question as to this organism forming in part, at least, Link's
original conception of this species. Later (1824) in Species Plan-
tarum, Tomus VI, page 70, Link11 redescribes P. glaucum and includes
in it the P. expansum of his Observationes. In this discussion Link
broadens his description of P. glaucum to include all green forms
found in decaying substances, upon the assumption that all such
forms are but a single species. It is evident that the earlier descrip-
tion P. expansum Link included this species with sufficient restriction

PENICILLIUM EXPANSUM. 27

to justify reviving the name /'. expansum Link and limiting it to the
penicillium i<>t <>l' apples alone.

/'. expansum Link (in part), the penicillium rol of apples, would
therefore stand as the type species of the genus.

The I cchnical rules for the establishment of type species arc in this
way satisfied. The kind of plant usa\ by Link is perfectly well
known. To sav jusl which forms he had in hand may be impossible,
hut the form we arc discussing was certainly one of them.

EXPLANATION OF DRAWINGS.

The drawings of the species described have been made with the
Bausch & Loml) camera lucid a in all cases except such as are marked
diagrammatic or partly diagrammatic.

For those with the magnification of 1 10, the lenses \i^h\ were the
Bausch & Loml) 1-inch ocular and the two-thirds objective; for those
of 900 the lenses were the Bausch & Lomb 1-inch ocular and the one-
eighth objective; for those of 1,400 the lenses were the Bausch X
Loinh 1-inch ocular and the one-twelfth objective: for those of 1,600
the Spencer No. 12 compensating ocular and the Zeiss 3 mm. apert.
L30 apochromatic. This apochromatic was also used in a few cases
at a magnification of 900; these are indicated in the legends. All
figures at magnification of 140 were drawn from the exposed surface
of the undisturbed colony in Petri-dish cultures. All other figures
were made from fluid mounts or hanging-drop cultures (for spore
germinations).

The series of sketches at 140 magnification are made with uniform
methods, so that comparison of species in culture can be much easier
than from figures made at different magnifications.

PENICILLIUM EXPANSUM Link.

P. expansum Link i in pari i, emended Thom=penicillium rol of apples and allied fruits.

Syn. Coremium glaucum Link. Observations, p, L9; [cones, V, 0i^. 31, 1809.
Floccaria glauca Greville, Scottish Flora, pi. 301, 6gs. 1—1.
Penicillium glaucum Link (in part), Species Plantarum, VI (1824), p. 70.
Coremium vulgart Corda (in part), Prachtflora, p. 54, PI. XXV, especially

figs. :;. I. 17. L8, L9, 20, and 21.
Possibly /'. thuiijiiiiiin Dierckx.

Colonies upon gelatin and potato or bean agar, green becoming gray-green and
slowly brown in several weeks (especially when exposed tolighl I, fioccose, with concen-
tric zones tufted with short, louse, coreni i uni-like aggregations of conidiophores, not
over! 2 mm. in height except in old cultures containing sugar, broadlj spreading with
broad white margin in growing colonies. Reverse somewhal brown. Conidiophores
either very shorj lateral branches of aerial byphse or verylongi I ami. or more), aris-
ing singly or grouped with others to form coremia. Conidial fructifications consist
of 1 to :; main branches bearing verticils ofbranchlets supporting crowded whorls
of conidiiferous cells, 130-200 by 50 <><».*' at base in cultures without sugar, with sugar

28

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

continuing for some weeks to produce great numbers of conidia which come to form
masses perhaps 1 mm. in thickness. Conidiiferous cells 8-10 by 2-3/f. Conidia
elliptical to globose 2 by 3.3/t or 3-3.4/c, green, homogeneous, persisting in chains
when mounted. Colonies begin to liquefy gelatin slowly after about 10 days and
continue until it is completely liquefied. Grows readily and rapidly upon all com-
mon media.

Occurs characteristically upon decaying apples-and other pomaceous fruits, where
old colonies often produce coremia 1 cm. or more in length and very large.

Collected at Ithaca and Geneva (Eustace), N. Y., at Middletown and Storrs, Conn.,
upon apples; upon pears and Mespilus at Hanover, Germany. Often appears as a
contamination in fungous cultures.

Fig. l.—Penicillium expansum Link: a, b,f, branching and arrangement of branches of conidial fructifi-
cation (X 900); c, d, e, conidiiferous cells and conidial chains (X 900); g, ft, j, k, I, sketches of fructifica-
tion (X 140); m, n, o, germination of conidia (x 900); r, s, sketches from photomicrographs, showing
in s loose aggregations of conidiophores beginning to develop in zones which become coremia like r
(coremium r was 1 mm. in height; X 35).

Many references in the literature to P. glaucum Link and P. crusta-
ceum (L.) Fries refer to this species. Unless, however, such citations
directly refer to the presence of coremia, or to the association of the
organism with the decay of pomaceous fruits, or both, there is no
means of fixing the application of such names to this species. Since
Link11 in 1824 lumped into his specie's P. glaucum every kind of green

PENICILLIUM ITALICUM. 29

penicillium for which he could find references, the myth of " i he
common green mold" seems to have had pretty general acceptance,
although we find here and there n protest against this view.

A cult are of this species can always be obtained from apples decay-
ing in storage, upon which usually well-developed coremia can be
found if proper search is made. The \\ ide distribution of t he organism
as noted above and as seen in the literature justifies belief in its
genera] distribution. Once carefully observed in cultures, the in-
vestigator will usually recognize the organism on sight when it ap-
pears in his cultures even as a contamination of other species. The
odor of this species i- so distinctive as to assist greatly in identifying

it in accidental cultures. It does not produce the yellow color in

the substratum as described by Lindau "' for P. glaucum, nor are its
spores globose from the first as Wehmer:!0 records them for that
species. It seems so very well characterized as to justify sharply
separating it from the other green forms.

I ULTUBAL DATA.

' olor, green or gray-green; color of reverse, yellowish to somewhal brown; color
in media, colorless or yellowish brown (milk).

< >dor, "fruity" in all or nearly all cultures.

Fifteen per cenl gelatin in water, good growth, with loose and ill-defined bu1 abun-
dant coremia; liquefaction slow, more or less liquid in 2 weeks; litmus reaction alka-
line or neutral. Potato agar and bean agar, characteristic colonies, gray-green with
more or less coremiform bundles of conidiophores. Potato plugs, characteristic

colony.

Kaolin's fluid agar, characteristic colony with concentric rings of broad coremia.
Kaolin's fluid, characteristic colony. Cohn's solution, slight growth, few coremia,
brownish below.

Synthetic fluid il>ox'si, carbon supplied as: Cane sugar, good coloring up to 50
per cent sugar, acid reaction, conidiophores in dense areas with few definite coremia.
Lactose '■'• per cent, slow development, about half normal. Levulose 'A per cent,

typical, alkaline, or neutral reaction. Galactose '■'< per cent, typical, acid reaction.

Glycerin '■'> per cent, slow development, increased greatly by adding sugar. Potato
starch '■'> per cent, fair growth, no coremia. Butterfat, rich typical growth.

Milk, typical colonies, coremia in a ring at glass; curdling (0.25 per cent calcium
chlorid added) in 1 week; digestion rather slow; color in milk yellowish brown.

At 37° C, do growth, culture grew when cooled ; at 20° < '.. good growth.

The coremiaof this species are especially characteristic <<i old cultures in which
drying has begun. In fluid cultures they are commonly attached to (he jjlass above
or at the very top of the fluid.

PENICILLIUM ITALICUM Wehmer.

Beitr. z. Kennt, einh. Pilze, Jena, 1895, p. 68, til.

Colonies on plain gelatin and potato or I nan agar bluish green, becoming graj -green
w heii old i bright bluish green on cane-sugar media and citrus fruit- 1, broadly spreading,
aerial port ion composed at t lie broad margin almost entirely of conidiophores, hut be-
coming slightly floccosein the center Reverse of colonies dark brownish, often almost

Mack in i lia containing .-iiLrur. ( lonidiophores from short i LOO/i to ^ erj lon^ ((iOO/t),

averaging perhaps 'Jot)", arising either directly from substratum or as branches of
aerial hyphse. I kmidial fructifications up to 300ft or more in lengt h, consisting usually

30

CULTURAL STUDIES OF SPECIES <JE PEKICILLIUM.

of a main branch and one lateral branch, each producing a whorl of branchlets bearing
crowded verticils of conidiiferous cells, 12-14 by 3^. Conidia breaking off in masses in
handling old cultures, which rise in clouds when shaken. Pronounced odor in cul-
tures containing cane sugar. Chains of conidia loosely divergent, long; conidia
2-3 by 3-5/t, cylindrical to elliptical or slightly ovate, clear green by transmitted
light, very variable in size but usually within the limits given. Masses of spores con-
tinue to increase from 2 to 3 weeks. Petri-dish colonies partially and slowly liquefy
gelatin (12 to 20 days). Numerous white sclerotia are produced upon the surface of

Fig. 2.—Ptnuittium italicum Wehmer: a, b, c, d, c, f, g, types of branching, formation of verticils of
conidiiferous cells and conidial chains (a, e, /, X 900; 6, c, d, g, X 1,400); j, k, sketches of conidial fructi-
fications (X 140); I, m, n, swelling and germination of conidia (X 900).

the medium after 2 to 3 weeks' growth, especially upon fruits and other acid media
rich in sugar.

Cosmopolitan; characteristic of decaying oranges, which become bright blue-green
with this mold as contrasted with the olive-green species which is often associated
with it upon the same fruit. These contrasting colors are illustrated by Wehmer.31

l'KMi II.LIC.M DIGITATUM. 31

In bis discussion of this fungus Wehmer S1 shows thai the blue-green
i -. * t of citrus fruits is a differenl species from the similarly colored
apple rot. His description seems to l>e the first recognition <>l this
species as differenl from the green molds occurring constantly upon
all kinds of food. This species has been discussed as P.glaueum in
recent papers i K. E. Smith,2' Powell20), \\ here its agency in the decay
of oil ins fruits is \ei\ fully considered.

Cultures were obtained by the writer from oranges in Hanover,
Germany, and identified by the describer, Dr. C. Wehmer. It has
since been repeatedly observed and collected in America. Pure cul-
tures <-an always l>e secured by finding decaying oranges in the mar-
ket which have the blue-green areas of rol just beginning to appear
upon them. These areas are usually blue-green in center surrounded
by white areas which are usually grouped into little white patche
toward the vegetative margin and the whole superficial colony sur-
rounded by an area of soft, watery rot. Xrry often such colonies
when older become much contaminated with the olive-colored rot,
given in this paper as P. digitatum.

CULTURAL DATA.

Color, dear bluish-green mi sugar media, shades of gray-greeD without sugar; reverse
of colony commonly brownish in areas; color in media none or slighl .

Oder distinct upon media containing cane Bugar, uone on lactose or media free from
sugar.

Fifteen percent gelatin in water, medium growth; Liquefaction, none until several
weeks old, then partial in acidified cultures; litmus reaction fairly alkaline. Potato
agar « Lth lactose, rather thin gray-green colonies, not vigorous, acid reaction, retain
plugs, pale but characteristic. Raulin's fluid, typical colonies. Cohn's solution,
germination onlj .

Synthetic fluid I 1 >e\ V , carbon supplied as: < lane sugar, good colonies up to 50 pei
■ ■en i wiih alkaline reaction. 1. act use :; per cent, slew growth bu1 typical, with acid
reaction. Lactic acid 0 9 per cent, small growth. Levulose ■'> per cent, not normal.
Galactose 3 per cent, medium growth, acid reaction. Glycerin 3 per ecu i . no growth;
growth began when sugar was added. Butterfal , growth slow.

Milk, good growth: curdling (0.25 per cenl calcium chlorid added ), ten days; diges-
tion partial and -low; color bIowIj brownish <>r yellow-brown.

At 37°C, killed; at 20° C, good growth.

PENICILLIUM DIGITATUM Saccardo.

The olive-green orange rot, /'. digitatum Sacc, in Mycotheca Italica, No. 986, Ber-
barium U.S. Department of Agriculture; in Sylloge Fungorum, Vol. [V, p. 79; in
Fungi [talici, No. 894.

= /'. olivaa urn Wehmer, Beitr. z. Kennt. einheim. Pilze, pp. 73, t. [I, Jena, I
"iMucor exspitosus 1... in Species Plantarum (1753), 11, p. list;, based upon

Micheli, tab. !U, lig. :'..
7 Manilla </i</ii(ihi Fries, Sj sterna Mycologicum, 1 1 1 , p. ill.

Colonies on .-agar gelatin ami potato or bean agar grayish olive, irregularly shaped

from the unequal growl h and branching of rather few hypha . aerial port ion consisting

only of very short conidiophores and conidia. Reverse of colony commonlj shows
brown to black colors. Conidiophores rising directrj from the substratum, 30 100 by

32 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

4-5/t, usually very short. Conidial fructification a few tangled conidial chains up to
160/t in length, borne upon conidiiferous cells 13-16 by S-4pt. Conidia cylindrical to
almost globose, 4-7 by 6-8/t (at times 6 by lO/i), often uneven in size and shape in the
same chain. Colonies do not liquefy sugar gelatin except at times partially in cultures
three weeks old or more. Litmus reaction acid. Grows readily on organic media, but
shows a very pronounced affinity for such media with high percentages of sugar, in
which it produces a strong odor. Refused to grow in synthetic media containing nitro-
gen as sodium nitrate.

Cosmopolitan upon citrus fruits, distinguished from P. italicum by the sharp con-
trast of its olive color with the blue of the other. Collected in Hanover and verified
by Dr. C. Wehmer. Received from Prof. P. H. Rolfs in Florida. Seen upon decaying
oranges everywhere. Pure cultures can always be secured from the common market
fruits.

Nomenclature. — Wehmer31 (1895) gives the first adequate discus-
sion of the decay of citrus fruits by the agency of species of Penicil-
lium in which the forms found were shown to be distinct species
associated constantly with these fruits instead of common green
species accidentally occurring upon these fruits. For the olive-
green form the name proposed, P. olivaceum, is descriptive, but had
already been used by Corda (Icones, III, p. 12, t. II, fig. 35) for
a species afterwards transferred to Ilormodendrum. The name
P. olivaceum Wehmer is therefore not tenable by present rules of
nomenclature.

P. digitatum is the name used by Saceardo, as is shown b}^ the
specimens distributed by him (Mycotheca Italica, No. 986) previous
to Wehmer 's work. In his description and synonymy he cites the
name from Fries (Systema Mycologicum, p. 411), where it appears as
Monilia digitata, transferred by Saceardo to Penicillium. But Fries
cites his use of the name from Persoon l0 (Synopsis Fungorum, p. 693),
who bases his description of Monilia digitata upon Micheli's 14 figure
and description (Nova Plantarum Genera, p. 213, pi. 91, fig. 3).
Here, under the name of Aspergillus, Micheli figures and describes as
No. 8 a fungus which may be taken for a Penicillium, which is said to
have been found upon semi-putrid lemons. This figure is cited by
Linnaeus in Species Plantarum (1753), II, p. 1656, as the basis of his
Mucor csespitosus.

If we accept Saccardo's citations as correctly giving the origin of
the name and tracing it back to Micheli, we must abandon the name
he uses and adopt the name given by Linnaeus — Mucor csespitosus =
P. csespitosum (L.). Careful scrutiny of Micheli's figure gives no
possible means of identification. Aside from identification from pre-
served specimens which are not cited by the authors in any case,
there is little possibility of showing what any of the authors com-
monly cited, for this species may have had until we come to the
description given by Saceardo in the Sylloge (IV, p. 79), and the
material distributed by him in Mycotheca Italica, which I have seen
and which is certainly this fungus. I have therefore continued the
use of Saccardo's name in this case because he has given the first

PEXICILLIUM DIGITATIM.

33

description based upon tangible material, but have rejected his
citations. The following are the principal citations which have hen
given a- referring to this species:

Mieheli, Genera Plantarum i L729), pi. 91, 6g. 3.

Linnaeus, Species Plantarum (1753), II. p. L656.

Per.-mHi, ( (lisorvations, p. -II.

Persoon, Synopsis Fungorum, p. 693, Monilia digitata.

Fries, Systema Mycologicum (1829), 111. p. 411, Monilia digitata.

Saccardo, Fungi Italici i L881 i, No. 894, and Sylloge, IV. p. 79, /'. digitatum.

Wehmer, beitr. z. Kennt. einheim. Pilze, p. 73, taf. II.

Fig. :>.. Penicillium digitatum Saccardo: ". whole conidlophore ami fructification; '».<-. <l, < . types of branch-
tag and formation of conidia; m,n,o, germination of conidia; all •

The only reasoD for retaining the name P. digitatum and ascribing

it- authorship to Saccardo is that the description cited is the first one
which we can ascribe definitely to material of this species. If, as
-aid above, his citation of literature proves to he correct, the name

he gives becomes untenable by present rules of nomenclature, and
must be changed to P. exspitosum as given by Linnaeus, although

there is no evidence that Linnaeua actually examined material of this
species. Since there is more or less doubt about usages previous to
Saccardo. and since the name given by Saccardo has become asso-
8108— Bull. 118—10 3

34 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

ciated with studies of this species in recent literature (Powell,20
Smith 22), this name is allowed to stand, at least until we have more
information.

CULTURAL DATA.

Color, shades of olive green; reverse brown or dark brown, especially in sugar media;
color in media, none or slightly yellowish.

Odor, associated with smell of rotting oranges, strongest upon cane-sugar media.

Fifteen per cent gelatin in water, weak growth, not adapted for this species; lique-
faction none, slowly accomplished in gelatin to which sugar is added; litmus reac-
tion acid. Potato agar and bean agar, colonies as described above, rather poor growth
which becomes enormously increased upon the addition of cane sugar. Potato plugs,
excellent growth. Raulin's fluid, colony colorless. Cohn's solution, germination
only.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, germinated only. Lac-
tose 3 per cent, germinated but no growth. Lactic acid, 0.9 per cent, germinated
only. Levulose 3 per cent, germination only. Galactose 3 per cent, slight growth.
Glycerin 3 per cent, germination. Potato starch 3 per cent, slight or no growth. But-
terfat, germination only. Conidia inoculated into this fluid with various sources of
carbon germinated but never developed normal colonies.

Milk. — This species never produced vigorous growth upon milk.

At 37° C, killed; check at 20° C. grew well.

PENICILLIUM ROQUEFORTI Thorn.

U. S. Department of Agriculture, Bureau of Animal Industry, Bui. 82, pp. 35-36,

fig. 2, 1906.
Report Storrs Agricultural Experiment Station, 1905, p. 111.

Syn. P. glaucum, various authors — not Link or Brefeld.
Colonies on potato agar or lactose gelatin quickly turning green, becoming a dirty
brown when old, velvety strict, indeterminately spreading by large main radiating,
branching hyphse, giving a somewhat uneven or indefinite margin, which gets a white,
fibrous, almost spider-web appearance from its alternation of submerged parts of
hyphse with short prostrate aerial loops. Reverse of colony yellowish white.
Conidiophores arising separately and in acropetal succession from the growing parts
of submerged hypha? (comparatively few aerial parts, but some), 200-300/i septate.
Fructification 90-120// or at times 160 by 30-60/z at broadest place, usually appear-
ing double by the divergence of the lowest branch; branchlets ("basidiophores")
irregularly verticillate, bearing crowded verticils of appressed conidiiferous cells
(basidia), 9-11 by 2.5/z with long, divergent chains of conidia. Conidia bluish green,
cylindrical to globose, smooth, rather firm walled, 4-5/* in diameter, germinating by
a straight tube. Colonies do not liquefy sugar-gelatin, though they soften it some-
what. Fungus on plain gelatin or potato agar changes litmus from red to blue very
rapidly and strongly almost from the beginning of the growth. Fruiting period short,
but one crop of spores upon the mycelium. Cosmopolitan and omnivorous, or nearly
so. Characteristic of Roquefort and related types of cheese.

The mold of Roquefort and related types of cheese has been com-
monly designated in dahy literature as P. glaucum Link. Such
citations refer to this fungus, though many times referring to it as
"the common green mold." Although it is not restricted in its
habitat to cheese, this species is so identified with the ripening process
of Roquefort cheese (in which pure cultures are used) that any one

PEXICILLIUM ROQUEFORTI.

35

desiring a culture of this species can always obtain it by purchasing
cheese of this type. It also occurs frequently in ensilage and is
often found as a contamination in laboratory cultures of other fungi.
The species is well marked by cultural characters from which it can he
readily recognized when once studied. Its agency in the ripening
of Roquefort, Gorgonzola, and Stilton cheeses is discussed in another
paper (Thorn85).

CULTURAL DATA.

Color, gray-green to clear green, becoming brownish when old if exposed; reverse
colorless or cream; color in media, none, or in some cases a Blight and evanescent
greenish 1 ingi

I >dor, none | unless accountable for some odor in Roquefort cheese).

WW lillW

j

Fig. i. Penicillium rnt/nttnrti Thorn: a, jciri of conidiophore and of base of fructification, highly mag-
Ing production of basidia on sides as well as a< apex of basidiophore; 6, c, other types of
branching; •/. young conidiophore jus! branching; <, /. basidia and the formation of conidia, highly
magnified; <j, U,j. diagrams of types of fructification as seen under low power i < 80); /.-. /, m, u, germina-
tion of conidia ana new conidia produced directly on the first hyphse. I From Bulletin 82, Bureau of
Animal Industry.)

Fifteen per cenl gelatin in water, good growth, n<>t heavy; liquefaction, none, or
partial after '1 to :'. weeke in acidified cult ures; lii mus reaction, alkaline. Potato agar
ami bean agar, gray-green, loose, becoming dense and deep green when sugar is added.
Potato plugs, characteristic. Raulin's fluid, \ ery dense, deep green colonies. Cohn'e
solution, Blighl growth.

Synthetic fluid i Dox's), carbon supplied a-: Cane sugar, Blow growing bul typical
colonies up to -'in per cent, persistently lti^h, not acid. Lactose '■'< per cent, weak

" Weidemann ' records the production of a red color in cane-sugar solution by this
species. The writer has nol obtained this color in his culture

36 CULTUEAL STUDIES OF SPECIES OF PENICILLIUM.

growth, evidently carbon deficiency. Lactic acid 0.9 per cent, better than lactose,
not typical colony, good color. Levulose 3 per cent, weak colonies. Galactose 3 per
cent, good growth, alkaline. Glycerin 3 per cent, weak growth. Alcohol, good
growth, becoming brown in age. Tartaric acid, very slight growth. Butterfat, rich
growth.

Milk, growth good, alkaline to litmus; curdling (0.25 per cent of calcium chlorid
added) in 10 days; digestion, fairly rapid; color in milk, none.

At 37° C, no growth; grew when cooled; check at 20° C., good colony.

PENICILLIUM PURPUROGENUM O. Stoll.

Beitr. z. morph. u. biol. Char. Penicillium, Wiirzburg, 1904, p. 32, t. I, fig. 6; t. Ill,
fig. 2; t. IV, fig. 3.

Colonies on lactose gelatin and potato or bean agar, gray-green to brown or olive,
deeper green upon cane-sugar media, closely floccose, almost velvety in surface
appearance, spreading slowly over the substratum and producing in the whole mass
of medium a red color. In acid media rich in sugar secondary floccose mycelium arises
white or with hyphae studded with yellow granules. Conidiophores 100-300 by 3.5/*,
arising separately or from portions of hyphse just above the surface of the substratum.
Conidial fructification 50-100/t in length, composed of one verticil of branches (some-
times with a secondary or partial secondary verticil), bearing whorls of conidiiferous
cells 11-12 by 2.5/x, narrowed abruptly to form sterigmata at the apices. Chains long,
divergent. Conidia elliptical, 3.4-3.8/( by 2-2. 5/x, green, granular, with from one to
several small highly refractive granules in each, in chains falling apart in fluid mounts.
Colonies liquefy sugar-gelatin slowly in 15 to 20 days.

Received from Krai in Prague.

The authority for the name of this species is attributed by Saccardo
to Otto Stoll, as here indicated, since his description, or rather dis-
cussion, of this fungus forms the basis of Saccardo's Latin diagnosis.
Stoll quotes the name from Krai, who gives the author as Alex. Fleroff,
in Warsaw. Stoll has given the first discussion that is in any way
adequate.

A closely similar organism has been found by Prof. F. D. Heald
upon corn (Zea mays) in Nebraska. A third form corresponding
closely in morphology and many cultural characters was sent from
Miami, Fla., by Professor Rolfs. Although distinguishable by some
characters, these forms resemble P. jyurpurogenum as described above
so closely in morphology and cultural characters as to justify including
them, temporarily at least, under this name. Neither of these forms
produces the purple color as rapidly or as purely as the original
race of P. imrpurogenum.

CULTURAL DATA.

Color gray-green, becoming dark green with the presence of cane sugar; reverse
yellow to reddish, or colorless; color in media, none to red to deep purple, almost black,
according to medium; odor, none.

Fifteen per cent gelatin in water, medium growth, characteristic fruiting; liquefac-
tion, partial in cultures 3 weeks old, or none, none in 10 to 12 days; litmus reaction,
slowly alkaline or often neutral. Potato agar and bean agar, good colonies but no
purple color, purple produced when sugar is added. Potato plugs, mycelium yellow

PENICILLIUM PINOPHILUM.

37

with granules, potato becoming purple. Uauliu's fluid, slofl development. Cohn'e
solul ion, germinal ion onlj -

Synthetic fluid I Dox'e . i arbon supplied as Cane Bugar LO per cenl in agar, rich
growth, deep red medium. Lactose 3 pei cent, very little growth, verj pale purplish.
Lactir acid 0.9 percent, medium growth, slowlj producing purple in medium. Levu-
lose 3 per cent, small colonies, fluid pale red. Galactose 3 per cent, small colonies,
acid reaction. Glycerin 3 per cent, no growth. Alcohol 3 to 5 per cenl in agar, ur« » ><1
growth bul Blow, medium becoming purple. Potato starch :'. per cent, good growth,
abundanl purple in medium. Tartaric acid, do growth. Butterfat, very slow growth
with purple color in fluid.

Milk. Blow development; curdling (0.25 per cenl calcium chlorid added) L3 days;
digestion slow and only partial; color in milk, shades of purple according to progress,
deepesl under colony.

(nuked apple, feeble growth.

Grew equally well at 37°C.and 20° C.

>v

> ;■ •'■ '■'

;

Fig. 5.- /'< nicillium purpurogt u I ». Btoll: a, b, c, conidial fructification showing branching and arrange-
ment i form with partial secondary verticil; d, e,f, g, conidiiferous cells and conidia i 1,400);
it, Z, 77i, sketches of whole fructifications (X 140).

PENICILLIUM PINOPHILUM Hedgcock momen novum i.

Syn. Penidllium annum Corda, emended Bedgcock, Mo. Bot. Gard. Rept. 17.
pp. L05 l<>7. pi. ii. figs. I 3.
Nol /'. """ a in Corda, Prachtflora, p. 38, t. XVIII.

Colonies on potato or bean agar and milk sugar gelatin, from green on agar through
shades of yellow -green to bright yellow and orange on media containing starch and cane
sugar. Superficial hyphae studded with yellow granules upon acidfied media. Reverse
of colony and substratum (upon these media) colored deep rich red. Surface growth
partly of simple conidiophores, partly aerial hyphae, and ropes of hyphae (which rarely
become vertical coremia) bearing conidiophores as lateral branches. Conidiophores
hiu 200/!. Cniiidial fructifications up to L20/< in Length, consisting of single verticils
of branches 10 L6by2 2.5ju, bearing whorls of conidiiferous cells L3 L5by2 2. 5/i taper-
ing into acuminate sterigmata bearing conidial chains which are parallel bul do not
form a column. Conidia elliptical, 3 3.6 by 2/t, smooth, pale green <>r yellowish green.

38

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

Colonies liquefy gelatin, but slowly and incompletely, and give a neutral or acid
reaction upon all litmus media. Under different conditions of culture and acidity the
discoloration of the medium varies from yellow to orange and deep red. Produces dis-
colorations upon commercial timbers. Habitat, pine wood, which is strongly colored
by it.

Culture received from the author, G. G. Hedgcock, of the Forest Pathological Lab-
oratory, Bureau of Plant Industry, United States Department of Agriculture.

Since the publication of his description of this fungus (1906) Hedg-
cock9 has reached the conclusion, concurred in by the writer, that
this species can not be regarded as identical with the species described
by Corda as P. aureum. He notes that this species is a common agent
in the discoloration of pine wood, hence proposes the name P. pino-
fliilum (here first published). Careful consideration of Corda's
figure and description would establish a strong presumption that

Fig. 6. — Penicillium pinophilum Hedgcock: a, young conidial fructification showing conidiiferous cells at
apex of central branch before all the branches appear (X 1,600); 6, a verticil of four branches, upon one
of which fruit appears (X 1,600); c, d, conidiiferous cells and conidia (X 1,600); c, rope of hyphse bearing
conidiophores sketched (X 140); /, g, forms of conidial fructification (X 140).

the form described by him would not now be considered a species of
Penicillium.

CULTURAL DATA.

Color, conidial areas green, vegetative mycelium colorless or studded with yellow
granules; reverse of colony red; color in media, red.

Odor, none.

Fifteen per cent gelatin in water, growth slow, surface growth of conidiophores
and green conidial fructifications only; liquefaction, none or very slow (in acidified
cultures only after several weeks); litmus reaction, acid. Potato agar and bean
agar, mycelium studded with yellow granules, conidial areas strict, green; reverse
of colony, red. Potato plugs, poor growth, not typical. Cohn's solution, spores
germinated only.

PENICILLIUM Krr.lMM AND I'. LUTEUM. 39

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, growth fair, hyphse yellow
with granules, acid reaction, conidia green, reverse, red. Lactose, growth bIow, nol
characteristic. Levulose 3 percent, fair growth, yellow mycelium, acid to Litmus.
Galactose 3 percent, fair growth, acid reaction. Glycerin, very slighl growth which

became typical when sugar was added. Potato starch, g 1 characteristic growth.

Butterfat, slow weak growth, with characteristic colors and red fluid.

.Milk, growth small; curdling (0.25 per cent calcium chlorid added), very slow;
digestion, none or very Blight; color in milk, red at top.
At :;7° ('., grew well, more rapidly than at 20° C.

PENICILLIUM RUBRUM O. Stoll.

Beitr. z. morp. u. biol. char. Penicillium, Wurzburg, 1904, p. 35, t. I, tiur. 7: t. III.
fig. 3; t. IV. fig. i.

Colonies upon lactose gelatin and potato or Ik -an agar, from green through ochraceous
to ochraceous red with varying conditions; consisting of green conidia with yellow
mycelium when cane sugar is added. Aerial portion velvety strict or very closely
floccose in media without sugar, becoming dense cushions of mycelium bearing suc-
cessive crops of green conidia in cane-sugar media; marginal growth continuous but
slow and not marked by a white border. Reverse and mycelium yellowish to red,
yellow in BUgar media; the .substratum also colored in old agar colonies. Conidio-
phores arising from substratum directly or as very short lateral branches of the felted
hyphse, mostly 15-30 by 3-3. 5/*, swollen at the apex, making a dense layer on the
surface. Conidial fructification usually massed into a heavy column with a broad
triangular base, 100-200,u in length, from a dense \erticil of branches of the conidio-
phore. each swollen at the apex. Conidiiferous cells 10-13 by 2-3//, with rat her abrupt
points from which the conidia are cut off. Conidia at first cylindrical, then elliptical
or even globose, 3.4 by 2/x, or 2.5-3.3/*, yellowish green to dark green when mature.
Colonies produce slow and only partial liquefaction of sugar gelatin.

A slow-growing fungus fruiting for several weeks and differing greatly in colors with
slight and undefined differences in the conditions. Sometimes producing a blood-
red color on the reverse of the colony.

Cultures received from Krai. So far not found native in America.

PENICILLIUM LUTEUM Zukal.

Sitzber. K. Akad. Wiss. (Vienna) Math. Naturw. Kb, XCVIII, p. 521, 1889.

Conidial form; Colonies on sugar gelatin and potato or bean agar, white or gray
or transiently yellow on media lacking sugars, with sometimes greenish areas of coni-
dial fructification showing shades of yellow (egg yellow) upon sugar media, later
passing over to reddish, especially with the formation of aerial wefts or balls of hyphse
producing asci (several weeks-; Burface rather dose floccose, spreading indefinitely
upon the substratum. Reverse of colon) more or less reddish, especially on sugar
media Conidiophores a thin and incomplete layer, scantily produced mostlj
lateral branches of aerial hyphse, 20-100/t (mostly 30 60) by 3/x. Conidial fructifica-
tion usually small up to SO/t in length, commonly \s it h a single lateral branch and but
two verticils of long acuminate basidia 13 10 by 3 I". Conidia elliptical to fusiform,
2.4 by 2.3/1, rather firm walled, greenish, swelling greatly and producing I or 2 tubes
in germinating.

This species characteristically produces yellow mycelium, from which, in a time
varying from a few days upon media rich in BUgar to several months upon plain potato

agar, ascigerous wefts of hyphse arise. As given by Wehmer,1 ascigerous concep-

tacles" are 0.5-2 mm. in diameter, globose, vitelline then red; asci reddish, globose
to fusiform 8.8 by 7-7.8/ij sporidia 1.8 by 3.3/(, transversely tricostate, hyaline to

40

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

reddish. Colonies do not liquefy, or only slowly and partially liquefy, sugar gelatin,
Litmus reaction neutral, the shades of purple found abowt the turning point of litmus,
Colonies on apple are brighter yellow and produce ascigerous masses in very few days.
Received from Professor Thaxter, identified from the ascigerous form by Dr. C.
Wehmer.

The author is convinced that Wehmer32 is in error in attributing
a large, degree of polymorphism to this fungus. Numerous cultures
watched under all sorts of conditions are evidence that this species
does not produce prominent coremia or large masses of green conidia.

CULTURAL DATA.

Color white, but mycelium studded with yellow granules in acid media, with some-
times reddish areas and conidial green areas; reverse of colony yellow to orange;
color in media, pink in potato.

Odor, none.

Fig. 7. — Penicillium rubrum O. Stoll: a, 6, c, d, e, Whole conidiophores and the branching of conidial
fructifications (X 1,400); /, g, conidiiferous cells and conidial formation (X 1,400); h, j, sketch and diagram
of habit of growth (X 140); fc, sketches of old conidial fructification in large size (X 140); m, diagrammatic
figure (the successive series of conidial fructifications are produced by new branches from hyphae over-
growing the earlier series). (Drawn from gelatin culture, but found with approximately the same
morphology upon potato agar.)

Fifteen per cent gelatin in water, medium growth; liquefaction none; litmus
reaction acid; potato agar and bean agar, mycelium transiently yellow, then color-
less or reddish or yellowish gray; potato plugs, white to gray, in parts yellow, potato
pinkish; Raulin's fluid, slow development, bright yellow colonies; Cohn's solution,
germination only.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, good growth, yellow, up
to 30 per cent sugar, no growth at 50 per cent. Lactose 3 per cent, very slight growth;
lactic acid 0.9 per cent, very slight growth. Levulose 3 per cent, very slight growth.
Galactose 3 per cent, typical, acid reaction. Glycerin 3 per cent, germination only.

l'INh 1LLIUM LUTEUM.

41

Fig H.—Penicillium lutcum Zukal: a, whole eonidiophore and fructification (X 900); 6, c, d, conidiiferous
cells, conidia and their arrangement ( ■ 900); g, h,J, k, I. m, n, Bketchea of fructifications (> 140); o, r,
swelling an<l germination of conidia, Bwollen conidia in the same chain with those which refuse to grow

(X900).

42

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

Potato Btarch 3 per cent, slow development but characteristic. Butterfat, little or
no growth.

Cooked apple, mycelium yellow, ascigerous masses produced most quickly (1 week).

Milk, grows poorly; curdling (with 0.25 per cent calcium chlorid) none; diges-
tion slight and slow.

At 37° C, grew well; at 20° C, more slowly.

PENICILLIUM DUCLAUXI Delacroix.

Bulletin de la Societe Mycologique de France, Tome VIII, 1891, p. 107, PI. VII.

Colonies grown upon gelatin and potato or bean agar clear dark green to olive when
old, consisting of short crowded conidiophores arising for the most part singly from
the substratum (strict), but sometimes producing short coremia. Long coremia are
produced abundantly upon orange, milk, potato, and all media rich in cane sugar.
Conidiophores very short, 10-50/*, either arising directly from the substratum or borne

Fig. 9. — Penicillium duclauxi Delacroix: a, b, conidial fructifications with young conidia smooth, from
potato-agar plate culture, simpler types (X 900); c, d, e, conidial fructifications from potato-agar plate
culture, more complex types (X 1,400); /, g, h,j, sketches of habit upon potato agar, showing the very
short conidiophores arising from the substratum ( X 140); k, ripe spores highly magnified to show delicate
markings (X 900, apochromatic); J, m, n, germination of conidia (X 900, apochromatic); s, t, coremia,
(sketch).

upon the upper third of the coremia, 1-2 septate, bearing a simple conidial fructifica-
tion or a terminal fructification and a divergent lateral branch with a whorl of coni-
diiferous cells. Conidial fructification often 100-160/£ in length consisting of a few eoni-
diiferous cells 10-12/t in length in a simple terminal whorl or less commonly in
secondary whorls. Conidia elliptical fusiform 3.6-4 by 2-2. 5/i, clear homogeneous green,
smooth when young, but rugulose when ripe. Colonies liquefy sugar-gelatin in Petri-
dish culture slowly from twelfth to twenty-fifth day and change red litmus to blue in 7
days. Produces a coloring agent in sugar media which is wine red in alkaline media
and yellow (bile-yellow) in acid media (acts as an indicator with neutral point very
near that of phenolphthalein).

This fungus is characterized by its enormous development of coremia upon milk,
orange, apple, and media containing cane sugar, while producing only very short
conidiophores in bean or potato agar and gelatin free from sugar.

Received from the author, George Delacroix, Paris, but one culture previously
obtained from P. H. Rolfs in Florida in the summer of 1905 was thought to be this.

I'KNK'Il.I.irM Cl.AVIloKMi:.

43

■

This species has not been found commonly in America, bul it has
been many times observed and recognized as a contamination of
other cultures in this laboratory since its introduction. Its identi-
fication is therefore easy.

CULTURAL DATA.

Color, bruiting areas olive to clear deep green, often brown when old; reverse, from
colorless to yellow and shades of red, according to medium; color in media, none in
media free from sugar, in sugar yellow at first, then slowly red (color acts as an indi-
cator acid when yellow, alkaline when red).

Odor, rather strong in Haulm's fluid, none in most cultures.

Fifteen percent gelatin in water, medium growth; Liquefaction, none in 15 days,
then alow in acidified cultures especially; litmus reaction, neutral slowly or weakly
alkaline. Potato and bean agar, green fruiting surfaces consisting of very short
crowded conidiophores slightly
tuberculate at times, bul no co-
remia. Potato plugs, abundant
coremia, deep green, reverse and
potato yellowish. Uaulin's fluid,

g I growth, many coremia, some

odor. Cohn's solution, germina-
tion only.

Synthetic fluid (Dox's), carbon
supplied as: Cane sugar 3 percent,
rich growth, coremia, poor colonies
at 30 per cent and (iO per cent.
Lactose 3 per cent, very weak
growth, tew very small coremia.
Lactic acid 0.9 per cent, good
growth, typical, coremia many,
remains acid. Levulose '■'> per

cent, weak colonies. Galactose

3 per cent, poorgrowth. Glycerin FlG w.—PenidHium daviforme Balnler: a, coremlum grown

:'. per cent, germination and slow, upon sugar media, showing branching stali with several

weak colonies Alcohol some small heads (if conidia; 6, coremium grown upon gelatin

,. -,-, , , . , free from sugar, showing typical unbranched stalk with a

growth. Potato starch, rich . , ... ,.,, . . .

6 ' single conidial mass splitting as it increases in size into

growth, coremia, reverse and fluid several columns composed of chains of conidia. (For full
yellowish Butterfat, slow but illustration of the structure of this species, see Bainier's

typical coremiform colony, fluid figures.)
yellow 'acid reaction).

Milk, typical coremiform colonies; curdling (0.25 per cent calcium chlorid added)
in 9 days; digestion slow; color, becoming yellow (acid) and later red (alkaline) in
very old culture.-.

At 37°C. no growth; check grew at 20° C.

PENICILLIUM CLAVLFORME Bainier.

Bulletin Trimestriel de la Soctete1 Mycologique de France, Tome XXI, 1905, p. 127,
PI. XI, figs. 8-11; Saccardo, Sylloge Fungorum, Vol. XVIII, p. 520.

Colonies on milk-sugar gelatin and potato agar, white or gray, with surface com-
piled of loosely (loccose liypha', bearing simple but definitely penicillate fructifica-
tions, between the bases of white or yellowish simple or variously branched coremia
1-2 cm. long, fertile only at the apices. Simple fructifications sparingly branched,
hearing small verticils of conidiiferous cells 9-10 by 2/<. Coremia! fructifications
consisting of closely branching and interwoven hyphse, producing vert icilsof conidiifer-

44 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

ous cells crowded into a false hymenium and producing chains of conidia adhering in
olive-green masses 1-3 mm. in height. Conidia elliptical, showing a connective,
4.2-4.6 by 3-3.3 /i, homogeneous green, remaining in chains in fluid mounts. Colonies
only partially liquefy gelatin media and give a weak alkaline or neutral reaction with
litmus.

Received from G. Bainier October, 1905.

A culture received from Reddick, Ithaca, N. Y., marked Whetzel
No. 2095, proved to be this species. It was found at Junius, N. Y.
A culture sent to Dr. C. H. Peck was not recognized. Although not
closely resembling other species of Penicillium it may best continue
under the name given by Bainier until closer affinities are found for it.

CULTURAL DATA.

Color, mycelium white or gray, conidial heads olive green; reverse of colony brown;
color in media brownish.

Odor, perceptible in media containing cane sugar, characteristic.

Fifteen per cent gelatin, growth, characteristic coremia, but mainly unbranched;
liquefaction slow, partial; litmus neutral. Potato agar and bean agar, typical
coremiform colonies. Potato plugs, vigorous typical growth with long coremia.
Raulin's fluid, characteristic. Cohn's solution, characteristic.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, typical growth with acid
reaction in concentrations from 1.5 to 30 per cent, coremia branching, no coremia at
50 per cent. Lactose 3 per cent, slight growth, poorly nourished colonies, very
delicate coremia. Lactic acid 0.9 per cent, germination only. Levulose 3 per cent,
slow-growing, poorly nourished colony. Galactose 3 per cent, typical growth, acid
reaction. Glycerin 3 per cent, not typical, no coremia, coremia produced after
addition of cane sugar. Malic acid 1 per cent, slight growth, few and very small
coremia. Butterfat, typical colonies.

Milk, good characteristic growth; curdling (0.25 per cent calcium chlorid added)
in 9 days; digestion slow but fairly complete; color in milk brown or reddish.

At 37° C, no growth in 6 days; culture grew when cooled to 20° C.

PENICILLIUM GRANULATUM Bainier.

Bui. Soc. Mycol. France, XXI, 1905, p. 127, PI. XI, figs. 6, 7.

Colonies upon plain gelatin and potato or bean agar yellowish green to gray or
grayish brown, superficially composed of crowded small coremia 1-3 mm. in height,
mixed with floccose hyphse and separate conidiophores, spreading indeterminately
upon the substratum. Reverse reddish orange (approaching "fulvous"), aerial
hyphse delicately granular or spinulose, which separates this from all other species
studied. Conidiophores 4-4. 5/i in diameter, short or very long, either separate or,
mostly, massed into very short, crowded coremia (less than 1 mm. in height). Conidial
fructifications usually 100-200,u in length, once or twice verticillate, with many
conidiiferous cells 9 by 2-2. 5/i, and long, loosely divergent chains of conidia. Conidia
at first cylindrical, then elliptical to globose, about 2.5-3 by 3-3.5 or 3/« in diam-
eter, yellowish green, granular, remaining in long chains in fluid mounts. Colonies
do not liquefy gelatin, litmus reaction slowly alkaline.

The delicately granular or spinulose hyphae as noted and figured
by Bainier are a valid and distinctive character. The species is also
easily recognized by its general appearance and habit. Obtained
from the type cultures of Professor Bainier, Paris.

PENICILLIUM i;i;i.\K Al'LE.

45

( I I.TUKAL DATA.

Color yellowish green to green and later \arious shades of hrown; reverse of colony
orange, or yellow to deep orange, or even red; color in media yellow to orange to
red in media containing starch or sugar.

< (dor, none.

Fifteen per cenl gelatin in water, characteristic growth; liquefaction, none, or

partial alter several weeks in acidified culture-; litmus reaction, strongly alkaline.

Potato agar and bean agar, yellow-green to In-own, yellow below, coremia not closely
crowded as in sugar media. Potato plugs, characteristic growth, potato stained yel-
low to ilcc]) brownish yellow. Etaulin's fluid, characteristic growth. < John's solution,
germination only.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, 3-30 per cent , characteristic
growth and colors, hut not normal at 00 percent, with acid reaction. Lactose 3 per
cent, characteristic structure but weak development. Lactic acid 0.9 per cent,
growth, hut not normal colonies. Levulose :5 per cent, slow development, small colo-
nies. Galactose 3 percent, characteristic. Glycerin :'. percent, slighl growth, grew
freely when cane sugar was added. Potato starch, characteristic. Butterfat, typical
growth, reverse and fluid orange; yellowish.

I i'.. n. i'i nirillium granulaium Balnier: a, b, c, branching of conidlal fructification showing granu-
lated echinulated cell walls (x 900); d, e, f, g, conidiiferous cells and conidlal chains ( < 1,400); h, j, k,
sketches of fructifications (X 140); m, sketches drawn from photomicrograph of coremia showing fertile
and sterile areas (X about 25).

Milk, rapid and characteristic growth; curdling, none; digestion, slow but complete
(fairly); color in milk, yellow to orange to deep red.
At 37 C, no growth, conidia grew when removed to Lower temperal ure; at 20° < '.,

excellent gTOWth.

PENICILLIUM BREVICAULE Saccardo.

Fungi Itali.i, No. 893, Mich., II, p. 547.

( 'clonics grown upon sugar gelatin grayish white, then yellowish brow n or chocolate,
consisting of Bhorl closely crowded conidiophores making powderj areas o\ ergrown by

loosely trailing floccose hyphse and ropes of hyphse, with broadly spreading indeter-
minate margin. Coridiophores, short , in :'()/< mostly, arising directly from the sub-
merged hyphse, or numerously and irregularly borne as lateral and perpendicular
branches of trailing aerial hyplue and rope- of hyphse. < 'on i dial fructifications either
simple chains terminating unbranched or sparingly branched conidiophores in young
colonies, or verticillately and irregularly twice verticillately branching systems bear-
ing numerous divergent chains often I ">< i" in length in old colonies. Conidiiferous

46

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

cells continuous with conidiophores 12-15 by 4/i tapering to slender sterigmata. Co-
nidia somewhat pear-shaped, slightly tuberculate at apex, with broad base, 6.5-7.5 by
7.5-9/i, in mass light brown to chocolate; at first smooth, then with thick tuberculate
walls, viable for many months, germinating by a single tube from the thin center of
the broad base into a bulbous enlargement from which mycelial hyphae about 2/t in
diameter arise. Mycelium very thin walled, narrow cells of varying length. Colonies
liquefy sugar gelatin and give a strong blue reaction in litmus media, but grow very
tardily, if at all, in potato or bean agar. Grows very rapidly upon neutral or alkaline
media, but very slowly or not at all in media acid to litmus. Digests milk. Refused
to grow after repeated inoculation into sterilized apple. Spores which refused to
germinate in the agar media used grew immediately when transferred to any of the
gelatin media.

Cosmopolitan, forms characteristic chocolate patches on Camembert cheese. Se-
cured from numerous brands of cheese and common in the laboratories of this station.

Fig. 12— Pen icillium brevicaule Saceardo: a, conidiophores and simple conidial chains with spores still
smooth (X 900); &,/, more complex conidial fructifications (X 900); c, two young conidial chains, show-
ing thick walls of spores (X 1,400); d, c, conidia after becoming echinulate (X 1,400); g, h,j, sketches of
forms and habit of conidial fructifications (X 140); g from an old cluture, sessile or almost so; h and j
show trailing hyphae and a rope of hyphae with lateral conidiophores; k, germinated conidium where
the old spore wall lies empty beside the growing cell (X 1,400).

The author does not believe that this species is closely related to
other species of this genus, but since it has been placed here by a
very liberal interpretation of descriptions it may perhaps remain
under this name until someone finds out its real affinities. The two
forms which follow as varieties are found in the same habitat, show
closely similar morphology, and give almost identical physiological
reactions. Their designation as varieties of P. brevicaule may there-
fore be justified, though one at least (var. glabrum) seems separate
enough to warrant proposing for it a specific name.

CULTURAL DATA.

Color, conidial surfaces clay-yellow to chocolate; reverse of colony,
colorless; color in media, none.
Odor, ammoniacal, used as a test for arsenic (Gosio et al.).

mycelium

PENICILLIUM BR KV I C A C LK.

47

Fifteen per cent gelatin in water, typical colony; liquefaction, rapid — 5-6 days;
litmus. Btrongly alkaline. Prefers alkaline, neutral, or only slightly acid media.
Potato agar, spores sometimes refuse even to germinate, bul grow when transferred
to gelatin; grows poorly on agar media in'*' from sugar, or peptone, or the by-products
of other fungus growth. < !ommon as a secondary growth in such plates. Potato plu
good colonies. Conn's solution, characteristic but slow growth. Raulin's fluid,
germinated, bu1 veryslighl growth.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar in agar 3 per cent, failed
to grow, same spores transferred to gelatin media grew promptly. Lactose K) percent,
plow, characteristic color to conidia. Lactic acid o.'.i per cent, germination only.
Levulose 2.5 per cent, growth characteristic, alkaline reaction to litmus. Galactose
3 per cent, characteristic growth, alkaline reaction to litmus. Glycerin 3 per cent.
,-poie- germinated only, grew well upon addition of cane sugar. Potato starch grew
well. Malic acid, germinated only. Butter fat, slow development, finally producing
drops of yellow oil which separate out.

Milk, rapid growth: curdling (0.25 per cent calcium chlorid added) in 10 days;
digestion, rapid; culm- in milk, none.

Cooked apple, failed to grow after repeated inoculation.

At 37 and 20 I '.. grew equally well.

I io. 13.- /'< niriiiuiu brevicauli , Tar. album.' ';. b, c, a", eonidlophores and fructification borne variously and
differently branching I < 900); e,f, ripe conidia; g, h, sketch of single hyphseand a rope of hyphae bearing
eonidlophores i X 140); j, germinated spore.

PENICILLIUM BREVICAULE Saccardo, var. ALBUM Thorn, n. var.

Colonic- upon Bugar gelatin white to cream-colored alike above and below upon all
media, stricl to sparsely floccose, with trailing hyphse and ropes of hyphse, indeter-
minately spreading. < 'onidiophoree either arising from Bubstratum directly or mostly
a- perpendicular branches of serial hyphse and ropes of hyphse 15-40/1 in length,
conidia! fructification varj ing from a Bingle chain to more or less complex penicillate
branching, mostly producing few chains of indefinite length and arrangemenl from
narrow tapering conidiiferous cells. Conidia pj riform tosubglobose, with basal collar,
9-10/', roughly tuberculate, white or slightly yellowish tinged, thick walled excepl at
the hase, the center of which remain- as a germ pore. ( lolonies rapidly liquefy sugar
gelatin with strong aminoniacal odor, and give an intensely alkaline reaction in litmus
media. Gives exactly the same reactions as /'. brevicaule Sacc. Differs from the
latter slightly, excepl in the color of the spore-.

Common upon imported Camemberl cheese. Found often upon domestic Camem-
berl and grow.- very readily in cheese cellars, where it becomes a nuisance.

48

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

CULTURAL DATA.

Same as P. brevicaule Sacc, except the following difference noted:

Conidia cream, somewhat larger than P. brevicaule.

Cohn's solution failed to produce characteristic colonies.

In Dox's solution, with butterfat as a source of carbon, it differs from P. brevicaule by-
failing to cause drops of yellow oil to separate out.

Agar-agar: In repeated cultures this organism has failed to grow well in agar media.
In some such cases the spores transferred from the agar to gelatin grew at once. Some
cultures upon agar grow slowly and in typical manner, but the development upon all
agar media has seemed uncertain. In synthetic solution cultures were obtained when
the inoculation of tubes of the same solution with 1.5 per cent of agar added to make a
solid substratum produced no growth.

PENICILLIUM BREVICAULE Saccardo, var. GLABRUM Thorn, n. var.

Colonies white or only slightly yellowish-tinged in all gelatin media, grow not at all
or with difficulty on agar of most formulae. Aerial portion consisting of short, closely
crowded conidiophores making a powdery surface overgrown by loosely trailing
hyphte and ropes of hyphse, spreading broadly over the substratum. Conidiophores,
short, mostly 10-30^, arising directly from submerged hyphse or numerously and irregu-

Fig. 14.— Penicillium brevicaule, var. glabrum: a, b, c, branching of conidial fructification (X 900); d, chain
of conidia (X 1,400); /, formation of conidium on young branch (X 1,400); g, h, sketch of appearance
in culture (X 140); m, n, o, germination of conidia.

larly borne as perpendicular branches of the superficial hyphse and ropes of hyphse.
Conidial fructifications from simple chains of spores to fairly complex penicillate
groups of branchlets resembling P. brevicaule, but mostly less complex. Conidia
obovate, pyriform 7-8 by 8-10/t or almost globose, 7-9//, smooth, white, rather thick-
walled and retaining their power to germinate for many months. In old potato and
other cultures black sclerotia are formed in the substratum but do not produce asci.
Liquefies gelatin rapidly (within one week), gives a strong alkaline reaction and
ammoniacal odor.

Habitat, found repeatedly on imported Camembert cheese and secondarily upon
domestic soft cheese, where it grows into prominent cottony patches indistinguishable
to the eye from the white variety of P. brevicaule. This fungus is separated from
P. brevicaule by its smooth white spores and the production of the black sclerotia in
the substratum.

I'KNh ll.Lir.M KOSEUM.

49

This form i- certainly closely related to P. brevicaul by its physiological reactions
and its general morphology. It was found in one case among the exsiccati in the
Earvard herbarium under the name of Monilia Candida. There is, however, ao possi-
bility of confusing this form with thai species as undersl 1 and described by more

recenl students, such as Hansen and Jorgenson.

Cl I n a \i DATA.

Exactly as in /'. bn i icauk , excepl for the following:
Conidia Bmooth, somewhal smaller.
< !olor more Dearly white.

Sclerotia black, found in very old potato-plug cultures or in agar cultures which
lia\ e grofl n several weeks or monl h
Cohn'e solution failed to produce a characteristic colony.

PENICILLIUM ROSEUM Link I ? .

Colonies on milk-sugar gelatin or potato agar white to pink or salmon in fruiting
areas, loose floccose with simple hyphse and ropesof hyphae, producing dense irregular

..;V.;S?f?..fi~'.-.';j.i.'

Fig. 15.— PenkUUum roaeum Link (?): o, 6, c, branching of conldlal fructlflcation, showing few cells In
ch verticil I < 900); d, e, conidiiferous cell and conidia ( x 900); g, h, j,k, sketches of ripe fructifications
showing agglutination of conidia Into slin < 1 10).

pinkish masses or Bclerotia up to I nun. or more in diameter in old culture. Coni-
diophoree borne as perpendicular branches of aerial h\ phse or ropes of h\ phas, 15 L25/i
Conidial fructification up to L40/i in length, once or twice irregularly alternately or
verticillately branched, with conidiiferous cells van ingfrom 12 by 2 :\n in the verticils
of •") or less to 17 by 2.::» when solitary, bearing conidia which becoi regated into

gelati nuns halls or masses. Conidia colorless (pink or rosy in mass), elliptical, 5 7 by
3 •">,'. slightly apiculate, Bmooth, appearing delicately granular within. Colonies
liquefy gelatin cultures rapidlj and give an alkaline reaction to litmus media.
Brought from Krai, in Prague, Bohemia.

8108— Bull. 118—10 4

50 CULTUEAL STUDIES OF SPECIES OF PENICILLIUM.

The same organism has been found once in accidental culture in this
laboratory; received once from a correspondent in Halle, Germany,
and later found under this name as No. 1179 in De Thumen's Myco-
theca Universalis; collected by Ravenel in South Carolina in 1876
upon leaves of Buxus; this and several other specimens were found
in the mycological collection of the Bureau of Plant Industry, United
States Department of Agriculture. The spores are the same length
as given by Saccardo, but slightly broader. The number of specimens
found under this name from widely different workers appears to
justify the belief that this is the organism described by Link under
this name. If the development of a mucilaginous mass enveloping
the conidia be regarded as a sufficient basis for separation of such
species under the generic name of Gliocladium, this species would
become Gliocladium roseum. (Link).

The form upon Buxus is cited by Saccardo, referring to it as UP.
roseum Cooke, non-Link," and held to be Verticillium buxi Auersw.
et Fleisch. Examination of the material would indicate that in
De Thumen's collection at least this species is more closely allied to
the other species of Penicillium than to Verticillium.

CULTURAL DATA.

Color white or shades of salmon pink; reverse cream or white; color in media,
none.

Odor, none.

Fifteen per cent gelatin in water, medium growth, white. Liquefaction, rapid.
Litmus reaction, alkaline, strongly. Potato agar and bean agar, good growth, but
white. Potato plugs, white colonies. Cohn's solution, slight growth.

In Dox's solution, with butterfat as a source of carbon, this species caused drops
of yellow oil to separate out.

At 37° C, killed; check at 20° C, good.

PENICILLIUM CAMEMBERTI Thorn.

Emended from U. S. Department of Agriculture, Bureau of Animal Industry, Bui.
82, p. 33, fig. 1, 1906.

Possible syn.: P. album Epstein (not Preuss), Archiv f. Hyg., Bd. 45, Hit. 4,
p. 360, 1902.
P. epsteini Lindau, Deutschl. Krypt. Flora, Pilze, VIII, p. 166.

Colonies on potato agar or lactose gelatin effused ; white (sometimes yellowish white),
changing in 5-8 days to gray -green (glaucous); surface of colony floccose, of loosely
felted hyphse about 5/< in diameter, reverse of colony yellowish white; conidiophores
300-800/i in length, 3-4/i in diameter, septate, cells thin-walled, often collapsing in
age, arising as branches of aerial hyphae; fructification sometimes 175/i in length, but
usually much less, consisting commonly of one main branch and one lateral branch,
sparingly branched to produce rather few conidiiferous cells which bear long loosely
divergent chains of conidia. Conidiiferous cells 8-11 by 2.4-3/z. Conidia at first
cylindrical, then elliptical, and finally globose when ripe, smooth bluish green by
transmitted light, thin-walled and commonly guttulate, 4.5-5.5/* in diameter, swelling
in germination to 8-10/t. Germ tubes one to several. Cells of mycelium about 5
by 20-40/*. Liquefies lactose gelatin only under center of colony. Produces a strong

PKXICILLU'M CA.MKMP.KKTI.

51

alkaline reaction in gelatin, free from sugar, bul in sugar media producee a in. .re or
less persisted acid reaction. Growing and fruiting period, about 2 weeks. Fruits
only upon exposed surfaces of the substrata; never produces spores in cavities not
broadly open.

Habitat, Camemberl and other sofl cheese

'Flic aarne /'. album Epstein, which is also P. epsteini Liiulau,1" is
inserted in the list of possible synonymy because i bis name is accepted
for tlii- mold by Maze |:; iii a recent paper. If Ave base identification
upon the cultural charac-
ters given for his mold
by Epstein, it could not
have Ween /'. carru mh, >fi.
The characterization, so
far as given by Epstein
and extracted from his
paper by Lindau, might
refer to the pure white
form (P. candidum) as
interpreted by Roger and
Maze better than to this
species. There is cer-
tainly at least a varietal
difference (spore color)
between P. camemberti
and the P. c<nt</i<hiin of
Roger and Maze, which
is designated a variety of
P. camemberti in this
paper.

The change in litmus
reaction in this descrip-
tion from thai previously
published is result of
proof that no acidity is
produced by this species

e\cel)t V hell sil'r-ir is Ol'es- ' "'• '''•— PenidUium camemberti Thorn: a, conidiophore show-

'~\ lug a common type of branching and the production of baslda

ent. A change from blue :ill,| conidla, highly magnified; &, a common form showing

to fed and black to blue much less branching; e, d, f, diagrams of large fructifications

. (X 80); q, i,j, germinating conidia. (From Bull. 82, Bu

indicates the production of Animal Industry.)

of acidity by fermenta-
tion of sugar followed by the neutralization of the acid so produced
by the alkaline by-products of proteolysis or by a further change
iii the acid. Therefore this statement is best omitted from the diag-
QOSl ic description.

52 CULTURAL STUDIES OF SPECIES OE PENICILLIUM.

CULTURAL DATA.

Color white, through cream to a shade of gray-green upon sugar media, without
sugar gray or drab, all becoming drab when old (exposure to light); reverse of colony,
uncolored, cream; media, uncolored.

Odor, none, except in very old cultures upon milk or cereal.

Fifteen per cent gelatin in water, moderate growth, not rich; liquefaction, none,
or slow and partial after 2 to 3 weeks in acidified cultures; litmus, strongly alkaline;
colonies slow, moderate growth, typical. Potato agar and bean agar and potato plugs,
fair, white to gray colonies lacking or nearly free from green color. Raulin's fluid,
very heavy growth, white through pronounced cream to green shades. Cohn's solu-
tion, slow growth, characteristic, but lacking in green color.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, characteristic, abundant
green color in solutions as high as 50 per cent, acid. Lactose 3 per cent, characteristic,
acid reaction. Lactic acid 0.9 per cent, slowly typical, with ultimately an alkaline
reaction to litmus. Levulose 3 per cent, slowly typical. Galactose 3 per cent, char-
acteristic growth, very slowly developing the typical green color. Alcohol about 5
per cent, characteristic. Potato starch, characteristic . Malic acid 1 per cent, fair
colony, colorless. Butterfat, slowly typical colony with a tinge of violet in reverse,
no color in fluid.

Milk, typical growth; curdling (0.25 per cent calcium chlorid added) in 10 days;
digestion, slow but fairly complete; color in milk, none.

Camembert cheese, capable of reducing moist curd soured by lactic organisms to a
semiliquid condition.

At 35° to 37° C, no growth; at 20° C, excellent.

PENICILLIUM CAMEMBERTI, var. ROGERI Thorn, n. var.

Syn. P. candidum of Roger and Maze, not Link.

Colonies grown upon sugar gelatin or bean or potato agar pure white, loosely and
evenly floccose to the very margin, where aerial and submerged hyphee grow with
equal rapidity. Reverse of colonies white or yellowish white (not discolored). Co-
nidiophores 3-5 by 100-800/.! varying greatly, mostly branches of aerial hyphse. Conidial
fructification 70-90/1 in length, loosely and irregularly branched and bearing rather
few basidia at unequal heights, with divergent chains of colorless conidia. Branch-
ing system of conidial fructification sometimes 75/* in length. Conidia smooth,
hyaline, 4^.5 or even 5.5/* in diameter, globose or nearly so when ripe. Sugar gelatin
is slowly liquefied under the center of the colony only, colonies never floating in a
pool of liquid. Reaction in the medium is acid to litmus at first, then changes to
alkaline.

This fungus has been found by the author only upon Camembert,
Brie, and Xeufchatel cheeses from western Europe.

This variety has been discussed by Maze 13 as P. candidum Link.
This seems an impossible application of the name P. candidum from
Link's description n or that given by Saccardo, since the spores are
stated to be 2-3// in diameter. Further, in such identifications no
account is taken of a paper by Morini/6 in which an ascigerous stage
is described for P. candidum Link. Under four years of cultivation
no signs of an ascigerous form have been produced. Stoll 2i has consid-
ered P. candidum to be only a colorless P. glaucum, but as the author
has so far failed to find a worker who will undertake to limit the name

PENN II.l.ir.M CAM I :.M I'.li; I 1.

53

P. glaucum to a special form, this does nol mend matters. Long
cultivation docs show, however, thai this organism is closely related
fco the one already described as P. camemberti.95 Since this is the
form given prominence in cheese studies by the work of Georges
Roger,21 it seems most natural to regard it as a variety of the former
species and designate it by Roger's name.

In Lafar's Handbuch der Technischen Mykologie, Professors
Weigmann88 and Wehmer84 refer to this fungus as probably iden-

Fra. 17. PeniciUium camemberti, vax. rogeri: o, 6, c, types of branching of conidial fructification ( < 900);
e, ripeconid J, h, j, k, sketches of conidial fructifications (> 140);

/, in. germinal ion '>i conidia by several tul

tical with P. camemberti. However this may appear from examina-
tion of the lit etat life alone no one actually familiar with the cultures
will claim such identity. If the possession of colorless spores be
regarded as a case of albinism, this form may perhaps be regarded
as a variety of P. camemberti. It has been kept separate and re-
mained constant in culture for several years, It would seem, there-
fore, equally proper to regard it as a different species were it not so
closclx associated with /'. camemberti in every other character.

54 CULTUKAL STUDIES OP SPECIES OF PENICILLIUM.

The name P. candidum was used by Link for a mold growing upon
decaying leaves, bulbs, and fungi, which was said to be common,
and Morini 16 later describes an ascigerous form of this fungus. The
spores are of different size (2-3/0. There appears to be no justifica-
tion for adopting this name for the fungus used by Roger in cheese
making.

CULTURAL DATA.

Different from P. camemberti only as follows:
Color of conidia persistently white.

Cohn's solution, failed to germinate. Same spores transferred to gelatin after 4
months grew normally.
Camemhert cheese, does not produce the same texture as the preceding species.

PENICILLIUM BLFORME n. sp.

Latin diagnosis.® — Coloniis in gelatina cultis, albis, lente glaucescentibus, densius
floccosis, margine sterili lata, aut, in agaro solani tuberosi cultis, albis glaucescentibus,
mox avellaneis vel fere olivaceis, parte aeria ex conidiophoris brevissimis et creberrimis
fructibus conidicisque composita; conidiophoris (sine ramis) 60-150/x in agaro, vel
longioribus ramosis ex hyphis floccosis in gelatina cultis; fructibus conidicis 100-200/t
longis, plerumque 1-2 ramosis alternates, ramis convergentibus vel divergentibus,
ramulis verticillatis basidiaapice verticillatagerentibus; basidiis 8-10 usque 13X3/*,
conidiis primum ellipticis vel cylindricis demum globosis, 3.2-3.5X4-4.3, vel 4/z
diam., in catenis manentibus submersis; coloniis copiosis in saccharo lactis, gela-
tinam in parte lente liquefacientibus, alkalinis lacmo, odore mucidis.

Habitat, in caseo, ex Gallia.

Cultivated in gelatin, white, slowly gray-green, densely floccose, with broad vegeta-
tive margin, spreading widely over the substratum; in potato agar white, then gray-
green, rapidly becoming yellowish-brown, drab, or almost olive, restricted in growth,
aerial portion consisting of very short densely crowded conidiophores and conidial
fructifications; conidiophores 60-150/* on agar or slightly longer when arising as
branches from the floccose aerial mycelium growing upon gelatin; conidial fructifi-
cations mostly once or twice alternately branched, branches convergent or divergent,
each bearing a verticil of branchlets crowned by verticils of conidiiferous cells with
chains of conidia, the whole 60-240/*, usually 100-200/*, in length; conidiiferous cells
8-10 or even 13 by 3/*; conidia elliptical or cylindrical, then globose, 3.2-3.5 by 4-4.3/*
or 4/z in diameter, adhering in chains in fluid mounts; grows luxuriantly in fluid offer-
ing milk-sugar as source of carbon, partially and slowly liquefies gelatin, with alkaline
reaction to litmus; odor, very strong, "moldy," characteristic.

This species was obtained from cheese sent from France by Georges
Roger. It was afterwards obtained from other French cheeses, but
does not seem to have any economic importance. It is closely
related by cultural characters as well as morphology to P. camem-
berti, with which it shares the ability to grow normally in fluid off ering
lactose as the source of carbon, but differs in its short conidiophores,
diverse habit upon potato agar and gelatin, and its strong charac-

aThe author is indebted to Prof. H. R. Monteith, of the Connecticut Agricultural
College, for much assistance in preparing the Latin diagnoses.

PENlCll.l.ir \l r.ll'nKMK.

55

teristic smell. It is perhaps intermediate in character between
P. camerriberti and the group of forms designated as /'. commune in
this paper.

CULTURAL HATA.

Color, white i" gray-green, gray, or drab in some media, all brown or < I ra 1 . when old;
reverse cream ; color in media, none.

Odor, very strong peculiar "moldy" odor, typical of this Bpecies under nearly all
conditions.

Fig. \$.—PeniciIlium hiforme: a,b,g, branching of conidial fructification from potato-agar culture ( 1,400);
c, d, e, /, conidiiferous cells an<l roni'li i > MOO); h,j, k, sketches of conidial fructifications on potato
agar (X 140); /, m, sketches of conidial (ructifications on sugar gelatin I < 140); o, r, germination of
conidia (X 900).

Fifteen per cent gelatin in water, typical floccose colony: liquefaction, partial in

old acidified culture-, none in L5 days; litmus read ion, strongly alkaline ( in nearly all
media). Potato agar and bean a_rar, colonies consisting only of very short conidiophores
green to drab in color, little or no floccose mycelium. Potato plugs, typical. < 'ohn's
solution. bIow, half normal growth, with odor.

Synthetic fluid ( Dux's i, carbon supplied as: Cane Bugar, typical up to 50 per cenl
sugar, with acid reaction. Lactose :i per cent, rich growth, typical. Lactic acid 0.9

56 CULTURAL STUDIES OP SPECIES OP PENICILLIUM.

per cent, typical, alkaline reaction when old, crystal drops. Levulose 3 per cent,
typical. Galactose 3 per cent, slow development, typical. Glycerin 3 percent,
t ypical. Potato starch 3 per cent, typical colony, crystal transpiration drops. Butter-
fat, typical colony, mycelial mass tinged violet.

Milk, rich growth; curdling (0.25 per cent calcium chlorid added), in 8 days; diges-
tion, rather slow; color in milk, none.

At :!7° ('., no growth, slowly green when cooled; check at 20 ° C., typical.

PENICILLIUM COMMUNE n. sp.

In examining numerous Petri-dish cultures made for the examina-
tion of milk by the bacteriologists of the Storrs Experiment Station,
large numbers of colonies of Penicillium have been studied. A very
large percentage of these colonies have a series of common characters
which are constant enough to mark out a species, or, perhaps better, a
group of races, between which differences are either minute or so
complicated by the occurrence of other races with overlapping char-
acters as to make their separation a matter of considerable doubt.
One of these has been selected as the basis of the following diagnosis.
This form is morphologically closely similar to P. expansum (see
figs. 1 and 19). It, however, lacks entirely the ability to form core-
mia and fails to discolor the substrata, but grows well in certain cul-
ture solutions which markedly restrain P. expansum, of which it might
possibly be regarded as a variety. From its abundance in the situa-
tions studied it has been designated as P. commune.

Latin diagnosis. — Coloniis in gelatina vel agaro Solani tuberosi aut phaseoli cultis,
viridibus, demum brunneolis, in substrato late crescentibus, zonatis; marginis cres-
centis parte aeria ex conidiophoris, centri atque ex hyphis plus minusve floccosis
composita; reverso et substrato incolorato; conidiophoris plerumque 300/i raro usque
700/.( longis; fructibus conidicis 100-200/j longis, cum ramis alternatis et verticillatis
confertis, basidiis 8-9X3/Z cum apicibus brevibus acutis, catenas conidiorum longas
parallelis gerentibus; conidiis primum cylindricis vel ellipticis, demum globosis,
3-4/t diam., 5-6/* incrassatis germinantibus, levibus, viridibus, in catenis manentibus
submersis; coloniis in gelatina in parte lente liquefacientibus; odore mucidis.

Habitat in lacte, caseo, etc., Storrs, Conn.

Colonies in gelatin or in potato or bean agar, dull green, becoming brown when old,
broadly spreading, zonate, with broad white growing margin composed only of conid-
iophores, in the older parts becoming floccose masses of interwoven hyphae; reverse
of colony and substratum never colored ; conidiophores commonly 300/« or less in length,
sometimes up to 700// ; conidial fructifications commonly 100-200/i in length, compact
at base and broadening above, variously branching with branches appressed, and verti-
cils of conidiiferous cells 8-9 by 3/<, abruptly narrowed to produce conidia; conidia
cylindrical to elliptical and finally globose 3-4//, becoming 4-5/t or larger in germinat-
ing, smooth, green, persisting in chains in fluid mounts; colonies liquefy gelatin slowly
or partially, softening rather than producing clear liquid, alkaline in media without
sugar but acid with either cane sugar or lactose, having a strong " moldy" odor.

Habitat, common in food, dairy products, etc., Storrs, Conn.

PENlcil.l.ir.M COMMUNE.

57

i l I li i; \1 DATA

Color green, becoming brown when old; reverse crea r white; color in media,

none.

i >dor, strong, " moldy."

Fifteen percent gelatin in water, g 1 groM th; liquefaction, nunc in 15 days, slow or

partial in older cultures; litmus reaction, Blowly alkaline. Potato agar and bean agar,
characteristic Potato plugs, characteristic. Raulin'a fluid, characteristic, with
odor "moldy." Cohn's solution, small colonies, trace of pink below.

Fig. 19. PenidHium commune: a, b, c. d, e, conidial fructification, branching, and production ofconidia
(X 900); /. <j, li, j. k, I, sketches <>f frucl iflcal ions in \ ai lous stages I • 1 10).

Synthetic fluid I Dox'e . carbon supplied as: Cane sugar, typical culture, transpired
drops crystal. Lactose 3 per cent, typical culture. Lactic acid O.i) per < •< -nt, good
growth. Levulose 3 percent, good culture, strong odor. Galactose 3 percent, good
culture, strong odor, alkaline reaction. Glycerin 3 per cent, typical, Btrong odor.
Butter Eat, typical growth.

Milk, good typical colony; curdling (<>.•_'•"> per cent calcium chlorid added i in 1 week;
digestion, slow; color in milk, none.

At 37° C, no growth, grew upon cooling; chock at 20° •'.. rich growth.

58 CULTURAL STUDIES OF SPECIES OP PENICILLIUM.

PENICILLIUM No. 22.

Colonies in gelatin or agar gray-green or glaucous persistently, or becoming gray,
not brown, otherwise appearing as P. commune; conidial fructifications more loosely
branching, with branches divergent; conidia larger and lighter color; odor, none or
indefinite; reactions as in P. commune.

Habitat: Isolated several times from domestic soft cheeses made in the State of New
York, 1904-5; found associated with P. camemberti; in appearance and color resembling
latter species, but in structure of colony, measurements of conidiophores, etc., resem-
bling P. commune. In pure culture this form has maintained its identity clearly for
four years.

CULTURAL DATA.

Color gray-green; reverse colorless; color in media, none.

Odor, none or very slight.

Fifteen per cent gelatin in water, good growth; liquefaction, none in 15 days, partial
in 2 to 3 weeks or more; litmus reaction, blue. Potato agar and bean agar, good typical
colonies, with alkaline reaction to litmus. Potato plugs, good growth, typical gray-
green colony with crystal drops of exuded water. Cohn's solution, medium growth,
trace of pink in media.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, good growth up to concen-
tration of 60 per cent, acid reaction. Lactose 3-10 per cent, typical. Levulose3 per
cent, good growth. Galactose 3 per cent, good growth, typical. Glycerin 3 per cent,
good growth. Butter fat, rich growth, typical.

Milk, typical colonies; curdling (0.25 per cent calcium chlorid added) in 8 days;
digestion, rather slow; color in milk, none.

At 37° C, no growth, grew when cooled; check at 20° G, typical.

PENICILLIUM CHRYSOGENUM n. sp.

Latin diagnosis. — Coloniis in gelatina vel agaro Solani tuberosi aut phaseoli cultis,
griseo-viridibus, demum brunneolis, in substrato late crescentibus, margine sterili
lata juvenilibus parte aeria ex conidiophoris et caespitibus sparsis hyphorum ad-
scendentium composita; reverso incolorato; conidiophoris plerumque singulatim
usque 300X4/£ orientibus, raro brevibus ex hyphis assurgentibus ramosis; fructibus
conidicis 100-200 longis cum 1-2 ramis alternatis et divergentibus ramulos 1-2 verti-
cillatos gerentibus; basidiis 8X2.5/4 verticillatis ex apicibus ramulorum, catenas
divergentes conidiorum gerentibus; conidiis primum cylindricis vel ellipticis, demum
globosis, 3-4 diam., pallide glaucis, magnis vacuolis; coloniis gelatinam liquefac-
ientibus, alkalinis lacmo; lactem, panem, gelatinam, auream colorantibr.s.

Habitat, in caseo, pane, etc., commune.

Cultivated in gelatin, or bean or potato agar, gray-green, becoming brownish when
old; aerial portion consisting of conidiophores with some tufts of trailing aerial hyphae;
broadly spreading in the substratum with a wide sterile margin when young. Reverse
of colony not discolored. Conidiophores mostly arising separately, up to 300 by 4/i,
partly arising as short branches of aerial hyphae; conidial fructifications 100-200/( in
length, with 1-2 alternate divergent branches, bearing alternate, verticillate or twice
verticillate branchlets. Conidiiferous cells 8 by 2. 5/* verticillate at the ends of branch-
lets bearing divergent chains of conidia. Conidia cylindrical or elliptical at first,
then globose, 3-4/t in diameter, pale green, with large vacuoles. Colonies liquefy
gelatin, with alkaline reaction to litmus and produce in milk, bread, gelatin, and
other substances a golden yellow color (from which the name).

Habitat, in bread, cheese, etc., apparently common and appearing in several
varieties which differ in the intensity of color production, in appearance in certain
cultures, but which are so far scarcely distinguishable by structural characters.

PEN K'll.I.ir.M (HRYSOGEN I'M.

59

The culture here named P. chrysogenum has been kept under
observation for more than four years without change. In this time
several forms have been collected or sent to this Laboratory which
also produce the golden color in the digestion of milk suggestive of

the name proposed. The cultural characters of these races differ

in some degree in several eases, in others substantial identity has
been observed. In reporting cultural data three numbers are

included for Comparison with this, viz, Nos. l'">, 35, and II. These

agree in the following characters: Spreading habit, surface mostly

l [Q 20.— Penicillium chrysogenum: a, l>, e, d, >, branching of conidial fructification from gelatin plates
(X 900); /, g, I'.j. I. m, sketches of conidial fructifications from potato-agar plates 140); n, o, germination
ofconidi .

of conidiophores averaging about 300/f in length, nearly the same
shade of green in color, similar morphology of conidial fructification,
digestion of milk, gelatin, etc., with the production of yellow or
golden color, liquefaction of gelatin progressive but slower than the
expansion of the colony in the subsl ratum so that a gro^ in r border
of submerged vegetative hyphffl extends into hard gelatin for con-
siderable time, in contrast to forms in which the colony becomes a
floating "island" in a pool of fluid within the first week. In spite
of these common characters colonies of these four races often show

60 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

great apparent differences in parallel culture in some specialized
media. It seems preferable to let this name stand in a broad enough
sense to include the forms having the common characters above
noted than to attempt any narrower delimitation of this group of
forms at this time.

Serial No. 57. — Another form which may be included temporarily
with this group does not produce a yellow color in the substratum
at all, but produces mycelium of orange color as seen from below.
The surface growth is clear green without definite differences from
the clirysogenum group, but the orange color in reverse remains as
a constant difference from that group, unattended by any coloration
of the medium.

CULTURAL DATA.

(Cf. Nos. 25, 35, and 44 in tables.)

Color gray -green, green, to brown when old; reverse colorless, or slight yellowish.

Color in media golden yellow in certain media, no color in others.

Odor, none.

Fifteen per cent gelatin in water, good growth, yellow color in gelatin; lique-
faction rapid — 6-10 days, more or less rapid and complete in all gelatin media used;
litmus reaction, alkaline. Potato agar and bean agar, good colonies, but no yellow
color. Potato plugs, typical, potato becoming yellow. Raulin's fluid, typical with
yellow transpired drops, but no yellow in fluid. Cohn's solution, growth, but not
normal.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, good growth up to 30 per
cent, reaction acid, fluid colorless. Lactose 3 per cent, fair growth, not heavy, fluid
yellow, with acid reaction. Lactic acid 0.9 per cent, some growth. Galactose 3
per cent, good growth. Glycerin 3 per cent, colony white to green, no yellow color
in fluid. Potato starch 3 per cent, good growth, yellow drops above, yellow in fluid.
Butter fat, rich growth, with fluid golden yellow.

Milk, rich growth ; curdling (0.25 per cent calcium chlorid added) in 1 week ; digestion
rapid ; color yellow.

PENICILLIUM RUGULOSUM n. sp.

Latin diagnosis.— Coloniis in gelatina vel agaro phaseoli cultis, flavo-viridibus, dein
viridibus, demum atro-viridibus, late crescentibus in agaro; parte aeria ex conidioph-
oris creberrimis et hyphis aereis et paucis composita ; reverso luteo et in parte aurantiaco
imprimis in tubere Solani; conidiophoris 100-200 X 2. 5-3/t, singulatim vel ex hyphis
aereis prope substratum orientibus; fructibus conidicis 100-150/( longis (in saccharo
niultolongioribus) exramisl0-15x2.5/u, compacte verticillatis, verticillos basidiorum,
vel ramulorum, vel ramulorum et basidiorum eodem verticillo gerentibus; basidiis
9-12X2/*, acuminatis, catenas longas et divergentes conidiorum gerentibus; conidiis
3.4-3. 8X-2.5-3,«, ellipticis, viridibus, uno apice incrassato, verruculosis maturis, in
catenis manentibus submersis, 5/t diam. incrassatis germinantibus; coloniis non (vel
solum in parte et lente) gelatinam liquefacientibus.

Commune in culturis, Storrs, Conn.

Cultivated in gelatin or bean agar, yellowish green, then green, at length dark green;
surface growth of densely crowded conidiophores with few aerial hyphse interspersed
at their bases; reverse of colonies yellow to orange in spots, especially upon potato
or upon sugar media; substratum not or slightly yellowed; conidiophores 100-200
by 2.5-3/(, arising separately or branching from aerial hyphse just above the substratum;
conidial fructifications 100-1 50/i in length, consisting of appressed, verticillate branches

l'KMt m.i.h.m i;n;ri.(isr.M.

61

10-1.") ] .%• 2.5/i, bearing \ erticils of conidiiferoue cells, of branchlets, or of conidiiferoue
cells and branchlets together; conidiiferous cells 9 L2 by 2/x, acuminate, bearing I
divergenl chains of conidia; conidia 3.4 3.8 by 2.5 :'>,», elliptical, green, mostly with
swelling at one end, verruculose when ripe, sw tiling to •">,"- and germinating by one
or two i ii 1 »• — : colonies do not or only partially liquefj gelatin.

Common in cultures, Storrs, Conn. Characterized by its verrucose spores and the
brilliant color of the mycelium viewed from below.

■ ; l Tl RAX DATA.

Color green or yellowish green; reverse yellow t" orange or reddish orange; color
in media, none.

Fifteen per cenl gelatin in water, bIow development; Liquefaction, none or slight
in old colonies; litmus read inn neutral. Potato agar, good growth, alkaline. Bean
agar • 5 per cenl cane BUgar, rich heavy growth producing conidia in heavy dark-
green masses, easily broken oft by shaking the tube. Potato plugs, green, reverse
bright orange.

Fig. 21. Pi » am: a,b, branching of conidiophore(Xl, 600); c,d,c, conidiiferous cells (basidia)

and conidia I ■ 1,600); f, fully ripe conidium, showing del rwalls(x 1,600); g,h,j, swell-

ing and germination of conid > 600); I, m, diagrams of conidial fructifications (X 260

Synthetic fluid (Dox'e . carbon supplied as: Cane sugar, growth in concentration
up to 60 per cent, acid. Lactose, 10 per cent, slight growth. Levulose, Pair growth,
acid reaction; galactose, good growth; glycerin, slow growth; butterfat, slo^ ly typical
colony, yellow belo? : fluid, slightly yellowish.

Milk, growth typical, reverse yellow to orange or reddish orange; curdling (0.25
per '''Hi calcium chlorid added) in 9 days; digestion bIow or .-li,Lrlii; color, none in
milk.

At37°C, no growth, grew when cooled; check at 20° C, typical.

PENICILLIUM CITRINUM n. sp.

Latin diagnosis. Coloniis in gelatins vel agaro Solani tuberosi aul pha eoli cultis,
aeruginoso-viridibus, demum fuligineis; fructibue \ iridibus usque ad marginem j^« •.- 1 i - ,

i. e. margine sterile angustissima; coloniis in gelatina rotundis, parvis, cito liqUefa-

62

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

centibus ; in agaro latioribus ; parte aeria ex conidiophoris et fructibus eonidicis creber-
riniis eomposita, interdum eaespitibus paucis hyphorum adscendentum in medio;
reverso incolorato; conidiophoris (sine ramis) non longioribus 150/<, singulatim orienti-
bus, aut paueis ex hyphis adscendentibus ramosis; fructibus eonidicis 3-5 ramorum
16-30X3,u, apice 5fi incrassatorum, in verticillo, basidia in verticillis compactis
gerentum; utroque verticillo catenis conidiorum in columno compacto 50-150/t
longos adhaerentibus; basidiis 6-7X2-3/*; conidiis globosis, 2.4-3/x raro 3.5/t diam.,
aeruginoso-glaucis, granulatis intus, in catenis manentibus submersis.

Coloniis, saccharo commixto, substrata citrina in colore efficientibus (ubi nomen).

Habitat, in caseo, pane, etc., commune (?).

Colonies grown upon gelatin and potato or bean agar blue-green when young,
becoming dark brown when old, with colored fruit borne almost to the very margin

mm

mm

FlG. 22. — Penicillium citrinum: a, b, e, /, </, branching of conidial fructification, showing number of
branches in each verticil and enlargement of ends of branches (a X 900, others X 1.600); c, d, n, eonidiif-
erous cells and the formation of conidia (X 1,600); h, j, k, I, m, sketches of conidial fructifications
(X 140); o, s, t, germination of spores (X 900).

of the colony, so that the white border of submerged mycelium and uncolored fruit
is very narrow; restricted in growth to a few millimeters in diameter upon gelatin,
but becoming much larger upon agar; aerial part of colony consisting of densely
standing conidiophores and conidia except in the center, where there arise a few tufts
of trailing aerial hyphae. Reverse of colony itself colorless or only yellowish. Conidio-
phores arising separately, rarely longer than 150/*, branching acropetally from sub-
merged hyphae radiating from the center of the colony, or branched from the hyphae
of the central aerial tuft. Conidial fructification a verticil 2 to 5 branches 16-30 by 3/z
enlarged at apex to 5/*, each producing a compact verticil of conidiiferous cells bearing
chains of conidia massed together into columns 50-150/* in length (usually 80-100/i).

PENICILLIUM NO. 37. 63

The whole fructification appears in this way double, t riplo, or quadruple or even
more complex by a secondary verticil from the central branch. Conidiiferous cells
6-7 by 2-3/1. Conidia globose when ripe, 2 I 3^ (even 3.5/a diam. in cane-sugar cul-
tures) in diameter, bluish-green, Blightly granular in contents, adhering in chains
in fluid mounts, losing vitality rapidly with change of color in old colonics. Colonies
liquefy gelatin rapidly, so thai they lie in pool.- of liquid within a week. Litmus
reaction in plain gelatin, Btrongly alkaline. Produces a lemon-yellow color soluble
in alcohol in media containing Bugars, milk, gelatin, bread, and potato.

Found in cultures from milk and cheese, probably cosmopolitan.

Could this be P. dtreo-niffrum Dierckx?

CULTUKAL DATA.

Color bluish green, becoming dark brown when old if exposed to light; reverse
colorless or yellowish; color in media lemon-yellow when cane sugar or gelatin or
peptone is present, none in some media.

< >dor, none.

Fifteen per cent gelatin in water, characteristic growth, becoming white by sec-
ondary sterile mycelium; liquefaction rapid, colonies floating in 5-(» days; litmus
reaction alkaline. Potato agar and bean agar, colonies spreading more than upon
gelatin, agar not colored or slightly colored. Potato plugs, typical growth, potato
colored yellow, with yellow drops transpired; Raulin's fluid, typical growth, very
slight yellow color. < John's solution, good growth, no yellow color.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, good growth with yellow
color up to 30 per cent, with acid reaction. Lactose 3 per cent, slow poor growth
giving a violet tinge instead of yellow color. Lactic acid 0.9 per cent, slight growth.
Levulose 3 per cent, small characteristic growth. Galactose 3 per cent, good growth,
reaction neutral or slightly acid. Glycerin 3 per cent, very small colonies, no color
in medium. Alcohol, 5 per cent, good growth, no yellow. Potato starch 3 per cent,
good growth, slight if any yellow. Tartaric acid, slow colorless colony. Butterfat,
rich growth, lemon-yellow fluid.

Milk, rapid growth; curdling (0.25 per cent calcium chlorid added) in 10 days;
digestion rapid; coloration pale yellowish.

At 37° C, slow growth, white colony, no color in medium; check at 20° C, rich
growth, green, yellow in medium, in bean agar with cane sugar.

PENICILLIUM No. 37.

(Var. of P. citrinumt Or allied to P. citrinumt)

Colonies in media without sugar, green, gray-green, or gray; with sugar persistently
green ; surface velvety strict, composed of short crowded conidiophores up to 100/x in
length, branching from closely woven mycelium partly submerged, partly aerial,
with margin narrow, nol widely spreading in the substratum; reverse of colony and
substratum nol colored or creamy ; coni dial fructifications sometimes a single verticil
of conidiiferous cells, sometimes 2 to 1 verticillate branches; chains of conidia from

each verticil forming a <olunm up to ;>(ii> i;niw( in length in sugar dia; branches

of fructification 13 II by 2 2.5^ enlarged a1 apex; conidiiferous cells 8 10 by 2+^
abruptly narrowed into sterigmata, usually ti It) in each verticil; conidia broadly
elliptical to globose, 2.5-3/* al first becoming I '><< before germinating, thin-walled,
smooth, pale yellowish green, germinating by a single tube; colonies liquefy gelatin
rapidly (6 to 7 days), with strong alkaline reaction to litmus.

Received from Prof. P. II. Rolfs, .Miami, I'la., in culture upon bean stems, 1905.

Allied to P. cilrinum by morphology and culture reactions, but differing in lacking
the power to color media yellow ami in its greater dependence for typical growth
upon the presence of cane sugar.

64

CULTUBAL STUDIES OF SPECIES OF PENICILL1UM.

CULTURAL DATA.

Color light blue-green, olive, or gray in various media; reverse, white or cream;
color in media, none or slightly yellowish.

Odor, none.

Fifteen per cent gelatin in water, small olive-green colonies; liquefaction, rapid —
6 to 7 days; litmus reaction, alkaline. Potato agar and bean agar, typical. Potato
plugs, typical. Raulin's fluid, typical. Cohn's solution, slow and weak-growing
colony.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, good growth even up
to 60 per cent, acid reaction. Lactose, 3 per cent, slow abnormal growth. Lactic
acid, 0.9 per cent, small but characteristic colony. Levulose, 3 per cent, small colo-

Fig. 2Z.—Penicillium No. 37: a, typical branched fructification, two verticils of conidiiferous cells (XI, 600);
6, c, conidiiferous cell and conidia (X 1,600); d, e, sketches of conidial fructifications from potato-agar
culture (X 140); /, g, sketches from an old culture on 3 per cent cane-sugar agar, showing simple (g) and
branched (/) forms (X 140); h, j, fc, branching of conidiophore, swollen ends of branches (X 1,600); I, m,
germination of conidia (X 1,600).

nies. Galactose, 3 per cent, typical. Glycerin, 3 per cent, small growth. Potato
starch, typical. Butterfat, slow growth, deep heavy green colony.

Milk, curdling (0.25 per cent calcium chlorid added), rapid;0 digestion rapid and
complete; color, none.

At 37° C, killed; check at 20° C, grew well.

PENICILLIUM No. 12.

This form differs from P. citrinum in producing no coloration of the medium and
in producing conidial fructifications in which the chains of conidia are more or less
divergent instead of aggregated into columns. In culture there is general corre-

ct The time of curdling is almost impossible to determine in cases where digestion
begins quickly and progresses rapidly.

I'l.Xh'lU.ir.M A.TRAMENTOSUM. 65

Bpondence in reactions, withoul identity; this form appears to be much more
dependent upon cane sugar for the production of typical color of the conidia and
growth than is P There is al eater tendency to the production

of a layer of mycelial hyphse jusl above the surface of the substratum, from which the
conidioph ires arise as aerial branchi

This form was received from Prof. C. E. Marshall, Agricultural College, Michigan
under i be uame of /'. glaucum.

(Mil i: Al DATA.

Color pale blue-green; reverse of colony cream, aol colored; color in media, none.

( (dor, uone.

Fifteen per cenl gelatin in water, rather small pale blue colonies, rapidly becoming
white by secondary sterile growth of hyphse; liquefaction, rapid- 6 days; Litmus

reaction, alkaline. Potato agar, as in gelatin. Potato plugs, very p ■ growth,

grayish or yellowish green. Raulin's Quid, slow bul typical colonies, delicate l>lue;
Cohn's solution, slow and restricted grow th.

Synthetic fluid (Dox's), carbon supplied as: Cane Bugar, grows well in concentra-
tions up i" 60 per cent. Lactose 3 per cent, small colonies Lacking nourishment.
Lactic acid 0.9 per cent, small colonies floating in fluid. Le> ulose 3 per cent, good
growth, alkaline reaction. Galactose 3 per cent, good growth, alkaline reaction.
Glycerin 3 per cent, slow-growing colonies, becoming gray-brown when old. Potato
starch, good colonies. Butterfat, b1o\s and ill nourished growth.

Milk, rapid growth; curdling (0.25 per cent calcium chlorid" added) in 7 days; diges-
tion rapid and very complete; color in milk, none.

At 37 C . killed in 6 days; at 20° C, good growth.

PENICILLIUM ATRAMENTOSUM n. sp.

Latin diagnosis. — Coloniis in gelatina vel agaro Solani tuberosi au1 phaseoli cultis,
viridibus, parte aeria plerumque ex conidiophoris singulatim orientibus, medio cum
hyphis aereis interspersis, margine alboexhyphis fertilibus angusta; reverso incolorato
vel parum ochraceo; substrato au< incolorato au1 in substratis saccharin is et in lacte
atrobrunneo tarde fere atro; conidiophoris 240 300 usque 400/i Longis; fructibus conidi-
cie LOO usque 200/i longis, ram is I 2 \ erticillatis 2-4 insequaliter Longis in verticillo in
apice incrassatis; basidiis v L0/i Longis, parallelis in verticillo; cutcnis conidiorum
eodem verticillo in columno compactis; conidiis ellipticis, 3.5 I (usque L.8) X2.5-3
usque 3.5/«, laevibus, viridibus, 6— 7/t incrassatis e1 uno tubo germinantibus; coloniis
gelatinam cito Liquefacientibus, alkalinis lacmo; odore in lacte proprio, in substratis
aliis aullo.

Ex caseo cull tun. Siori ' onn., 1905.

AHine P. citrino.

Colonic- upon gelatin or upon potato or bean agar bright green, aerial pan mostly
of simple conidiophores, mixed in older parts with branching aerial hyphse bul nar-
rowly spreading al the margin by new conidiophores onlj . Reverse of colonies shows
a slight production of yellow (ochraceous) color. Conidiophores 240 100/t, averaging
about 300// in length. Conidial fructification up to 200/i in length, usuallj LOO/t or
le.-.-. verticillatelj or twice verticillately branched; branches 2 I in a verticil di-
vergent, unequal in length, swollen al ends, bearing conidiiferous cells. Theconjdial
i bains from each verticil form a dene column, which diverges more or Less from the
other columns when old. Conidiiferous cells 8 LO/t in length, closely parallel. Conidia
elliptical, varying from 3.5 [ft by 2.5 3/i on omewhal Larger in gelatin cultures,

up to L8 bj 3 5/i, smooth, homogeneous green with a slight yellowish made when seen
in mass, swelling to 6 7<< in diameter and germinating bj a single tube. Mycelial
cell- 5 7/< in diameter and up to 30// or more in Lengt h. « oloniee Liquefy BUgar-gelatin

8108— Bull. 118—10 5

66

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

rapidly, give an alkaline reaction to litmus, digest milk, and color potato agar con-
taining high percentage of sugar a deep black.

Found upon Camembert cheese imported from France. Closely related morpho-
logically to P. citrinum, from which it is separated by the longer conidiophores and
larger spores as well as the black discoloration of sugar media.

CULTURAL DATA.

Color deep (blue) green to brown when old; reverse uncolored, or brown in some
media; color in media, none, or brownish to almost black.

Odor, none.

Fifteen per cent gelatin in water, deep green, brown when old, rich growth; lique-
faction, 7 days, varies from 6 to 12 days in other gelatin media; litmus reaction, alka-
line. Potato agar and bean agar, typical, no color below. Potato plugs, dark green,
potato blackened. Raulin's fluid, rather weak growth. Cohn's solution, germinated
only.

Efc

% llgJr

ill*

a ill

771.

Fig. 24. — Penicillium atramentosum: a, b, c, d, branching of conidial fructification showing unequal
length of branches, swollen ends (X 900); e, /, conidiiferous cell and chain of conidia(X 900); g, h, j,
sketches of conidial fructifications (X HO); i, coriidia (X 1,600); m, n, o, r, germination of conidia
(X 900).

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, good growth up to 50 per
cent, with acid reaction in 50 per cent solution. Lactose 3 per cent, small and slow
growth. Lactic acid 0.9 per cent, no growth. Levulose 3 per cent, slowly typical.
Galactose 3 per cent, typical, with alkaline reaction. Glycerin 3 per cent, germina-
tion. Potato starch 3 per cent, good growth, no color in fluid or reverse of colony.
Butterfat, typical, green colonies with reverse brown, and fluid uncolored.

Milk, curdling (0.25 per cent calcium chlorid added) in 9 days; digestion, rapid,
fairly complete; color, brownish to almost black.

At 37° C, no growth, grew when cooled; check at 20° C, typical.

PENICILLIUM No. 24.

(Related to P. atramentosum'!)

Colonies upon gelatin and potato or bean agar blue-green, becoming brown rapidly
when old, or smoky with very dense velvety surface consisting of conidiophores aris-
ing in the substratum or just above its surface, with a very abrupt narrow white

PENIdLLIUM NO. 24.

67

margin of unripened fruit and submerged mycelium during the growing period.
Reverse of colony and mm •ilium colorless under all conditions studied. < kmidiophores
from 100 i" H"1". a> eraging about 250//, in length, either arising separately or as lateral
1 warn In-- of hyphae just above the substratum. Conidial fructification up to 200/j in
length produced by various branching from the conidiophores in which cadi primary
1. ranch is often divergent to produce separate massof conidia. Conidiiferou8cells7 10
by '■[■' Conidia globose 3.3 l/t, homogeneous blue-green, Bmooth, seeming to lose
vitality rapidly under laboratory conditions. Colonies liquefy gelatin in 7 to 12 days
so that they lie in pels of liquid. Litmus reaction strongly alkaline.

Found in the culture- from Camemberl cheese in the laboratories at Men--. Conn.
Differs from the preceding and from P. citrinum by its longer conidiophores, the
alternate branching of its fructification, the Bize of its spores, and by failure to color
the substrata. The relative value of ellipticity of conidia as a diagnostic character
appear- to be questionable. This form is therefore presented under cultural number

— ■ ■

Fig. 25.— Penicillium No. 24: a. h, branching and arrangement of branches in conidial fructification! X no);
c, d, (. conidiiferous cells and conidia < < 1,400); g, h.j, sketches of form and arrangement of conidiophores
(X 140); m, n, germination of conidia (X9O0 .

only, whereas the preceding bas been identified from accidental cultures more fre-
quently, and hence j~ given oame and description as a species.

Mill RAX DATA.

Color deep green (blue-green), becoming brown when old and exposed; reverse
white or cream; color in media, none.

( »dor, cone.

Fifteen per cent gelatin in water, characteristic colony; liquefaction, rapid -
11 days or even less; litmus reaction alkaline, potato plugs, deep blue-green, crystal
drops. Etaulin's fluid, weak hut characteristic growth. Cohn's solution, slow hut
characteristic development .

Synthetic fluid I Dox's), carbon supphed: Cane sugar, good growth up to 30 per
cent, acid reaction. Lactose, ■'< per cent, slow development, uol typical. Lactic
acid 0.9 per cent, good colony. Levulose 3 per cent, small colonies. Galactose :'•

68

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

per cent, good growth, strongly alkaline. Glycerin 3 per cent, weak growth. Potato
starch, characteristic colony.

Milk, curdling (0.25 per cent calcium chlorid added) in 8 days; digestion medium
rapid; color in milk, none or ?

At 37° C, no growth, grew when cooled; check at 20° C, good typical.

PENICILLIUM STOLONIFERUM n. sp.

Latin diagnosis. — Coloniis in gelatina vel agaro Solani tuberosi cultis, viridibus vel
flavo-viridibus, demum griseo-viridibus vel griseis in agaro sine saccharo, cum sac-

W,«i: I://////..-* ,'A J

Fig. 26. — Penicillium stoloniferum: a, b, c. e,f, the types of branching at the tips of the ''stolons" by which
this species spreads in substrata (6, c, e,J, x 900); d, conidial fructification (X 900); h, j, k, I, sketches
oi conidial fructifications of various ages, h and j being characteristic shapes (X 140); g, formation of
conidia (X 900), i, ripe conidia showing minute granulation (X 1,600); m, n, germination of conidia
(X 900); o, rough diagram of habit.

charo viridibus, floccosis, in culturis juvenalibus stolonibus aereis citius quam hyphis
submersis crescentibus, re verso incolorato vel in parte flavo; conidiophoris brevibus
ex hyphis adscendentibus ramosis, usque 100/( longis, aut singulatim orientibus (sine
ramis) plus minus 300/i longis; fructibus conidicis 40-80/* raro usque 170/z longis, ex
ramis brevibus compactis, et basidiis verticillatis, in baside confertissimis, catenas
conidiorum late divergentes gerentibus compositis, interdum ramus infimus tarn
divergens ut fructus duplex videatur; basidiis 10X 3/z; conidiis ellipticis vel paene
globosis, 2.8-3.4/* diam., pallido flavo-viridibus laevibusque; coloniis gelatinam cito
liquefacientibus, alkalinis lacmo.

PENICELLIUM FTJNICTTLOSUM. 69

Habitat, in fungie putreacentibus, Boletis, Polyporis; Storre, Conn.; Paris, Gallia.

Cultivated in gelatin or potato agar, green or yellowish green, becoming gray-green
or gmy when old (remaining green in sugar media . H spreading more rapidly

in young cultures by aerial stolons than by submerged hi phse (i. e., the submerged
mycelium Beems to arise from the aerial rather than vice versa ; reverse of colony nol
colored or partly yellow; conidiophores arising as short branches i LOO/i or less in length l
from aerial hyphse, or arising separately 300/i or more in length especially at the mar-
gins of older colonies; conidial fructification 10 sn more rarely up to 170". in Length,
composed of Bhort appressed branches and numerous conidiiferous cells densely
crowded at the base bearing very loosely divergenl chains of conidia; Bometimes the
lowesl branch diverges bo thai the fructification appears double; conidiiferous cells
10 by ■>;•; conidia Blightly elliptical or globose, 2.8 3.4/i, Bmooth, yellowish green in
mass, almost hyaline by transmitted light; colonies liquefy gelatin \ ery rapidly, with
u strong alkaline reaction to litmus.

Habitat, decaying fungi, Boleti, Polypori; cultures from milk and ensilage. Col-
lected repeatedly at Storrs, Conn.; once upon decaying Boh tus scaber at the Jardin des
Plantes in Paris, hence probably widely distributed. The stolon-producing character
is so easily seen and so characteristic of this species as to seem adequate to distinguish
it from all other species studied. This has been observed upon a decaying Boletus
with a hand lens.

CULTURAL DATA.

Color white to yellowish green, deep green becoming yellowish brown or gray in
old cultures; reverse, not colored (or slight ly yellow i; color in media, none or slight.

< >dor, none.

Fifteen percent gelatin in water, good growth, yellowish green; liquefaction, rapid
in all gelatin media; litmus reaction, strongly alkaline. Potato agar, good growth,

pale green to pray. Bean agar, g 1 growth, pale green to gray. Potato plugs, good

growth, deep green, transpired drops brown. Raulin's Quid, slow but characteristic
lie- Cohn's solution, typical growth.

Synthetic fluid (Dox'a i, carbon supplied as: < lane sugar, good growth up to 50 per
cent, conidial areas persistently preen (viridis to atro-viridis), acid reaction. Lactose
3 per cent, slow abnormal colonies, weak. Lactic acid 0.9 per cent, small colonies be-
conum: alkaline. Levulose 3 per cent . t ypical colonies, alkaline reaction. Galactose
3 per cent, typical, alkaline reaction. Glycerin 3 per cent, pood growth, not heavy.
Potato starch 3 per cent, typical, drops yellow, fluid colorless.

Milk, typical colonic-; curdling (0.25 per cenl calcium chlorid added) in 1 week;
digestion, rapid; color in milk, little or none.

At 37° C, killed; -heck at 20° C, g 1 colony.

PENICILLIUM FUNICULOSUM n. sp.

Latin diagnosis. Coloniis in gelatina vel agaro Solani tuberosi aut phaseoli cultis,
atro-viridibus, late crescentibus, floccosie; parte aeria ex hyphis decumbentibus,
ramosis, csespitosis, late intricatis, e1 fasciculatis, conidiophoros breves gerentibus
interdum hyphos secundarias albas floccosas lente evolvente; reverao rubescente
demun atro-vinoso; aubstrato-aut lacte-aut gelatina, \ inoso; conidiophoris (sine rami- -
2() vn u-'p.e loop longis, plerumque ex hyphis repentibus vel fasciculatis, interdum
Bingulatim orientibus, fructibus conidicis usque L25 L60/i longis, cum 1 , 2 rands alter-
natis, dein ramulis vert icillatis, basidia in vert icilloe densos eaten is coiddiorum paral-
lelis gerentibus; basidiis Hi 1 1 • _' 3/t, parallelis in verticillo, acuminatis; conidiis
primum cj lindricis, demum fusiformibusv el ellipticis, 3-4X2-3/4, viridibus; conidi-
orum catenis solventibus submerais; coloniis gelatinam non liquefacientibus, acidis
lacmo, siccantibus senescentibusque interdum coremiis paucis e\ olventibus.

In cultura, Storre, Conn., 1905; communicavi 1 Dr. E. A. ..Miami, l-'la., 1908.

70

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

Cultivated in gelatin or potato or bean agar, deep green, broadly spreading, surface
closely floccose with procumbent hyphse, tufts and ropes of hyphse bearing lateral
conidiophores; reverse becoming red, purple, or very dark purple, almost black, with
the whole mass of medium colored; conidiphores short, 20-80 or lOO/i, mostly perpen-
dicular branches from trailing hyphae, sometimes arising separately from the sub-
stratum; conidial fructification up to 125 or 160/t in length, with 1 or 2 alternate
appressed branches bearing verticillate branchlets and dense verticils of parallel coni-
diiferous cells 10-14 by 2-3/z; conidia at first cylindrical, then elliptical or fusiform,
3-4 by 2-3jn, green, in chains which break up completely in fluid mounts; colonies
not liquefying gelatin in 2 weeks, with acid reaction to litmus.

Fig. 27. — Penicillium funiculosum: a, b, c, d, c,f, conidial fructifications with conidiiferous cells and conidia
(X 900, except e, 1,600); g, h,j, k, I, m, n, sketches of fructifications, separate and borne upon hyphse
and ropes of hyphse (x 140). o, r, germination of conidia (X 900).

Found in accidental culture, Storrs, Conn., 1905; also received from Dr. E. A.
Bessey, Miami, Fla., 1908. Easily recognized in culture.

CULTURAL DATA.

Color, deep green with secondary floccose masses of mycelium in some cultures;
reverse and color in media, red to very dark red, or, colorless in certain media.

Odor, none.

Fifteen per cent gelatin in water, thin, widespread but characteristic growth; lique-
faction, none or very slight; litmus reaction acid. Potato agar and bean agar, typical

PENICILLIUM DBCUMBENS.

71

colonics with red i olorin medium. Potato plugs, tj pical, reverse of colony and potato
both deep red. Raulin's fluid, good growth, bul uo color in fluid. < John's solution.
germination only.

Synthetic fluid (Dox'e , carbon supplied as: Cane sugar 3 per cent, good growth, but
no red color. Lactose 3 per cent, very little growth. Lactic acid 0 9 per cent, little
growth. Levulose 3 per cent, little growth. Galactose 3 per cent, little growth.
Glycerin 3 per cent, germinated only, grew when sugar was added. Potato starch 3
percent, good colonics, bu1 no red color. Butterfat, rai her small colonies with many
delicate coremia, reverse of colonies red, with uo color in fluid.

Milk, good growth with scattered coremia in old culture-; curdling (0.25 per cent
calcium chlorid added) very Blow about I weeks; digestion slow or slight, no clear
fluid; color in milk, colony deep red below, milk deep red I \ Inosus) at top, shading to
white below, very slowly colored.

Grew about equally well at 37° C. and 20° C.

PENICILLIUM DECUMBENS n. sp.

Latin diagnosis. — Coloniis in gelatina pura vel agaro Solani tuberosi aut phaseoli
cultis, i:risco-glaucis,grisei8, demunbrunneolis-sparsis; in saccharo ofbeinaro commixto
denrior, glaucescentihus; parte aeria ex
hyphis decumbentibus vel stoloniformibus
conidiophoros brevissimos gerentibus,
denum csespitulis albis densis hypharum
sterilium Becundariarum, conspersis ; re-
verso incolorato; conidiophoris 20-100X3,u,
basidiis 7-9X2-3/1, in uno verticillo denso
gerentibus; fructibus conidicis ex catenis
conidiorum primum in columno usque 100/t
longo, mox, incapituloconglutinato solutis;
conidiis globosie, 2.5-3/(, vacuolatis, lsevi-
bus, primum pallide glaueis demum brun-
neolis; coloniis gelatinam non liquefa cien-
tibus, alkalinis lacmo, saccharophilis,
odorem in saccharo evolventibus.

Communicavit, Prof. P. II. Polls, Miami,
Fla., 1905.

( lultivated in gelatin or potato agar, white
to gray, gray-green ultimately yellowish
brown, green in cultures with cane sugar,
surface growth consisting of trailing or stolon-
like hyplue sparseh de\ eloped and so close

to the Bubstratum as to appear only as fer-
tile hyphse, hearing the conidiophores as
short branches 20-100/t in length, in old
colonies with dense tufts of sterile second-
ary mycelium scattered upon the surface;
conidial fructifications consisting of single
verticils of crowded conidiiferoue colls,

7-9 by 2 3/i, bearing conidial chains firsl in loose columns up to LOO/u in length bul soon
becoming enveloped and broken up in the drops of fluid secreted abundantly from the
mycelium (Gliocladmin-like) ; conidia globose 2.5 :;», vacuolate, Bmooth, pale green
then brownish in mass; colonies do not liquefy gelatin; give a weakly alkaline reac-
tion to litmus; produce a definite odor in cultures containing cane sugar.
Contributed by Prof. P. II. Rolfs from Miami, Fla., 1905.

! 1 1, . .'-. PeniciUium decumbent: a, b,c,d, conidial
fructification, a single verticil >>f conldllferona
cells i ■ 900); ftJ.Jt, sketches of conidial fructifi-
cations, wiih diagram ol habit and appearance
of young culture on potato-agar (X 140).

72 CULTUEAL STUDIES OF SPECIES OF PENICILLIUM.

CULTURAL DATA.

Color gray, gray -green, often gray or gray-brown when old; reverse white; color in
media, none.

Odor, distinct in cane-sugar media.

Fifteen per cent gelatin in water, medium growth, gray -green to brown when old;
liquefaction, none; litmus reaction neutral. Potato agar and bean agar, rather small
colonies, weak growth, grayish green to yellow-brown. Potato plugs, white to yellow-
ish brown colonies, very weak growth. Raulin's fluid, rich growth, bright green, dis-
tinct odor. Cohn's solution, germination only.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, rich growth up to 30 per
cent. Lactose 3 per cent, slight growth. Lactic acid 0.9 per cent, medium colony,
light green. Levulose 3 per cent, small greenish colonies. Galactose 3 per cent,
growth, faintly alkaline reaction. Glycerin 3 per cent, germination only. Potato
starch 3 per cent, very slight growth. Butterfat, weak colonies.

Milk, not adapted to this species, colonies grow very slowly; curdling (0.25 per cent
calcium chlorid added) slow — about 4 weeks; digestion, little or none; color in milk,
none.

At 37° C. some growth; check at 20° C. better than 37° C.

PENICILLIUM DIVARICATUM n. sp.

Latin diagnosis. — Coloniis in gelatina vel agaro phaseoli cultis, avellaneis, nun-
quam viridibus, in substrato late crescentibus; parte aria ex hyphis fertilibus intri-
catis, demum fere pulverulenta; re verso incolorato; hyphis fertilibus septatis, ple-
rumque brevibus, repentibus vel adscendentibus; fructibus conidicis aut terminalibus
aut lateralibus ex hyphis fertilibus repentibus ex verticillis sessiles ramorum et basid-
iorum, irregulariter in hyphis fertilibus orientibus; basidiis 15-20X3//, sterigmatibus
longis acuminatis, in baside confertis, apice late divergentibus, catenas longas coni-
diorum gerentibus; conidiis ellipticis vel fusiformibus, 5-7X2.5-3^, avellaneis, 10/z
incrassatis 2-3 tubis germinantibus; coloniis gelatinam non liquefacientibus, alka-
linis lacmo.

Legit, C. Thom, Storrs, Conn.

Cultivated in gelatin or bean agar, yellowish brown (avellaneous), never green,
broadly spreading in the substratum; superficial growth consisting only of closely
woven fertile hyphae, becoming powdery in appearance when mature; reverse of colony
not discolored; fertile hyphge septate, usually short, mostly creeping; conidial fructi-
fications either terminal or on short branches of creeping or partially erect hyphse, con-
sisting of separate conidiiferous cells, of verticils, or of series of verticils of branchlets
and conidiiferous cells irregularly distributed along the fertile hyphae; conidiiferous
cells 15-20 by 3/x, with long acuminate sterigmata, broadly divergent at the apices
and bearing long chains of conidia; conidia elliptical or fusiform, 5-7 by 2.5-3/*, yel-
lowish to brownish, swelling in germination to 10/i and producing 2 or more tubes;
does not liquefy gelatin; litmus reaction alkaline.

Unmistakable when once seen in culture. Found in a mucilage bottle, Storrs,
Conn., 1904. Later contributed by Prof. G. F. Atkinson from North Carolina.

In common with several other forms included in the genus Peni-
cillium, the conidial fructifications of this species are not strictly peni
cillate and terminal. Every gradation is found from fruiting systems
typical of the genus to simple chains of conidia borne by single cells or
basidia upon prostrate or even submerged hyphse. It partakes, how-
ever, of the cultural character of the species of the genus, as shown by
its copious growth upon many different substrata.

PENICILLIUM LILACINUM.

73

i i M i 1; \i. D \ i \

Color light clay to chocolate or yellow (avellaneous, nearl) i to darker, approaching
brownish yellow, never green ; reverse uncolored ; color in media, none.

Odor, none.

Fifteen per cenl gelatin in water, good growth; liquefaction, none; litmus reaction,
acid. Potato agar, bean agar, and potato plugs, typical. Raulin'e fluid, typical.
Conn's solution, germination only.

Synthetic fluid carbon supplied as: Cane sugar, g 1 growth up to 30 per

cent, acid reaction. Lactose 3 per cent, slight growth. Lactic acid 0.9 per cent,
medium growth, not fully normal. Levulose 3 per cent , small development . < ialac-

Fig. 20. —PeniciUium dirarica/um: a, d, e, f, oonidiiferous cells, conidia and their arrangement (X 1,600);
b, c, Irregular typi ngement; g, h, k, sketches of conldial fructification ( < 200); m, //. germination

ol conldfa (x 900).

tose 3 per cent, alow growth, reaction neutral. < Uycerin 3 per cent, germinated only;
grew when sugar was added. Potato starch 3 per cent, typical. Butter fat, weak
growth, bul characteristic fruiting.
Milk, .-low and weak colonies; curdling (0.25 percent calcium chlorid added I slow —

1 1 days or more; digestion, Blow and slight; color in milk, none.

Colonies grew better al 37° C. than at 20° C.

PENICILLIUM LILACINUM n. sp.

Latin diagnosis.- Coloniis in gelatina vel agaro phaseoli cult is, albi.-, vol alias demnm
paliide lilacinie imprimis in saccharo oflicinaro commizto, floccosis; hyphia aereis
ramosis, adscendentibus, eeptatis, 3/z cr., ramos fertiles 1 >r« ■ \ i--ii,iisgerentibus;reverso

74

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

incolorato; fructibus conidicis usque 100 longis, e basidiis sessilibus, solitariis vel verti-
cillatis, aut, e ramis brevissimis vel apicibus hyphorum aeriorum, 1, 2, 3 verticillos
ramulorum et basidiorum, catenas longas et divergentes conidiorum gerentum; basidiis
basidibus incrassatis, apicibus acuminatis et divergentibus, 7-10 longis; conidiis
2.5-3X2/* ellipticis, lsevibus, pallide lilacinis.

Coloniis gelatinam lente liquefacientibus, alkalinis lacmo.
Comm., Prof. G. F. Atkinson et C. W. Edgarton, Ithaca, N. Y.

Cultivated in pure gelatin or bean agar white, white to pale lilac in cultures con-
taining sugars, more or less loosely floccose with hyphse branched, septate, ascending, 3/i
in diameter, producing conidial masses upon very short branches irregularly distrib-
uted, or becoming conidiophores toward the apex; reverse of colony not discolored;
conidial fructifications up to 100/x in length, consisting of solitary, sessile conidiiferous
cells, or verticils of conidiiferous cells, or short branches bearing 1, 2, or 3 verticils of
branchlets and conidiiferous cells with long, tangled chains of conidia. Conidiiferous

cells flask-shaped, divergent at
the apices, acuminate, 7-10/* in
length ; conidia elliptical, smooth,
2. 5-3 by 2fi, thin walled, pale
lilac. Colonies slowly liquefy
gelatin, with strongly alkaline
reaction.

Received from Prof. G. F. At-
kinson and C. W. Edgarton,
Ithaca, N. Y.

A relationship of this spe-
cies to the common green
forms is very doubtful. The
chains of conidia produced
break up so quickly and
completely in mounting in
fluid for examination that it
is often difficult to find even
a single conidium attached
to its sterigma. The hyphae
with branches and basidial
cells, aside from the produc-
tion of long conidial chains,
might readily be placed in any one of several hyphomycete genera.
The form of conidial fructification varies from a single conidiiferous
cell or basidium with a chain of conidia upon an aerial hypha to a
single verticil, or a branch with two or three successive verticils and
even to a terminal fructification allying it with the typical penicillate
forms.

CULTURAL DATA.

Color white to a characteristic lilac shade; reverse of colony white; color in media,
none.

Odor, none.

Fifteen per cent gelatin in water, fair growth, not heavy, white; liquefaction
rather slow — 14-16 days, litmus reaction strongly blue Peptone milk sugar gela-

FiG. 30. — Penicillium lilacinum: a, b, c, short conidiophores
and verticils of conidiiferous cells showing the various branch-
ing and arrangement of cells (x 1,600); d, conidiiferous cell,
solitary and sessile on an aerial hypha, not uncommon in
this species (X 1,600); e, conidia (X 1,600); /, g, ft, sketches of
conidial fructifications, varying from a single chain to a typi-
cal penicillate form (X 260).

PENIt'ILLUM INI l!I( A I I'M.

75

tin (Conn's), liquefied in 2 week.-, white colonies. Potato plugs, white colony;
Culm'- solution, verj weak growth but typical lilac color.

Synthetic fluid (Dox's), carbon applied as: Cane sugar 3 percenl to 30 percent,
good growth with typical lilac color, alkaline reaction, do fermentation. Lacto
percent, germination only, slighl growth. Lactic acid 0.9 per cent, germination only.
Levulose 3 per cent, slow growth. Galactose 3 per cent, Blow development with
alkaline reaction. Glycerin 3 per cent, nol characteristic, little more than ger-
mination. Butterfat, typical colon) gh ing brownish color to fluid and causing drops
of j ellow oil to separate oul .

Milk, curdling, Blow ; digestion, Blow; color in milk. none.

At 37 C, l»-i growth; at 20 ' C, g 1 growth.

PENICILLIUM INTRICATUM n. sp.

Latin diagnosis. Coloniie in gelatina vel agaro phaseoli cultis, albis, griseis, griseo-
glaucis, demum griseis, lente fere fuligineis, floccosis; zonatis; parte aeria usque 1-3
mm. cr., exhyphis aereis ramosis
dense intricatis; reverso incolo-
vel sulphureo interdum
lente avellaneo; substrato buI-
plmreo colorato; conidiophoris
interdum terminalibus plerum-
que ex hyphis aereis brevibus
30-50/1 ramosis; fructibus conid-
icie 50 loo,, usque 1 tO/i longis —
muli" longioribus in substratis
Baccharinis — ex verticillo basidi-
orum, vel ex 1 '■'> verticillis ba-
Bidiorum in rami- divergenti-
bus, vel ex verticillis ramulorum
i'l basidiorum eodem verticillo,
catenis conidiorum saepe co-
lumns laxe convergentibus; ba-
sidiis8-10X- 2.5/t, paucis(4-10)
verticillo, cum catenis basidi-
orum divergentibus; conidiis el-

lipticisvel globosis, hyalinis, vel pallide glaucis, 2.5-3// diam., Isevibus, leptodermibus,
intue granulosis, in catenis manentibus Bubmersis; coloniis gelatinam nonliquefacien-
tibus, alkalini- lacmo.

Culturae ex bumo, Prof. W. M. Eaten, Storrs, Conn., 1907.

I I roii up'. n gelatin or bean agar, white, gray, greenish gray, when eld gray or
Bmoky, floccose, becoming a mass of interwoven byphse and ropes of hyphse 1 '■'■ mm.
in thickness; reverse of colony and substratum uol colored in bean agar, more or less
sulphur yellow or even brownish in sugar media; conidiophores sometimes terminal,
more commonly branches of aerial byphse 30 50/* in length; conidial fructifications
">o loo ii] > to 1 Hi" in length, or much longer in old sugar cultures, consisting of simple
verticils of conidiiferous cellB, or of I '■'• verticil upon divergent branchlets, or of
branchlets and conidiiferous cells in the same verticil; conidiiferous cells 8 L0 by
2 2.5ft, few i l to 10) in each \ ert icil, bearing more ot less divergenl chains of conidia
frequently aggregated into a loose column; conidia elliptical or globose hyaline or
pale greenish, 2.5 3/i diameter, smooth, thin walled, granular within, remaining in
chains in fluid mounts; colonies alkaline to litmus, Dot liquefying gelatin.

Found in cultures from soil, Storrs, Conn., by Prof. \\ . M. Eaten, l!t()7.

Fig. 31. — Penicillin in intricatum: a, 6, c, conidial fructification,
conidiiferous cell, conidial chain (x l.f.00); d, e, I, sketches ol
conidiophores, branching and arrangement (X 2i

76

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

CULTURAL DATA.

Color white or greenish gray — not green, grayish to brown or drab; reverse and
medium uncolored or sulphur-yellow in some media; litmus reaction strongly alka-
line. Potato and bean agar, typical. Potato plugs, weak growth, not adapted to
this species. Cohn's solution, weak growth, yellowish-green colonies.

Synthetic fluid (Dox's) carbon supplied as: Cane sugar, grew well up to 30 per cent.
Lactose 10 per cent, good colonies. Levulose 3 per cent, good growth, yellowish
mycelium, fluid yellowish, alkaline. Galactose 3 per cent, typical alkaline reaction.
Glycerin 3 per cent, slow growth. Butterfat, good growth, reverse yellow, fat little
changed.

Milk, fruiting areas upon glass, mycelium in contact with milk sulphur-yellow;
curdling (0.25 per cent calcium chlorid added) very slow — 3 weeks; digestion very
slow in normal or acid milk, rapid when alkaline; color in milk, none.

At 37° C. grew more rapidly than at 20° C.

PENICILLIUM SPINULOSUM n. sp.

Latin diagnosis. — Coloniis in gelatinavel agaro phaseoli cultis,atro-viridibus, demum
fere atris, cito et late in substrato crescentibus, margine sterili lata juvenilibus; parte

aeria ex conidiophoris
et ex hyphis floccosis
sparsis composita ; re-
verso incolorato ; conidio-
phoris 105-300 X 3-3. 5/i,
vel longioribus, apice
b/i incrassato, verticillum
basidiorum 9.5-11X2-3^
gerente; fructibus coni-
dicis in columno denso
300 usque 500X15-30/*
ex catenis conidiorum
compositis ; conidiis pyri-
formibus vel globosis,
3.2-3.5X3.6-4/(, lepto-
dermibus, primum lsevi-
bus demum ministissime
spinulosis; coloniis gela-
tinam lente liquefacien-
tibus, acidis lacmo.

In cultura in labora-
torium, Hannover, Ger-
mania.

Cultivated upon gelatin or bean agar, deep green, spreading broadly in the sub-
stratum with broad sterile margin when young; aerial portion consisting of conidio-
phores and scattered aerial hyphoe; reverse of colony not discolored'; conidiophores
150-300/t or longer by 3-3.5^, with apex enlarged to 5/i in diameter, bearing a single
verticil of conidiiferous cells 9.5-11 by2-3u; conidial fructification a close column of
conidial chains up to 300 or eve,n 500/i in length by 15-30/*; conidia pyriform to glo-
bose, 3.2-3.5 by 3.6-4/i, very thin walled, smooth at first then delicately spinulose
or verrucose, yellowish green then almost smoky; liquefying gelatin slowly, with
strongly acid reaction.

Found as a contamination of another species of Penicillium obtained in Doctor
Wehmer's laboratory at Hanover, Germany. Easily recognized and cultivated.

Fig. H2.— Penicillium spinulosum: a, b, conidial fructifications consisting
of single verticils of conidiiferous cells (X 900); c, conidiiferous cell with
chain of young conidia smooth (X 900); d, f, ripe conidia, delicately
echinulate (X 900); e , swollen end of conidiophore bearing conidiiferous
cells (X 900); g, h, sketches of conidial fructifications (X 1,400).

PENICILLIUM NO. 28.

77

X?

i I Ml I! \l. LATA.

Color deep 'lull green; reverse cream, or Blight traces of pink or violet; color in
media, none.

( (dor, aone.

Fifteen per cen! gelatin in water, .-low but typical; liquefaction, rather slow and
variable; litmus reaction, acid. Potato and beau agar and potato plugs, typical,
producing a very heavy layer of dark-green conidia when cane Bugar is added. Etau-
1 in's fluid, typical. Cohn's solution, germination only.

Synthetic fluid (Dox'g . carbon supplied as: Cane sugar, grows well up to 50 per
cent solution with acid reaction. Lactose 3 percent, weak growth. Levulose 3 per
cent, tj pical. < rlycerin 3 per cent .
half normal growth. Potato starch .'r-?'
:'. per cent, typical. Butterfat, rich
growth.

Milk, colonies crow verj slowly;
curdling (0.25 per cent calcium
chloric! added i very Blow; digestion,
very slight ; color, none.

At :i7° C crew hotter than chock
at 20° C.

PENICILLIUM No. 28.

Colonies upon sugar gelatin and
potato or bean agar, gray-green with
broad white border when growing,
floccose, tangled tufts of hyphse
and rope- of hvph;e spreading inde-
terminately upon the substratum,
i -e j ellow or tan on media con-
taining sugar, conidiophorea arising
direct from the substratum as short
lateral branches from 38 L60ju in
length, '.'>». in diameter, swelling to

5/i at apex, from aerial hyphse or ropes of hyphse. Conidial fructification a simple
column. :!()()/( or even ">(>(>" in length by 10-15/i in diameter, produced from a single
whorl of conidiiferous cells at the apex of the conidiophore. Conidia elliptical to
globose, 2 3/zor2 2.4/1 by 3 3.3/i in diameter, light yellowish green in mass, smooth.
Colonic- liquefy Bugar gelatin. Litmus reaction strongly acid.

(irow - readily upon all common media.

Pound at Storrs, I onn . upon decaying mushroom. Very characteristic and read-
ily recognized from others with related morphology.

H

I'n,. 33.— Penicillium No. 28: n, b, conidial fructifications
each a single verticil (X 900); g, h, j. sketches of conidio-
phores and fructification (X 140); /.', tip of conidiophore,
swollen at apex, bases of lowesl two conidiiferous colls
(X L,600 .

i l III RAL OATA.

Color gray-green; reverse yellow, or tan when sugar is present; color in i lia,

more or less yellow, according to media.

Fifteen per cenl gelatin in water, good growth, (dear croon; liquefaction Blow — 2
weeks or more; litmus reaction acid. Potato agar, typical growth, acid reactions.
Bean agar, typical growth, acid reactions. Raulin's Quid, good growth, edges pink,
fluid brownish fluorescent. Cohn's solution, weak growth.

Synthetic fluid (Dox's), carbon supplied as: Cane .-near, good growth up to 50
percent, acid reaction. Lactose 3 per cent, slow hut fairly typical growth. Lactic

78 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

acid 0.9 per cent, fair growth, not good. Levulose 3 per cent, good but slow colo-
nies, acid reactions. Glycerin 3 per cent, medium growth, pink tinge to fluid. Potato
starch 3 per cent, good growth, yellow drops of transpired fluid, fluid tinged yellow
at top only. Butterfat, rich growth, fluid reddish brown.

Milk, good growth, acid in litmus milk; curdling (0.25 per cent calcium chlorid
added) in 8-9 days, good curd; digestion slow, incomplete; color, pale yellow in
digested fluid.

At 37° C, killed; check at 20° C, good.

SPECIES FORMING PINK SCLEROTIA.

Four races have been found in which pink sclerotia are regularly
formed in culture. These sclerotia are elliptical to globose and
from 200 to 500/f in diameter. They begin to be formed within the
first week in richly nourished cultures. Although examined repeat-
edly, no trace of ascus formation has yet been found. These forms
are included here under their serial numbers, 29, 30, 31, and 32,
rather than with specific names. The descriptions and figures
introduced will, it is thought, identify these organisms clearly in
their penicillium form, but the uniformit}^ of sclerotium "production
makes ascus production so probable under proper conditions that
it seems best not to give specific names to this imperfect form when
some of them may be already recognizable by others or by further

investigation.

Penicillium No. 29.

Colonies grown upon gelatin and potato or bean agar white to gray-green, sometimes
partly clear green, becoming zonate with rings of pink sclerotia in age, sparsely or
loosely floccose, indeterminate broadly spreading margins persistently white, slightly
yellow below. Conidiophores 80-200// or even 400/t by 3-5/z commonly 150-200/t
in length as branches, usually perpendicular, from hyphse 4-5/i in diameter. Co-
nidial fructification a single verticil of rather few (about 12-15) conidiiferous cells
9 by 2/i, producing chains of conidia in a loose column 150-250/* or even 400/t by 20-
30/i. Conidia elliptical, 3-3.6 by 2.3-2.8/t, smooth, very pale blue (transmitted
light) . Sclerotia in loose networks of mycelium, numerous, pink, elliptical to globose,
150-300/z in diameter. These begin to appear in one week in gelatin cultures. Colo-
nies liquefy sugar gelatin slowly but completely in 10-12 days. Give a strong acid
reaction with litmus media.

Characterized by the production of large numbers of pink sclerotia with compar-
atively small quantities of conidia, whereas the next form (Penicillium No. 30) pro-
duces few sclerotia and great quantities of conidia.

Collected at Storrs, Conn., on decaying mushroom. Probably not closely related
to the common species of Penicillium, but its occurrence in culture and ready adap-
tation to all media tried, in numerous cultures, justify its inclusion with these species.

CULTURAL DATA.

Color white to gray or green, with many pink sclerotia; reverse colorless or slightly
salmon; color in media slightly yellow in some media.

Odor, none.

Fifteen per cent gelatin in water, good growth; liquefaction, 15 days; litmus reac-
tion alkaline. Potato agar and bean agar, typical colonies, white or gray, with. few
green areas and abundant pink sclerotia. Raulin's fluid, typical. Colonies upon

SPECIES FOBMING PINE SCLEROTIA.

79

media with cane sugar produce Bclerotia more aumerously and more quickly (5 da
than without sugar. Butterfal as a sourceof carbonin Dox'sfluid, typical colonies.
Milk, typical; curdling (0.25 per cenl calcium chlorid added I in 9 days; digestion,
Blow; color in milk, none.

Pen n m X". 30.

Colonics on sugar gelatin and potato or bean agar gray-green or green persistently,
Burface growth mostly of crowded conidiophores and producing pink Bclerotia al the
surface or parti) embedded in thesubstratum L50 300u in diameter, broadly spreading.
Conidiophores from 240 525>, usually aboul 300/i, in length either arising separately
from the substratum or as branches verj close to its surface. < lonidiaJ fructification
a single verticil of conidiiferoue cells bearing conidia in a close column up to500/i in
length by L"> 30/z. Conidia elliptical or Bubglobose 2.5 by '■[■■ oiZfi, withaslighl green-
ish color. Colonics liquefy sugar gelatin rather -lowly, and give an acid reaction
with litmus.

Fig. M.—Penicilliuin No. 29: a, b, conidlophore and verticil of conidiiferous cells (X 'J00); c,d, e, germina-
tion of conidia i < 900);/, g, h,j, sketches of conidia! fructifications i < L40); /.. diagrammatic sketch
from photomicrograph showing relations of sclerotia and conidial fructifications.

Apparently related to No. 29, bu1 differing in the length and density of its column
of conidia, in the position of the sclerotia. in habit, in culture, and in its acid reaction.
Collected at Storrs, Conn., upon decaying Laeiariua vellereus, September, li>04.

CULTURAL HA l \

Color green or grayish green, persistently, with abundanl pink sclerotia; reverse

UUColored; color in media, none.

( >dor, none.

Fifteen percent gelatin in water, typical; Liquefaction rather -low; litmus reac-
tion acid. Potato agar and bean agar, typical. Potato plugs, typical. Cohn's
solution, germination only.

Synthetic fluid (Lox'si, carbon supplied as: Cane sugar, rich growth. Lactose 3
per cent, slowly typical colonies, acid reaction. Lactic acid 0.9 per cent, weak
growth. Levulose 3 per cent, medium growth. Galactose 3 per cent, typical. Gly-
cerin :; per cent, very weak growth. Butterfat, slow , bul characterisl ic colonies.

Milk, curdling (0.25 per cent calcium chlorid added I in 9 days; digestion, slow but
complete; color in milk, none.

At 37° C, no growth; grew when cooled; check al 20° C, typical.

80

CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

Penicillium No. 31.

Colonies upon gelatin and potato or bean agar from white to gray to gray-green
mostly white, with few areas of green conidia sprinkled with pink sclerotia, sparsely
floccose, broadly spreading. Conidiophores branching from aerial hyphae, very short
to 380/* in length, commonly 150-240/*, conidial fructification with a single verticil
or once branched with branch, conidiiferous cells and chains of conidia divergent, up
to 140/* in length, but usually much less. Conidia 2.5-3// globose, smooth, rarely
found in quantity to color the colony. Sclerotia elliptical or globose, 160-330/*, pink,
developed in 10-15 days. No asci have been secured. Colonies liquefy sugar gelatin
rapidly and give a stongly alkaline reaction to litmus in the same cultures.
Grows readily in conidial transfers upon all common media.

Collected upon decaying Clavaria at Storrs, Conn., September, 1904. Identical

culture sent from Cambridge, Mass., by
Dr. A. F. Blakeslee in culture obtained
from fruit imported from Porto Rico.

CULTURAL DATA.

Color white or gray, conidial areas gray-
green, very numerous pink sclerotia; re-
verse colorless or with yellow areas; color
in media, none or slightly yellowish.
Odor, none.

Fifteen per cent gelatin in water, typ-
ical white or gray colonies; liquefaction
rapid; litmus reaction alkaline. Potato
and bean agar, typical, cultures with sugar
added become distinctly greener than
others. Potato plugs, typical, white or
gray with greenish areas, sclerotia, and
crystal drops of transpired fluid. Rau-
lin's fluid, some growth, not entirely
typical. Cohn's solution, weak develop-
ment but characteristic.

Synthetic fluid (Dox's), carbon sup-
plied as: Cane sugar 1.5-20 per cent, typ-
ical growth. Lactose 3 per cent, weak growth. Lactic acid 0.9 per cent, no growth.
Levulose 3 per cent, typical. Galactose 3 per cent, typical, alkaline. Glycerin 3
per cent, slight growth. Butterfat, typical growth.

Milk, curdling (0.25 per cent calcium chlorid added) in 9 days; digestion com-
plete; color, none.
At 37° C, no growth, grew when cooled; check at 20° C, typical.

Penicillium No. 32.

Colonies upon milk-sugar -gelatin and potato or bean agar gray-green; floccose, but
with aerial part mostly long conidiophores and few vegetative hyphse, slightly yel-
lowish to pronounced salmon color below; broadly spreading; developing elliptical to
globose sclerotia 150-200/* in diameter at the surface of the substratum in 2-3 weeks.
Conidiophores 200-500/* by 3-4/*. Conidial fructification a verticil of 3-5 branches
10-17/* by 2-3/* rarely a secondary verticil, each bearing a dense verticil of conidiiferous
cells, 8-10/* by 2/* producing long, parallel, or slightly divergent chains of conidia.
Conidia elliptical or fusiform, 3.5-4/* by 2-3/*, green, granular within, smooth, swelling
in germination to 6/* and producing from one to several germ tubes. Colonies slowly
liquefy milk-sugar-gelatin and produce purple or neutral colors in litmus media.

Sent by Prof. P. H. Rolfs from Miami, Fla., upon portion of pineapple, March, 1905.

Fig. 35.— Penicillium No. 31 : a, b, branching of conid-
iophore(X 900); c, germination of conidia (X 900);
d, e, f, sketches of conidiophores.

COMPAKATIVl. ( II.ITKAI. DATA.

81

I I 111 I: A I. DATA.

Color gray-green, with scattered while to pink Bclerotia; reverse, sulphur-yellowish
to pronounced salmon; color in media reddish or yellowish in special cases, others
none. < >dor, none.

Fifteen per cent gelatin in water, typical; liquefaction, none in 15 days, later very
slow liquefaction; litmus reaction neutral, leaves both acid and alkaline media
purple-blue. Potato agar and bean agar, typical, Blightly thinner than gelatin cul-
tures, gray-green without sugar, clear green with cane sugar. Potato plugs, typical,
transpires yellow drops which become very dark yellow (balsam). Ranlin's fluid,
good colonies becoming rosy below. Cohn's solution, small colonies, fluid slightly
yellow.

Synthetic fluid (Dox's), carbon supplied as: Cane sugar, grew in solutions up to 30
percent with acid reaction. Lactose 3 percent, very slow growth of small character-
istic colonies. Lactic acid oil per cent, good growth, light green. Levulose 3 per

Fig. 30.— rcnicillium No. 32: a, b, (/.branching of conidial fructifications (a X 1,400, 6 and d,X 900);
c, a single secondary verticil (xl,400); g, h, j, k, sketches of fructifications of various ages
(X 140); m, n, o, germination of conidia (X 900).

cent, very alow-growing but heavy colonies. Galactoses percent, typical. Glycerin,
very small colonic-. Lutterfat, typical colonies.

Milk, curdling (0.25 per cenl calcium chlorid added) very slow; digestion, very
slow; color in milk, none.

At 37° I ., grew more rapidly than check at 20° C.

COMPARATIVE CULTURAL DATA.

A summary <>!' accessory cultural data has already bees given for
each species in connection with the descriptions. Many scries of
culture- have been made with numerous media to obtain data as to
the ability of the species studied to grow upon particular media or
under particular conditions. It has been possible thus to determine
the relative activity of single species and groups of species.
Although particular species in these cultures have shown unique
differences which assist in their differentiation, the most valuable

8108— Bull. 118—10 6

82 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

result of comparative culture is found in the separation of the series
into groups of races or species which resemble each other closely in
their metabolic activities. Part of these experiments are tabulated
in Tables 2, 3, 4, 5, and 6, and will be discussed in the following
sections.

Since the complex composition of the media in common use for
cultural work makes the analysis of the data obtained impossible,
it was first necessary to determine the reactions of these species to
some of the individual substances of which these media are composed.
It has already been noted that gelatin alone in distilled water sus-
tains growth in the large majority of the species of this genus.
These cultures in certain species lack green color, which, how-
ever, becomes present on using the peptones and sugars added to
gelatin in most formula?. Such media are still too complex to make
close analysis of results possible.0

Man}' of the determinations given were made in duplicate and in
some cases the entire series was repeated one to several times. In
inoculating cultures for this work conidia were transferred to the
tubes in large numbers, so that their presence could be detected by
examination with a lens. Where species failed to grow, the doubts
of inoculation were commonly dispelled by the addition of cane
sugar, which permitted the conidia to develop normally if present
and still viable. The presence of germinated conidia upon the sur-
face of a medium is good evidence of proper inoculation. The data
given are believed, therefore, to represent with a fair degree of
accuracy the comparative cultural reactions of the species used.

In reporting these series of comparative cultures, the data have
been tabulated as far as possible for convenient comparison of the
relative activity of the different forms. (See tables beginning on
p. 98.) The names as far as determined are given, together with the
cultural number, in Table 1 . In the remaining tables the numbers
are repeated without the names. In studying the tables a reference
to cultural numbers will quickly locate the forms discussed.

CULTURES IN DISTILLED WATER.

To determine the possibilities of growth from food stored in the
conidia, cultures were made in distilled water. Of forty-four strains
under cultivation but six showed clearly descernible germination.
None produced more than germ tubes hanging down into the fluid.

a This work was carried on in cooperation with Mr. A. W. Dox, who has studied the
metabolism of the species concerned with cheese ripening (P. camemberti and P.
roqueforti), as well as a few other species, under many conditions of culture, while the
writer has conducted comparative studies of a large number of forms under more
limited cultural conditions. All chemical questions arising throughout this work
have been passed upon by Mr. Dox.

COMPARA I l\'i: CULTURAL DATA. 83

\(. LR- \<. Ai: A> A SOI i:< K OF POOD.

Tubes of I .5 percent agar-agar in distilled water were inoculated
with eighteen species of PenicUlium. Sixteen of these produced
growth. In all cases the colonies produced were very small, some
of them barely discernible to the naked eye. Not one of them was
distinctly colored by conidia] masses, but in ncarh even ease some
conidial fructifications were found. These cultures show that the
species tested were able to obtain from the medium sufficient nourish-
meiit for very slight growth.

AGAR-AGAR AS A SOURCE <>F CARBON.

Agar-agar is a carbohydrate and might serve as a source of carbon
if other nutrients were supplied. One and one-half per cent of agar
was therefore introduced into Mr. Dox's synthetic fluid, already con-
taining all essential elements except carbon. Thirty-seven races of
PeniciUium were inoculated into this medium, and nearly every
species produced some growth. Examined with the microscope,
conidial fructifications were found in nearly every case, but in no
case was the colony large enough or definite enough to affect the
observer's estimate of results if such growth were added to or sub-
tracted from the colonies upon nutrients really adapted to sustain
the species studied. Dox's stock solution, with or without the addi-
tion of agar, was in this way shown to be a safe medium for the study
of the metabolic reactions of these species to changed sources of
carbon.

The possibility of error in the introduction of agar was shown in
the following manner: One and one-half per cent of agar was intro-
duced into Dux's fluid and 1.1 normal lactic acid added in quantity
to make the whole 0.5 per cent acid. The medium was then auto-
claved. After this treatment the agar refused to solidify. Tubes of
tins fluid were inoculated with nine different species of Penicillin m.
All except P. breoicauL grew well and produced colonies recognizable
by their cultural characters. In introducing agar in such work it is
therefore accessary to guard against the introduction of acid before
dissolving the agar, since this changes the agar itself into other car-
bohydrates assimilable by fungi. Although parallel cultures were
commonly made with agar, the studies of metabolism recorded in this
paper were made in t nbe culture of the fluid nutrients only, to avoid
possibilit ies of error.

VA1MOI s SOURCES OP CARBON.

Cane sugar ( Tables ', and 5). Cane sugar was added to Dox's fluid
in the following percentages: 1 ..>, :;, 10, 20, 60, and 75. Of the spe-
cies used, but one — P. digitatum of Saccardo failed to grow typ-
ically. This, together with other work, indicates that this species is

84 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

incapable of assimilating nitrogen from the sodium nitrate of this
solution. Four more forms — P. brevicaule and its closely related
varieties, and P. roseum Link — reached normal appearance slowly.
Other cultures indicate that these forms assimilate nitrogen in this
form less readily than in organic combinations. This medium con-
taining cane sugar in amounts as great as 20 per cent proved well
adapted for all other species tried. The medium as used was neutral
or slightly alkaline in reaction. Thirty-three of the forms inoculated
into it produced pronounced acid reactions to litmus in a few days, i. e.,
were able to ferment this form of sugar. In prolonged culture with
smaller amounts of sugar, some of these forms finally reduced the
acidity and even brought about an alkaline reaction again; others
remained acid as long as observed. Twelve forms failed to produce
acidity. This number includes the four forms in which growth was
delayed and P. italicum, P. luteum,, P. jiurpurogenum, P. roqueforti,
and P. duclauxi.

In cultures at a concentration of 60 per cent cane sugar, five forms
produced typical colonies at once; six others slowly reached normal
proportions ; a few more grew fairly well ; but fully half the species tried
produced germination with but little further growth. Water in
amount approximately to reduce the concentration to 35 per cent was
added to the cultures that failed to produce normal colonies, and this
was followed by the prompt recovery of several species which quickly
reached normal development. Critical examination of the data
obtained showed that closely related types responded in exactly the
same manner to changed percentages of cane sugar. As a means of
separating closely related forms, further determination seemed fruit-
less. Exact determination of the maximum percentages of cane
sugar tolerated by particular species has not been completed. The
inhibition is not a stoppage of all development at a definite critical
concentration, but rather a gradual reduction of activities with the
increasing concentration of the medium. Specific maxima and
minima are therefore almost impossible to define. All determina-
tions would therefore rest upon the judgment of the observer rather
than upon fixed standards.

Lactose (Tables 4 and 5). — Lactose was added to Dox's fluid in
percentages up to 10 per cent. Prompt and normal development was
determined in eight forms ; nine more forms reached typical appearance
more slowly. Only twelve of these produced definite acid reactions to
litmus. Among the forms included in the seventeen, four groups of
closely related organisms were found, namely, the camemberti group,
Nos. 5, 6, 39 ; the common green group, Nos. 22, 23, 40 ; the chrysogenum
group, Nos. 25, 26, 35, and 44; and the brevicaule group, Nos. 2, 3,
and 4. With lactose as with cane sugar closely related forms give

COMPARATIVK ( T LIT UAL DAI \. 85

approximately the same reaction in most cases. Three of these
groups show acid and the fourth alkaline tests with litmus. A few of
the remaining species continued to grow until after several weeks
they reached almost normal development. In such cases the assimi-
lation of carbon from lactose seems to be \ery slowly and with diffi-
culty accomplished by a large proportioE of the species studied, and
to be practically impossible to some species.

Comparison of t he litmus reactions produced by the various species
with cane sugar and with lactose accounts for striking differences in
this reaction when litmus is introduced into gelatin or agar media
containing these two forms of sugar. A species which will ferment
cane sugar and not ferment lactose will produce an acid reaction with
one and an alkaline reaction in media containing the other.

Lactic acid (Tables 4 and 5). — Tubes were prepared containing
Dox*s fluid to which 0.9 per cent of lactic acid was added. Thirty
forms were inoculated into this medium. Of these forms nine pro-
duced normal and typical colonies, showing but slight inhibiting
effect from the acid. As manymore cultures slowly became typical
colonies. Nearly every form germinated and produced slight growth.
In the camemberti group (Nos. 5, 6, 39) and some others the litmus
reaction became alkaline. It was thus shown that a series of species
could secure carbon from lactic acid and in doing so dest roved the acid
character of the medium. The species which grew most rapidly
in lactic acid were those which had developed best in the lactose
solutions.

Levulose (Tables 4 and 5). — Tubes were prepared into which 2.5 per
cent of levulose was introduced as the source of carbon. The re-
sults as tabulated may be seen to group together about the same
species as the previous experiments, except that one or two forms
were found to grow well in levulose thai failed to grow with lactose or
lactic acid as a source of carbon. Fourteen forms produced typical
colonies without inhibiting effects, while five more slowly reached
typical development.

Galactose (Tables 4 mid 5). — A series of cultures was made in the
same way with '.-> per cent galactose as a source of carbon. Galactose
proved much better adapted to supply carbon than either lactose or
Levulose. Twenty-five forms produced typical growth, and others
grew more or less readily. This form of sugar therefore proved of
but small assistance in the separation of species.

(•hjeerin (Tables 4 and 5). — Cultures offering carbon in the form of
3 per cent glycerin produced much less growth than those containing
sugars. Eleven forms eventually reached fairly typical growth; four
only of these showed no restraining effect of t he medium. Of these,
three are probably closely related if not merely races of a single
species — the one most common in general cultural work in this

86 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

region. Apparently glycerin presents a form of carbon much less
available for assimilation by species of this genus than the sugars.

Butterfat (Table 4.). — To test the ability of these fungi to assimilate
fat, butter was melted, strained, filtered through filter paper, and
added to Dox's fluid. Although not chemically pure, perhaps, it
is believed that the amounts of other nutrients would be too small
to affect results. Only one form (P. digitatum) failed to grow.
P. luteum gave only slight growth. The majority of forms, although
growing slowly, produced typical or fairly characteristic colonies.
The masses of fat were visibly much changed, becoming incrusted
with a white substance in most cultures. In a few cultures the
action of fungus caused the separation of the various fats, so that
drops of yellow oil separated out from the remaining nonliquid
matter.

Potato starch. — In one series twenty-seven forms were cultivated
in Dox's fluid, containing 3 per cent of potato starch. All of the
common species of the genus were found to grow normally upon a
medium containing starch as the source of carbon. The characters
in this medium were approximately the same as in the stock agar
or gelatin cultures. Two species (P. decumbens and P. digitatum).
which failed to grow well have since been shown to depend upon
the presence of cane sugar for vigorous growth and green color to
their spores. Similarly the same species grown upon plugs of potato
failed to produce strong colonies of pronounced green color.

Malic and succinic acids. — Series of cultures were made with
1 per cent malic acid and with 1 per cent succinic acid as sources
of carbon. All species germinated, but no species reached fully
typical development in either series. Some few species produced
slowly colonies of half or more of the normal size with conidial
masses of typical color. Many of the species grew sufficiently to
produce a few conidial fructifications recognizable with the hand
lens. These two series emphasize the observation already made
that species inoculated into a medium ill adapted to their nourish-
ment will nevertheless grow and produce small amounts of fruit
under widely different conditions even where normal growth is
impossible.

CULTURES IN RAULIN's FLUID AND COHN's SOLUTION.

The comparative data for. cultures in Raulin's fluid and in Cohn's
solution are shown in Tables 3 and 5. Raulin's fluid, as given by
Smith, is a highly acid medium and has been found very well adapted
for the growth of certain species. The solution is, however, too
complex to make analysis of cultural results upon it readily possible.
It contains carbon in three different forms — tartaric acid, potassium
carbonate, and magnesium carbonate — and nitrogen in two forms

COMPARATIVE CULTURAL DAI \. 87

iii ammonium nitrate. Aside from data as to growth or failure to
grow, culture in such a medium is fruitless.

Cohn's solution is also an acid medium, but its acidity is due to
the presence of monopotassium phosphate (KHaP04). In a series
tit' cult i ncs wit 1 1 Dox's formula it was shown thai the monopotassium
phosphate had practically do different effeel upon cultures of these

species than the dipotassium phosphate (K2HP04).

Very few of the species were found to gro\* typically upon Cohn's
solution, however, although nearly all germinated. Comparative
work with Dox's formula shows that bul few of the species studied
are capable of normal assimilation of carbon from forms of tartaric
acid. Only three of the species experimented with failed to show
distinct germination. One of these inoculated with V. camemberti
var. rogt ri, remained four months without exhibiting any germination
of the conidia. The conidia were then transferred with a platinum
needle to a pet ri dish of gelatin; under these conditions the same
spores developed into typical colonies without showing any ill
effects of four months' immersion in the fluid which they were unable

to assimilate.

In solutions nontoxic in character many species exhibit definite
selective preferences for nutrient elements in particular chemical
combinations. This selective adaptability to particular forms of
nutrients differs greatly for different groups of species. Some
forms of wide distribution seem adapted to produce typical growth
upon quite varied chemical solutions. Other species (e. g., the rots
of citrus fruits ), equally w ide in t heir dist ribution, are closely depend-
ent upon particular forms of food, whereas between these extremes
are many forms exhibiting preferences as to nutrients yet capable
of development, although more slowly, in media containing nutrients
in forms assimilated with difficulty. These experiments have offered
no tangible evidence of rapid adaptation in media found unadapted
at first to development. Such power of gradual adaptation is not
excluded, however, by these experiments.

COLOB IN CONIDIAL AREAS.

In dealing with all species it has been found by repented experi-
ment that the green color of the spores is dependent upon the proper

assimilation of the carbon element. Many of these species produce

a brighter green when canesugar is present than with any other
form tried. Grown upon gelatin, or upon potato agar or bean agar
(free from sugar), several species produce green masses of conidia
which rapidly become some shade of gray, brown, or almost black
in media without sugar, but when cane sugar is present they are
deep green and remain unchanged for much longer periods. Certain
other species entirely lack green color except when sugar is present.

88 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

A color determination is therefore dependent for its value upon
knowledge of the form of carbon presented by the medium. These
same species grown with lactose as a source of carbon respond exactly
as they do in potato or bean agar or in gelatin free from sugar.
Another series of forms are little if at all changed in color by changing
the form of carbon presented, provided only that they are able to
grow readily in the medium as presented. It seems therefore certain
that very many widely distributed species are capable of assimilating
carbon in very widely different chemical forms and to produce in
such cases normal and typical growth and colors, whereas other
species are entirely dependent for normal growth and color upon
the presence of particular chemical combinations or groups of com-
binations. Study of the forms so responding likewise shows that
the ubiquitous species are capable of assimilating carbon in the most
varied combinations, whereas the forms lacking this power are
mostly less common and more specialized.

EFFECT OF CONCENTRATED MEDIA.

Similarly great differences in the mass of the growth produced
are directly attributable to the presence of nutrients easily available
or in greater concentration. The formulas usually recommended con-
tain the nutrient used, in extremely dilute proportions. The deter-
minations already given with different percentages of cane sugar
show that these species are able to assimilate sugar in widely different
concentrations. With the majority of forms studied there seems to
be no deleterious effect from increasing the concentration of the nutri-
ents offered until the solution has attained an osmotic pressure suffi-
cient to inhibit growth by plasmolyzing the cells, or until the reduced
percentage of water gradually reduced the rate of fungous growth. In
one experiment nine species were inoculated into a medium contain-
ing 5 per cent cane sugar, 10 per cent Witte's peptone, and 5 per cent
Liebig's extract. All species grew luxuriously. The mass of myce-
lium produced was much in excess of the results with ordinary culture
media. One form normally producing sclerotia produced a very rich
growth of mycelium, but no sclerotia. In media of higher concen-
trations the amount of growth — the mass of mycelium and conidia —
is, however, so greatly increased as to render the discrimination of
the character of the species more difficult by mechanical interference
due to the quantity of material. For purposes of stud}', therefore,
the usual formulae really produce the more satisfactory growth in
nearly every species, although from the standpoint of fungus devel-
opment such media must be recognized as far below the optimum
concentration for the species of this genus.

It must further be noted that in cultures containing cane sugar
many species continue to produce conidia for a much longer period

COMPAKATIYL CULTURAL DATA. 89

than in solutions Lacking sugar or some nutricni equally assimilable.
In these cultures the quantity of conidia produced increases enor-
mously, often becoming a layer over the whole surface of the colony
half a millimeter or more in thickness. The presence of such masses
of spores greatly complicates the stud} of the structure of the colo-
nies, hut is especially characterisl ic of such species.

THE GROUPING OF SPECIES.

Analysis of the cultural tables presented in the light of many series
of comparative cultures makes possible the grouping together of par-
ticular races or species which possess common cultural characters.
It is comparatively a simple matter to single out first the unique
forms those which are never green, those which produce a particular
form of sclerotia, those which regularly produce prominent coremia,
or those which produce striking colors in the substrata, even those
associated with particular substrata. There remain, however, the
Large number of green forms which lack these striking characters.
This large group comprises probably most of the forms which have
masqueraded under the name P. glaucum. Lines of differentiation
among these forms are more or less obscure. Kept in continuous
culture races are easily differentiated with the eye by shades of color
or habit, but characters of easily recognizable diagnostic value in
written descriptions are more difficult to find. Inspection of cul-
tural data shows, however, that there is a well-marked group of these
forms which are able to ferment lactose as well as cane sugar. These
comprise the cdmemberti group (Nos. 5, 6, 39), the chrysogenum
group (Xos. 25, 26, 35, 44), and what we may call the "commune"
group (Xos. L'L\ I'.'!, 40). Among those not causing an acid reaction in
lactose cultures is a series of rapid liquefiers of gelatin which have
many characters in common (Xos. 12, l.">, LM. i!7, 3S). All of these
forms show special adaptability to growth in cane-sugar media. No.
L5, P. dtrinum, produces brilliant lemon color, especially in sugar
media; No. 38 is given as P. atramentosum, from its blackening of
the substrata in sugar media and in milk; No. 37 by developing green
color when sugar is present, which color is Lacking or evanescent with-
out sugar. These forms differ in their color reactions in the medium,
in the size ami shape of their conidia, in the length and origins of their
conidiophores, and in the arrangement of the elements in the conidial
fructifications.

Another marked habit difference which holds t rue throughout many
series of cultures is the tendency of colonies of certain species to
spread rapidly over the whole surface of the substratum, whereas
others are quite restricted in their habit of growth. ]\\ the first the
developing margin is almost uniformly broad and while growing
white, e. g., P. roqueforti, P. italicum, P. chrysogenum, P. divaricatum,

90 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

P. spinulosum , P. expansum, P. hrevicaule. and such floccose forms as
P. camemberti and P. biforme; the species of restricted habit with
narrow growing border are represented by P. citrinum and P. atra-
mentosum and their allies. The designation "spreading," "restricted,"
"with broad margin," or "with narrow margin" is descriptive for
colonies of such species.

ODORS.

Although many persons seem to detect the presence of mold by its
odor, very many of the species give but little definite odor. A few
forms, some of them (e. g., P. biforme) always, others upon special
media, produce difinite odors by which they, can be recognized or
placed in particular groups. Grown upon gelatin media, P. brevi-
caule produces a strong ammoniacal odor. A piece of moistened lit-
mus paper held over such a colony will promptly give the alkaline re-
action. This organism is recorded as emitting arsin from cultures con-
taining arsenic in any form and it is said to be a very delicate test for
the presence of that element a (Gosio) . The two forms here described
as varieties of P. brevicaule give exactly the same odor. In nearly
every medium used P. expansum (the apple rot) gives a strong odor,
which suggests decaying fruit to some. Once well distinguished, the
presence of this organism can be detected even as a contamination
wherever it occurs, by the odor alone. Another given here as P.
atramentosum produces a very characteristic odor while digesting milk,
defined by one as the odor of rancid walnuts, to another it suggested
mice. A series of forms when grown upon cane sugar produce a very
characteristic odor — an ester, according to the chemists to whom it
was submitted — recognizable to the sense of smell, but not definable.
The olive-colored orange rot (P. cligitatum) gives this most strongly, but
it is also given by P. italicum, P. decumbens, and No. 13. The odor
of P. claviforme is found under nearly all cultural conditions, and
would readily identify it were its big coremia not already very distinc-
tive. The common and undefinable green group contains a series of
races or forms, many of which give what is popularly called the smell
of mold. In others of this group this odor is scarcely distinguishable.

ANAEROBIC CULTURES (WITH CARBON DIOXDd).

The possibility of some species developing under anaerobic con-
ditions was tested as follows: Vials were prepared with Dox's fluid
having 5 per cent cane sugar as a source of carbon. This had already
been shown to be an excellent medium for the growth of nearly all the
species used. These vials, containing all the species herein described,
were packed in a crate or test-tube basket and put into a Novy jar.
The jar was then given in charge of Mr. Dox. The air was exhausted

o Gosio. Azione di alcune muffe sui composti fissi d'arsenico. Rivista d'Igiene
e Sanita Pubblica, Rome, 1892. See page 201.

INCUBATION EXPERIMENTS. 91

as completely as possible. Carbon dioxid washed through water and
through sulphuric acid was carefully introduced. The exhaustion
was repeated and the gas introduced a second and a third time. The
cultures were then permitted to stand one week and examined. A cul-
ture of Oidium lactis among the species of PenicHUium was found to
have grown some. No species of P< nidUium had produced a colony.
The carbon dioxid was then three times exhausted and replaced by
air and the jar permitted to stand a second week. Normal colonies
of every species were produced. Clearly, therefore, no one of the
species of P< niciUium under experimenl was capable of grow Lug in an
atmosphere of carbon dioxid. and no species was killed by exposure
to such at mosphere.

INCUBATION TESTS.

The importance of temperature in determining the distribution of
fungi in nature, and in controlling their presence in the household, t be
dairy, and the storage room, made the determination of the limits of
temperature for the growth of these species desirable. The following
incubation experiments were therefore made. (See Table 6.)

1 . Incubation at 20° C. Tbis was repeated several times with fully
checked records.

2. Incubation at 37° C. (range of variation 35° to 38° C.) for six
days; cultures then examined, recorded, and the incubator cooled to
20° C. for the succeeding six days.

3. Use of the iee thermostat.0 Four series of cultures were made
in bean-agar with 5 per cent cane sugar — a medium adapted to pro-
duce typical colonies of all species in a minimum of time. The tem-
perature of the incubator was recorded eight times a day at three-
hour intervals. The range of temperatures in the compartments
used and the average of fifty-five observations taken in the first seven
days were as follows:

Compart menl I, average 1.05° C, range 0.5° to 2° C.
Compartmenl 2, average 4° C, range 3.2° to 6° C.
Compartment 3, average 7° C, range 6° to 10° C.
Compartment 5, average 8.7° ('., range 7° to 10.5° C.

Observations upon the cultures were made by the writer at 3 and

7 days and repeated, by the kindness of Miss Lucia McCullough,
of the Bureau of Plant Industry, at 15, 23, and 29 days. For con-
venience of comparison, the notes from all these observations have
been reduced as fairly as possible to a decimal code, iii which ger-
mination of conidia w it houl furl her growth is given as 0. 1 and typical
colonies at l.Oj fractions from 0.1 to 0.7 represent vegetative
mycelium without colored conidia] areas, and from 0.7 to 1.0 the

" I '>> the courtesy of Dr. Envin 1'. Smith the ice ilicnuostat of the Bureau of Plant
Industry of the United States Department of Agriculture was placed at my disposal.
It was iced and regulated under his instructions.

92 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

completion of the typical colony. The figures are brought together
in Table 6.

Data at 20° C. — The data at 20° C, as given in the first column of
Table 6, are regarded as typical for the species studied and given as 1 .0.
Numerous series of cultures with all these forms under close observa-
tion in the incubator at 20° C. and in the laboratory where the tem-
peratures used ranged from 15° to 25° C. or slightly higher have given
approximately the same results. Within these limits, rise or fall in
temperature affects the amount of growth or the stage of develop-
ment of the colonies within a specified time without affecting the
character of such growth. The differences between cultures grown at
different temperatures within these general limits are quantitative,
not qualitative. Unless made for a specific purpose cultures of these
fungi may be safely grown outside the incubator without affecting
their character, since the conditions in the ordinary working room are
approximately those furnished to these forms by nature.

Data at 37° C. — At 37° C. thirteen forms showed normal develop-
ment. Of these seven grew better at 37° C. than at 20° C, this
number including but one well-known species — P. luteum. At the
same temperature the spores of seven species were killed, including
among these P. italicum and P. digitatum, the species destructive of
citrus fruits. Of the green forms abundantly found, only one grew
well at 37° C. — P. chrysogenum. The numerous green forms studied
were not killed, but simply prevented from growing by the heat.
Every form except those noted as killed developed normally in the
same tubes as soon as cooled to 20° C.

Ice thermostat. — In compartment 1 of the ice thermostat, ranging
from 0.5° to 2° C, 20 of the forms experimented with either produced
germ tubes only or failed even to germinate in twenty-nine days. Of
the remaining 18, only 6 produced colored conidial areas in that time.
Several other species produced considerable masses of white mycelium.

In compartment 2, ranging from 3.2° to 6° C, with an average
slightly above 4° C, 16 or 17 still showed germination only or com-
plete inhibition; 11 showed colored conidia; several additional forms
had germinated or produced distinguishable mycelium.

In compartment 3, with an average temperature about 7° C. and a
range from 6° to 10° C, 16 forms produced colored fruit. Of these,
9 had reached the typical appearance of mature colonies of the species
within the 29 days. Nine only remained without showing some
mycelial growth in addition to germination.

In compartment 5, averaging about 9° C. and ranging from 7° to
10.5° C, all species had germinated and all but one had produced
mycelium; 25 forms had developed colored conidia; 17 had produced
colonies of typical appearance.

DISCUSSION OF INCUBATION TESTS. 93

Study of the figures given shows a progressive increase in growth
from tin- coldest to the warmest compartment. Examination of the
cultures showed thai at the lower limits of growth very many species
produce colored fruit verj slowly at temperatures at which vegetative
mycelium is still developed quite rapidly. There often results,
therefore, in cold temperatures a disproportionate growth of white
mycelium and a tardy development of colored conidia, which often
a I Fret'- the appea ranee of the resulting colony considerably. Cultures
grown under such conditions would be difficult to identify in many
cases. The forms which grew most rapidly at .".7° C. either failed to
grovi at the colder temperatures or responded very slowly. The large
majority of the species, and especially those most commonly found in
food materials, are seen to begin fairly rapid growth at temperatures
within a few degrees of the freezing point. When taken from the ice
thermostat all cultures which had failed to produce normal colonies
grew quickly to typical appearance. There was, therefore, no injury
attributable to continuation for 29 days at Low temperature. The
general effect of low temperatures upon these species is the suspension
of or the reduction of the rate of development. In many cultures
the beginnings of growth were difficult to detect and in most eases an
exactly critical temperature is not determinable, since cultures not
showing any growth in one week seem gradually to adjust them-
selves to conditions and produce mycelium in the succeeding weeks.

Eustace8 has recorded that one of these species (P. expansum
Link of this paper) will produce rot in storage apples where the
temperature of the room as recorded does not rise above 32° F.
(0° C). When the time was extended to two months Petri-dish
cultures under the same conditions produced small colonies. The
experiments here recorded tend to suggest that very little growth
will occur in most species at temperatures nearer than 2° C. to the
freezing point, although many of them will germinate. Unpub-
lished records of the temperatures of apples in storage, furnished
by Mr. ('. I). Jarvis, of this station, showed that the flesh of apples
in storage was constantly from 1 to 2 degrees at least above that of
the room. Allowing for the conductivity <>f the thermometer itself,
the difference is probably somewhat greater. Both series of data
indicate, however, that storage1 temperature to exclude fungous
grow th must be close to the freezing point. It is clear that low tem-
peratures (above freezing) merely restrain growth, not entirely
prevent it. It is also clear thai by restraining the production of
colored fruit many colonies would be rendered inconspicuous (al-
though widely growing), thus accounting for the complaint so often
heard with reference to dairy products taken from the refrigerators,
that they turn green with mold very quickly.

94 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

SUMMARY OF DATA FROM COMPARATIVE CULTURE.

1. Species closely related in morphology and general appearance
give closely similar reactions in culture under most conditions, but
commonly show a few well-marked differences in special media or
under special conditions.

• 2. Certain species will grow in media of widely differing composi-
tion; others require particular media for normal development.

3. Cane sugar in low concentrations is readily assimilated by
every species studied.

4. Butterfat was attacked by nearly every species.

5. Lactose, galactose, levulose, and glycerin are assimilated by
some species and not by others.

6. Potato starch produced normal growth in most species.

7. Very few species grew at 37° C; i. e., very few species could
be parasitic to wTarm-blooded animals. Few species were killed at
37° C, and species not killed grew normally when the medium was
cooled to 20° C.

8. Incubation at low temperatures shows that growth in some
species will begin at temperatures very close to the freezing point,
but that only a few species will actually develop in cold-storage
temperatures.

9. None of the species was found to grow in an atmosphere of
carbon dioxid, but no species was killed by such atmosphere.

10. In most species failing to grow in a particular medium a
change of concentration or the addition of a missing element will
permit normal growth unless killed by osmotic pressure or definitely
toxic agents.

KEYS TO CULTURAL IDENTIFICATION OF SPECIES.

Media: Prepare the following media —

1. 15 per cent gelatin0 in distilled water.

2. 15 per cent gelatin in distilled water plus 3 per cent cane sugar.

3. Either bean or potato decoction plus 1.5 per cent agar.

4. Bean or potato agar plus 3 per cent cane sugar.

Litmus solution may be added if desired when cultures are made.

Prepare Petri dishes with 10 c. c. of each of the media used and
allow them to cool. Inoculate two or more Petri dishes of each
medium with spores of the species under examination. Incubate
at 20° C. (the temperature of the working laboratory is usually

aIn most of these studies the "gold-label" gelatin imported by Bausch & Lomb
from Germany has been used. Culture of a species in a solution of gelatin in water
has two uses in this paper — the detection of the production of enzyms capable of
liquefying this medium, and the estimation of the ability of the species to grow in a
medium free from carbohydrates. Neither of these tests seems to be vitiated by
media made up at different times from materials of different origin. Such differences
as are induced by the differences in the gelatin are quantitative, not qualitative.

CULTURAL IDENTIFICATION OF SPECIES. 95

satisfactory). Examine at intervals of three days OX less, making
observations with the naked eye from above and below, with the
hand lens and with the compound microscope, using 16 nun. and
8 mm. or 6 nun. objectives to determine details of structure and
fruit formation from growing colonies. A drop of litmus solution
upon the margin of the colony will test acidity or alkalinity. Ex-
amine 1 and 2 for liquefaction, 2 and 1 for coremium and sclerotium
formation. Sclerotium formation will be found to call for continued
examination for at Least two weeks.

Two separate keys are presented in the following pages: (1) A gen-
eral key to all the forms discussed in this paper, based upon cultures in
the media referred to above, and (2) a key to those species for which
presence upon a particular substratum establishes a presumption of
ident it \ .

The species of most economic importance are found in key 2.
Key 1, however faulty, is an endeavor to analyze the data from
many series of comparative cultures in such a way as to simplify
the identification of the forms included as far as possible. If the
complexity of the data offered proves a barrier to identification it
may be hoped that it will also deter those who fail to identify species
clearly from usinir specific names in discussing work done with forms
of this genus.

Key 1.- Analysis oi Species i\ Ci ltures upon Gelatin and Agar.

A. Species fruiting typically by coremia (vertical and definite).

a. < Joremia long (3 15 mm.).

1. Conidial masses strictly terminal, olive green, fragrant. . /'. claviforme.

l'. CTpper third of coremia fertile, conidia green P. duelauxi.

aa. Coremia -mall:

1. Goremia definite, densely crowded, colony orange below,

/'. granulatum.

2. Goremiform character indicated in cultures by clustering of

conidiophores, definite coremia only in old cultures, becom-
ing large and definite upon apples P. expansum.

AA. Species not (or rarelj ' producing coremia in culture.

B. Species constantly producing sclerotia or ascigerous masses.

b. Producing ascigerous masses, yellow or reddish P. luteum.

hh. Sclerotia appearing as white masses in old cultures P. italicumfi

bbh. Sclerotia reddish or pink, globose or i Uiptical, 500/i or Less in diameter.

c. Conidial fructification a column:

1. Column dense, long, Bclerotia partially buried in Bubstratum,

/'. No. 30.

2. Column Formed of loose chains, scler tia numerous, exposed,

P. No. 29.
cc. Conidial fructification of divergenl chains:

1. Rapid liquefier, spores glob i '■[■' /'. No. 31.

2. Slow liquefier, spores elliptical, 3.5-4X2.5-3// /'. No. 32.

"In its earlier development /'. italicum Wehmer will be usually thrown under the
head kk, on account of its habit of growth.

96 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

BB. Sclerotia not (or rarely) produced (under special conditions).
(Use gelatin cultures (1) and (2), compare agar cultures).

C. Rapid liquefiers (abundant liquid in 5 to 12 days).

D. With definite, strong ammoniacal odor:

1. Yellowish brown-avellaneous spores rough P. brevicaule.

2. White or cream, spores rough P. brevicaule, var. album.

3. White or cream, spores smooth P. brevicaule, var. glabrum.

DD. Without ammoniacal odor.

E. With yellow coloration of liquefied gelatin (not of mycelium in reverse).

1. Colonies small, conidiophores 100-150// in length P. citrinum.

2. Colonies broadly spreading, conidiophores 250-300,u. .P. chrysogenum.
EE. Without yellow color in liquefied gelatin (or slight traces only).

e. Colonies white to pink or salmon P. roseum.

ee. Colonies some shade of green.

/. Colonies floccose, margin spreading by stolons P. stoloniferum.

ff. Colonies velvety-surface growth of fruiting hyphae only.
g. Conidiophores very "short (100-200/t):

1 P. No. 12.

2 P. No. 37.

gg. Conidiophores longer (200-400^):

1. Conidiophores variously branched, reverse always colorless. .P. No. 24.

2. Conidiophores each with a verticil of branches — each branch

bearing a columnar fructification — reverse and medium dark-
ened in sugar media P. atramentosum.

CC. Liquefaction of gelatin none or slower than 10-12 days, or only partial.
G. Colonies never green.

h. Colonies yellowish brown, spores elliptical P. divaricatum.

hh. Colonies white to lilac, slow liquefier, 14-16 days P. lilacinum.

hhli. Colonies floccose white or creamy:

1 . Conidiophores long, typical penicillate branching,

P. camemberti, var. rogeri.

2. Conidial chains borne upon short branches of floccose hyphae,

P. No. 33.
GG. Colonies some shade of green.

H. Surface with hyphae definitely in ropes or trailing, bearing numerous conidio-
phores as short branches distinctly traceable to their origin in such
hyphae .
i. Colonies usually red below and reddening the substratum.

1. Fruiting areas dark green P. funiculosum.

2. Fruiting areas mixed yellow and green P. pinophilum.

ii. Colonies not producing red color:

1. Colonies gray rarely greenish, very loose floccose P . intricatum.

2. Colonies green, conidial chains in simple compact columns. . P. No. 28.

3. Colonies gray to green, hyphae scattered, creeping P. decumbens.

CULTUKAL IDENTIFICATION OF SPECIES. 97

IIII. Surface hyphae qoI in well-defined ropes, nor trailing.

Surface hyphae woven Qoccose, course of hyphse no1 traceable.

1 . < rray-green, I< >hlt conidiophores, do odor /'. cam* mberti.

2. Gray-green, shorter conidiophores, strong odor /'. biforrrie.

jj. Surface growth al margin simple conidiophores, in older parts both floc-

■• hyphae and conidiophores.
1. Gray-greenish, branching of conidiophore rather loose, odor

Don • Blighl P. X... 22.

i'. Green, conidial fructifications rather compact, odor definite,

"moldy" P. commune.

X. B. There are probably a number of races in this group.
. Fruiting Burface velvety of simple conidiophores or conidiophores I mine
lose to surface of substratum as to appear simple:
fc. Conidial ma - a dense column of conidial chains.

1 . ( iolumn from a single verticil of basidia P. spinulosum.

2. Column from ;i verticil of brachlets with verticillate cells and

chains /'. rubrum.

hh. Elements of conidial fructification do1 in a column.

I. Conidia smooth.

1. Green, broadly spreading, ripe conidia globose, 4-5/t P. roqueforti.

2. Green, less spreading, conidia elliptical, medium, commonly

purpled :.....". purpurogenum.

3. Gray or oliA e green, conidia, 5 -7 by 3-5/* /'. digitatum.

II. < onidia delicate rugulose /'. rugulomm.

Key 2.— Species Determinable from Substrata.

these species the substratum establishes a presumption of identii

< !h e < lamemberl and Brie i:

1. Floccose, white unchangeable, no odor P. camemberti var. rogeri.

2. Floccose, white to gray-green, no odor 1' . cmnemberti.

:'.. Powdery, yellowish white, spores smooth, ammoniacal odor,

P. brevieaule var. (/labrum.
I. Powdery, yellowish white, spores tuberculate, ammoniacal odor,

P. hr< vieaule \-.\v. allium .
5. Forming yellowish-brown areas, spores rough, ammoniacal odor. . /'. brevieaule.

< h e I Roquefort :

I . < Ireen streaks inside the cheese P. roqueforti.

< litrua fruits:

1 . Colonies of mold, blue-green /'. italicum.

2. Colonies of mold, olive-green P . digitatum=olivaceum.

Pomaceous fruits (apples, peai 3, etc. i:

I. Blue-green colonies finally producing coremia /'. expansum.

Polj poracese - Boleti, Polypori, etc. i:

1. ('nli. nil'- green (yellowish green) spreading by stolons /'. stoloniferum.

Wood I pine):

Producing orange i" red stains in pine wood /'. pinophilum.

slds Bull. US— 10 7

98

CULTURAL STUDIES OF SPECIES OF PENICTLLIUM.

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100 CULTUKAL STUDIES OF SPECIES OF PENICILLIUM.

Table 2. — Gelatin and color reactions.

Species.

No.

1
2

3

4
5

6

7
8
9

10

II

12

13

14

15

16

17

18
19

20
21

22

23
24
25

26

27

28

29

30
31

32

33

34

35
36
37

38
39
40
41
42
43
44
45

46

47
'49

51

54

56

Name.

pinophilum.

brcvicauh . .

var. glabrum

var. album . .

camembcrti

var. rogeri .
clan forme. . .

lilacinum

granulatum.

itaUcum .
luteum..

expansum .
citrinum. .

digitatum

pur purogenum .

roqucforti.
roseum...

duclauxi.
rubrum.

commune .

cf. 26

chrysogenum.
st olonif crura .

duaricatum.

cf. 26

decumbens . .

atramentosum .
biforme

= 19

funiculosum...

cf. 26

spinulosum.

rugulosum..

intricatum.

=38.

15 per cent gelatin in water.

Liquefaction.

Very slow, 6

weeks.
Rapid, 5 to 6

days.

....do

do

None or very

slow.

do

Slow

14 to 10 days

None

None or slow.

None

6 days

None in 15 days.
Partial, 14 days .

Rapid, 4 to 6

days.
None in 15 days.
....do

....do

Rapid, 8 days.

None in 15 days.
(?)

Partial, 15 to
20 days.

do

Rapid StolOdays
do

.do.

Rapid, 6 to 8

days.
Very slow, 20 -f

days.
Rapid

Very slow.
Rapid

Partial, slow,

15+ days.
None

.do.

Rapid

None or ?

Rapid, 6 to 7

days.
Rap id, 6to 10 days
None in 15 days.

do "...

Rapid

Litmus.

Acid

Alkaline .

.do.
.do.
.do.

do...

Neutral . .
Alkaline .
....do...

Faint alkaline

Acid

Alkaline

.do.
.do.

.do.

Alkaline .
do...

See color

(?)
Slow alkaline

Alkaline .
....do...
....do...

....do...

do...

Acid

Alkaline .

Acid

Alkaline.

Neutral?.

Acid

....do...

Alkaline.
Neutral?.
Alkaline .

None.
None or ? . . .

Rapid

Partial, slow

None or ? . . .

Rapid.
None . .

None or ? slow.

Partial and

slow.
Rapid

.do.
.do.
.do.
.do.

Acid

Neutral?.
Alkaline.
Acid

Neutral?.

Alkaline.
do...

.do.
.do.
.do.

Color, a

In conidial
mass.

Green.

Drab to choco-
late.
White or cream
...do

Gray-green

In medium.

Yellow to red.
None

White ,

Olive

White or lilac .
Yellowish

green.
Bluish green. .

Green, small
areas.

Pale b 1 u e-
green.

Deep green

Green to gray-
green.

Green

.do.
.do.
.do.

do

Brownish

None

Yellow to

orange.
None

None or ? .
None

.do.
.do.

Olive.
Green.

....do

White or sal-
mon.

Olive or green.

Green or vari-
ous.

Gray-green

Green

Deep green.
Green

Lemon yellow.

None

Red or no ...

None . .
do.

(Yellow, acid.

\Red, alkaline.

(?)

None

.do.

Yellowish
green.

Gray-green to
green.

White, some
green.

Green

Gray or green-
ish.

Gray-green

White or

cream.
Yellowish

brown.

Gray or green.
Gray to blue-
green.

Deep green

Gray-green

Green.

(?)

Green

Light olive

(?)
Green

.do.

(?)
Gray to green-
ish.
Yellowish
green.
(?)

(?)

do

do

Yellow in cer-
tain media.
None or golden

None or ?

Yellow! violet!

None or slight .

None

None or yel-
lowish.

None or red-
dish.

None

.do.

None . .
do.

None or brown

None

do

(?)
Red or none...
Red

(?)
None

None or yel-
lowish.

(?)
None

.do.

(?)
(?)

Reverse of
colony.

Red.

None.

Do.
Do.

Cream.

Do.
Brownish.
None.
Yellow to

orange.
Brownish

areas.
Yellow to

orange.
None.

Do.

None or brown-
ish.

None or yel-
lowish.

Brownish.

Yellow to red
or none.

None.

White or
cream.

Yellow, acid.

Red, alkaline.

Reddish.

or

yel-
yel-
yel-

White
cream.
Do.
Do.

White or
lowish.

None or
lowish.

Cream or
lowish.

None or yellow-
ish.
Do.

None.

None or yel-
lowish.

Yellowish t o
salmon.

Cream.

None.

White.
Do.

None or brown.
Cream.
Do.

(?)
Red or none.
Orange to red.

Cream or vio-
let.

Yellow to
orange red.

White or sul-
phur.
Uncolored.

a -'None" means the absence of a definite color, varying from hyaline to cream at times.

COMPARATIVE CULTURAL DATA.
Table 3.— Comparativt cultures.

101

No.

Conn's

solution.

Raulin's

fluid.

-1
5

6

7
B
9

10

11

V2

13

14

16

17

In

19

!

21

24
25

J 7

28

31
32
33
34

36
37

411
41
42

41

■I.",
46
47
19

51

54

56

Germ!

Slow, b u t

typical.
( ; ermlnated

only.

Fair, white. .

Gennl

.do

\ ery heavy

growth do. .

Typical.

Weak, lyp-

Leal.
i; ermlnated Typical —

. .do

slow, yellow
Slight... slow, typ-

ical.

Milk.

Milk +< ':ii Is,
curdling.

slow, l 5

da
in days

-

.do.

....do.

in .1 iys..

te of
tlon.

Very slow.
Rapid...

.do

Slow...

- .do

..do

Coloration.

Natural
substratum.

W I

ing.

R< i ..

None Sofl -

do... .

Data at 37° C.

.do.

.do.
.do.

...do

..do

Reddish
brown.

None

Slowly red .

Yellowish .

days.

Very slow growth in milk.
Rapid... None.

do

Came

lo.. .

Citrus fruits.

Small

no yel-
low.
QermJ
i.
Slight growth

do

i term!

Fair growth
.Small colo-

Slowly typ-
ical.

Not i j

Typical

Weak growth

Good....
Typical.

Good, some
yellow.

Some growth
rowth.

Richly typ-
ical.

.do

todays Slow .do

..do Yellow Ish

brown.
Rapid... Slighl yel

low.
poor growth in milk

Apples.

13 days.
8 days.

9 days.

13 days.

Typical. .

S|..w, typical s days

Typical.

Slowly typ

leal.
Tvpi.al...
.....do....
i ! ermlnated
Slowly up

Small colo i lood growth

iwth.. . Typical

do.

i

.do.

Not i

8 to 9 days.

9 days...
do..

.. .do

.do.

Very slow. .

From top

down.
Slow

Slow Slight red.

Rapid... None, or

escenl
greenish.

Oranges.

Roque fort
cheese.

....do.

Slow..

.do.
.do.
.do.

None
Yellow to
red.

.do

Rapid.

.do.

.do.

.do.

.do.

Slow . .

do.

do.

Rapid.

Slow..
Slight.

Gennl

Lnated .

Half typical.
do

Germinated .
No growth...

Germinated .

■rowth

Small colo-
nies.

Richly typ-
ical.

al

Weak

'.I days.

Typical .

... .
. . .do

9 days

Very slow. .

9 daj s.
7 days.

Very slow. . .
9 days

Slow, 3

weeks.
None, 4

weeks.

1 1 days

Little or

none.
Med i u m

rapid.
Little or

■ ne.
Fairly rapid.
Rapid

Rather slow.

do

. ..do

Little

Slow.

Rapid

Pale yellow

Golden yel-

low.
Slight,ifany Fleshy fungi.

Pale yellow

None

....do

....do

do..

....do.
do.

Yellow.

None. .

....do

Reddish
brown.

... . .
.do.. .
.do

Red where
touched.

Not colored
Golden yel-
low.

Slow.. . None

Slight do..

Slow do

Very slow do

do do.

slow.. .. ..do.

Gre'w

Grew well.

iwth.

Do.
Do.

Do.
Do.

;rowth.

No growth.

Killed.
Besl growth.

Killd.

No grow th.
Do.

S 1 i g b
growl h.

Killed.

i irew well.
None.

Killed.
No growth.

Do.
Do.

Do.

Slow growth.
Good growth.

Killed.

Do.

■will.

Do

Crew liest.
Killed.
drew best.

Slight

L'l'im th.
Killed.
No grow ill

Do.
Do.

Grew well.

Do.

Crew best.
No growth.

Crew i

No growth.

102 CULTURAL STUDIES OP SPECIES OF PENICILLIUM.

Table 4. — Comparative cultures in synthetic fluid (Dox's).

Carbon supplied as-

No.

9

10
11
12
13
14
15

16

17
18
19

20

21
22
23
24

25

26
27

28
29
30

31
32
33

34

35
36

37

38
39

40
41
42
43
44
45

46

47
49
51

54
56

Cane sugar.

1 to 20 per cent.

Typical.

Slowly typical

....do..
....do..

Typical.

.do.
.do.

.do.

.do.
.do.
.do.
.do.
.do.
.do.
.do.

60 percent.

No growth

Germi n a t e d
onlv.

Slight

....do

Slowly typical..

do -

Nocoremiabut

acid.
Slight

75 percent.

Not typical . .

Typical
do..

Not typical..

Typical.

do..

....do..

.do.
.do.
.do.

.do.

Typical .

Not typical .

Typical

....do

.do.

.do.
.do.

.do.

.do.
.do.

....do

Slowly typical

Typical

do

Not typical

Slow, typical...

No growth

Good, acid

....do

....do

Weak, no yel-
low.

Germ i n a t e d
only.

....do

Not typical . . .

No growth

Germ i n a t e d
only.

Slowly good ..

do

Weak

Good . .
....do.
....do.

.do.

Green, no

sclerotia.

....do

Weak

Germ i n a t e d

only.
Slight

Good

Very weak .

Slowly typical.

.do.
.do.

.do.
.do.

.do.

.do.
.do.
.do.

.do.
.do.

....do

Weak

....do

....do

Slowly typical.
....do

Slight growth .

do.

do.

Slow . .

Slight.

Lactose.

Growth.

Not typical . . .
Slow, typical..

....do..

do..

Typical .

No growth .

Germinated...
No growth ....
Germinated. . .

Slow

....do

Not typical . . .
Weak, hall typ

ical.
No growth i Germinated

....do

Very weak

growth.
Germ inated

only.

Slow, weak

Slight

....do

Weak, typical.
Slow, typical..

....do:

Slow growth. .

Litmus.

No effect.
....do...

....do.
....do.

Acid . .

....do...
No effect.

....do...

....do

Weak

No growth .
....do

....do

Very poor

....do

Germinated,
small.

Good

....do

....do

Fair

No growth .

Slight

No growth .
....do

.do.

Fair, not typ-
ical.

Typical

Weak

Very small col-
onies.

Typical

Sfowlytypical..

Good growth. .
Pinhead colo-
nies.
Hall typical...

Very weak .
Slight

....do

No growth

....do

....do

Good growth...
Pinhead colo-
nies.
Germinated

Weak

No growth . —
Pinhead colo-
nies.

Good

Weak

....do

Weak

Slight

Germinated.

....do

Rich growth..

....do

Small growth.

Typical

Slowly typical.
Weak..:

Typical

Slight

Slowly typical.

Slight.

do.

do.

....do...

'vcid

No effect.
....do...

....do...
....do...
....do...

.do.

Typical .
Sl'ight...

Slowly growing

Slowly typical..
Typical

....do..

Slight...
....do..
....do..
Tvpical .

Slight...

.do.

Good

....do

Germinated.

Slight

Slow, typical.

.do.

.do.
.do.
.do.
.do.

....do...

Acid

....do...

No effect.

Acid

....do...
No effect.

Acid

No effect.
Acid

No effect.
....do...
....do...

.do.

Acid

No effect.

.do.

....do.
Acid . .

do...

No effect.

do...

do...

Acid

No effect.

do...

Acid

No effect .
do...

.do.
.do.

0.9 per cent lac-
tic acid.

Germinated.

Typical, alka-
line.
Do.
Germinated.

Slight.

Not tvpical.
Slight.

Do.
Weak.
Good.

Do.
Slight.

Germinated.

Slow.
Weak.

Good, acid.

Good.
Do.

Typical.

Small colo-
nies.
Not typical.

Weak.

No growth.
Good.

Fair, not typ-
ical.

Do.

Small colo-
nies.

No growth.

Typical, alka-
line.

Little growth.
Do.

COM I'Ai; AT1VK CULTURAL DATA. 103

Table 4. -Comparativi cultures in synthetic fluid (Dox's Continued.

Carbon sti]

No.

1

.'

3

4

5

•

7

8

9

10

11

12
13

14
15
16
17

Is

19
20

::

24

26
27
28

30
31

33

34

39

4D
41
42

i:

41

45

i -
47
19
51

2.5 per ..'in ].-\ u-

i. acid

Fair, alkaline.

Typical...

slow . small. . . .

J

Slighl

i lood, alkaline.

Small colonies.

Germinal

Small

Weak.

Slight.

:: per cenl galao-

! . . . .

dine.

I ypical

do

] . acid . .
dine.

Typical

i

l . . . .

.do

Small, arid
i olonies

cenl glyc-
ei in.

< terminated only

..do

slight colony

typical

i

emia

Slight

•Will

i terminate L. ...
Slow, not typical.

Small

rowth.

•In.

Butterfat.

Grow 'I,.

Typical, slow.
Slow

do

slow , t \ ; .

do.

do

do

do

do

Slight.

Good.,
do.
.do.

Fair.

il.
Slowly typical.

J growth. .

ly typical.

I \ pi al

do

Little growth.

do

.do.

Slow .

' 'Will.

ical

da

do

do

Little growth.
....do

Very

"do.
Very small colo-
nies.
Slowly typical. . .
Germinated

( terminated.

Slighl

Germinated.

il

( terminated.
....do

Typical.
Good..

Half

typical.

( ...... i .

Hall" typical.
. Slow.'.

Typical.

do. .

Slaw

Slow ly typical.

Typical

low.

Very slow, typical

do

Slowly typical. . .

Weal..
Slow ....

do

al... .

Coremiform colo-
nics.
Slow . typical

Typical

slowly typical.

Color ill i Hum.

Fair

Small, typical.

Yellow to red.

. .1.
Tinged brownish.

A isll.

None.

Do.

Do.

Do.

Lemon yellow.

Slowl J

Do.
Yellow i a.

Do.
Do.

Reddish brown.
None.

Do.

Do

Do.

Slightly rusty.
None. '

Do.
Do.
Do.
Do.
Do.

Do.

yellow ish.

Nolle.

Slighl yellowish.

Nolle.

104 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

Table 5. — Decimal summary of comparative cultures in synthetic fluid (Dox's).

Carbon supplied as

Cane

sugar.

Lactose.

is
o

it

<D

w

_o

"3
>

3

O
H

tjC

a

8

3

A

U

o

ex

ID
m

O

-^

o

03
"el

o

o

u

■6
'a
c3
a
'-+3
o
03

.c
o

H

2"

w
C

03

43

%

O

M

C
O

"3

w

a

0

0

c

Name.

Up to 20
per cent.

c A

t< o
o

c

s- O

OJ —
t~

A

is

o

o

2

<

0
•0
0
0

'•&
09

No.

o

o

2

'3

<

OJ

+3

O

03

i

S3
O

1

pinophilum

brevicaule

1.0
-1.0
-1.0
-1.0

1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0

(?)

1.0
1.0

-1.0
1.0

1.0
0.0
0.0
0.0
1.0
1.0
1.0

(?)

1.0
0.0
0.0
1.0
1.0
1.0
1.0

(?)

0.0
0.0

0.0

(?)

0.1
0.1
0.1
0.1
-1.0
-1.0
0.2
0.1
0.3
-1.0
0.1
0.1
1.0
1.0
0.5
0.1
0.1
0.3
0.0
0.1

0.0
0.1
0.1
0.1
0.5
0.5
0.2
0.0
0.1
0.0
0.1
0.6
0.8
0.4
0.5
0.0
0.0

0.2

0.0
0.0

0.0

-1.0

-1.0

-1.0

1.0

1.0

0.2

0.1

0.5

0.3

0.2

0.7

-1.0

-1.0

-0.6

0.1

0.1

0.5

0.2

0.1

0
0
0
0

1
1

0
0
0

?1

0
0
0

(?)

0

(?)

0

0
0
0

1.0
0.8

1.0
1.0
0.4
0.4
0.5
0.5
0.2
1.0
1.0
1.0
0.6
0.1
0.3
0.5
0.2
0.2

0.2
0.1
0.1
0.3
-1.0
-l.C
0.3
0.2
0.4
0.3
0.1
0.4
-1.0
-1.0
0.6
0.1
0.0
0.4
0.2
0.2

0.8
0.8

0.1
-1.0
0.1
0.1
0.8
0.0
0.2
-0.7
0.1
0.1
0.1
0.2
0.4
0.5
0.7
0.1
0.1
0.2
0.2
0.1

0

9

0.1

0.1
0.1
0.1
+ 1.0
+ 1.0
1.0

1.0
1.0
-0.7
-1.0
1.0
1.0
1.0
0.4
0.6
1.0

1

S

var. glabrum .

var. album. . .
camemberti

var. rogeri

claviforme

lilacinum

1

4

1

5

6

7
8

1.0
1.0
0.9
0.8
1.0
0.9
1.0
1.0
1.0
1.0
1.0
0.4
0.4
0.8

1.0
1.0
0.1
0.3
0.6
0.3
0.3
0.4
0.8

0.4
0.1
0.6
0.4

0
0

1

0

9
10

granulatum

italicum

0

1

11

luteum

0

1?

0

13

1

14
15

expansion

citrinum

1

0

16

digitatum

1

17
18

purpurogenum . . .
roqueforti

0
0

19

0

90

duclauxi

0.3

0.8

1.0

(?)

?1

99

1.0
1.0
1.0

1.0
1.0
1.0
1.0

1.0
1.0
1.0
1.0
1.0
1.0
1.0

-1.0

-1.0

0.4

0.7

0.7

1.0

1.0

0.0

0.4

0.5

0.3

0.1

0.1

1.0

0.2

0.5

0.7

-0.9

-0.9

0.1

0.1

0.7

-0.8

-0.6

0.3

-1.0

0.3
0.6
0.3
0.7
0.7
-1.0
-1.0
0.2

0.2
0.0
0.0
0.0
0.7
0.5
0.8
0.3
0.4
0.4
0.0
0.0
0.8
0.5
0.1
0.0
0.8

-1.0
-1.0
0.6
1.0
-1.0
0.6
1.0

1

1

0

1

1

0

1

1.0
1.0
0.6
1.0

1.0
-1.0

-1.0

-1.0

0.5

0.7
0.7
0.9
0.9

1.0
1.0
1.0
1.0

0.8
0.5
-1.0
0.5
0.5
1.0
0.6

0.1
-1.0
0.6
0.0
0.1

(?)

23
94

commune

1.0
1.0
1.0

1.0
-1.0

1.0

-1.0

1.0

1.0

0.5
0.8
1.0
1.0

1

0

?5

(?)

9fi

chrysogenum

stoloniferum

0

27

98

1.0

0.5
0.5

0

(?)

99

0

30

1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0

1.0
1.0
1.0

(?)

1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0
1.0

-1.0
0.2
0.2
0.2
0.2
1.0
0.2
0.5
0.9
1.0
1.0
0.2
0.2
1.0
0.2
0.2
1.0
0.2

1

0
0

(?)

0

1

0
0
0

1
1

0
0

1

0
0

o

0

1.0
1.0
-1.0
0.3
0.3

0.2
0.2
0.3
-1.0
0.1

1.0
1.0
1.0
1.0
0.4

0.4
0.0
1.0

0.7

0

31

0

39

0

33

0

34
35

divaricatum

cf 26

0

36
37

decumbens

0.3
0.5
0.9
1.0
1.0
0.2
0.2

1.0
1.0
1.0

0.1

0.5

0.1

-1.0

0.8

0.1

0.1

0.5

0.6

-1.0

-1.0

0.1

0.4
1.0
1.0
1.0
1.0
0.3
0.3

0.7
0.7
0.0
1.0

1.0
1.0
0.5

0.1
0.3
0.1

0.7
0.7
0.1
0.0

1

0

38
39

atramentosum

biforme

0

1

40

1

42
43

funiculosum

cf. 17

0.3
0.3

1.0

0
0

44

cf. 26

45
46

spinulosum

rugulosum

intricatum

1.0

0.1

0

0.9
1.0
1.0

0

49
51

0.3

0

Explanation of decimals: 1.0 denotes normal development, or present; —1.0, slowly typical; 0.1, ger-
mination of spores only; decimals from 0.1 to 1.0 denote estimated amount of development between mere
germination and typical development; +1.0 denotes very rich growth.

COMPARATIVE CULTURAL DATA.
Table 6. Incubation experiments.

105

Species.

Name.

pinophilum

brevtcauli

nr.glabrwm.

var. album. .
camemberti

\ ir. rogiri. . .

clnrifurmt

lilucinum..

granulatutn

// ■llicum

lull um . .

apansum

citrinum

digit ilium

purpurogt num

roqveforti

roai um

iluclauii

Tubrum

1

2

3

•1

5

6

7

8

9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25

'.'7
28

30
31

33 .

34 ilirariculum..

35 cf.26

36 dtcti mlii us._ .

atra

biformt

cf. 23

-19

funiculosum.

cf. 17

cf. 26

s/iiniiliisum... .
ruijulnsum

commune

Chrysogt num...

■■/■■It r ii in .

20° C.

38

411
41
42
43
44

46

47

40 intrica

54

38

o

In

:

l.i'

1.0
1.0
1.0
1.0
1.0
1.0

1.(1

1.0
1.0
1.0

1.(1

1.0
1.0
1.0
1.0
1.0
1.0
1.0

t.o

1.0

1.0
1.0
1.0

1.(1

1.0

1.0
1.0
1.0
1.0
1.0
1.0
1.0

1
I
1

at37°(\

Compartment l. Compartment 2,

0.5 2° ('. 3.2 (i ( ...i\. 1 ( .

ant 5,
,av. i.5° C, av.

7°+ I . 8.7° C.

o

.0
1.0

l.o
l.ii
l.o

1.0
1.0
1.0
1.0
1.0

1.0
1.0

+ 1.0

1.0
0.0

0.0
0.0

(i o
0.0

1.0
0.0
O.ll

+ 1.0
0.0
0.0

0.0

0.3

O.ll
1.0

0.0

0.0

11.11

0.0
0.0
0.8

l.o
0.0
0.0

'6.6

0 i

+ 1.0
O.ll

+1.0

Growth, periods Growth, periods Growth,] Growth, periods

in daj ^. in m in da In da

0.1 I'.
o.oo.
o.oo.
0.0

0.0
O.I

0.0

O.l

1.1

0.1

0.00.

o.oo.

I.

1.00.
O.OO.

: i. 7. l.-,.

; l:..

lo.i o.l o.l o.l 0.1? 0.1? 0.1? ,0.1 0.1? 0.1? 0.1? 0.1 0.3 0.5

00.0 I

oo.o o.O 0.0

2 0.:.

Oil. 1

1 0.8

00.0

2 0..".
I o. 1
1 0.0
1 0. 1

1.0

o.o

0.0
(l.o
0.0

1.0

0.0
0.0

0.0

1.0
0.0

0.2

0.0
0.0
0.0
0.0
0.0

l.ol

1.0
-0.4

O I,

+ 1.0

o.o

0.0

0.0
1.0

0.0

0.0

1.0

o.o
0.0
0.0

0.0

O II

0.0
0.0
0.0
0.0

o. l o
0.0 o

0.0 o

0.3

0.0

0.6
0.5
0.6

o.o
0.6
0.9

0.1?

0. s
0.4
0. 1?
o.l
o. 1'.'

0.1

||. I

o ii
o :,

0.8

0.7

0.1
0.1

o.o

0.1

0.5

0.3
0.1
0.1

o. 1
o.l
0.6

0.6

o. 1

0 5
0.6

0.0 0.0

o. lo. l?0. I?
o. 1 o. 1 0.3

0.4 0.4 0.7

0.00.0

0. 0 0. 0

0.0

0.0

o.oo. l'.'o. 1
0.20. 1 o.l

0. 0 0. 0

0. 1 0. 1

o.o
o.l

0.0 0.0 o.il
0.0 (1.0 0.0

o.o o.o o. i o.2 o.;; 0.40.5
o.o o.o 0.2 o. 1 o. 1 o

0.5

o. I
0.4
o.o
0.6
0.8
0.0
o. I?

5 0.7
2 0.7
l?0. 1'.'

i. o.
0

0.0

o. I?

0.5

0.9

o. I?
ii I?

0.2

0. IV
O.l

0. 1?

0.0

II. 1
(1. 1

0.4

0.0

o.l

0.1

0.2

0.6

0.5

0.7

0.7

0.5

0.4

0.1

o. (IV
0.5

0. 1

o.l
0. I?
0. 1

0 J

o.o

0.0

(1. 1

0. 1

o.;;
o. l
0.7

i. i

0.6
o.l
o.l
0.2

0.7

0. s
i. :,
0.3
0.6

0.8

o. 1

o 1 o. 1

0.2

o. r.
o.l
0.5

0.1

o.r.

0.2

o 2

0.0

o.O

0.0
0.0

0.9
0.9

o.r.

0.3

0 o

(I..-, (I.

O.ll
il. 0

0.4

o. :;
o. 1

0.0

o l
o. r
o. i
0 ii

0 6

0 I

0.4
0.5
o. l
o. l
o.o

0.5

0.2 0. 1
0. I?

0.7 0.2
0.8 o 5

o. I o.o
0 l'.'o. I

0.3

0.;;

ii i
o. I

II O O 1

0.0

II. 0
0.7

o.ii
o..-,
o .;

0.4
0.8

0.0

0.3 0. 1

0.6
0.5

0.7 o.
o.r, o.

ii. l?0.
0.6 o.

o.o I
o. l'.'o.
o.ii o
0.0 1.

0.6 o

0. 1 0

I. l'.'o.
1.0 1.

0.0 o

0.0 (I.

O. III.

0.60.
.070.8 1.

l'.'o. I o.
.0 o. L'o,

0.0 1.
0.0 o,
0.2 0.
0.5 0.

'.i

3

7

l?|0.3|0. 1

20.

o 6

0.7
0.7

1.0
1.0

0.5

1.0

1.0
0.3

o _•
l.o

o

o. 1

1.0
1.0

0.8
0.5

0. 1
o. .",
0.7
1.0

0.0

1.0

1.0

ii 7

1.0

1.0

1.0

0.6
0.6
0.8

1.0

o.l

0.4

o.o
o.o

0.0

(1. 1

0.4

l.o
0.9

o 3 o.-". 0.7

ii.:.

o :,

0.7
0.5

0.7

o 6

o. .;

0.3

0.4

o.r, 1

0.6

0.0

0.2

o. 1
0.8
o 6

o l.o
0.4

0.8
1.0

1.0

1.0

l.o 1.0
0.0 1.0

0.6 0.8
0.7 0.8

0.7 o. s

0.2 0.3

o.7 0.7

0.3 0.3

o.o 1.0

ii. o 1.0

o. 10.7 0.8
o. l 0.3 oi

6

o.7 1.0
0.7 o 8

0. 1 0. 3 0. 6
7? 0.8

l.o
l.o
1.0
o 6
1.0

1.0

o.:,
0.2

l.o
1.0

0.3

0.1
1.0
0.6

o.l

1.0
1.0

1.0
1.0
0.9

o.O

0.3

0.8
0.3

1.0
1.0

0.9

1.0
1.0

0.6

l.o?

Explanation of decimals: 0.1 d< mtnatiort of conidia only; decimals op to 0.7. growth without

tin' formation ofcoli 0.7 to 1.0, 1 rtth colored conidia; 1.0 denotes typical colony.

+ 1.0 denotes growth more rapid at 37° than at .'0° C.

106 CULTURAL STUDIES OF SPECIES OF PENICILLIUM.

REFERENCES TO LITERATURE.

1. Bessey, E. A. Spore forms of Spegazzinia ornata Sacc. Journal of Mycology,

13, 43-45, 1907.

2. Brefeld, Oscar. Botanische Untersuchungen iiber Schimmelpilze. Heft 2:

Die Entwicklungsgeschichte von Penicillium. Plates. 98 pp., Leipzig, 1874.

3. Clements, Frederic E. Anecologic view of the species conception. Botanical

Society of America. Publication 34, pp. 253-264, Baltimore, May 30, 1908.

4. Conn. Herbert W. Bacteria in milk and its products. Illustrated, 306 pp.,

Philadelphia, 1903. See p. 268.

5. Dierckx, R. P. Un essai de revision du genere Penicillium Link. Annales de la

Societe Scientifique de Bruxelles, annee 25, 1900-1901, 1st fascicule, pp. 83-89,
Louvain, 1901.

6. Eustace, H. J. Investigations on some fruit diseases. I. Apple rots in cold

storage. New York Agricultural Experiment Station, Bulletin 297, Geneva,
1908.

7. Fries, Elias. Systemamycologicum. Vol. 3, Gryphiswaldse, 1829. See p. 407-

8. Greville, Robert K. Scottish cryptogamic flora, or colored figures and descrip.

tions of cryptogamic plants, belonging chiefly to the order fungi. Vols. 1-7,
Edinburgh, 1823-1828.

9. Hedgcock, George G., and Spaulding, Perley. A new method of mounting

fungi grown in cultures for the herbarium. Journal of Mycology, vol. 12, No.
84, p. 147, Columbus, 1906.

10. Lindau, G. Deutschl. Krypt. Flora. Pilze, vol. 8, p. 166.

11. Link, H. F. Linne species plantarum. Editio quarta, tomus 6, p. 70.

12. Observationes in ordines plantarum naturales. Magazin fiir die Neusten Ent-

deckungen in der Gesammten Naturkunde, Jahrg. 3, p. 16-18, 1809.

13. Maze, P. Les microbes dans l'industrie fromagere. Annales de l'lnstitut Pas-

teur, tome 19, No. 6, pp. 378-403, June 25; No. 8, pp. 481-493, August 25, Paris,
1905.

14. Micheli, Petro A. Nova plantarum genera. Illustrated, 234 pp., Florence,

1729. See plate 91, figures 1-4.

15. Milburn, Thomas. Ueber Aenderungen der Farben bei Pilzen und Bakterien.

Centralblatt fin Bakteriologie, abt 2, band 13, No. 5/7, pp. 129-138, Oct. 7; No.
9/11, pp. 257-276, Oct. 21, Jena, 1904.

16. Morini, Fausto. Sulla forma ascofora del Penicillium candidum Link. Mal-

pighia, anno 2, fascicule 5/6, pp. 224-234, Messina, 1888.

17. Oudemans, C. A. J. A., and Kontng, C. J. Prodome d'une flore mycologique

obtenue par la culture sur gelatine preparee de la terre humeuse du Spanders-
woud, pres de Bussum. Archives Neerlandaises des Sciences Exactes et
Naturelles, ser. 2, tome 7, No. 2/3, pp. 266-298. La Haye, 1902. See p. 288.

18. Peck, Charles H. Penicillium pallidofulvum, n. sp. New York State Museum,

Bulletin 67, Report of the State Botanist for 1902, p. 30, Albany, 1903.

19. Persoon, D. C. II. Synopsis methodica fungorum. Part 2, p. 693. Gottingen,

1801.

20. Powell, G. Harold, Stubenrauch, A. V., Tenny, L. S., Eustace, H. J.,

Hosford, G. W., and White, H. M. The decay of oranges while in transit from
California. U. S. Department of Agriculture, Bureau of Plant Industry, Bulle-
tin 123, Washington, 1908.

21. Roger, Georges. (Article in) Revue Hebdomadaire, vol. 7, p. 334, Paris.

22. Smith, Ralph E. The "soft spot" of oranges. (Penicillium digitatum.) Botan-

ical Gazette, vol. 24, No. 2, pp. 103-104, Chicago, 1897.

23. California Agricultural Experiment Station, Bulletin 184. Report of the Plant

Pathologist to July 1, 1906. Sacramento, 1907.

REFERENCES TO LITERATURE. 107

24. Stoll, Otto. Beitrage zur morphologischen unci biologischen Charakteristik von

Penicilliumarten. Inaugural Dissertation. Plates, 56 pp., Wurzbiirg, 1904.

25. Thom, Charles Fungi in cheese ripening: Camemberl and Roquefort. U. S.

Department of Agriculture, Bureau of Animal Industry, Bulletin 82, Wash-
ington, L906

26. Storrs Agricultural Experimenl Station, L7th Annual Report, L905, pp. , '■ I L5,

L90

27. Some suggestions from the study of dairy fungi. Journal of Mycology, vol.

1 1. No. 77, pp. 1 L7 I'-'l. Columbus, L90
28 Wehmer, Carl. Beitrage zur Kenntnis einheimischer Pilze. Hefl L: Zweineue
Schimmelpilze als Erreger einer Citronensaure-garung. Plates, 92 pp., Ban-
over and Leipzig, l"1 1

29. Zur morphologic und entwickelungsgeschichte des Penidllium luteum Zuk.,

cine- uberaus haufigen griinen Schimmelpilzes. Berichte der Deutschen
Botanischen Gesellschaft, Band II. Heft8,pp. 199 516, tafel 25, Berlin, L893.

30. Eine neue sklerotien-bildende Penicillium-species (P. italicum). Bedwigia.

Organ fur Kryptogamenkunde, Band 33, Eefl 4, pp. 211 -214, Dresden, L894.

31. Beitrage zur Kenntnis einheimischer Pilze. licit 2: (Jntersuchungen iiber

die FaulnisderPruchtc Plates, S4 pp., .Icna. 1*95. Seep. 68, plate 2.

32. Kleinere mykologische Mitteilungen. Centralblatl fur Bakteriologie, abt. 2,

Band 3, No. 6, pp. 1 17 153, Jena, Mar. 20, 1897. See p. I 1!).
(Teberdie Lebensdauer eingetrockneter Pilzkulturen. Berichte der Deutschen
Botanischen Gesellschaft, Band 22, licit 8, pp. 176-478, Berlin, 1904.

34. Morphologie, Physiologic und Systcmatik einiger technisch wichtiger adherer

Ascomyceten und verwandter Formen. Lafar's Bandbuch der Technische
Mykologie, Band I, Lieferung 11, pp. 192-238, Jena, L906. Sec p. 226.

35. Weidemann, Carl. Morphologische und physiologische Beschreibung einiger

Penicillium-arten. Centralblatl fur Bacteriologie, Abt. 2, Band 19, No. 21 23,
pp. 675 690, Nov. 11, and No. 24 25, pp. 755-770, Nov. 26, Jena, 1907.

36. Weigmann, II. Die Garungen der Milch und der Abbau ihrer Bestandteile. La-

far's Bandbuch der Technischen Mykologie, Band 2, Lieferung 9 and 1l', pp.
48-189, Jena, 1905-1906. See p. 18G.

INDEX OF SPECIES.

/'. nicillium — Page.

allium Epstein 50

titnim* ntosum

auri um I orda, <• aded Hedgcock 37

hn vicaule Saccardo 15

hr, vicault Saccardo, \ ar. album n. var 17

brevicauL Saccardo, var. glabrum n. var 18

camemh rli Thorn 50

ram, inhi rli Thom, \ ar. rogeri n. var 52

candidum Link 52

chrysogi nam n. sp 58

citrinum a. sp 61

claviformt Bainier 13

commune a. >\> 56

decumbt ns d. sp '1

digitatum Saccardo 31

</m aricatum a. sp

dud a a n Delacroix l-

ep«fc irai Lindau 50

. pansum Link, emended 27

funiculosum a. sp 69

glu a cam Link 26

jgrunuhit am Bainier II

intricatum d. sp 75

italic, im W. •Inner 29

lilacin, im n. sp

/»/< a m Zukal 39

um Wehmer 31

pinophilum Bedgcock, n. qov 37

purpurogenum 0. Stoll 36

roqueforti Tin mi 3 I

rost "m Link i '.' 1"

rubrum 0. Stoll 39

rugulosum a. -\> 60

spinulosum a. sp 76

stoloni/i rum n. Bp 68

L09

o

QK623.P4'T515BO,,n,C"°",,*nL"""1'
Th0|Tl 9.harles/Cul,ural studies of specie