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Journal of Experimental Medicine

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TI-IE S P I R O C H E T A L

FLORA OF THE NORMAL MALE
GENITALIA.

BY HIDEYO NOGUCHI, M.D.

(From the Laboratories of The Rockefeller Institute for Medical Research.)
Pr~s

30 To 32.

(Received for publication, April 3, 1918.)
An a c c u r a t e knowledge of the varieties of spirochetal organisms
which n o r m a l l y inhabit the s m e g m a or the m u c o u s m e m b r a n e s of theurogenital region has b e c o m e i m p e r a t i v e for the e s t a b l i s h m e n t of an
etiological relationship b e t w e e n a spirochete a n d a disease in which
the organism m a y be found in the urine.
The classic work of Inada, Ido, Hoki, and others 1 on the presence of Leptospiraicterohmmorrhagice in the urine of convalescents from infectious jaundice has introduced a new procedure by which the disease may be easily diagnosed, and it isnatural that a similar procedure should be followed in the search for an etiologicalagent in other diseases of infectious origin.
Attention has been directed by Martin, ~ Nankivell and Sundell,* and Patterson* to the urine in eases of trench fevers. In fact, Nankivell and Sundell
early demonstrated minute spirochetes in specimens of urine from soldiers suffering from so called trench fever. Of 26 patients, most of them suffering from a"five-day fever," 99 specimens were examined, with 29 positive findings. Spirochetes were found in 12 out of 15 typical cases, while none of the 8 controls showeda spirochete (Fig. 32). The investigators considered the possibility of contamination of the urine from the smegma or from preputial sources, but it seemed to
11nada, R., Ido, Y., Hoki, R., Kaneko, R., and Ito, H., Etiology, mode of in.
fection, and specific therapy of Weil's disease (spirochmtosis icteroh~emorrhagica),
J. Exp. Med., 1916, xxiii, 377.
Martin, C. J., quoted by Nankivell and Sundell. a
3 Nankivell, A. T., and SundeU, C. E., On the presence of a spirochmte in theurine in cases of trench fever, Lancet, 1917, ii, 672.
4 Patterson, S. W., Preliminary note on spiroch~etes occurring in the urine
in cases of "P. U. 0.," J. Roy. Army Med. Corps, 1917, xxix, 503; Spirocha~tes
occurring in the urine in cases of "pyrexia of unknown origin," Brit. Med. 3"., 1917,
ii, 418.
667

668

SPIROCHETAL FLORA OF NORMAL MALE GENITALIA

them improbable that the spirochetes came from these sources, since the urine
usually contained the minute spirochetes unaccompanied by the coarse Spiro,nema refringens, and in case of contamination the presence of the large varieties was to have been expected. Moreover, in some positive instances no sp~ochetes could be discovered in the smegma. The occurrence of spirochetes in theurine was not constant, that is, not detectable on successive occasions, but wasrecurrent at irregular intervals. A certain relation seemed to exist between theappearance of spirochetes in the urine and the height of pyrexia, spirochetur~aoccurring usually 24 hours after the height of fever. I n still other cases they
appeared on the 14th to the 16th day. They were actively motile, averaged
8.15 # in length and 0.3 # in width, with an average of five curves; i.e., varying
from two and a half curves in 5 ~t to ten curves in 12.5 #. The spirals varied in
depth. The extremities tapered to sharp points with a flagellum at one or bothends. The organism differed from Treponema pallidum in its shortness and its
fewer spirals.
Patterson, using the Fontana, Wilmaers-Renaux,5 and India ink methods, examined specimens of urine from various groups of trench affections, 3 cases oftrench nephritis, 1 case of pyelonephritis, with abscess of the lungs, 15 cases ofrelapsing type of pyrexia of unknown origin, 1 case of myalgia following pyrexia
of unknown origin, and 5 cases of appendicitis (?)~not yet diagnosed, finding
spirochetes with the following features: They were about one to one and a halftimes the diameter of a red blood corpuscle, very thin, with tapering extremities,some having five to eight more or less regular curves, some being straight, somebowed, or lying in a semicircle (Fig. 33). The spirals were not so fine as thoseof Treponema pallidum or so coarse as those of Spironema recurrentis. The organisms took Giemsa's, Leishman's, or Romanovsky's stain poorly but were
easily demonstrated by the Indian ink method. Patterson depicts the spirochetes found in the abdominal type of cases as rather closely wound, short, thickforms, and those found in the relapsing type as much more tightly coiled. Little
attention was given to control cases.
The main objection to the work of the British investigators has been the possibility of an accidental contamination of the urine by unclean surroundings.
Many have insisted upon the necessity of collecting specimens by catheterization. The investigation of Stoddard 6 brought out an unsuspected source of
spirochetes in the periurethral as well as the intraurethral region of the male
genitalia. After examining 50 healthy soldiers and 50 miscellaneous hospitalpatients without history or symptoms of relapsing fever (trench fever), Stoddard
drew the conclusion that (1) spirochetes are not uncommon organisms in theurethra of men without history or symptoms of relapsing fever; (2) many dif5Wilmaers, L., and Renaux, E., Quarante-sept cas de Spiroch6tose icteroh6morragique, Arch. rr~l. Belges, 1917, lxx, 115, 207.
e Stoddard, J. L., Occurrence of spirochetes in the urine, Brit. M d . J., 1917,
ii, 416.

mDEYO NOOUCHI

669

ferent varieties are found; (3) some of the varieties seen are morphologically closely
similar to pathogenic varieties; (4) the spirochetes occur so definitely within
the urethra that they are an obvious source of contamination in uncatheterized
specimens of urine; (5) they are a sufficiently dangerous source of error even incatheterized specimens to deserve attention in careful work; and that finally(6) it is possible that a staining reaction or some other morphological charactermay be discovered to differentiate microscopically the common and harmlessfrom the pathogenic spirochetes. Of 50 hospital cases 56 per cent showed spirochetes, of which 46 per cent were not Spironema refringens. Of 50 American
soldiers spirochetes in the urethra occurred in 22 per cent, 2 per cent of whichshowed Spironema refringens also. Films from periurethral parts contained
more of the coarse refringens type.
The spirochetes found by Stoddard measured from 3.75 to 22 #, more commonly 6.75 to 9 #, but 11 # was not uncommon. They were either moderately
thick or extremely slender. The ends tapered or were blunt and formed a hook.
The spiral length varied from 0.5 to 1/z. Flattened and longer spirals also occurred, averaging 1.5 to 3.3 # and as long sometimes as 4 #. Stoddard states thata type with about eleven curves in 7.5/z occurs frequently, but I have not beenable to verify this finding. In some cases the spirals were exceedingly close andfine and almost impossible to count. They were often irregular, the deep narrow,deep wide, flat narrow, and fiat wide types of spirals sometimes occurring simultaneously in the same organism. The middle portion sometimes had looser
coils or none at all. In. one film many different varieties were often present,
including frequently organisms similar to Leptospira icterohcemorrhagice.7
Obviously it is n o t simple to interpret w h a t one sees in the richspirochetal m a t e r i a l as described b y Stoddard, who sees in it m a n ydifferent varieties, including the leptospira type. B u t , as this article is intended to show, a critical analysis of the spirochetal flora
reduces the n u m b e r of varieties to n o t m o r e t h a n three, or a t m o s tfour; namely, Spironema refringens, Treponema calligyrum, a n d Tre-

ponema minutum, n. sp.
Since the t i m e of S c h a u d i n n a n d H o f f m a n n a coarse spirochete
designated b y t h e m Sp#ochceta refringens h a s been k n o w n to i n h a b i t
the genital region, b u t no p a r t i c u l a r a t t e n t i o n has been given to thepossibility of the existence of other varieties. I t was n o t until the subject was t a k e n u p n o t only f r o m the morphological b u t also f r o m thecultural s t a n d p o i n t t h a t some interest c a m e to be a t t a c h e d to these7 Noguchi, H., Spirochceta icteroh6emorrhagi6e in American wild rats and its
relation to the Japanese and European strains. First paper, J. Exp. Med., 1917,
xxv, 755.

670

SPIROCHETAL FLORA OF NORMAL MALE GENITALIA

spirochetes. I n the present work a strain of Spironema refringer~s
w a s isolated a n d its morphological and cultural features studied,thus establishing its e n t i t y as a species. L a t e r a strain of spirochete (Treponema calligyrum) was o b t a i n e d f r o m a condyloma, which
resembled Treponema pallidum on the one h a n d a n d Spironema
refringens on the other, being an i n t e r m e d i a r y organism in its m o r p h o logical and cultural characteristics.
S u b s e q u e n t observations h a v e
led m e to regard th~s p a r t i c u l a r species as one of the m o s t c o m m o nvarieties t h a t are found in the flora of the s m e g m a or of the u r e t h r a lregion. I n fact, Treponema "calligyrum is m o r e frequently m e t w i t h
t h a n the b e t t e r k n o w n coarse Spironema refringens. T h e r e is, inaddition to these two varieties, another, m u c h smaller spirochetein the genital flora, which will be described in a subsequent p a r a graph. T h e s e three, the m i n u t e , m e d i u m , a n d coarse types, constitute the spirochetal flora which a t first glance present such acomplex aspect.
T h e s m e g m a and u r e t h r a l films f r o m six soldiers who were adm i t t e d to the H o s p i t a l of T h e Rockefeller I n s t i t u t e for t r e a t m e n t forp n e u m o n i a h a v e been examined. 8 T h e specimens were examined in
fresh condition u n d e r the dark-field microscope and also as stainedp r e p a r a t i o n s . F o r staining m e t h o d s G i e m s a ' s stain, F o n t a n a ' s silver
impregnation, 9 a n d occasionally Benians' Congo red negative im8 These specimens were obtained through the courtesy of Captain Henry T.
Chickering.
9 (a) Fix the air-dried film in Solution 1, which consists of
C6.

Glacial acetic acid . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
8Formalin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
20
Distilled water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
100
for 1 minute and wash well with water. (b) Mordant with Solution 2, which
consists ofTannin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
5 gm.
Phenol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1 cc.
Water . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
100 "for 1 minute over a gentle flame to the point of steaming, then wash thoroughly
in water. (c) Treat in a 0.25 per cent Silver nitrate solution to which one dropof ammonia is added to 40 cc. of the solution. The film turns brown in a few
minutes. Wash in water and then (d) cover with the mordant and warm it
over a flame until it begins to steam. Then wash the film in water and dry.

HIDEYO NOGUCHI

671

pression method were employed;1° also a mordant staining recommended by me for various spirochetes, includingTreponema pallidum.
~he method is similar to that advanced by Wilmaers and Renaux,b u t seems to give a better color value on account of the use ofgentian violet instead of fuchsin. The film is fixed in methyl alcohol for 15 minutes, then after being washed in water is covered witha solution of mordant (5 per cent tannin plus 1 per cent phenol)and held over a gentle flame for 1 minute, during which time it
begins to steam. I t is again washed in running water, covered
with a strong aqueous solution of gerltian violet to which 1 per centphenol has been added, and steamed briefly over a flame, then washedwell in water, air-dried, and examined. This method gives excellent results also with the pallidum. Care must be taken not tomake too thick a film.
The number of cases examined was small, but the finding wassuch that it was sufficient to determine the average flora in male
genitalia. The varieties of spirochetes encountered in most of thesmegma were the same as those found in one. All contained Treponema calligyrum and Treponema minutum, n. sp., u and most ofthem Spironema refringens, although the latter was absent in somecases.
No spirochetes were found in the films made from the urethralmucosa by means of a platinum loop. Just where the fault in thetechnique lay I am unable to explain. The finding was uniformly
negative also with the specimens of urine from ten soldiers. With
the idea that in nephritis cases there might be more possibility ofencountering spirochetes in the urine, ten different specimens fromacute as well as chronic cases of nephritis were subjected to a careful examination, but with no positive finding as yet. 12 In Table I
10A few drops of a 2 per cent Congo red solution (filtered) are mixed with adrop of the material suspected of containing a spirochete and spread over a cleanslide to form a film, The slide after being air-dried is immersed in a jar of absolute alcohol containing 1 per cent hydrochloric acid. In a few minutes the redcolor of the film turns to a bluish tint. The slide is then removed from theacid alcohol and air-dried.
lz Noguchi, H., Morphologicalcharacteristics and nomenclature of Leptospira(Spirochaaa) icterohcemorrhagice(Inada and Ido), J. Exp. Med., 1918, xxvii, 575.
13The specimens used in these tests were obtained through the courtesy of
Dr. W. W. Palmer of the Presbyterian Hospital.

672

SPIROCHETAL FLORA OF NORMAL MALE GENITALIA

are recorded some of the results obtained in the present study. There
are at least three different varieties distinguishable in the photomicrographs or under the dark-field microscope, a minute (minutum),a medium (calligyrum), and a large (refringens) type. Their biometriccharacteristics, as encountered in twenty-five specimens of each type,are given in Table I.
TABLE I.
Length.
Thickness. Spiral amplitude
Av- Exand intervals.
erage, tremes.

Spiral depth.

No. of spirals

Minute type.

7-10# 3-14#i 9.25-0.3# 0.9-1#. Fairly regularintervals.

0. 2-0. 5~t.
Some mayreach I# in
penultlmate
spirals.

7-10 spirals;vary
according tolength.

Medium type.

9-12# 4--14# 0.35-0.4~ 1.75#. Usually fairly
regular;that is, agiven amplitude iswell maintained in a
specimen.

0.5-1#. Often
flattenednear
themiddle instainedspecimens.
Reaches
1.5~
in
some.

5-8, varyinl
accordingto the splral amplitude; someonly 3.

Large type.

[12-16# 7-22~ ~0.7~

3-3;
quite
0.3-1.5#.
variable;
Almost conexceptionstant;inlive specially 8 in avery longmens changing the pospecimen.
sition of the
waves. In
stainedspecimens
often
irregularly
flattenedout.

T. pallddum.

8-14# 6-18~ O. 25-0.3# 1#

a~pe.

01" W a v e S .

I

2-3~. Usually moreor less regular.

0.8-1#

8-14;
16.

some

HIDEYO NOGUCHI

673

Some of the dark-field, as well as the ordinary photomicrographs,
representing the minute, the medium, and the large types are shownin Figs. 1 to 14. The minute type is decidedly smaller than Treponema pallidum and has a larger number of shallower spirals in proportion to its length (Figs. 1 and 5). There are also many shortspecimens such as are never found among the pallida. The medium
type has an aspect like that of atypical specimens of the pallidum.
The spirals are fairly deep but not so deep as those of a typicalpallidum, while the intervals between them are wider (Figs. 2, 5, 6,
9, and 10). All appear somewhat thicker than the pallidum (Fig.
21) when seen under the dark-field microscope. This does not
apply to the specimens stained by mordanting techniques (Fontana's
and the writer's), in which there occurs often an uncontrollable uneven heavy deposit of the dyes, due to various external factors(Figs. 5 to 14, 17, 20, and 22). Among organisms of the medium
type are noticed two forms, one with more closely set spirals and theother with wider ones, but this is due to certain temporary conditions and may be made to disappear or reappear by regulation ofcultural conditions. For example, there will be more of the wide,
flat spiral forms when the medium is more fluid. The large type ismuch heavier, comparatively short, with few spirals, and constantly
changes its curves (Figs. 4 and 8). The spirals of the minute type
become readily obliterated after the death of the organism (upperorganism in Fig. 8).
When fresh they all exhibit moderately active movements, rotary,lashing, and forward and backward locomotion. The large type isthe most energetic and the minute variety the least so. In many ofthe large type there is a distinct double contour effect upon examination under the dark-field microscope. All are provided with a terminal filament or flagellum at one or both ends.
As has been noted before, not all smegmata contain a spirochete,
and the varieties present may all belong to one or two of the threegroups. As a rule, however, all three types are present, the medium
type usually predominating.

674

SPIROCHETAL FLORA OF NORMAL MALE GENITALIA

Cultural Characteristics.
By selecting the smegma specimens which were rich in the typedesired, a culture of each of the three types described was obtained.
The technique employed was similar to that previously used for thecultivation 'of Spironema refringens 13 and Treponema calligyrum. 14
All require strict anaerobiosis (addition of fresh tissue to the media),
the presence of suitable body fluid (ascitic fluid), and an optimal
temperature (37°C.). The growth in the fluid medium, consistingof ascitic fluid and a piece of fresh rabbit kidney and a layer ofparaffin oil, is invisible, while in a solid medium, consisting of 2 partsof the neutral agar and 1 part of ascitic fluid with a piece of the fresh
rabbit kidney at the bottom, a faint haze appears to develop near
the tissue, gradually extending upward within a fortnight. No discrete, circumscribed, sharp colonies have so far been observed. In
this respect all the strains obtained are analogous to the culturesof various anaerobic treponemata and spironemata? n None produced a putrefactive or offensive odor, the absence of odor from theculture of the minute type serving to distinguish it from either
Treponema microdentium ~6 or Treponema mucosumY Carbohydrates
added to the culture media exert neither a favorable nor a retarding
influence upon growth, and no visible alterations of the media result
from their presence.
In young fluid cultures, whether of the minute, medium, or large
type, the organisms are short and active, b u t as they grow older (2weeks) the longer forms, some in chains, and some in tangled masses,
predominate, their motility meanwhile being considerably redaced.
The spirals are quite regular (Figs. 23 to 31). Very short forms dot, Noguchi, Pure cultivation of Spirochceta refringens, J. Exp. Med., 1912, xv,
466.
14Noguchi, Cultivation of Treponema calligyrum (new species) from condylomata of man, J. Exp. Meal., 1913, xvii, 89.
is Noguchi, Experimental research in syphilis with especial reference to Splroch~ta pallida (Treponema pallidum), J. Am. Med. Assn., 1912, lvlli, 1163.
16Noguchi, Cultural studies on mouth splroch~etab Treponema microdentium
and macrodentium, J. Exp. Med., 1912, xv, 81.
1~Noguchi, Treponema mucosum (new species), a mucin-producing spiroch~eta
from pyorrhea alveolaris, grown in pure culture, J. Exp. Med., 1912, xvi, 194.

HIDE¥O NOGUCHI

675

not appear in the solid media, the organisms appearing to attainaverage length within a short time. The spirals are remarkably
regular in solid media and so deep, in the case of the medium type,as to simulate a pallidum (Fig. 29). In older cultures two, three, andfour individuals in chains have occasionally been encountered (Fig.
27). Division in all three types is brought about b y transverse andperhaps also b y longitudinal fission.

Identification.
The morphological and cultural characteristics of the large typeshow it clearly to be a Spironema refringens, those of the medium typeidentify it with Treponema calligyrum. The latter type may be thesame organism as that described by Levaditi and Stanesco ts in 1909
as Spirochceta gracilis, found in a case of balanitis, but, as pointed out
previously, these authors used a name already designating anotherspironema from an ulcerating jaw, which is very different from thepresent medktm type. The name Treponema calligyrum was given toa non-pathogenic spirochete cultivated from the surface of a condyloma, b u t subsequent studies on the spirochetal flora of the genitalia
have convinced me that this type is one of the most commonly met
inhabitants of the genital region.
The minute type is not unlike the minute spirochete of the mouth,
Treponema microdentium, but its cultural characteristics differentiate
it from the latter. Treponema microdentium produces a peculiar
odor, especially when freshly isolated, and in a fluid medium the colorof the fresh tissue is made grayish within about 10 days and thefluid somewhat faintly opalescent. The minutum produces no odor
and remains without any perceptible action upon the culture medium,though in dimension there is a general resemblance.
In order to determine whether these two closely similar organisms
are immunologically related to each other, agglutination tests were
undertaken in which the action of a microdentium antiserum (rabbit)
was tested on both types. It was found that the serum caused amarked agglutination of Treponema microdentium in 1:500 dilutionb u t only a slight one with two different strains of the minutum, evenx8Levaditi, C., and Stanesco, V., Culture de deux spirochetes de l'homme(Sp. gracilis et Sp. balanitidis), Compt. rend. Sac. biol., 1909, lxvii, 188.

676

S P I R O C H E T A L F L O R A OF N O R M A L

M_ALE GENITALIA

in a dilution of 1: 20. In this connection it may be mentioned thata calligyrum serum (rabbit) gave a eopio,us agglutination with thecultures of the meciium type in a 1:200 dilution, but only a slight
one with those of the minute type in 1:20. There was a partial
reaction, but rLot marked enough to render the differenti&tion of thetwo types difficult.
In all probability, the minute smegma spirochete has been repeatedly observed by investigatbrs, but no special attention seems tohave been given to its identity. I have been accustomed to pass it
over as probably identical with Treponema microdentium. Now that
this type has been found to constitute an independent group, differentiated by several well defined features, it may well be known
under a separate name, Treponema minutum.
In the spirochetal flora of male smegma, therefore, only the threeforms, Spironer~a refringens, Treponema calligyrum, and Treponema
minutum, were recognized.
DISCUSSION AND SUMMARY.

The varieties of spirochetes enumerated and photomicrographedfrom the male smegma flora represent practically every form hitherto
described by Nankivell and Sundell and by Patterson in the specimens of urine from trench fever cases (Figs. 32 and 33). The urethral
flora, as studied by Stoddard, seem to contain more varieties, but,
except those of his more detailed morphological descriptions, every
form observed by him is among those found in the smegma. Stoddard saw certain forms with hooked ends suggestive of the Leptaspira icterohoemorrhagiceof infectious jaundice, but the resemblanceends with this one feature, and differentiation should always be
possible under the dark-field microscope, by means of which the leptospira reveals its highly characteristic minute elementary spirals,presenting the appearance of a chain of dots (Fig. 18). Fig. 19
shows that a very favorable fixation with the osmic acid vapor followed by Giemsa's staining may also bring out the elementaryspirals. Of all the spirochetes, none has so closely set spirals as thejaundice leptospira, the distance between two..spirals being only
0.5 ~. Various methods, including Fontana's, Benians', the mordantgentian violet stain, or Burri's India ink method, are inadequate

HIDEYO NOGUCHI

677

t o differentiate the leptospira from other spirochetes (Figs. 12, 14,
15, 16, 17).
W h y a positive spirochete finding with the films from the u r e t h r aand in the specimens of urine was not obtained, is difficult to explain,
except on the grounds of the p a u c i t y of specimens examined. A t all
events, the recent negative results reported b y Fiessinger 19 withF r e n c h soldiers a n d invalids after cleansing of the u r e t h r a and glansseem to be in h a r m o n y with m y results.
I n conclusion it m a y be stated t h a t Spironema refrlngens, Treponema calligyrum, and Treponema minutum represent practically all
the spirochetal forms observed in the male smegma flora. A leptospira has never been conclusively shown to be present in the specimens of normal urine or smegma. For the satisfactory microscopic
demonstration of a leptospira a dark-field illuminator is indispensable.
EXPLANATION OF PLATES.
PLATE 30.
Magnification, × 1,000.
FIGS. 1 to 4. Dark-field views of the spirochetes in a male smegma. Fig. 1
represents Treponema minutum, Fig. 2 Treponema calligyrum, Fig. 3 Spironema
refringens, and Fig. 4 a Splronema refrlngens (below) and a Treponema mlnutum.
FIGS. 5 to 11. Various types of spirochetes in smegma, stained by Fontana's
method.
FIG. 5. Two specimens of Treponema minutum.
FIG. 6. A specimen of Treponema minutum and two of Treponema calligyrum,of varying lengths.
FIG. 7. A group of Treponema calligyrum, with two specimens of Treponema

rninutum.
FIG. 8. A group of Spironema refringens from a sample of male smegma.
FIGS. 9 to 11. Treponema calligyrum from two different specimens of malesmegma.
FIGs. 12 and 13. Treponema calligyrum in preparation stained by the mordantgentian violet method. In Fig. 12 there are two specimens without distinct
spirals which closely resemble Leptospira icterohcemorrhagi~e in similar stained
preparations. Further study by means of a dark-field microscope is necessary
to determine whether they are leptospira or calligyrum.
FIG. 14. A group of Treponema calligyrum type from a specimen o~ malesmegma, stained by Giemsa's method. The organisms appear much thinnerhere than in specimens stained by other methods. A hooked spirochete resembling strongly the leptospira is seen near the left upper corner.
19Fiessinger, N., Apropos des Spirochetes du m~at et de l'urine de l'homme
normal, Compt. rend. Soc. biol., 1918, Ixxxi, 38.

678

S P I R O C H E T A L FLORA OF NORMAL MALE G E N I T A L I A

FIGS. 15 to 19. Leptospira icterohcemorrhagice under various conditions (forcomparison).
FIG. 15. Four specimens of Leptospira icteroh~emorrhagice stained by themordant gentian violet method. They appear blunt and curved and without
any indication of the minute elementary spirals which are the characteristicfeature of this genus. As they appear here they are indistinguishable from thestretched forms of the calligyrum type.
FIG. 16. A few leptospira~ as demonstrated by Benians' Congo red method,
Here, too, they do not show their elementary spirals.
FIG. 17. A group of Leptospira icterohcemorrhagi~z from a culture, stained by
Fontana's method. They fail to show their elementary spirals by this staining.
FIG. 18. A leptospira viewed under the dark-field microscope, showing its
minute elementary spirals.
FIG. 19. A number of Zeptospira icterohcemorrhaglce, fixed with osmic acidvapor and stained by Giemsa's stain, showing the elementary spirals.
FIGS. 20 to 22. Treponema pallidum under different conditions (for comparison).
FIG. 20. Treponema pallidum when stained by the mordant gentian violet
method.
FIG. 21. Treponema pallidum under the dark-field microscope.
FIG. 22. Treponeraa pallidum as stained by Fontana's silver impregnation
method.
PLATE 31.
Magnification, X 1,000.
FzGs. 23 and 24. Dark-field view of a culture of Treponema ~ninutum from amale smegma.
FIG. 25. Treponeraa minutum from a culture. Stained by the mordant gentian violet method.
FIG. 26. Similar specimens stained by Fontana's method.
FIG. 27. Dark-field view of a culture of Treponema calligyrum from a malesmegma.
FzG~ 28. A culture of Treponema calligyrum stained by the mordant gentian
violet method.
FIG. 29. Similar specimens stained by Fontana's method.
FIG. 30. Dark-field view of a culture of Spironema refringens from a malesmegma.
FzG. 31. Similar specimens stained by Fontana's method.
PLATE 32.
FIG. 32. Photographic reproduction of the photomicrographs of spirochetes
in Nankivell and Sundell's article on the spirochetes in the urine in trench fevercases. 3
Fla. 33. Photographic reproduction of the schematic drawings by Patterson
in his article. 4

THE JOURNAL OF EXPERIMENTAL MEDICINE VOL. XXVII.

PLATE 30.

(Noguchi: Spirochetal flora of normal male genitalia.)

THE JOURNAL OF EXPERIMENTAL MEDICINE VOL. X X V l h

PLATE 31.

(Noguchi: Spirochetal flora of normal male genitalia.)

THE JOURNAL OF EXPERIMENTAL MEDICINE VOL. X X V I I .

PLATE 32.

FIG. 32.

/
I. Spirochetes of Type I, abdominal P. U. O.
II. Type 2, relapsing P. U. O.
IIL $piri|lar form from urethra°Fie. 33.
(Noguchi: Splrochetal flora of normal male genitalia.)