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MEDICAL SCHOOL
LIIB1RAIKV




Gift of
Glanvilie Y. Rusk, M.D.



A TEXT-BOOK



OF



ACTERIOLOGY



BY

GEORGE M. STERNBERG, M.D., LL.D.

SURGEON- GENERAL U.S. ARMY

EX-PRESIDENT AMERICAN PUBLIC HEALTH ASSOCIATION; HONORARY MEMBER OP THE EPIDEMIOLOGICAL

SOCIETY OP LONDON, OP THE ROYAL ACADEMY OP MEDICINE OP ROME, OP THE ACADEMY

OP MEDICINE OF RIO DE JANEIRO, OP THE SOCIETE FRANCAISE D'HYGIENE,

ETC., ETC.



ILLUSTRATED BY HELIOTYPE AND CHROMO-LITHOGRAPHIC PLATES
AND TWO HUNDRED ENGRAVINGS.



NEW YORK
WILLIAM WOOD AND COMPANY

1896



COPYRIGHT BY

.1.1 AM WOOD & COMPANY,
189C.



PREFACE.



THE writer's Manual of Bacteriology, published in 1892, has been
very favorably received both in this country and abroad, but its use-
fulness has no doubt been to some extent restricted by the size and
expense of the volume. The following is an extract from the preface
of the Manual :

" A Manual of Bacteriology, therefore, which fairly represents the
present state of knowledge, will consist largely of a statement of facts
established by experimental data, and cannot fail to be of value to
physicians and to advanced students of bacteriology as a work of
reference. The present volume is an attempt to supply such a man-
ual, and at the same time a text-book of bacteriology for students
and guide for laboratory work. That portion of the book which is
printed in large type will, it is hoped, be found to give an accurate
and sufficiently extended account of the most important pathogenic
bacteria, and of bacteriological technology, to serve as a text-book for
medical students and others interested in this department of science.
The descriptions of non-pathogenic bacteria, and of the less important
or imperfectly described species of pathogenic bacteria, are given in
smaller type."

For the benefit of students of medicine and others who do not care
especially for the detailed descriptions of non-pathogenic bacteria and
the extensive bibliography contained in the Manual, this TEXT-BOOK
OF BACTERIOLOGY is now published. It comprises that portion of the
Manual above referred to as printed in large type, revised to include
all important additions to our knowledge of the pathogenic bacteria
since the original date of publication.



TABLE OF COISTTEOTS.



PART FIRST.

CLASSIFICATION, MORPHOLOGY, AND GENERAL BACTERIOLOGICAL

TECHNOLOGY.

PAGE

I. HISTORICAL, 3

II. CLASSIFICATION, 10

III. MORPHOLOGY, 20

IV. STAINING METHODS, 25

V. CULTURE MEDIA 37

VI. STERILIZATION OF CULTURE MEDIA, 50

VII. CULTURES IN LIQUID MEDIA, 60

VIII. CULTURES IN SOLID MEDIA, 67

IX. CULTIVATION OF ANAEROBIC BACTERIA, 78

X. INCUBATING OVENS AND THERMO-REGULATORS, . .86

XI. EXPERIMENTS UPON ANIMALS, 94

XII. PHOTOGRAPHING BACTERIA, 101



PART SECOND.
GENERAL BIOLOGICAL CHARACTERS.

I. STRUCTURE, MOTIONS, REPRODUCTION, 115

II. CONDITIONS OF GROWTH, 122

III. MODIFICATIONS OF BIOLOGICAL CHARACTERS, . . . .126

IV. PRODUCTS OF VITAL ACTIVITY, 130

V. PTOMAINES AND Tox ALBUMINS, .... . . 143

VI. INFLUENCE OF PHYSICAL AGENTS, .149

VII. ANTISEPTICS AND DISINFECTANTS (GENERAL ACCOUNT OF THE

ACTION OF), 160

VIII. ACTION OF GASES AND OF THE HALOID ELEMENTS UPON BAC-
TERIA, .168

IX. ACTION OF ACIDS AND ALKALIES, .... .176

X. ACTION OF VARIOUS SALTS,

XI. ACTION OF COAL-TAR PRODUCTS, ESSENTIAL OILS, ETC., . 193

XII. ACTION OF BLOOD SERUM AND OTHER ORGANIC LIQUIDS, . 204

XIII. PRACTICAL DIRECTIONS FOR DISINFECTION, . . . .208



ri TABLE OP CONTENTS.

PART THIRD.
PATHOGENIC BACTERIA.

PAGE

I. MODES OF ACTION, 221

1 1 . CHANNELS OP INFECTION 229

III. SUSCEPTIBILITY AND IMMUNITY, 233

IV. I'VOOENIC BACTERIA 275

V. I:\.-TKIM \ IN CROUPOUS PNEUMONIA, 300

VI. PATHOGENIC MIOROCOCCI NOT DESCRIBED IN SECTIONS IV.

AND V., 322

VII. THE BACILLUS OF ANTHRAX, 339

VMI. THE BACILLUS OF TYPHOID FEVER, .... .349

IX r.v< iKitiA IN DIPHTHERIA, 371

X. BACTERIA IN INFLUENZA 387

X I . BACILLI IN CHRONIC INFECTIOUS DISEASES 392

XII. BACILLI WHICH PRODUCE SEPTICAEMIA IN SUSCEPTIBLE ANI-
MALS 428

X 1 1 [. PATHOGENIC AEROBIC BACILLI NOT DESCRIBED IN PREVIOUS

SECTIONS, 461

XIV. PATHOGENIC ANAEROBIC BACILLI, . ... 531

XV. PATHOGENIC SPIRILLA, .... .... 549

XVI. BACTERIA IN INFECTIOUS DISEASES, .... 571



PART FOURTH.
SAPROPHYTES.

I. BACTERIA IN THE AIR 623

1 1 BACTERIA IN WATER, 636

III i'.ACTERIA IN THE SOIL, 652

IV. BACTERIA OF THE SURFACE OF THE BODY AND OF EXPOSED

Mucous MEMBRANES 658

v BACTERIA OF THE STOMACH AND INTESTINES, .... 668
vi I:\.TKIM.V OF CADAVERS AND OF PUTREFYING MATERIAL FROM

VARIOUS SOURCES 674

vil. BACTERIA IN ARTICLES OF FOOD, 677

INDEX, . 68 3



LIST OF ILLUSTRATIONS.



PAGE

1. Staphylococci, . . . . . . . . . 21

2. Zoogloea 21

3. Ascococcus, 21

4. Streptococci, 21

5 Tetrads, . . 22

6. Packets sarcina, . . . 22

7. Bacilli, 23

8. Involution forms, ........... 23

9. Chains formed by binary division, ........ 23

10. Spirilla, 24

11. Cladothrix 24

12. Flagella, 24

13. Platinum wire in glass handle, ........ 25

14. Flask for drawing off blood serum, . 38

15. Method of forcing blood serum into test tube, 38

16. Suction pipette, 38

17. Hot- water funnel, 42

18. Karlinski's agar filter, . . . . 44

19. Unna's agar filter, 45

20. Glass dishes for preserving potato cultures, 48

21. Test tube for sterilizing potato, . 48

22. Shape of potato for test-tube culture, 48

23. Hot air oven, . 52

24. Koch's steam sterilizer, . 53

25. Koch's steam sterilizer, 53

26. Arnold's steam sterilizer, ... 54

27. Miincke's steam sterilizer, ......... 54

28. Koch's apparatus for coagulating blood serum, ..... 55

29. Miincke s steam sterilizer and coagulator, 56

30. Pasteur Chamberlain filter, 57

31. Pasteur- Chamberlain filter without metal case 58

32. Modified Pasteur-Chamberlain filter, 59

33. Erlenmeyer flask 6 *

34. Flask used by Pasteur, 61

35. Platinum wire loop, 62

36. Platinum needle, 63

37. Sternberg's bulb,

38. Fermentation tube, 66

39. Method of making stick culture, 6 ?



i ji LIST OF ILLUSTRATIONS.

PAGE

o. 68

40. Sloping surface of culture medium,

40A. Growth of non-liquefying bacteria in gelatin stick cultures, .

41. Growth of same along line of puncture, ... ^

42. Growth of liquefying bacilli, ... ^

43. Colonies of bacteria, 73

44. Apparatus for gelatin plates, . . ^
Ksmarch roll tube

46. (See Fig. 15). 7S

47 Mode of development of a facultative anaerobic bacillus, .

48. Mode of development of strict anaerobic in long stick culture, . . 78

49. Exhausted-air flask for liquid media,

50. Method of displacing air with hydrogen,

51. Salomonson's tube

52. Frftnkel's method of cultivation

58. Sternberg's method of cultivation

54. Sternberg's method of cultivation

55. Buchner's method of cultivation,

56. Hydrogen generator,

Hydrogen apparatus for plate cultures .

58. Incubating oven

59. Thermo-regulator for gas

60. Moitessier's pressure regulator,

r,i. Mica screen for flame,

62. Koch's device for cutting off flame

63. Heichert's thermo regulator,

64. Bohr's thermo-regulator

65. MQncke's thermo- regulator

66. Sternberg's thermo-regulator

67. Gas valve for the same

68. D'Arsonval's incubating apparatus,

69. Roux's incubating oven and thermo-regulator,

70. Koux's thermo-regulator

71. Koch's syringe

7J. Sternberg's glass syringe

78. Pringle's photomicrographic apparatus, .

74. Sternberg's photomicrographic apparatus for gas,

75. Spores of bacilli

76. Method of germination of spores,

77. Apparatus for cultivating anaerobic bacilli l^->

Bacillus of mouse septicaemia in leucocytes from blood of mouse, . ~~><>

79. Staphylococcus pyogenes aureus,

80. Gelatin culture of Staphylococcus pyogenes aureus, . . . 27 :

81. Vertical section through a subcutaneous abscess caused by inoculation

with staphylococd in the rabbit 281

88. Pus containing streptococci, 2*7

88. Streptococcus of erysipelas in nutrient gelatin, 288

84. Section from margin of un erysipelatous inflammation, showing strepto-

cocci in lymph spaces, 289

85. Gouococci 2J.">

86. Gonococcus in gonorrhoea! pus 2JW

87. Gonorrhoea 1 conjunctivitis, second day of sickness, . . . 298



LIST OF ILLUSTRATIONS. i x

FIG. PAGE

88. Friedlander's bacillus, 308

89. Friedlander's bacillus; stick culture in gelatin, 309

90. Micrococcus pneumonise crouposae, 313

91. Micrococcus pneumonias crouposae, 313

92. Micrococcus pneumoniae crouposae, 313

93. Micrococcus pneumoniae crouposae, showing capsule, . . . 314

94. Single colony of Micrococcus pneumoniae crouposae upon agar plate, . 315

95. Micrococcus pneumoniae crouposae in blood of rabbit inoculated with

sputum, 319

96. Microcoecus of progressive tissue necrosis in mice, .... 328

97. Micrococcus of pyaemia in rabbits, .* 334

98. Micrococcus tetragenus, 326

99. Streptococcus of mastitis in cows, 333

100. Bacillus anthracis, showing development of long threads in convoluted

bundles, 340

101. Bacillus anthracis, showing formation of spores, 341

102. Culture of Bacillus anthracis in nutrient gelatin, . . . . . 342

103. Colonies of Bacillus anthracis upon gelatin plates, .... 343

104. Bacillus anthracis in liver of mouse 346

105. Bacillus anthracis in kidney of rabbit, 347

106. Bacillus of typhoid fever; colonies in stained sections of spleen, . . 353

107. Bacillus of typhoid fever; colonies in stained sections of spleen, . . 352

108. Bacillus typhi abdominalis, 358

109. Bacillus typhi abdominalis, 358

110. Bacillus typhi abdominalis, showing flagella, 359

111. Single colony of Bacillus typhi abdominalis in nutrient gelatin, . . 360

112. Bacillus typhi abdomiualis; stick culture in nutrient gelatin, . . 360

113. Section through wall of intestine, showing invasion by typhoid bacilli, 364

114. Bacillus diphtheriae, 375

115. Colonies of Bacillus diphtheriae in nutrient agar, ..... 376

116. Bacillus tuberculosis, .......... 394

117. Bacillus tuberculosis in sputum, ........ 395

lib. Section through a tuberculous nodule in the lung of a cow, showing

two giant cells, 397

119. Tubercle bacilli from surface of culture upon blood serum, . . . 400

120. Culture of tubercle bacillus upon glycerin-agar, 402

121. Limited epithelioid- celled tubercle of the iris, 408

122. Section of a recent lepra nodule of the skin, 414

123. Bacillus mallei, 417

124. Section of a glanders nodule, ......... 417

125. Section through a glanders nodule in liver of field mouse, . . . 420

126. Migrating cell containing syphilis bacilli, ...... 423

127. Pus from hard chancre containing syphilis bacilli, 423

128. Bacillus of rhinoscleroma in lymphatic vessels of the superficial part of

tumor, ............. 425

129. Bacillus septicaemiae haemorrhagicae in blood of a rabbit, . . . 439

130. Bacillus septicaemiae haemorrhagicae; stick culture in nutrient gelatin, . 431

131. Bacillus of Schweineseuche, 431

132. Colonies of bacillus of swine plague, ....... 431

133. Bacillus of Schweineseuche in blood of rabbit, 433

134. Bacillus of hog cholera, ... ... . 435



OF ILLUSTRATION-.



PAGE



185. Bacillus of mouse septictemia in leucocytes from blood of mouse,

186. Bacillus of rouget,

Ilacillus of mouse septicaemia ; culture in nutrient gelatin, .

188. Bacillus of mouse septicaemia; single colony in nutrient gelatin, .
m Section of diaphragm of a mouse dead from mouse septicaemia, .

140. Bacillus cavicida Havaniensis,

141. Bacillus crassus sputigenus

Proteushominiscapsulatus, * 5 *

148. Bacillus capsulatus,

144. Bacillus hydrophilus fuscus, ...

145. Culture of Bacillus hydrophilus fuscus in nutrient gelatin, .

146. Bacillus coli communis,

1 5:u illus coli communis in nutrient gelatin, 46*

148. A port ion of the growth shown in Fig. 147, 465

149. Bacillus lactis aSrogenes, 472

150. Bacillus acidiformuns 4 ^ 4

151. Culture of Bacillus acidiformans in nutrient gelatin, .... 4^4

152. Bacillus cuniculicida Havaniensis 4^5

158. Colonies of Bacillus cuniculicida Havaniensis, 476

154. Colonies of Bacillus cuniculicida Havaniensis 476

155. Bacillus pyocyanus, 479

156. Proteus vulgaris 485

157. " Swarming islands" from a culture of Proteus mirabilis, . . . 490

158. Spiral zooglcea from a culture of Proteus mirabilis, .... 490

159. Bacillus gradlis cadaveris 516

160. Colonies of B;u ill us gracilis cadaveris, . 516

161. Tetanus bacillus 532

162. Tetanus bacillus, 532

168. Culture of Bacillus tetani in nutrient gelatin, 533

164. Bacillus ocdematis maligni, 538

165. Bacillus cedematis maligni, 538

166. Cultures of. Bacillus cedematis maligni in nutrient gelatin, . . . 539
Hi?. Bacillus cadaveris, 541

168. Bacillus cadaveris 541

169. Bacillus of symptomatic anthrax, 542

170. Bacillus of symptomatic anthrax, ........ 542

171. Culture of bacillus of symptomatic anthrax, 543

172. Spirillum Obermeieri, 550

178. Spirillum Obermeieri, 550

174. Spirillum choleras Asiatic, 552

175. Spirillum cholerae Asiaticae, :>.YJ

176. Colonies of Spirillum cholera Asiaticae 553

177. Spirillum cholerae Asiaticae, 553

178. Cultures of Spirillum cholerae Asiatic;r in nutrient gelatin, . . . 554

179. Spirillum cholera; Asiaticae, 554

180. Colonies in nutrient gelatin of Spirillum cholerae Asiaticae, Spirillum

tyrogenum, and Spirillum of Finkler and Prior, .... 555

181. Section through mucous membrane of intestine from cholera cadaver, 559

188. Spirillum of Finkl.-r -md Prior 562

188. Colonies of Spirillum of Finkler and Prior, 5r,-j

184. Spirillum of Finkler and Prior; culture in gelatin, .... 562



LIST OF ILLUSTRATIONS. xi

FIG- PAGE

185. Spirillum tyrogenum, , 553

186. Colonies of Spirillum tyrogenum, 563

187. Spirillum Metschnikovi, 564

188. Penicillum glaucum, .......... 624

189. Miquel's aeroscope, 625

190. Hesse's aeroscope, 627

191. Miquel's flask, 629

192. Straus and Wiirtz's soluble filter, 629

193. Petri's sand filter, 630

194. Sugar filter, 631

195. Sedgwick and Tucker's apparatus, 631

196. Sternberg's vacuum tube, 637

197. Lepsius' apparatus for collecting water at various depths, . . . 638

198. Koch's plate method, 639

199. Smear preparation from liver of yellow-fever cadaver, .... 675

200. Bacillus cadaveris grandis, 675



PART FIRST.



CLASSIFICATION, MORPHOLOGY, AND GENERAL
BACTERIOLOGICAL TECHNOLOGY.

I. HISTORICAL. II. CLASSIFICATION. III. MORPHOLOGY. IV. STAINING
METHODS. V. CULTURE MEDIA. VI. STERILIZATION OP CULTURE
MEDIA. VII. CULTURES IN LIQUID MEDIA. VIII. CULTURES
IN SOLID MEDIA. IX. CULTIVATION OF ANAEROBIC BAC-
TERIA. X. INCUBATING OVENS AND THERMO REGU-
LATORS. XI. EXPERIMENTS UPON ANIMALS.
XII. PHOTOGRAPHING BACTERIA.



PAET FTEST.



I.

HISTORICAL.

IT is probable that Leeuwenhoeck, " the father of microscopy/'
observed some of the larger species of bacteria in faeces, putrid in-
fusions, etc., which he examined with his magnifying glasses (1675),
but it was nearly a century later before an attempt was made to de-
fine the characters of these minute organisms and to classify them
(O. F. Miffler, 1773).

In the absence of any reliable methods for obtaining pure cultures,
it is not surprising that the earlier botanists, in their efforts to classify
microorganisms, fell into serious errors, one of which was to include
under the name of infusoria various motile bacteria. These are now
generally recognized as vegetable organisms, while the Infusoria are
unicellular animal organisms.

Ehrenberg (1838), under the general name of Vibrioniens, de-
scribes four genera of filamentous bacteria, as follows :

1. Bacterium filaments linear and inflexible ; three species.

2. Vibrio filaments linear, snake-like, flexible ; nine species.

3. Spirillum filaments spiral, inflexible ; three species.

4. Spirochcete filaments spiral, flexible ; one species.

These vibrioniens were described by Ehrenberg as " filiform ani-
mals, distinctly or apparently polygastric, naked, without external
organs, with the body uniform and united in chains or in filiform
series as a result of incomplete division/'

Dujardin (1841) also placed the vibrioniens of Ehrenberg among
the infusoria, describing them as "filiform animals, extremely slen-
der, without appreciable organization, and without visible locomotive
organs."

Charles Robin (1853) suggested the relationship of Ehrenberg's
vibrioniens with the genus Leptothrix, which belongs to the algae ;
and Davaine (1859) insisted that the vibrioniens are vegetable organ-



HISTORICAL.

nearly allied to the algae. His classification will be found
in the " Dictionnaire Encyclop. des Sciences Medicales," art. " Bac-
teriej* " (1868). This view is also sustained by the German botanist
Cohn and is now generally accepted.

Spallan/ani, in 177>, endeavored to show by experiment that the
generally received theory of the spontaneous generation of micro-
organisms in organic liquids was not true. This he did by boiling
putrescible liquids in carefully sealed flasks. The experiment was
n <>t always successful, but in a certain number of instances the
liquids were sterilized and remained unchanged for an indefinite
Period. The objection was raised to these experiments that the oxy-
gen of the air was excluded by hermetically sealing the flasks, and
it was claimed, in accordance with the views of Gay-Lussac, that
free admission of this gas was essential for the development of fer-
mentation.

This objection was met by Franz Schulze (1836), who admitted air
to boiled putrescible liquids by drawing it through strong sulphuric
acid, in which suspended microorganisms were destroyed. He thus
demonstrated that boiled solutions, which, when exposed to the air
without any precautions, quickly fell into putrefaction, remained un-
changed when freely supplied with air which had been passed through
an agent capable of quickly destroying all living organisms con-
tain. M! in it.

Schwann (1839) demonstrated the same fact by another method.
Air was freely admitted to his boiled liquids through a tube which

-> heated to a point which insured the destruction of suspended
microorganisms. The same author is entitled to the credit of hav-
ing first clearly stated the essential relation of the yeast plant
lioromyces cereuisice to the process of fermentation in saccha-
rine liquids, which results in the formation of alcohol and carbonic
acid.

Helmholtz, in 1843, repeated the experiments of Schwann with
calcined air, and arrived at similar results i.e., he found that the
free admission of calcined air to boiled organic infusions did not pro-
<luct3 fermentation of any kind.

It was objected to these experiments that the air, having been
^'ihject.-d t" ;| hitfh temperature, had perhaps undergone some chem-
i - .ll change which prevented it from ina HI;- lira ting processes of fer-



This objection was met by Schroder and Von Dusch (1854) by a

\ simpl,. device which has since proved to ho of inestimable value

in bacteriological researches, These observers showed that a loose

plug of cotton, through which free communication with the external

air is maintained, excludes all suspended microorganisms, and that



HISTORICAL. 5

air passed through such a filter does not cause the fermentation of
boiled organic liquids.

The experiments of Pasteur and of Hoffman, made a few years
later, showed that even without a cotton filter, when the neck of the
flask containing the boiled liquid is long drawn out and turned down-
ward, the contents may be preserved indefinitely without change.
In this case suspended particles do not reach the interior of the flask,
as there is no current of air to carry them upward through its long-
drawn-out neck, and they are prevented by the force of gravity from
ascending.

Tyndall showed at a later date that in a closed chamber, in which
the air is not disturbed by currents, all suspended particles settle to
the floor of the chamber, leaving the air optically pure, as is proved
by passing a beam of light through such a chamber.

Notwithstanding the fact that the experimenters mentioned had
succeeded in keeping boiled organic liquids sterile in flasks to which
the oxygen of the air had free access, the question of the possibility
of spontaneous generation heterogenesis still remained unsettled,
inasmuch as occasionally a development of bacterial organisms did
occur in such boiled liquids.

This fact was explained by Pasteur (1860), who showed that the
generally received idea that the temperature of boiling water must
destroy all living organisms was a mistaken one, and that, especially
in alkaline liquids, a higher temperature was required to insure ster-
ilization. His experiments showed that a temperature of 110 to
112 C. (230 to 233.6 F.), which he obtained by boiling under a
pressure of one and a half atmospheres, was sufficient in every case.
These experiments, which have been repeated by numerous investi-
gators since, settled the spontaneous-generation controversy ; and it
is now generally admitted that no development of microorganisms
occurs in organic liquids, and no processes of putrefaction or fermen-
tation occur in such liquids, when they are completely sterilized and
guarded against the entrance of living germs from without.

Pasteur at a later date (1865) showed that the atmospheric or-
ganisms which resist the boiling temperature are in fact reproduc-
tive bodies, or spores, which he described under the name of " corpus-
cles ovoides " or " corpuscles brillants." Spores had been previously
seen by Perty (1852) and Robin (1853), but it was not until 1876 that
the development of these reproductive bodies was studied with care
by Cohn and by Koch. The last-named observer determined the
conditions under which spores are formed by the anthrax bacillus.
Five years later (1881) Koch published his valuable researches relat-
ing to the resisting power of anthrax spores to heat and to chemical
agents.



6 HISTORICAL.

The development of our knowledge relating to the bacteria,
stimulated by the controversy relating to spontaneous generation
and by the demonstration that various processes of fermentation
and putrefaction are due to microorganisms of this class, has
depended largely upon improvements in methods of research.
Among the most important points in the development of bacterio-
logical technique we may mention, first, the use of a cotton air
filter (Schroder and Von Dusch, 1854) ; second, the sterilization of
culture fluids by heat (methods perfected by Pasteur, Koch, and
others) ; third, the use of the aniline dyes as staining agents (first
recommended by Weigert in 1877) ; fourth, the introduction of
solid culture media, and the "plate method " for obtaining pure cul-
tures, by Koch in 1881.

The various improvements in methods of research, and espe-
cially the introduction of solid culture media and Koch's "plate
method" for isolating bacteria from mixed cultures, have placed
bacteriology upon a scientific basis, and have shown that many of
the observations and inferences of the earlier investigators were
erroneous owing to the imperfection of the methods employed.

Since it has been demonstrated that certain infectious diseases of
man and the lower animals are due to organisms of this class, phy-
sicians have been especially interested in bacteriological researches,
and the progress made during the past fifteen years has been largely
due to their investigations. It was a distinguished French physi-
cian, Davaine, who first demonstrated the etiological relation of a
microorganism of this class to a specific infectious disease. The an-
thrax bacillus had been seen in the blood of animals dying from this
disease by Pollender in 1849 and by Davaine in 1850, but it was sev-
eral years later (1863) before the last-named observer claimed to
have demonstrated by inoculation experiments the causal relation of
the bacillus to the disease in question.

The experiments of Davaine were not generally accepted as con-
clusive, because in inoculating an animal with blood containing the
bacillus, from an infected animal which had succumbed to the
disease, the living microorganism was associated with material
ti"in the body of the diseased animal. This objection was subse-
quently removed by the experiments of Pasteur, Koch, and many
>t IHTS with pure cultures of the bacillus, which were shown to have
the same pathogenic effects as had been obtained in inoculation ex-
periments with the blood of an infected animal.

The next demonstration of the causal relation of a parasitic mi-



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