b. Coccidioidomycosis 
Direct microscopic examination of wet, un- 
stained clinical material should be made, but, 
regardless of the direct findings, cultures 
and/or animal inoculations are indicated. 
Safety precautions: Petri plate cultures 
should never be made because they may lead 
to laboratory infection. Cultures should be in 
large test tubes which give broad agar sur- 
faces for inoculation. Specimens held for any 
length of time, or shipped to the laboratory, 
should be in tubes containing an antibiotic 
to retard growth of contaminating organisms. 
If the laboratory worker does not have the 
proper experience or equipment for handling 
the highly infectious cultures of C. immitis, 
the animal inoculation or “indirect method” 
for demonstration of the fungus should be 
used (see below). 
( 1 ) Specimen collection for laboratory 
examination 
(a) Merthiolated serum from serial 
blood specimens should be ex- 
amined by the complement fix- 
ation test. Add merthiolate to 
retard growth of contaminants. 
(b) Sputum, pus, pleural fluid, and 
gastric washings treated with the 
antibacterial antibiotics, chlor- 
amphenicol or a combination of 
penicillin and streptomycin, may 
be inoculated directly into mice or 
cultured on a selective medium 
such as that containing cyclohexi- 
mide and chloramphenicol. Pre- 
liminary identification of cultures 
by microscopic examination must 
be confirmed by animal inocula- 
tion. Unless safety hoods are 
available, however, it is advisable 
to use the animal inoculation or 
“ indirect method ” for the demon- 
stration of C. immitis. By this 
method, the handling of the highly 
infectious mycelial phase of this 
fungus is avoided. 
(2) Animal inoculation of cultures sus- 
pected of being C. immitis and the “in- 
direct method” for demonstration of 
C. immitis from the clinical materials. 
Mice may be inoculated intraperitone- 
ally and guinea pigs intratesticularly 
with culture suspensions, or directly 
with clinical materials which have been 
treated with antibacterial antibiotics. 
The male guinea pig is the animal of 
choice in determining whether a 
mycelial culture is or is not C. immitis, 
or whether the clinical material con- 
tains elements of this fungus. Intra- 
testicular injection of these animals with 
C. immitis gives rise to an orchitis and 
examination of fluid withdrawn from 
the testes will reveal the tissue form of 
C. immitis. If guinea pigs are not avail- 
able, inject mice intraperitoneally and 
examine lesions or lymphatic exudates 
for the fungus. 
c. Histoplasmosis 
If possible, culture media should be inocu- 
lated immediately following collection of 
clinical material. If the specimens are to be 
shipped or held for any length of time, they 
should be refrigerated because these organ- 
isms die out rapidly when exposed to higher 
temperatures. An antibiotic such as chlor- 
amphenicol should be added to specimens 
that are to be shipped.* Recovery of H. 
capsulatum by direct culture from clinical 
materials is not always difficult, but the 
chances of recovery are greatly increased if 
mice are inoculated and sacrificed for culture 
at the end of two weeks. 
( 1 ) Specimen collection for laboratory ex- 
amination 
(a) Blood smear examination is un- 
profitable except in acute, dissem- 
inated cases. 
(b) Citrated blood is of value for 
culture and animal inoculation in 
acute, disseminated cases. 
(c) Merthiolated** serum from serial 
blood specimens, taken at 3- to 4- 
week intervals, should be tested 
by complement fixation and agar 
gel precipitin tests. 
(d) Sputum specimens*** should be 
collected in sterile bottles to which 
chloramphenicol has been added. 
A minimum of six specimens 
should be collected from each 
patient before histoplasmosis is 
ruled out. 
* See appendix II, page xx 
** See appendix II, page xx 
* * * Sputum brought up from bronchial areas by having the 
patient use correct postural drainage procedures. 
18 
