development within the animal before the FA test may be made or isolation begun.
Plague bacteriophage may be used to differentiate between plague and pseudotuberculosis organisms.
Since the routine cultivation of the etiological agent
in syphilis, Treponema pallidum, is as yet impossible,
the diagnosis rests at present on indirect methods
involving serologic tests except in the primary
disease. In this case, the organism may be demon-
strated microscopically in the dark field. The
laboratory diagnosis of syphilis, therefore would
hardly seem to fall within the scope of this manual.
The collection and handling of blood specimens for
serologic tests involves no more than observance of
aseptic technic to prevent contamination of the
specimens supplemented by storage and shipment
under such conditions as will prevent hemolysis
and breakage in transit.
Application of FA technic in syphilis
No special requirements have to be met in collect-
ing and handling specimens for FA staining in the
diagnosis of syphilis. Any specimen of blood that
has been collected aseptically in a sterile tube
and which has neither hemolyzed nor contains an
excess of chyle when received in the laboratory
will yield a satisfactory serum for FA staining.
Tuberculosis
Since M. tuberculosis may invade any organ of the
body, such varied specimens as sputum, pus, urine,
or spinal fluid may be sent to the laboratory for
examination. Sputum is the specimen most fre-
quently submitted. Patients must be taught the
difference between saliva and sputum and told that
an early morning specimen is generally most productive.
They should also be cautioned to deposit sputum in
the container and not contaminate the exterior,
which creates a hazard for all who handle the
specimen.
Pooling of sputum for several days is not desirable
because it may be toxic to tubercle bacilli present
and the degree of contamination may be increased.
Shipment of specimens through the mails must be in sterile screw-capped glass jars having resilient rubber liners in the caps and packaging must be in double mailing containers. The need for examination of repeated specimens should be stressed and the results of a single negative specimen should never be accepted as conclusive evidence of the absence of diseases. Specimens submitted in unsterile containers, in pill boxes, fruit jars, ointment jars, or in paper envelopes or on pieces of gauze are utterly useless and will not be examined by the laboratory.
Pathogenic fungi capable of causing pulmonary mycosis are sometimes encountered during culture work for tuberculosis. While certain saprophytic fungi also survive the processing of specimens, none of the fungi isolated should be casually discarded as harmless contaminants but should be referred to a competent mycology laboratory for identification.
Specimens to be collected for isolation of the agent
(1) Sputum, the thick yellowish-green exudate from the lungs and not saliva, should be collected in whatever type of sterile container is furnished by the laboratory and no other. Be sure the sputum is deposited within the container without soiling the exterior. Make sure the closure is leak-proof and forward the specimen to the laboratory in the double mailing container.
Gastric washings, collected in the morning on a fasting stomach, should be transported to the laboratory as promptly as feasible. Recovery of the tubercle bacillus from a gastric lavage is in direct proportion to the promptness of examination.
Urine may be contaminated with acid-fast
saprophytes present on the external genitals
so a catheterized specimen is preferable.
If catheterization is impractical, a mid-
stream sample, collected in a sterile
container, after careful washing of the
external genitals, may be acceptable.
Other materials such as samples of spinal, pleural or synovial fluid, and pus, may be
collected for diagnostic examination. Aseptic
collection in sterile tubes and, where indicated,
with an anticoagulant such as ammonium oxalate,
is essential to maintain the specimen in a
fluid state.
The laboratory diagnosis of tularemia may be accomplished by either a) the agglutination test or b) cultural examination. The extreme infectivity of Pasteurella tularensis makes routine culture inadvisable, however, and agglutina tion tests are the method of choice in the diagnostic laboratory.
Specimens for isolation of the agent
Culture of P. tularensis from any one of the natural
hosts such as ground squirrels, wild rabbits, wild
mice, quail, or other animal should not be attempted
except in a reference laboratory. If such an animal
is received for shipment, the body should be wrapped
in cloth soaked in cresol, placed in a container with
dry ice, and forwarded by air express to the laboratory.
Recognition of the organism in smears made from the
blood or tissues of the host is not possible; but
cultures may be made from any of the organs or from
the sputum, blood or exudates in human cases.
Specimens for serologic tests
Blood should be drawn aseptically, allowed to clot,
and the serum removed for agglutination tests.
Care should be taken to see that the blood is
not hemolyzed due to exposure to excessive heat
or cold in transit to the laboratory. Preferably,
at least two blood specimens should be obtained,
the first during the acute stage of the disease,
and the second about three weeks later, to show
a rise in antibody titer. Precipitin tests are
also possible; but these, together with FA tests,
probably will only be available in a reference
laboratory.
Infections with Enteric Bacteria
The specimens submitted for laboratory examina-
tion in typhoid fever and salmonellosis are blood,
feces, and urine for culture. Blood serum for
agglutination tests is not recommended. Blood cultures and serologic tests are not done in shigellosis. The multiplicity of fluids suggested as transport media for specimens to be cultured indicates that none is ideal and, therefore, isolation of the agent should be attempted in the nearest competent laboratory.
Specimens to be collected for isolation of
Salmonella typhosa
(a) Blood is the specimen of choice early in
the course of illness but the probability
of recovery of the organism decreases
rapidly after 7 to 10 days. Ten to
fifteen ml. of blood should be drawn
directly into a "B-D" blood culture
medium bottle and be examined for
growth after 3, 5, 7, and 14 days'
incubation. If the blood is not
drawn directly into the culture medium,
an anticoagulant should be added.
(b) Stool examination is accomplished by
adding approximately two grams, or a
portion the size of a small marble, of
formed stool to one ounce of Sachs 30%
glycerol in buffered physiological-
saline or Hajna's "SP" transport solu-
tion in a two-ounce, screw-cap bottle.
Shake the bottle vigorously to emulsify
the specimen before shipment. If the
stool is liquid, add 2 ml. of the speci-
men to the preservative in the bottle.
Be sure to include in the specimen any
bits of mucosa or mucus present in the
stool. Before shipment of the specimen,
make sure there is no leakage from the
bottle.
Urine is collected as cleanly as con-
venient, after cleansing the penis with
soap and water and wiping the external
meatus with 70% alcohol following by
sterile saline, and placed in a tightly
closed container for shipment or taken
to the laboratory promptly for immediate
culture. In typhoid fever, stool examination
is found to be more profitable than urine
culture.
(2) Specimens for serologic tests for S. typhosa
For agglutination tests, acute and convalescent
phase serum specimens should be collected.
The results of serologic tests in typhoid fever
and other salmonelloses are often difficult to
interpret so persistent efforts to obtain an
isolation of the offending organism should be
made. A few negative serologic results should
not be accepted as conclusive. Vi hemagglutination
cannot be used to diagnose typhoid but Vi hemaggluti-
nation appears to be a good screening procedure
for the detection of chronic typhoid carriers.
Food poisoning may be caused by certain members of
the Salmonella group. Preliminary epidemiological
investigation should reduce to a minimum the number
of suspected foods likely to be responsible in any
given outbreak, and indiscriminate collection of
samples is unwarranted. Such inquiries will indicate
the food consumed in common by those made ill and
although such evidence is not infallible, it is at
least presumptive.
(1) Specimens to be collected for isolation of
the agent.
If food from sealed containers is suspect,
the original container should be obtained,
if available, or a representative sample
transferred to a sterile sample bottle or
ice cream carton and refrigerated during
transit to the laboratory.
Blood specimens for culture from patients
in food poisoning are of no value.
Blood cultures are of value in the severe
enteric, typhoidal, and septicemic types
of salmonellosis. In such cases 10 to 15
ml. of blood may be drawn into a tube con-
taining an anticoagulant or a bottle of
blood culture medium.
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