Page 1457 - Small Animal Internal Medicine, 6th Edition

CHAPTER 91 Laboratory Diagnosis of Infectious Diseases 1429

some infectious agents constitutes a definitive diagnosis. It
is no longer recommended to use morphologic appearance
VetBooks.ir and Gram stain of bacteria to aid in the selection of empiric
antibiotics while waiting for results of culture and antimi-
crobial susceptibility testing as the results are not accurate
(Lappin et al., 2017).
For demonstration of most infectious agents, thin smears
are preferred. Blood can be prepared as follows: 1 drop of
blood approximately the size of a match head is placed at one
end of a clean microscope slide. The short edge of another
slide (i.e., spreader slide) is placed against the slide at a
30-degree angle and pulled back until the blood and the
spreader slide make contact. After the blood spreads across
FIG 91.4 the width of spreader slide, the slide is smoothly and quickly
Giardia cysts after zinc sulfate flotation. The cysts are pushed away from the blood across the length of the slide
approximately 10 × 8 µm. (“push” smears). For materials other than blood, the spreader
slide is laid gently on top of the material; the slides are then
smoothly and rapidly pulled apart on parallel planes (“pull”
smears). Cells in airway washings, prostatic washings, urine,
aqueous humor, and cerebrospinal fluid (CSF) should be
pelleted by centrifugation at 2000 g for 5 minutes before
staining. Multiple slides should always be made, if possible.
After being placed on the microscope slide, the material is
air-dried at room temperature, fixed if indicated by the pro-
cedure used, and stained. Slides that are not stained imme-
diately should be fixed by dipping in 100% methanol and
air-dried.
Cytologic specimens can be stained with routine stains;
immunocytochemical techniques for certain pathogens are
available (see Immunologic Techniques, p. 1432). Stains rou-
FIG 91.5 tinely used for the diagnosis of infectious diseases in small
Aelurostrongylus abstrusus larvae in an airway washing animal practice include Wright-Giemsa stain, Diff-Quik,
collected by bronchoalveolar lavage. (Courtesy Dr. Timothy Gram stain, and acid-fast stain. Immunocytochemical tech-
Hackett, Colorado State University, Fort Collins.) niques (e.g., fluorescent antibody staining of bone marrow
cells for feline leukemia virus) are only performed in refer-
ence or research laboratories (see Immunologic Techniques,
Preservation of Feces p. 1432). The laboratory should be contacted for specific
Feces should be refrigerated, not frozen, until assayed. If specimen handling information.
present, refrigerated Toxoplasma gondii oocysts will not
likely sporulate and become infectious. In addition, refriger- Bacterial Diseases
ated feces have less overgrowth of yeast, leading to fewer If bacterial disease is suspected, materials are collected
false-positive results. If a fecal sample is to be sent to a diag- aseptically and handled initially for culture (see Culture
nostic laboratory for further analysis and will not be evalu- Techniques, p. 1431). After slides are prepared for cytologic
ated within 48 hours, it should be preserved. Polyvinyl evaluation, one is generally stained initially with Wright-
alcohol, merthiolate-iodine-formalin, and 10% formalin Giemsa or Diff-Quik stain. If bacteria are noted, Gram stain
preservation can be used. Ten percent formalin is commonly of another slide is performed to differentiate gram-positive
used because of its routine availability; the clinician should and gram-negative agents. If filamentous, gram-positive rods
add 1 part feces to 9 parts formalin and mix well. are noted, acid-fast staining can help differentiate Actino-
myces (not acid fast) from Nocardia (generally acid fast).
CYTOLOGY If macrophages or neutrophils are detected, acid-fast stain-
Cytologic evaluation of exudates, bone marrow aspiration, ing is indicated to assess for Mycobacterium spp. within
blood smears, synovial fluid, gastric brushings, duodenal the cytoplasm; Mycobacterium spp. can often be seen on
secretions, urine, prostatic washings, airway washings, fecal Diff-Quik or Wright-Giemsa stained slides (see Fig. 74.2).
smears, tissue imprints, and aspiration biopsies is an inex- Bacteria can be present in small numbers or can be intra-
pensive and extremely valuable tool for the documentation of cellular (Bartonella spp.), so failure to document organisms
infectious agents; see individual chapters for cytologic find- cytologically does not totally exclude the diagnosis. Bacterial
ings for select infectious agents. Cytologic demonstration of culture of all samples with increased numbers of neutrophils