Clinical Laboratory - Microbiology

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Microbiologic Testing

Preanalytic Variables that determine the quality and yield of Microbiologic Testing

Normal Microflora

Microbiologic organisms are everywhere in the environment, and any body surface that contacts or connects to the environment may harbor these organisms. This includes many bacterial species and some fungi. Examples of sites include skin, vagina, upper respiratory tract above the epiglottis, and gastrointestinal tract. Colonization is the presence of organisms on a body surface or in a lumen, but not producing disease. Such organisms can be beneficial in generating nutrients (vitamin K) and in suppressing overgrowth of harmful organisms (such as Clostridium difficile). The organisms that form the normal flora of a body site are non-pathogenic under normal circumstances.

Contaminants in samples and the effect on culture results

Presence of normal flora complicates interpretation of culture results, because you must be aware of the possibility of contamination of a specimen in the collection process (e.g., did you have aseptic technique when the blood culture was drawn?

If the gram-stain and/or culture has multiple different organisms, and organisms that could be normal flora at the site of collection, and each organism in small amounts, then the possibility of contamination becomes more likely.

Is the sputum specimen mostly oral flora? If the sputum sample has many squamous epithelial cells, which come from the oral cavity, and no alveolar macrophages, which come from the distal aveoli, then it is unlikely that the sample represents lower respiratory tract.

How was the urine sample obtained? It could be contaminated by skin flora,

Some of the organisms most often to be part of normal flora (but might be pathogens under certain circumstances) include: Staphylococcus spp., Streptococcus spp., Lactobacillus spp., "diptheroids", Neisseria, various Enterobacteriaciae such as E. coli, Clostridium spp., and the ubiquitous fungus Candida.

For molecular diagnostic methods, collection technique is important to prevent contamination by even tiny amounts of genetic material. Testing of the sample should occur in a location separate from where the specimen was collected.

Sample collection techniques, specimen transport media, timing, and storage conditions

Sample collection requires using the appropriate device (swab, syringe, catheter, etc) and actually contacting the potentially infected site, not just waving the collection device somewhere in the vicinity of the patient.

Every laboratory has a manual or guide that provides information about collection, transport, and methodologies to assist in making sure that the appropriate samples are sent to the laboratory under the appropriate conditions.

The transport media must be appropriate to sustain the micro-organisms. For example, a blood culture requires both aerobic and anaerobic collection. For some organisms such as Neisseria gonorrheae specific media (chocolate agar) must be used. The shorter the transport time, the better the yield. Transport must be appropriate to organism requirements for temperature (not too hot, not too cold). Collection yield may be increased by timing to patient signs and symptoms, such as fever from sepsis.

Sample volume and identifying pathologic organisms in normally sterile sites

Diagnostic yield is improved with larger samples, because there is a greater chance that viable organisms will be present in the sample. For example, blood culture yield is improved by drawing a greater volume of blood.

Blood culture yield is also improved by drawing a series of samples at different sites and at different times, based upon variability in organism location, growth, and release.

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