Melissa B. Miller - Cases in Medical Microbiology and Infectious Diseases

The same RPR serologic assay is being used in a hypothetical population of octogenarian nuns, none of whom are or have been sexually active in at least 6 decades.

In this patient population, there is only one true case of syphilis, presumably acquired many years previously. The specificity of the test in this patient population is the same as it is in the individuals attending the STI clinic (in reality, it is likely to be different in different populations and also in different stages of syphilis). Because there is one case of syphilis, and 169 of the positive RPR results are false-positive test results, the positive predictive value in this patient population is only 0.6%. Clearly, this is a patient population in which the decision to test for syphilis using the RPR assay is not cost-effective.

In making a decision to order a specific test, the physician should know what he or she will do with the test results—essentially, how the results will alter the care of the patient. In a patient who the physician is certain does not have a specific disease, if the test for that disease has an appreciable rate of false-positive results, a positive test result is likely to be false positive and should not alter clinical care. Conversely, if the physician is certain that a patient has a disease, there is no good reason to order a test with a low sensitivity, as a negative result will likely be false negative. Tests are best used when there is uncertainty and when the results will alter the posttest probability and, therefore, the management of the patient.

Each laboratory test has three stages.

1 1. The preanalytical stage: The caregiver selects the test to be done, determines the type of specimen to be collected for analysis, ensures that it is properly labeled with the patient’s name, and facilitates rapid and proper transport of this specimen to the laboratory.

2 2. The analytical stage: The specimen is analyzed by the laboratory for the presence of specific microbial pathogens. The remaining sections of this chapter describe various analytic approaches to the detection of pathogens.

3 3. The postanalytical stage: The caregiver uses the laboratory results to determine what therapies, if any, to use in the care of the patient.

The preanalytical stage is the most important stage in laboratory testing!If the wrong test is ordered, if the wrong specimen is collected, if the specimen is labeled with the wrong patient’s name, or if the correct specimen is collected but is improperly transported, the microbe causing the patient’s illness may not be detected in the analytical stage. As a result, at the postanalytical stage, the caregiver may not have the appropriate information to make the correct therapeutic decision. The maxim frequently used in laboratory medicine is “garbage in, garbage out.”

Specimen selection is important. A patient with a fever, chills, and malaise may have an infection in any one of several organ systems. If a patient has a urinary tract infection and if urine is not selected for culture, the etiology and source of the infection will be missed. Careful history taking and physical examination play an important role in selecting the correct specimen.

Continuing with the example of a patient with a fever due to a urinary tract infection, the next phase in the diagnosis of infection is the collection of a urine specimen. Because the urethra has resident microbiota, urine specimens typically are not sterile. A properly collected urine specimen is one in which the external genitalia are cleansed and midstream urine is collected. Collection of midstream urine is important because the initial portion of the stream washes out much of the urethral microbiota. Even with careful attention to detail, clean-catch urine can be contaminated with urethral microbiota, rendering the specimen uninterpretable at the postanalytical stage.

An important concept when considering the transport of clinical specimens for culture is to recognize that they contain living organisms whose viability is influenced by transport conditions. These organisms may be killed by changes in temperature, drying of the specimen, exposure to oxygen, lack of vital nutrients, or changes in specimen pH. Transport conditions that support the viability of any clinically significant organisms present in the specimen should be established. It should also be noted that the longer the transport takes, the less likely it is that viability will be maintained. Rapid transport of specimens is important for maximal accuracy at the analytical stage.

If the correct test is selected, the proper specimen is collected and transported, but the specimen is labeled with the wrong name, the test findings might be harmful to two different patients. The patient from whom the specimen came might not receive the proper therapy, while a second patient whose name was mistakenly used to label the specimen might receive a potentially harmful therapy.

Once a specimen is received in the clinical laboratory, the first step in the determination of the cause of an infection is to examine it. Frequently, infected urine, joint, or cerebrospinal fluid specimens will be “cloudy” because of the presence of microorganisms and white blood cells, suggesting that an infectious process is occurring. Occasionally, the organism can be seen by simply looking for it in a clinical specimen or by looking for it on the patient. Certain worms or parts of worms can be seen in the feces of patients with ascariasis or tapeworm infections. Careful examination of an individual’s scalp or pubic area may reveal lice, while examination of the anal region may result in the detection of pinworms. Ticks can act as vectors for several infectious agents, including Rocky Mountain spotted fever, Lyme disease, and ehrlichiosis. When they are found engorged on the skin, physicians may remove and submit these ticks to the laboratory to determine their identity. This is done because certain ticks (deer ticks) act as a vector for certain infectious agents ( Borrelia burgdorferi , the organism that causes Lyme disease). Knowing the vector may help the physician determine the patient’s diagnosis.

Because most infectious agents are visible only when viewed with the aid of a light microscope, microscopic examination is central to the laboratory diagnosis of infectious diseases. Microscopic examination does not have the overall sensitivity and specificity of culture or the newer molecular diagnostic techniques. However, microscopic examination is very rapid, is usually relatively inexpensive (especially when compared with molecular techniques), is available around the clock in at least some formats in most institutions, and in many clinical settings, but by no means all, is highly accurate when done by highly skilled laboratorians. The organisms can be detected either unstained or by using a wide variety of stains, some of which are described below. Microbes have characteristic shapes that are important in their identification. Morphology can be very simple, with most clinically important bacteria generally appearing as either bacilli ( Fig. 1a) or cocci ( Fig. 1b). The bacilli can be very long or so short that they can be confused with cocci (coccobacilli); they can be fat or thin, have pointed ends, or be curved. The arrangement of cocci can be very helpful in determining their identity. These organisms can be arranged in clusters (staphylococci), pairs or diplococci ( S. pneumoniae ), or chains (various streptococcal and enterococcal species).