Chapter

Interpretation for Case Study 12-1:

Norovirus cannot be cultured. Laboratory tests include electron microscopy, serology, and RT-PCR. Electron microscopy and immune electron microscopy require specialized equipment and technical expertise. Detection of serum antibodies to the virus is not always straightforward, as most of the adult (but not child) population has serum antibodies to this virus. Furthermore, it can take several days after exposure to develop detectable IgM antibodies. RT-PCR, therefore, is the method of choice for detection of this RNA virus. The gene target is the viral RNA polymerase. Using this target, a broad spectrum of noroviral types can be detected. The results indicated that the virus was present in salad lettuce served at the hotel. Lettuce sampled directly from the distributor did not carry virus, indicating that the contamination occurred at the hotel. This assumption was supported by the discovery of viral RNA in hotel employees who had prepared the food. Direct sequencing of the RT-PCR products revealed identical sequences for all positive specimens, confirming that the guests, workers, and food source shared the same viral strain. Interpretation for Case Study 12-2:

Results from the culture of the isolates were consistent with MRSA. The results from the PFGE analysis indicated that all except one of the isolates from the students were the same strain. One isolate exhibited two differences from the others, indicating that it was closely related to these S. aureus isolates. Resistance to oxacillin/methicillin results from the expression of an altered penicillin-binding protein encoded by the mecA

gene. All five isolates shared the type IV mecA gene, associated with MRSA. PVL, also found in these isolates, is thought to be responsible for tissue necrosis in MRSA infections.

Further investigation into the cases revealed that all the students had participated in a wrestling meet at one of the high schools. Passage of the organism during this event was the likely source of the infection. The meet location was thoroughly cleaned according to CDC recommendations, and students were encouraged to always wash their hands and maintain good hygiene. Interpretation for Case Study 12-3:

The fact that the patient's viral loads were gradually increasing was a sign that the virus was developing resistance to the antiviral drugs. Slight variations of viral quantity within 0.3 log10 units are considered normal. This patient, however, was seeing significant increases in viral replication over the last 6 months.

This patient's virus has a mutation in the reverse transcriptase gene that has made the virus resistant to AZT. The patient's drug treatment needs to be changed immediately, with AZT being replaced by another reverse transcriptase inhibitor that would be unaffected by this mutation, such as didanosine or lamivudine. Viral load measurements should be taken regularly to make sure that the change in drug therapy causes a decrease in the viral load over the next few months. Genotyping should be performed again if the viral load starts to trend up.

1. Which of the following genes would be analyzed to determine whether an isolate of Staphylococcus aureus is resistant to oxacillin?

a. mecA

2. Which of the following is a genotypic method used to compare two isolates in an epidemiological investigation?

c. Ribotyping

3. For which of the following organisms does caution need to be exercised when evaluating positive PCR results because the organism can be found as normal flora in some patient populations?

b. Streptococcus pneumoniae

4. Which of the following controls are critical for ensuring that amplification is occurring in a patient sample and that the lack of PCR product is not due to the presence of inhibitors? d. Amplification control

5. A PCR assay performed to detect Bordetella pertussis on sputum obtained from a 14-year-old girl who has had a chronic cough had two bands, one consistent with the internal control and the other consistent with the size expected for amplification of the B. pertussis target. How should these results be interpreted?

b. The girl has clinically-significant B. pertussis infection

6. Which of the following is a disadvantage of molecular-based testing?

a. Results stay positive longer after treatment than do cultures

7. A molecular-based typing method that has high typing capacity, reproducibility, and discriminatory power, moderate ease of performance, and good to moderate ease of interpretation is:

c. PCR-RFLP

8. A patient has antibodies against HCV and a viral load of 100,000 copies/mL. What is the next test that should be performed on this patient's isolate?

d. Inno-LiPA HCV genotyping

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