Guide to the Application of Genotyping to Tuberculosis Prevention and Control

Applying Genotyping Results to Tuberculosis Control Practices

Suspected False-Positive Culture Investigations

Goal

The goal of a suspected false-positive culture investigation is to confirm (or refute) the suspicion that one or more of the patients in a genotyping cluster has been falsely diagnosed with TB on the basis of a false-positive culture result.

Steps

The first step of an investigation of suspected false-positive cultures is to gather information to verify or refute that suspicion. Clues that are helpful in deciding if a false-positive culture occurred fall into two categories. The first type of clue comes from an analysis of the path that the suspect specimens took from collection through the final laboratory processing step to identify possible common collection or processing points and common times that could have resulted in cross-contamination. The second type of clue comes from reviewing medical records to identify patients in the cluster who, despite a diagnosis of TB, do not fit the typical clinical picture of the disease. The TB program should alert providers of any patient with a suspected false-positive culture result in order to determine the patient’s clinical status and if the patient is receiving anti-tuberculosis treatment.

Possible sources and locations of cross-contamination of clinical specimens include bronchoscopes, sputum collection areas, and laboratory processing steps. The laboratory that reported the suspect culture should be contacted and asked to provide information on all M. tuberculosis isolates from any specimens collected or processed on the same day or within a few days of the suspect isolate. If a contaminated bronchoscope or other instrument or a sputum collection booth is implicated in cross contamination, the respective health-care facility should be contacted and asked to provide information on other patients who were examined with the same instrument or had sputum collected in the same location. Information on all implicated specimens should be recorded and compared to identify potential overlap that could have resulted in cross-contamination. Realize that M. tuberculosis can remain viable in certain environments for days.

Clues to patients who may have been misdiagnosed with TB include patients diagnosed with pulmonary TB but who have normal chest radiographs, patients who were diagnosed with a different condition before the suspect M. tuberculosis culture results were reported, patients who have not been started on treatment for TB or who were started only after the culture results were reported, and patients who have had multiple specimens evaluated for M. tuberculosis but only one positive specimen. Finally, if genotyping results of isolates from suspected false-positive cultures were not the basis of initiating the investigation, those results should be obtained as soon as possible. All M. tuberculosis isolates that were collected or processed at the location or during the time that the cross contamination might have occurred should be genotyped.

Deciding whether to request RFLP analysis on isolates identified as part of a false-positive culture investigation depends on the strength of the available evidence. Experience has shown that if the laboratory or the health-care provider suspected a false-positive culture before the PCR genotyping results indicated a match, the PCR results are sufficient to confirm the presence of a false-positive culture. If, on the other hand, the PCR cluster results were the first indication of a problem, RFLP analysis of the clustered isolates should be requested.

Outcome

A suspected false-positive culture result should be considered “confirmed” as being false if a) all three genotyping methods show a match with the presumed source of the false-positive culture (or, if a previous suspicion of a false-positive culture existed and the PCR genotyping methods show a match), b) the investigation confirmed that the suspect isolates were processed at the same time or collected in the same location or with the same instrument, c) there is no other likely explanation for the findings, and d) the presumed misdiagnosed patient does not have a clinical picture consistent with TB. If critical specimens are unavailable for genotyping but all the other criteria are met, a suspected false-positive culture result should be considered “likely.”

A suspected false-positive culture result should be considered “unlikely” to be false (i.e., it is likely that the culture results of M. tuberculosis are correct) if the genotyping results do not show a match between the isolate from the suspected false-positive culture and other isolates processed at the same time or collected at the same place.

If the investigation leads to the conclusion that a false-positive culture result is confirmed or likely, the next steps are a) to identify which patients actually have TB and which patient or patients were misdiagnosed on the basis of false-positive culture results, b) to alert the involved health-care providers so that they can correctly diagnose and treat the misdiagnosed patient, and c) to alert the involved clinical laboratory or health-care facility so that the cause of the false-positive culture can be determined and corrected.

Identifying Sources of Error

Although it is possible to determine if a suspected false-positive culture result is confirmed, likely, or unlikely without first identifying the precise mechanism that led to the problem, it is obviously important to document this mechanism so that it can be corrected. Identifying the precise nature of the problem also aids in our understanding of how these types of errors can occur and the importance of adhering to procedures that will prevent them.

The laboratory or health-care facility that was involved should be contacted and provided the results of the investigation and the preliminary interpretation. In a collaborative fashion, the investigation should be finalized, and steps should be taken to describe as thoroughly as possible the precise mechanism that led to the false-positive culture result. Necessary procedural changes should be described and instituted, including updated quality-control and quality-assurance procedures. Technical assistance may be required and should be offered by state and national reference laboratories.

Communicating Results of Investigation

Once a final determination is made about the likelihood of a culture result being false and the likely source of error, the TB program should communicate the results of the investigation to the appropriate persons. The health-care providers of the patient or patients who were misdiagnosed need to receive this information immediately. The facility or laboratory that was determined to be the source of the error should also receive the final report of the investigation. Finally, there should be a formal reporting process for collecting and analyzing the results of false-positive cultures so that TB programs can monitor their frequency and track problems that can be remedied.

Possible Additional Steps

• Send a fact sheet to each implicated laboratory during the investigation describing the risk of M. tuberculosis laboratory cross-contamination and steps to prevent it.
• Assess and educate clinical laboratories when a false-positive culture event is identified.
• Provide local and statewide education to laboratory staff regarding prevention steps and early detection of persons who have false-positive specimens.
• Provide local and statewide education to local program staff regarding early detection of laboratory cross-contamination.
• Report the number of false-positive events, number of persons treated, number of months treatment was given, and basic characteristics of laboratories where these events occurred.
• Summarize these data on a national level for use in recommendations for mycobacteriology laboratory practice.
• Review findings periodically to determine whether specimen contamination has been reduced or whether contamination is suspected more frequently.