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Dr Richard W Gilpin’s Blog

Legionella Dx by Real-Time PCR, Etc. More Reliable than Culture

[Question: Why does CDC push culture when it is becoming clear that it is inadequate?]

Improvement of Legionnaires’ disease diagnosis using real-time PCR assay:
A retrospective analysis, Italy, 2010 to 2015 - Maria Luisa Ricci et al. - Eurosurveillance, Vol 23, Issue 50, December 13, 2018.

“According to LD case definition, culture, a fourfold raise in Lp sg1 antibodies and urinary antigen test (UAT) are the only laboratory methods considered reliable for LD case confirmation. While serology has been nearly abandoned, UAT has almost completely replaced culture, representing 82% and 97% of diagnosis in Europe and in the United States (US), respectively.”

“Our results highlight a higher sensitivity of PCR compared with culture and a higher diagnostic efficiency compared with UAT. Furthermore, as recently stressed by other authors, it is important to perform more than one diagnostic assay in order to properly diagnose LD. Five of the eight LD cases with negative UAT results would have been missed if PCR assays, able to detect all Lp serogroups, had not been performed. Although in some instances UAT can incidentally detect non-1 Lp serogroups, they are designed to specifically detect Lp1 antigen, therefore, negative UAT results do not completely rule out LD infection. In addition to the aforementioned five cases (negative for UAT and for culture), three more culture-negative cases, resulted positive for Lp DNA by PCR. For these three, clinicians had only requested cultures and did not request UAT. Overall the eight additional cases show that even with a negative diagnosis but in presence of pneumonia, LD infection should be suspected and all available tests performed to investigate it.”

In this study, the use of real-time PCR resulted in an increment of eight (18.2%) identified LD cases and therefore is an objective improvement in the diagnosis of LD. Real-time PCR has been considered a poorly reliable method due to the risk of cross-contaminations, however, the introduction of automated procedures for DNA extractions and also for PCR set up, has resulted in a consistent improvement in preventing this PCR drawback. Therefore, after an appropriate validation of their own molecular tests, clinical microbiology laboratories can adopt PCR assays to detect Legionella in respiratory samples.

The reliability of PCR in diagnosing LD is recognized by the scientific community. Recent studies demonstrated a better performance of PCR compared with other diagnostic assays [culture, urine antigen]. PCR can also detect the presence of all Legionella species which are difficult to isolate by culture.

New approach to environmental investigation of an explosive legionnaires´ disease outbreak in Spain: early identification of potential risk sources by rapid Legionella spp immunosensing technique
Fernando Cebrián, et al. BMC Infectious Diseases201818:696. 27 December 2018.
htps://doi.org/10.1186/s12879-018-3605-8
“The culture method has been traditionally used to detect Legionella in a microbiological investigation. This method facilitates the characterization of the bacterial genome by sequence analysis and, if applicable, it allows establishing the relationship between strains isolated from environmental sources and those isolated from affected patients. Despite its advantages, either the overgrowth of accompanying organisms or the transition of Legionella cells into a viable but nonculturable state (VBNC) could compromise the culture results. In this VBNC state, cells are unable to form colonies on standard medium but they remain alive and infective.”

”Protocolized and immediate intervention in an outbreak is a crucial issue to reduce their effects on public health. For this, identification and control of the suspicious sources able to disseminate the bacteria and cause the illness is required. Rapid analytical techniques like IMS method based on the whole bacterial cell detection are shown as excellent tools to investigate all the potential sources of risk.”