Normalized real-time PCR for diagnosis of H. pylori infection

Abstract EN

Objectives : There is increasing interest in the use of quantitative PCR (q-PCR) for diagnosis of H.

pylori infection.

However, the assay remains largely unstandardized, making comparison between studies unreliable.

The objective of this study was to assess accuracy of a normalized q-PCR assay for diagnosis of the infection.

Subjects and methods : Seventy-six fresh gastric biopsy specimens were collected from patients undergoing upper gastrointestinal tract endoscopy and examined by rapid urease test (RUT), culture, and a commercial TaqMan q-PCR assay targeting the ureA gene.

Counts obtained from the latter assay were normalized to the human ACTB gene.

A subject was considered to be infected if two or more assays were positive.

Results : The detection rates were 42.1 %, 52.6 %, and 78.9 % by culture, RUT and q-PCR, respectively.

Bacterial density ranged 0.005 to 4800 bacteria per 100 human cells.

Because q-PCR showed low initial specificity (45.7 %), the cutoff value for the assay was recalculated as 1 bacterium 2 per 100 human cells, using ROC curve analysis.

Accordingly, the sensitivities and specificities were 79.5 % and 97.3 %, respectively, for culture ; 94.9 % and 91.9 %, respectively, for RUT ; and 94.9 % and 94.6 %, respectively, for q-PCR.

By gold standard, 39 of the dyspeptic patients (51.3 %) were found to be infected.

Conclusions : With the identified cutoff value, the q-PCR assay diagnosed H.

pylori infection with an accuracy slightly superior to that of RUT.

However, the possibility that low counts detected only by q-PCR represent true infections warrants further investigation.

Normalization of bacterial counts for standardization of q-PCR H.

pylori assays is recommended.