Comparison of PCR and clinical laboratory tests for diagnosing H. pylori infection in pediatric patients

Abstract

Background: Histology and/or culture are generally considered the gold standard for the detection of H. pylori infection. Especially in children, these tests may result in a false negative outcome because of patchy distribution of the organism in the stomach mucosa. We have developed a PCR assay utilizing nested primer pairs directed against a subunit of the H. pylori urease gene (ureA). As part of a prospective evaluation of diagnostic tests to aid in detecting H. pylori infection in children, the aim of this study was to compare our PCR and Western blot assays with results obtained from histologic examination of biopsy specimens, rapid urease tests, and an FDA approved serologic assay and published PCR results to determine if we could validate the assays for diagnostic use on our patient population.

Results: Gastric biopsy specimens obtained from 101 pediatric patients were evaluated for the presence of H. pylori using histologic techniques, rapid urease (CLOtest) test and the PCR assay. Serum samples from each patient were assayed using both ELISA and Western Blot for antibodies to H. pylori. A total of 32 patients tested were positive by at least one of the methods evaluated. Thirteen patients had positive histology, 13 had a positive CLOtest, and 17 patients had positive H. pylori PCR. Out of the 13 CLO positive patients, 12 were positive by histologic analysis and all 13 were positive by PCR. Results of serologic tests on the same population did not correlate well with other assays. Twenty-eight patients showed serologic evidence of H. pylori infection, of which 9 were both CLO and histology positive and 12 were positive by PCR. Of the seropositive patients, 26 were ELISA positive, 13 were positive by Western blot, and 11 by both serologic methods.

Conclusions: The results obtained suggest that our nested PCR assay has the specificity and sensitivity necessary for clinical application when compared to standard histologic examination and rapid urease test. In addition, we found the current commercially available approved ELISA method appears unable to accurately detect H. pylori in this population. The Western blot assay yielded better concordance with CLOtest and histology, but not as good as the nested PCR assay.

Figure 1

Results of patient testing for H. pylori. Results of all testing (including biopsy-based methods and serology) are shown for the thirty-four samples included in the study which gave at least one positive result. Samples 20 and 21 were biopsies collected during endoscopic examinations performed 4 months apart on the same patient. Matched serum samples for this patient were obtained at each visit. Sample 31 was a biopsy collected from the antrum of a patient undergoing endoscopy, while sample 32 was from the body of the stomach of the same patient. Only one serum sample from this patient was received for analysis. The remaining sixty-seven patients (71 samples) were negative for all assays performed (data not shown). The BIOPSY column includes all specimens for which at least one test result was positive (CLOtest, PCR, or histology). The SEROLOGY column includes all serum samples considered reactive by either H. pylori ELISA (IgG and/or IgA), Western Blot (IgG and/or IgA), or both.

Figure 2

Sensitivity and specificity of the test methods evaluated for detection of H. pylori infection. The sensitivity and specificity of each test method was compared to our accepted biopsy gold standard – patients were considered to be infected with H. pylori if both rapid urease and histology tests were positive.