Lottspeich et al. evaluated the novel Helicobacter pylori ClariRes assay (Ingenetix, Vienna, Austria) using feces of symptomatic children (2). This test, being a modified version of a biprobe 23S rRNA gene real-time PCR assay published by Schabereiter-Gurtner et al. (5), allows the detection and clarithromycin susceptibility testing of Helicobacter pylori in gastric biopsy and stool specimens. In the study of Lottspeich et al., the sensitivity of the ClariRes assay was found to be only 63%, differing considerably from that shown in the previous publication. The authors suggested that this could be due to the use of frozen instead of fresh samples and also to differences between the gastrointestinal tract of children and that of adults. However, in the study of Schabereiter-Gurtner et al. as well as in another recent study (4), stool specimens were also frozen until they were used for DNA extraction. This latter study, also performed using stool specimens from pediatric patients, showed a high degree of concordance between the ClariRes assay and the monoclonal stool antigen test; using the urea breath test (UBT) and serology as the reference methods, real-time PCR was shown to be at least as sensitive as the antigen test but more specific by far.
It is well known that preanalytic handling, and thus the quality of the sample, is of crucial importance with regard to sensitivity for any test aimed at the detection of microbial DNA in clinical samples. Thus, prolonged storage or transportation of feces at room temperature and repeated thawing of the sample or the DNA extract may cause a loss of sensitivity of the ClariRes assay. Unfortunately, this kind of information is not provided at all in the manuscript of Lottspeich et al. For example, it would be of interest to know how many times these stool specimens had been unfrozen in the past prior to examination by real-time PCR. Was the DNA used for PCR directly after extraction or had it been stored prior to the reaction? Was the stool antigen test performed at the same time as the real-time PCR or some time before? Were the samples used for each of the tests equivalent in quality?
Throughout the manuscript, Lottspeich et al. repeatedly refer to the study of Schabereiter-Gurtner et al., where samples were run in duplicate in the PCR. In 7% of the cases with positive H. pylori status, only one of the two reactions was positive when stool samples were analyzed. For pediatric patients, however, this percentage was shown to be considerably higher (22.5%). If the authors did not run samples in duplicate (this information is not provided in the manuscript), this would definitely be one of the main reasons for the poor sensitivity of the ClariRes assay.
Thus, as for many other tests, including nonmolecular ones (e.g., H. pylori culture tests, the sensitivity of which has been shown to range between 55.9% and 100% in recent literature) (1, 3), appropriate preanalytic and laboratory practice is of crucial importance for the accurate performance of the H. pylori ClariRes assay with regard to sensitivity.
REFERENCES
- 1.Kullavanijaya, P., D. Thong-Ngam, O. Hanvivatvong, P. Nunthapisud, P. Tangkijvanich, and P. Suwanagool. 2004. Analysis of eight different methods for the detection of Helicobacter pylori infection in patients with dyspepsia. J. Gastroenterol. Hepatol. 19:1392-1396. [DOI] [PubMed] [Google Scholar]
- 2.Lottspeich, C., A. Schwarzer, K. Panthel, S. Koletzko, and H. Rüssman. 2007. Evaluation of the novel Helicobacter pylori ClariRes real-time PCR assay for detection and clarithromycin susceptibility testing of H. pylori in stool specimens from symptomatic children. J. Clin. Microbiol. 45:1718-1722. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Monteiro, L., A. de Mascarel, A. M. Sarrasqueta, B. Bergey, C. Barberis, P. Talby, D. Roux, L. Shouler, D. Goldfain, H. Lamouliatte, and F. Megraud. 2001. Diagnosis of Helicobacter pylori infection: noninvasive methods compared to invasive methods and evaluation of two new tests. Am. J. Gastroenterol. 96:353-358. [DOI] [PubMed] [Google Scholar]
- 4.Puz, S., Z. Kovách, A. M. Hirschl, M. Häfner, A. Innerhofer, M. Rotter, and A. Makristathis. 2006. Evaluation of the novel Helicobacter pylori ClariRes real-time PCR assay for detection and clarithromycin susceptibility testing of H. pylori in stool specimens and gastric biopsies; comparison with the stool antigen test. Helicobacter 11:396. [Google Scholar]
- 5.Schabereiter-Gurtner, C., A. M. Hirschl, B. Dragosics, P. Hufnagl, S. Puz, Z. Kovách, M. Rotter, and A. Makristathis. 2004. Novel real-time PCR assay for detection of Helicobacter pylori infection and simultaneous clarithromycin susceptibility testing of stool and biopsy specimens. J. Clin. Microbiol. 42:4512-4518. [DOI] [PMC free article] [PubMed] [Google Scholar]