Application of PCR and Microscopy to Detect Helicobacter pylori in Gastric Biopsy Specimen among Acid Peptic Disorders at Tertiary Care Centre in Eastern Nepal

Nayanum Pokhrel, Basudha Khanal, Keshav Rai, Manish Subedi, Narayan Raj Bhattarai, Nayanum Pokhrel, Basudha Khanal, Keshav Rai, Manish Subedi, Narayan Raj Bhattarai

Abstract

Background: Helicobacter pylori infection is most prevalent in developing countries. It is an etiological agent of peptic ulcer, gastric adenocarcinoma, and mucosal-associated lymphoid tissue (MALT) lymphoma. Despite the development of different assays to confirm H. pylori infection, the diagnosis of infection is challenged by precision of the applied assay. Hence, the aim of this study was to understand the diagnostic accuracy of PCR and microscopy to detect the H. pylori in the gastric antrum biopsy specimen from gastric disorder patients.

Methods: A total of 52 patients with gastric disorders underwent upper gastrointestinal endoscopy with biopsy. The H. pylori infection in gastric biopsies was identified after examination by microscopy and 23S rRNA specific PCR. The agreement between two test results were analysed by McNemar's test and Kappa coefficient.

Result: H. pylori infection was confirmed in 9 (17.30%) patients by both assays, 6.25% in antral gastritis, 22.22% in gastric ulcer, 100% in gastric ulcer with duodenitis, 50% in gastric ulcer with duodenal ulcer, and 33.33% in severe erosive duodenitis with antral gastritis. Out of nine H. pylori infection confirmed patients, 3 patients were confirmed by microscopy and 8 patients by PCR. In case of two patients, both microscopy and PCR assay confirmed the H. pylori infection. The agreement between two test results was 86.54% and disagreed by 13.46% (p value > 0.05).

Conclusion: We found that PCR assay to detect H. pylori is more sensitive than microscopy. However, we advocate for the combination of both assays to increase the strength of diagnostic accuracy due to the absence of the gold standard assay for H. pylori infection.

References

    1. Maeda S., Mentis A. F. Pathogenesis of Helicobacter pylori infection. Helicobacter. 2007;12(1):10–14. doi: 10.1111/j.1523-5378.2007.00529.x.
    1. Moss S. F., Malfertheiner P. Helicobacter and gastric malignancies. Helicobacter. 2007;12(1):23–30. doi: 10.1111/j.1523-5378.2007.00539.x.
    1. McGee D. J., Mobley H. L. T. Pathogenesis of Helicobacter pylori infection. Current Opinion in Gastroenterology. 2000;16(1):24–31. doi: 10.1097/00001574-200001000-00005.
    1. Cheng H., Hu F., Zhang L., et al. Prevalence of Helicobacter pylori infection and identification of risk factors in rural and urban Beijing, China. Helicobacter. 2009;14(2):128–133. doi: 10.1111/j.1523-5378.2009.00668.x.
    1. Salih B. Helicobacter pylori infection in developing countries: the burden for how long? Saudi Journal of Gastroenterology. 2009;15(3):201–207. doi: 10.4103/1319-3767.54743.
    1. Testerman T. L., Morris J. Beyond the stomach: an updated view of Helicobacter pylori pathogenesis, diagnosis, and treatment. World Journal of Gastroenterology. 2014;20(36):12781–12808. doi: 10.3748/wjg.v20.i36.12781.
    1. Axon A. Helicobacter pyloriand public health. Helicobacter. 2014;19(1):68–73. doi: 10.1111/hel.12155.
    1. Hunt R. H., Xiao S. D., Megraud F., et al. Helicobacter pylori in developing countries. World gastroenterology organisation global guideline. Journal of Gastrointestinal and Liver Diseases. 2011;20:299–304.
    1. Fasciana T., Scarpulla G., Giammanco A., et al. Resistance to clarithromycin and genotypes in Helicobacter pylori strains isolated in Sicily. Journal of Medical Microbiology. 2015;64(11):1408–1414. doi: 10.1099/jmm.0.000163.
    1. Fasciana T., Capra G., Cala C., et al. Helicobacter pylori and epstein-barr co-infection in gastric disease. Pharmacologyonline. 2017;1:73–82.
    1. Thapa N., Shah K. B., Bhandari B. B., et al. Association of Helicobacter pylori infection and stomach cancer. Medical Journal of Shree Birendra Hospital. 2013;12(2):36–41. doi: 10.3126/mjsbh.v12i2.12927.
    1. Public Health England. Standards for microbiology investigations. Investigation of Gastric Biopsies for Helicobacter pylori. 2014;B55(6):1–21.
    1. Promega. Wizard® Genomic DNA Purification Kit. Madison, WI, USA: Promega; 2017. .
    1. Schabereiter-Gurtner C., Hirschl A. M., Dragosics B., et al. Novel real-time PCR assay for detection of Helicobacter pylori infection and simultaneous clarithromycin susceptibility testing of stool and biopsy specimens. Journal of Clinical Microbiology. 2004;42:4512–4518. doi: 10.1128/jcm.42.10.4512-4518.2004.
    1. Saiki R. K., Scharf S., Faloona F., et al. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science. 1992;230(4732):1350–1354.
    1. SPSS Inc. SPSS 11.5 for Windows. Chicago, IL, USA: SPSS; 2002.
    1. R Core Team. R: A Language and Environment for Statistical Computing. Vienna, Austria: R Foundation for Statistical Computing; 2018.
    1. Russmann H., Kempf V. A., Koletzko S., et al. Comparison of fluorescent in situ hybridization and conventional culturing for detection of Helicobacter pylori in gastric biopsy specimens. Journal of Clinical Microbiology. 2001;39(1):304–308. doi: 10.1128/jcm.39.1.304-308.2001.
    1. Trebesius K., Panthel K., Strobel S., et al. Rapid and specific detection of Helicobacter pylori macrolide resistance in gastric tissue by fluorescent in situ hybridisation. Gut. 2000;46(5):608–614. doi: 10.1136/gut.46.5.608.
    1. Ramis I. B., de Moraes E. P., Fernandes M. S., et al. Evaluation of diagnostic methods for the detection of Helicobacter pylori in gastric biopsy specimens of dyspeptic patients. Brazilian Journal of Microbiology. 2012;43:903–908. doi: 10.1590/s1517-83822012000300008.
    1. Jalalypour F., Farajnia S., Somi M. H., et al. Comparative evaluation of RUT, PCR and ELISA tests for detection of infection with cytotoxigenic H. pylori. Advanced Pharmaceutical Bulletin. 2016;6:261–266. doi: 10.15171/apb.2016.036.
    1. Khalifehgholi M., Shamsipour F., Ajhdarkosh H., et al. Comparison of five diagnostic methods for Helicobacter pylori. Iranian Journal of Microbiology. 2013;5:396–401.
    1. Siavoshi F., Saniee P., Khalili-Samani S., et al. Evaluation of methods for H. pylori detection in PPI consumption using culture, rapid urease test and smear examination. Annals of Translational Medicine. 2015;3:p. 11.
    1. Roy A. D., Deuri S., Dutta U. C. The diagnostic accuracy of rapid urease biopsy test compared to histopathology in implementing “test and treat” policy for Helicobacter pylori. International Journal of Applied and Basic Medical Research. 2016;6(1):18–22. doi: 10.4103/2229-516x.174003.
    1. Rotimi O., Cairns A., Gray S., et al. Histological identification of Helicobacter pylori: comparison of staining methods. Journal of Clinical Pathology. 2000;53(10):756–759. doi: 10.1136/jcp.53.10.756.
    1. Laine L., Lewin D. N., Naritoku W., et al. Prospective comparison of H&E, Giemsa, and Genta stains for the diagnosis of Helicobacter pylori. Gastrointestinal Endoscopy. 1997;45(6):463–467. doi: 10.1016/s0016-5107(97)70174-3.
    1. Khadangi F., Yassi M., Kerachian M. A. Review: diagnostic accuracy of PCR-based detection tests for Helicobacter Pylori in stool samples. Helicobacter. 2017;22(6) doi: 10.1111/hel.12444.
    1. Zhen-Hua Z., De-Qiang H., Yong X., et al. Characterization of 23S rRNA gene mutation in primary and secondary clarithromycin-resistant Helicobacter pylori strains from East China. Turkish Journal of Gastroenterology. 2013;24(1):5–9. doi: 10.4318/tjg.2013.0525.
    1. Archampong T. N., Asmah R. H., Aidoo E. K., et al. Helicobacter pylori cagA and vacA genes in dyspeptic Ghanaian patients. BMC Research Notes. 2017;10(1):p. 231. doi: 10.1186/s13104-017-2542-8.
    1. Ruparelia J. R., Sodagar N. R., Patel J. S., et al. Comparison of conventional diagnostic modalities with PCR for detection of Helicobcter pylori infection in symptomatic patients. Asian Journal of Medical and Pharmaceutical Researches. 2013;3:105–110.
    1. Sugimoto M., Wu J. Y., Abudayyeh S., et al. Unreliability of results of PCR detection of Helicobacter pylori in clinical or environmental samples. Journal of Clinical Microbiology. 2009;47(3):738–742. doi: 10.1128/jcm.01563-08.
    1. Chisholm S. A., Owen R. J., Teare E. L., et al. PCR-based diagnosis of Helicobacter pylori infection and real-time determination of clarithromycin resistance directly from human gastric biopsy samples. Journal of Clinical Microbiology. 2001;39(4):1217–1220. doi: 10.1128/jcm.39.4.1217-1220.2001.
    1. Patel S. K., Pratap C. B., Jain A. K., et al. Diagnosis of Helicobacter pylori: what should be the gold standard? World Journal of Gastroenterology. 2014;20(36):12847–12859. doi: 10.3748/wjg.v20.i36.12847.
    1. Shetty V., Ballal M., Balaraju G., et al. Helicobacter pylori in dyspepsia: phenotypic and genotypic methods of diagnosis. Journal of Global Infectious Diseases. 2017;9(4):131–134. doi: 10.4103/jgid.jgid_52_17.
    1. Lim C. Y., Lee K. H., Cho M. J., et al. Detection of Helicobacter pylori in gastric mucosa of patients with gastroduodenal diseases by PCR-restriction analysis using the RNA polymerase gene (rpoB) Journal of Clinical Microbiology. 2003;41(7):3387–3391. doi: 10.1128/jcm.41.7.3387-3391.2003.
    1. Lorusso F., Caleca M. P., Bellavia C., et al. The EBV-DNA can be used as a diagnostic and follow-up parameter of the Rhinopharyngeal tumors in the non-endemic population of western Sicily. Indian Journal of Otolaryngology and Head & Neck Surgery. 2018 doi: 10.1007/s12070-018-1427-z. In press.

Source: PubMed

3
Tilaa