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

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.

Genetic diversity of Leishmania donovani that causes cutaneous leishmaniasis in Sri Lanka: a cross sectional study with regional comparisons.

BACKGROUND: Leishmania donovani is the etiological agent of visceral leishmaniasis (VL) in the Indian subcontinent. However, it is also known to cause cutaneous leishmaniasis (CL) in Sri Lanka. Sri Lankan L. donovani differs from other L. donovani strains, both at the molecular and biochemical level. To investigate the different species or strain-specific differences of L. donovani in Sri Lanka we evaluated sequence variation of the kinetoplastid DNA (kDNA). METHODS: Parasites isolated from skin lesions of 34 CL patients and bone marrow aspirates from 4 VL patients were genotyped using the kDNA minicircle PCR analysis. A total of 301 minicircle sequences that included sequences from Sri Lanka, India, Nepal and six reference species of Leishmania were analyzed. RESULTS: Haplotype diversity of Sri Lankan isolates were high (H d = 0.757) with strong inter-geographical genetic differentiation (F ST > 0.25). In this study, L. donovani isolates clustered according to their geographic origin, while Sri Lankan isolates formed a separate cluster and were clearly distinct from other Leishmania species. Within the Sri Lankan group, there were three distinct sub-clusters formed, from CL patients who responded to standard antimony therapy, CL patients who responded poorly to antimony therapy and from VL patients. There was no specific clustering of sequences based on geographical origin within Sri Lanka. CONCLUSION: This study reveals high levels of haplotype diversity of L. donovani in Sri Lanka with a distinct genetic association with clinically relevant phenotypic characteristics. The use of genetic tools to identify clinically relevant features of Leishmania parasites has important therapeutic implications for leishmaniasis.

PCR and direct agglutination as Leishmania infection markers among healthy Nepalese subjects living in areas endemic for Kala-Azar.

OBJECTIVE: To compare a PCR assay and direct agglutination test (DAT) for the detection of potential markers of Leishmania infection in 231 healthy subjects living in a kala-azar endemic focus of Nepal. METHODS: The sample was composed of 184 (80%) persons without any known history of KA and not living in the same house as known kala-azar cases (HNK), 24 (10%) Healthy Household Contacts (HHC) and 23 (10%) past kala-azar cases which had been successfully treated (HPK). RESULTS: PCR and DAT positivity scores were, respectively: HNK, 17.6% and 5.6%; HHC, 12.5% and 20.8%; HPK, 26.1% and 95.7%. The ratio PCR-positives/DAT-positives was significantly higher in HNK (ratio = 3.1) than in HHC (ratio = 0.6, P = 0.036) and in HPK (ratio = 0.2, P = 0.012). The ratio PCR-positives/DAT-positives did not significantly differ between HHC (ratio = 0.6) and HPK (ratio = 0.2, P = 0.473). The positive agreement index between PCR and DAT in HNK was 5%; in HHC, 0%; in HPK, 43%. CONCLUSIONS: Our study highlights the specific character of PCR and DAT for the exploration of Leishmania asymptomatic infections. PCR is probably more informative for very recent infections among HNK, while DAT provides more information among HHC and HPK, a feature likely related to the power of serology to track less recent infections.