Differentiating Entamoeba hystolytica, E. disparandE. moshkovskii in stool sample from rural community of Nepal using nested-PCR

Abstract

Background: Salmonella enteric serovar Typhi is a gram-negative, rod-shaped facultative anaerobe that only infects humans which is major causative agent for typhoid fever. Enteric fever is a systemic infection of reticuloendothelial system caused by the human adapted pathogens Salmonella enterica serotype Typhi and Salmonella enterica serotype paratyphi A, B. However, the prevalence of Salmonella enterica B and C infection has not been reported in Nepal so for. Methods and materials: A total of 270 faecal samples were collected from south eastern terai region of Nepal after the informed consent form. The samples were processed for microscopic examination of Entamoeba species along with other intestinal parasites. Additionally, Nested- PCR was carried out for characterisation of microscopically positive fecal samples containing the E complex. Results: Of the 270 stool sample from diarrheal patients, 67.3% were male whereas only 32.7% were female. Microscopy-positive fecal samples containing the E complex (n = 56) were subject to PCR for confirmation of the diagnosis of E complex whereas PCR products were detected in 46 samples. Surprisingly, out of 46 PCR-positive samples, only 3 samples were reported to contain E. histolytica. Likewise, among total sample, viral diarrheal was most significant form of diarrhoea found in 76.67% of patients. Among different organisms, As. Lumbricoids and E. histolytica, G. lambia and H. nana were identified in most of the patients accounting for 11.11%, 8.52%, 2.59% and 1.11% respectively. However, Lumbricoids, G. lambia, Teniasoliumand E. histolytica were present in an individual patient while two patient was found with both As. Lumbricoids and G. lambia.Among several symptoms, diarrhoea seems to be the common symptoms infecting all of the patients which is followed by fever and vomiting which accounts for 55.1% and 46.2% correspondingly. Whereas, nausea appears to be the least common symptoms infecting only 14.4% of patients. Conclusion: Molecular techniques are indeed promising tools for epidemiological studies, particularly in discriminating the pathogenic from the non-pathogenic species, which is beneficial for proper diagnosis, immunological assay and drug testing. This is the first study in Nepal using molecular techniques to determine the true incidence of E complex and highlights the molecular methods to differentiate among the species.