Molecular diagnosis of bacterial and viral diarrhoea using multiplex-PCR assays: An observational prospective study among paediatric patients from India.

PURPOSE: In developing countries, the aetiology of diarrhoea goes undiagnosed as only microscopy, stool culture or enzyme immunoassay are done to find the causative agent. The present study aims to detect common paediatric viral and bacterial diarrhoea pathogens by microscopy, stool culture for bacteria, and multiplex polymerase chain reaction (mPCR) for bacteria and virus detections.

MATERIALS AND

METHODS

Diarrheal stool samples (n ​= ​109) received at the laboratory from paediatric patients aged one month to 18 years were included in the study. They were cultured for common bacterial pathogens and simultaneously subjected to two multiplex PCRs one for the detection of Salmonella spp., Shigella spp., Enteroinvasive E.coli and Enteropathogenic E.coli, another for the detection of adenovirus, astrovirus, rotavirus and norovirus.

RESULTS

Of the 109 samples cultured for bacterial aetiology, 0.9% (1/109) grew Salmonella enterica ser.Typhi and 2% (2/109) Shigella flexneri. By mPCR, 16% of samples (17/109) were positive for Shigella spp., 0.9% (1/109) for Salmonella spp., and 21% (23/109) for rotavirus. One sample (0.9%) had rotavirus and Shigella spp., which indicates mixed aetiology.

CONCLUSIONS

Shigella spp. and rotavirus are the prime causative agents of childhood diarrhoea in our region. The rate of detection of bacterial aetiology by culture was poor. Isolation of pathogens by conventional culture helps to know the species, serotypes and antibiotic susceptibility of the pathogens. Virus isolation is cumbersome, time-consuming, and not available for routine diagnostic use. Therefore, real-time mPCR would be a better choice for early detection of pathogens, thereby ensuring timely diagnosis, treatment, and a reduction in mortality.