Assessing the utility of using a next generation sequencing platform to develop a Shigella spp threshold and emerging pathogens to profile faecal samples from individuals with suspected diarrhoea

BackgroundGlobally, it is estimated that there are 2 billion cases of diarrhoeal disease each year, with 1.9 million children under the age of 5 years, dying from diarrhoea. This also affects 1 in 5 people in the UK each year. Rapid diagnosis, appropriate treatment and infection control measures are, therefore, particularly important. Currently, Public Health Wales and England Microbiology Division test for five key bacterial GI pathogens Escherichia coli O157 (VTEC), Shigella dysenteriae, Salmonella spp., Campylobacter spp. and Clostridioides difficile. There is, however, poor success rate for identification of these pathogens, leaving the patient at risk from untreated infections. This study has developed tools that are effective and reliable, with a high positive outcome for patient diagnosis of diarrhoeal infection. ResultsThe study blindly analysed 592 samples, with the most abundant species being Shigella sonnei at 15%, and the top genera Bacteroides at 26% and Shigella 15%. Campylobacter spp. had an abundance of 4% and Clostridium difficile 3%, and Salmonella spp. 0.2%. There were also significant differences in abundance at genera level, between the Flemish Gut project and diarrhoea samples, with respect to Shigella (0.2%) and Campylobacter (0.1%). The project introduced a novel threshold of 5.32% to determine if the Shigella spp. (Escherichia) were from a healthy or unhealthy community. A DMBiome model was developed to integrate the 5.32% threshold of Shigella spp., the Public Health laboratory tested pathogens, and two emerging enteropathogens. The overall positive outcome was that 89% of all samples were diagnosed with diarrhoea infections, leaving 11% unknown.ConclusionAdvances in illumina Miseq 16S rRNA gene amplicon sequencing has the potential to improve the detection of pathogens associated with diarrhoea. One of the advantages is the ability to detect microorganisms directly from a sample without prior knowledge. Comparison of the healthy and unhealthy communities in this study, resulted in the discovery of a biomarker to detect which community the Shigella spp. came from. The 5.32% threshold, and introducing new emerging pathogens into the DMBiome model, provides an enormous benefit in improving the detection rate of diarrhoeal infections, which is currently low at approx. 10%.