Use of Deep-Amplicon Sequencing (DAS), Real-Time PCR and In Situ Hybridization to Detect and Other Pathogenic Helicobacter Species in Feces from Children.

BACKGROUND

Detecting in fecal samples is easier and more comfortable than invasive techniques, especially in children. Thus, the objective of the present work was to detect in feces from children by molecular methods as an alternative for diagnostic and epidemiological studies.

METHODS

Forty-five fecal samples were taken from pediatric patients who presented symptoms compatible with infection. HpSA test, culture, real-time quantitative PCR (qPCR), fluorescence in situ hybridization (FISH), direct viable count associated with FISH (DVC-FISH), and Illumina-based deep-amplicon sequencing (DAS) were applied.

RESULTS

No colonies were isolated from the samples. qPCR analysis detected in the feces of 24.4% of the patients. In comparison, DVC-FISH analysis showed the presence of viable cells in 53.3% of the samples, 37% of which carried 23S rRNA mutations that confer resistance to clarithromycin. After DAS, -specific 16S rDNA sequences were detected in 26 samples. In addition, DNA from was identified in 10 samples, and DNA was detected in one sample.

CONCLUSION

The results of this study show the presence of , , and in children's stools, demonstrating the coexistence of more than one species in the same patient. The DVC-FISH method showed the presence of viable, potentially infective cells in a high percentage of the children's stools. These results support the idea that fecal-oral transmission is probably a common route for and suggest possible fecal-oral transmission of other pathogenic species.