Background A low-cost targeted solution to profiling gut microbial diversity is sequencing of the 16S rRNA gene; however, it is often insufficient to gain species level resolution due to high homology across different bacteria. Therefore, we developed a first-of-its-kind targeted sequencing solution that supplements 16S gene targets, with highly species-specific primers for a cohort of 73 bacteria associated with research in diabetes, cancer and its response to immunotherapy, gastrointestinal and auto-immune disorders. This assay performs at 100% sensitivity and specificity for the species-level detection (Ion AmpliSeq Microbiome Health Research Kit: www.thermofisher.com/ngsmicrobiome) of these bacteria and is hence better suited for gut microbiome profiling in the context of the above phenotypes, as compared to other existing solutions.
Methods To assess the utility of the panel in cancer immunotherapy research, we sequenced DNA from 15 stool samples from subjects with Non-Small Cell Lung Carcinoma (NSCLC) undergoing immunotherapy, and compared their microbiome profiles to 26 healthy stool samples collected internally. With our post-sequencing workflow on Ion Reporter™, we automatically generate a report with taxonomic classifications, sample diversity metrics through QIIME2 integration, and relative abundance visualizations for bacteria across multiple samples.
Results We identified significant microbiome composition differences between the healthy samples and cancer/treated samples, as evidenced by (i) a clear separation between the two cohorts based on a beta diversity principal coordinate analysis (PCoA) plot, driven by large abundance changes in Clostridium, Lachnoclostridium, Subdolinigranulum and Oscillibacter (P < 0.05), (ii) grouping into distinct classes based on overall microbiome profiles (Analysis-of-Similarities ANOSIM P = 0.003), and (iii) differences in abundances of specific bacteria with anti-tumor effects1 such as F. prausnitzii (P = 0.02).
Conclusions We have created a highly multiplexed, sensitive and specific assay for robust characterization of gut microbiota, with compatibility on both (i) the Ion GeneStudio S5™ with a 48-hr sample-to-result turnaround, and (ii) the new Ion Genexus™ System, a fully integrated platform featuring a hands-off, automated sample-to-report workflow that delivers results in a single day. It enables an efficient and affordable means for conducting extensive analyses of the human microbiome having applications in the study of phenotypic variability, and the potential relationship to disease.For research use only. Not for use in diagnostic procedures.
Ma J, Sun L, Liu Y. et al. Alter between gut bacteria and blood metabolites and the anti-tumor effects of Faecalibacterium prausnitzii in breast cancer. BMC Microbiol 2020; 20:1–19.
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