Abstract B14: Flow cytometry for targeted culturomics of gut commensal species and rapid overview of microbiota composition

Abstract

Abstract An ever-increasing number of studies report the importance of the composition of the intestinal microbiota in the treatment of cancers. The mechanisms involved are varied. The reciprocal interaction of drugs with bacteria can have an effect on the efficacy as well as the toxicity of treatments. In addition, some species or strains are able to stimulate CD8+ T cell production and the effectiveness of immune checkpoint blockers (ICBs). As part of these treatments, there is therefore a potential interest in being able to isolate and cultivate bacterial species associated with the response to ICBs in different cancers. The objective is to better understand the mechanisms involved and possibly develop complementary therapeutic and/or diagnostic solutions. However, this approach is still limited by the fact that a large number of commensal bacterial species in the intestine are difficult to cultivate, due to specific nutritive requirements, extreme oxygen sensitivity (EOS), or under-representation in the whole bacterial population. In an attempt to circumvent these limitations, we developed flow cytometry (FCM) and cell sorting in anaerobic conditions. A series of polyclonal antibodies specific to commensal species known to be beneficial to the host such as Faecalibacterium prausnitzii, Akkermansia muciniphila, Christensenella minuta, and Eubacterium hallii have been produced and used in combination with viability and Gram staining to rapidly detect, sort, and culture species of interest. This strategy has enabled a rapid collection of strains to be built up from the stools of healthy volunteers, demonstrating the usefulness of the approach. Developments are under way to apply reverse genomics strategies to identify sequences encoding immunogenic proteins in order to produce antibodies even in the absence of already cultivated strains, which is still the case for approximately 70% of the human gut microbiota species. In conclusion, our work confirms that FCM is well adapted for complex bacterial microbiota studies. When used in conjunction with appropriate staining, it gives a general overview of microbiota composition and variations in longitudinal studies, including bacterial load, which is an important piece of information. In addition to sorting and cultivating target species of interest, it can be used for a quick and easy assessment of microbiota composition, either before starting a drug treatment or for monitoring the effects of the treatment on commensal microbiota. Citation Format: Samuel Bellais, Mélanie Nehlich, Aurore Duquenoy, Maryne Ania, Jan Baijer, Ilia Belotserkovsky, Vincent Thomas. Flow cytometry for targeted culturomics of gut commensal species and rapid overview of microbiota composition [abstract]. In: Proceedings of the AACR Special Conference on the Microbiome, Viruses, and Cancer; 2020 Feb 21-24; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2020;80(8 Suppl):Abstract nr B14.