Based on these observations, the concept of Next Generation Probiotics (NGP) gained interest. A range of disorders was also shown to be associated with altered gut microbiota composition ( Lin and Zhang, 2017). Diverse and specific to each individual, gut microbiota can be influenced by different factors such as age, sex, diseases, drug treatments, environment or nutrition ( Lozupone et al., 2012). The analysis of gut microbiota led to the concept of enterotypes that divide human microbiota based on the enrichment of specific taxa ( Arumugam et al., 2011 Wu et al., 2011). Bacteria are the most studied microorganisms with around 10 11 bacteria per gram of stool and a predominance of Firmicutes and Bacteroidetes ( Qin et al., 2010). Human gut microbiota are composed of a wide variety of microorganisms including bacteria, archaea, fungi, viruses and yeasts ( Lozupone et al., 2012 Dieterich et al., 2018). It is assumed to be applicable as an efficient and fast approach to improve the control of processes linked to a wide range of ecosystems or known communities of bacterial species in both research and industrial contexts. The flow cytometry staining method that was developed has the potential to facilitate the analysis of complex ecosystems by highlighting changes in bacterial communities’ dynamics. This combination of staining methods makes it possible to follow-up evolutions of complex microbial communities, supporting its future use as a rapid analysis tool in various applications. After validation on both aerobic/anaerobic facultative and strictly anaerobic bacteria, the staining methods were applied on complex ecosystems, revealing differences between culture conditions and demonstrating that minor pH variations have strong impacts on microbial community structure, which was confirmed by 16S rRNA gene sequencing. In addition, we tested the use of a combination of labeled vancomycin and Wheat Germ Agglutinin (WGA) lectin to discriminate Gram-positive from Gram-negative bacteria in complex ecosystems. This sorting of “viable” fraction demonstrated that 10–80% of identified “viable” cells of pure cultures of strictly anaerobic bacteria were culturable. In this study, we evaluated the culturability of strictly anaerobic bacteria that were stained with a classical Live/Dead staining, and then sorted using flow cytometry under anaerobic conditions. In addition, it can potentially be used to select, isolate and cultivate specific bacteria of interest. Flow cytometry overcomes these disadvantages, providing key information on communities within hours. Whereas sequence-based metagenomics profiling is widely used for microbial ecosystems characterization, it still requires time and specific expertise for analysis. In this context, there is a major interest to develop analysis tools allowing simple and cost-effective population pattern analysis of these complex ecosystems to follow changes over time. Over the past years, gut microbiota became a major field of interest with increasing reports suggesting its association with a large number of human diseases.
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