pr-2017-00461v_0001
Publication
Evaluating in Vitro Culture Medium of Gut Microbiome with Orthogonal Experimental Design and a Metaproteomics Approach

Leyuan Li, Xu Zhang, Zhibin Ning, Janice Mayne, Jasmine I. Moore, James Butcher, Cheng-Kang Chiang, David Mack, Alain Stintzi, Daniel Figeys

In vitro culture based approaches are time- and cost-effective solutions for rapidly evaluating the effects of drugs or natural compounds against microbiomes. The nutritional composition of the culture medium is an important determinant for effectively maintaining the gut microbiome in vitro. This study combines orthogonal experimental design and a metaproteomics approach to obtaining functional insights into the effects of different medium components on the microbiome. Our results show that the metaproteomic profile respond differently to medium components, including inorganic salts, bile salts, mucin, and short-chain fatty acids. Multifactor analysis of variance further revealed significant main and interaction effects of inorganic salts, bile salts, and mucin on the different functional groups of gut microbial proteins. While a broad regulating effect was observed on basic metabolic pathways, different medium components also showed significant modulations on cell wall, membrane, and envelope biogenesis and cell motility related functions. In particular, flagellar assembly related proteins were significantly responsive to the presence of mucin. This study provides information on the functional influences of medium components on the in vitro growth of microbiome communities and gives insight on the key components that must be considered when selecting and optimizing media for culturing ex vivo microbiotas.
1-s2.0-S1046202318303360-gr4
Publication
RNA-Seq analysis of the T3SA regulon in Shigella flexneri reveals two new chromosomal genes upregulated in the on-state.

Navoun Silué, Endrei Marcantonio, Francois-Xavier Campbell-Valois 

Shigella spp. are enterobacteria that invade human colonic mucosal cells using their Type Three Secretion Apparatus (T3SA). Shigella spp. possess a large plasmid that encodes most of its virulence factors and has been the focus of seminal work that defined the T3SA regulon. Thus, a global assessment of the transcriptional response regulated by the T3SA has been lacking. Herein we used RNA-Seq to identify genes that are differentially expressed when the T3SA is active (on-state) versus inactive (off-state). The quality of the RNA-Seq dataset was validated by its correlation with a prior microarray study. Using novel insights about the expression of non-coding regions, bioinformatic tools and experimentations, we demonstrated the existence of six operons and evidence that ipaH2.5 is a pseudogene. In addition, 86 chromosomal genes were downregulated in the on-state including several non-coding transcripts corresponding to short antisense RNA embedded in the 16S and 23S RNA genes, and 40 coding transcripts, whose cognate proteins were highly connected at the genetic and biochemical levels. Finally, we identified two novel chromosomal genes dubbed gem1 and gem3, which were upregulated in the on-state similarly to genes belonging to the T3SA regulon. The latter findings were validated on biological triplicates by droplet digital PCR. To our knowledge gem1 and gem3 are the first chromosomal members of the T3SA regulon that have no homologs on the plasmid. Our approach provides a path to optimizing RNA-Seq studies in case of bacterial models that had previously been the subject of medium to large scale studies.
Keystone2019_poster_Leyuan_Li_20190228

Conference poster presentation

Functional Screening Reveals Individual Gut Microbiome Responses to Xenobiotics

Keystone Symposia: Microbiome: Chemical Mechanisms and Biological Consequences (C3), Montreal, QC, Canada, March 10 to March 14, 2019

Leyuan Li  

Xenobiotics can affect the microbiome, we developed an approach, Rapid Assay of Individual Microbiome (RapidAIM), to screen xenobiotics against individual microbiomes. Using a metaproteomic approach, we showed that tested compounds significantly affected overall microbiome abundance, microbiome composition and functional pathways at multiple taxonomic levels.  Compounds such as berberine, metformin, diclofenac, FOS and most antibiotics led to consistent functional responses across microbiomes. In contrast, some compounds induced individually distinct functional responses. Other drugs had little effects on the microbiomes. Our workflow offers a solution to systematically study the functional effects of compounds on individual microbiomes.

[Mercurio]OISB_poster

Conference poster presentation

Identifying genes required for Saccharomyces cerevisiae growth in mucin

Ottawa Institute of Systems Biology Conference, Cornwall, Canada, May 6, 2019

Kevin Mercurio, Dylan Singh, Kristin Baetz