Microbiology - Journal Articles
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Item Isolation and characterization of a novel lytic Parabacteroides distasonis bacteriophage φPDS1 from the human gut(Taylor & Francis, 2024) Cortés-Martín, Adrián; Denise, Rémi; Guerin, Emma; Stockdale, Stephen R.; Draper, Lorraine A.; Ross, R. Paul; Shkoporov, Andrey N.; Hill, Colin; Science Foundation IrelandThe human gut microbiome plays a significant role in health and disease. The viral component (virome) is predominantly composed of bacteriophages (phages) and has received significantly less attention in comparison to the bacteriome. This knowledge gap is largely due to challenges associated with the isolation and characterization of novel gut phages, and bioinformatic hurdles such as the lack of a universal phage marker gene and the absence of sufficient numbers of homologs in viral databases. Here, we describe the isolation from human feces of a novel lytic phage with siphovirus morphology, φPDS1, infecting Parabacteroides distasonis APCS2/PD, and classified within a newly proposed Sagittacolavirus genus. In silico and biological characterization of this phage is presented in this study. Key to the isolation of φPDS1 was the antibiotic-driven selective enrichment of the bacterial host in a fecal fermenter. Despite producing plaques and lacking genes associated with lysogeny, φPDS1 demonstrates the ability to coexist in liquid culture for multiple days without affecting the abundance of its host. Multiple studies have shown that changes in Parabacteroides distasonis abundance can be linked to various disease states, rendering this novel phage-host pair and their interactions of particular interest.Item Harnessing the endogenous Type I-C CRISPR-Cas system for genome editing in Bifidobacterium breve(American Society for Microbiology, 2024-02-06) Han, Xiao; Chang, Lulu; Chen, Haiqin; Zhao, Jianxin; Tian, Fengwei; Ross, R. Paul; Stanton, Catherine; van Sinderen, Douwe; Chen, Wei; Yang, Bo; Dudley, Edward G.; National Key Research and Development Program of China; National Natural Science Foundation of China; Project 211; Collaborative Innovationcenter of Food Safety and Quality Control in Jiangsu ProvinceBifidobacterium breve, one of the main bifidobacterial species colonizing the human gastrointestinal tract in early life, has received extensive attention for its purported beneficial effects on human health. However, exploration of the mode of action of such beneficial effects exerted by B. breve is cumbersome due to the lack of effective genetic tools, which limits its synthetic biology application. The widespread presence of CRISPR-Cas systems in the B. breve genome makes endogenous CRISPR-based gene editing toolkits a promising tool. This study revealed that Type I-C CRISPR-Cas systems in B. breve can be divided into two groups based on the amino acid sequences encoded by cas gene clusters. Deletion of the gene coding uracil phosphoribosyl-transferase (upp) was achieved in five B. breve strains from both groups using this system. In addition, translational termination of uracil phosphoribosyl-transferase was successfully achieved in B. breve FJSWX38M7 by single-base substitution of the upp gene and insertion of three stop codons. The gene encoding linoleic acid isomerase (bbi) in B. breve, being a characteristic trait, was deleted after plasmid curing, which rendered it unable to convert linoleic acid into conjugated linoleic acid, demonstrating the feasibility of successive editing. This study expands the toolkit for gene manipulation in B. breve and provides a new approach toward functional genome editing and analysis of B. breve strains.Item Linking human milk oligosaccharide metabolism and early life gut microbiota: bifidobacteria and beyond(American Society for Microbiology, 2024-01-11) Lordan, Cathy; Roche, Aoife K.; Delsing, Dianne; Nauta, Arjen; Groeneveld, Andre; MacSharry, John; Cotter, Paul D.; van Sinderen, Douwe; Rey, Federico; FrieslandCampinaHuman milk oligosaccharides (HMOs) are complex, multi-functional glycans present in human breast milk. They represent an intricate mix of heterogeneous structures which reach the infant intestine in an intact form as they resist gastrointestinal digestion. Therefore, they confer a multitude of benefits, directly and/or indirectly, to the developing neonate. Certain bifidobacterial species, being among the earliest gut colonizers of breast-fed infants, have an adapted functional capacity to metabolize various HMO structures. This ability is typically observed in infant-associated bifidobacteria, as opposed to bifidobacteria associated with a mature microbiota. In recent years, information has been gleaned regarding how these infant-associated bifidobacteria as well as certain other taxa are able to assimilate HMOs, including the mechanistic strategies enabling their acquisition and consumption. Additionally, complex metabolic interactions occur between microbes facilitated by HMOs, including the utilization of breakdown products released from HMO degradation. Interest in HMO-mediated changes in microbial composition and function has been the focal point of numerous studies, in recent times fueled by the availability of individual biosynthetic HMOs, some of which are now commonly included in infant formula. In this review, we outline the main HMO assimilatory and catabolic strategies employed by infant-associated bifidobacteria, discuss other taxa that exhibit breast milk glycan degradation capacity, and cover HMO-supported cross-feeding interactions and related metabolites that have been described thus far.Item Impact of a phage cocktail targeting Escherichia coli and Enterococcus faecalis as members of a gut bacterial consortium in vitro and in vivo(Frontiers Media S.A., 2022) Buttimer, Colin; Sutton, Thomas D. S.; Colom, Joan; Murray, Ellen; Bettio, Pedro H.; Smith, Linda; Bolocan, Andrei Sorin; Shkoporov, Andrey N.; Oka, Akihiko; Liu, Bo; Herzog, Jeremy W.; Sartor, R. Balfour; Draper, Lorraine A.; Ross, R. Paul; Hill, Colin; Science Foundation Ireland; Janssen BiotechEscherichia coli and Enterococcus faecalis have been implicated as important players in human gut health that have been associated with the onset of inflammatory bowel disease (IBD). Bacteriophage (phage) therapy has been used for decades to target pathogens as an alternative to antibiotics, but the ability of phage to shape complex bacterial consortia in the lower gastrointestinal tract is not clearly understood. We administered a cocktail of six phages (either viable or heat-inactivated) targeting pro-inflammatory Escherichia coli LF82 and Enterococcus faecalis OG1RF as members of a defined community in both a continuous fermenter and a murine colitis model. The two target strains were members of a six species simplified human microbiome consortium (SIHUMI-6). In a 72-h continuous fermentation, the phage cocktail caused a 1.1 and 1.5 log (log10 genome copies/mL) reduction in E. faecalis and E. coli numbers, respectively. This interaction was accompanied by changes in the numbers of other SIHUMI-6 members, with an increase of Lactiplantibacillus plantarum (1.7 log) and Faecalibacterium prausnitzii (1.8 log). However, in germ-free mice colonized by the same bacterial consortium, the same phage cocktail administered twice a week over nine weeks did not cause a significant reduction of the target strains. Mice treated with active or inactive phage had similar levels of pro-inflammatory cytokines (IFN-y/IL12p40) in unstimulated colorectal colonic strip cultures. However, histology scores of the murine lower GIT (cecum and distal colon) were lower in the viable phage-treated mice, suggesting that the phage cocktail did influence the functionality of the SIHUMI-6 consortium. For this study, we conclude that the observed potential of phages to reduce host populations in in vitro models did not translate to a similar outcome in an in vivo setting, with this effect likely brought about by the reduction of phage numbers during transit of the mouse GIT.Item Impact of bacteriocin-producing strains on bacterial community composition in a simplified human intestinal microbiota(Frontiers Media S.A., 2023) Ríos Colombo, Natalia S.; Pérez-Ibarreche, Mariana; Draper, Lorraine A.; O’Connor, Paula M.; Field, Des; Ross, R. Paul; Hill, Colin; Horizon 2020; Science Foundation Ireland; European Research CouncilBacteriocins are antimicrobial peptides that have been studied for decades as food bio-preservatives or as alternatives to antibiotics. They also have potential as modulators of the gut microbiome, which has been linked to human health. However, it is difficult to predict a priori how bacteriocins will impact complex microbial communities through direct and indirect effects. Here we assess the effect of different bacteriocin-producing strains on a Simplified Human Intestinal Microbiota (SIHUMI) model, using a set of bacteriocin-producing strains (Bac+) and otherwise isogenic non-producers (Bac−). Bacteriocins from different classes and with different activity spectra were selected, including lantibiotics such as lacticin 3147 and nisin A, and pediocin-like bacteriocins such as pediocin PA-1 among other peptides. SIHUMI is a bacterial consortium of seven diverse human gut species that assembles to a predictable final composition in a particular growth medium. Each member can be individually tracked by qPCR. Bac+ and Bac− strains were superimposed on the SIHUMI system, and samples were taken at intervals up to 48 h. The genome copy number of each SIHUMI member was evaluated using specific primers. We establish that the composition of the community changes in response to the presence of either broad- or narrow-spectrum bacteriocin producers and confirm that there are significant off-target effects. These effects were analyzed considering antagonistic inter-species interactions within the SIHUMI community, providing a comprehensive insight into the possible mechanisms by which complex communities can be shaped by bacteriocins.