A major function of the gut microbiota is to provide colonization resistance, wherein pathogens are inhibited or suppressed below infectious levels. However, the fraction of gut microbiota required for colonization resistance remains unclear. We used culturomics to isolate a gut microbiota culture collection comprising 1,590 isolates belonging to 102 species. This culture collection represents 34.57% of the taxonomic diversity and 70% functional capacity, as estimated by metagenomic sequencing of the fecal samples used for culture. Using whole-genome sequencing, we characterized species representatives from this collection and predicted their phenotypic traits, further characterizing isolates by defining nutrient utilization profiles and short-chain fatty acid production. When screened with a coculture assay, 66 species in our culture collection inhibited Clostridioides difficile. Several phenotypes, particularly, growth rate, production of SCFAs, and the utilization of mannitol, sorbitol, or succinate, correlated with C. difficile inhibition. We used a combinatorial community assembly approach to formulate defined bacterial mixes inhibitory to C. difficile. We tested 256 combinations and found that both species composition and blend size were important in inhibition. Our results show that the interaction of bacteria with one another in a mix and with other members of gut commensals must be investigated to design defined bacterial mixes for inhibiting C. difficile in vivo. IMPORTANCE Antibiotic treatment causes instability of gut microbiota and the loss of colonization resistance, thus allowing pathogens such as Clostridioides difficile to colonize and causing recurrent infection and mortality. Although fecal microbiome transplantation has been shown to be an effective treatment for C. difficile infection (CDI), a more desirable approach would be the use of a defined mix of inhibitory gut bacteria. The C. difficile-inhibiting species and bacterial combinations identified herein improve the understanding of the ecological interactions controlling colonization resistance against C. difficile and could aid in the design of defined bacteriotherapy as a nonantibiotic alternative against CDI.
Waterborne Escherichia coli are a major reservoir of antimicrobial resistance (AMR), including but not limited to extended-spectrum beta-lactamase (ESBL) and Klebsiella pneumoniae carbapenemase (KPC) mechanisms. This study quantified and described ESBL- and KPC-producing E. coli in Northern Colorado from sewer water, surface water, and influent and effluent wastewater treatment sources. Total detected bacteria and E. coli abundances, and the percentages that contain ESBL and/or KPC, were compared between water sources. Seventy E. coli isolates from the various waters had drug resistance validated with a panel of 17 antibiotics using a broth microdilution assay. The diverse drug resistance observed across E. coli isolates was further documented by polymerase chain reaction of common ESBL genes and functional relatedness by PhenePlate assay-generated dendrograms (n=70). The total E. coli abundance decreased through the water treatment process as expected, yet the percentages of E. coli harboring ESBL resistance were increased (1.70%) in surface water. Whole-genome sequencing analysis was completed for 185 AMR genes in wastewater E. coli isolates and confirmed the presence of diverse AMR gene classes (e.g., beta-lactams and efflux pumps) in isolate genomes. This study completed surveillance of AMR patterns in E. coli that reside in environmental water systems and suggests a role for integrating both phenotypic and genotypic profiling beyond ESBL and KPC mechanisms. AMR screening via multiple approaches may assist in the prevention of drug-resistant E. coli spread from waters to animals and humans.
The aim of this study was to generate a reference set of Salmonella enterica genomes isolated from wildlife from the United States and to determine the antimicrobial resistance and virulence gene profile of the isolates from the genome sequence data. We sequenced the whole genomes of 103 Salmonella isolates sampled between 1988 and 2003 from wildlife and exotic pet cases that were submitted to the Oklahoma Animal Disease Diagnostic Laboratory, Stillwater, Oklahoma. Among 103 isolates, 50.48% were from wild birds, 0.9% was from fish, 24.27% each were from reptiles and mammals. 50.48% isolates showed resistance to at least one antibiotic. Resistance against the aminoglycoside streptomycin was most common while 9 isolates were found to be multi-drug resistant having resistance against more than three antibiotics. Determination of virulence gene profile revealed that the genes belonging to csg operons, the fim genes that encode for type 1 fimbriae and the genes belonging to type III secretion system were predominant among the isolates. The universal presence of fimbrial genes and the genes encoded by pathogenicity islands 1–2 among the isolates we report here indicates that these isolates could potentially cause disease in humans. Therefore, the genomes we report here could be a valuable reference point for future traceback investigations when wildlife is considered to be the potential source of human Salmonellosis.Electronic supplementary materialThe online version of this article (10.1186/s13099-017-0213-x) contains supplementary material, which is available to authorized users.
The gut microbiome plays an important role in the immune system development, maintenance of normal health status, and in disease progression. In this study, we comparatively examined the fecal microbiomes of Amish (rural) and non-Amish (urban) infants and investigated how they could affect the mucosal immune maturation in germ-free piglets that were inoculated with the two types of infant fecal microbiota (IFM). Differences in microbiome diversity and structure were noted between the two types of fecal microbiotas. The fecal microbiota of the non-Amish (urban) infants had a greater relative abundance of Actinobacteria and Bacteroidetes phyla, while that of the Amish (rural) counterparts was dominated by Firmicutes. Amish infants had greater species richness compared with the non-Amish infants' microbiota. The fecal microbiotas of the Amish and the non-Amish infants were successfully transplanted into germ-free piglets, and the diversity and structure of the microbiota in the transplanted piglets remained similar at phylum level but not at the genus level. Principal coordinates analysis (PCoA) based on Weighted-UniFrac distance revealed distinct microbiota structure in the intestines of the transplanted piglets. Shotgun metagenomic analysis also revealed clear differences in functional diversity of fecal microbiome between Amish and non-Amish donors as well as microbiota transplanted piglets. Specific functional features were enriched in either of the microbiota transplanted piglet groups directly corresponding to the predominance of certain bacterial populations in their gut environment. Some of the colonized bacterial genera were correlated with the frequency of important lymphoid and myeloid immune cells in the ileal submucosa and mesenteric lymph nodes (MLN), both important for mucosal immune maturation. Overall, this study demonstrated that transplantation of diverse IFM into germ-free piglets largely recapitulates the differences in gut microbiota structure between rural (Amish) and urban (non-Amish) infants. Thus, fecal microbiota transplantation to germ-free piglets could be a useful large animal model system for elucidating the impact of gut microbiota on the mucosal immune system development. Future studies can focus on determining the additional advantages of the pig model over the rodent model.
Background Antimicrobial resistance (AMR) of bacterial pathogens is an emerging public health threat. This threat extends to pets as it also compromises our ability to treat their infections. Surveillance programs in the United States have traditionally focused on collecting data from food animals, foods, and people. The Veterinary Laboratory Investigation and Response Network (Vet-LIRN), a national network of 45 veterinary diagnostic laboratories, tested the antimicrobial susceptibility of clinically relevant bacterial isolates from animals, with companion animal species represented for the first time in a monitoring program. During 2017, we systematically collected and tested 1968 isolates. To identify genetic determinants associated with AMR and the potential genetic relatedness of animal and human strains, whole genome sequencing (WGS) was performed on 192 isolates: 69 Salmonella enterica (all animal sources), 63 Escherichia coli (dogs), and 60 Staphylococcus pseudintermedius (dogs). Results We found that most Salmonella isolates (46/69, 67%) had no known resistance genes. Several isolates from both food and companion animals, however, showed genetic relatedness to isolates from humans. For pathogenic E. coli , no resistance genes were identified in 60% (38/63) of the isolates. Diverse resistance patterns were observed, and one of the isolates had predicted resistance to fluoroquinolones and cephalosporins, important antibiotics in human and veterinary medicine. For S. pseudintermedius , we observed a bimodal distribution of resistance genes, with some isolates having a diverse array of resistance mechanisms, including the mecA gene (19/60, 32%). Conclusion The findings from this study highlight the critical importance of veterinary diagnostic laboratory data as part of any national antimicrobial resistance surveillance program. The finding of some highly resistant bacteria from companion animals, and the observation of isolates related to those isolated from humans demonstrates the public health significance of incorporating companion animal data into surveillance systems. Vet-LIRN will continue to build the infrastructure to collect the data necessary to perform surveillance of resistant bacteria as part of fulfilling its mission to advance human and animal health. A One Health approach to AMR surveillance programs is crucial and must include data from humans, animals, and environmental sources to be effective. Electronic supplementary material The online version of this article (10.1186/s12917-019-1864-2) contains supplementary material, which is available to authorized users.
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