Characterization of wild and captive baboon gut microbiota and their antibiotic resistomes

Antibiotic exposure results in acute and persistent shifts in the composition and function of microbial communities associated with vertebrate hosts. However, little is known about the state of these communities in the era before the widespread introduction of antibiotics into clinical and agricultural practice. We characterized the fecal microbiota and antibiotic resistomes of wild and captive baboon populations to understand the effect of human exposure and to understand how the primate microbiota may have been altered during the antibiotic era. We used culture-independent and bioinformatics methods to identify functional resistance genes in the guts of wild and captive baboons and show that exposure to humans is associated with changes in microbiota composition and resistome expansion compared to wild baboon groups. Our results suggest that captivity and lifestyle changes associated with human contact can lead to marked changes in the ecology of primate gut communities.Environmental microbes have harbored the capacity for antibiotic production for millions of years, spanning the evolution of humans and other vertebrates. However, the industrial-scale use of antibiotics in clinical and agricultural practice over the past century has led to a substantial increase in exposure of these agents to human and environmental microbiota. This perturbation is predicted to alter the ecology of microbial communities and to promote the evolution and transfer of antibiotic resistance (AR) genes. We studied wild and captive baboon populations to understand the effects of exposure to humans and human activities (e.g., antibiotic therapy) on the composition of the primate fecal microbiota and the antibiotic-resistant genes that it collectively harbors (the “resistome”). Using a culture-independent metagenomic approach, we identified functional antibiotic resistance genes in the gut microbiota of wild and captive baboon groups and saw marked variation in microbiota architecture and resistomes across habitats and lifeways. Our results support the view that antibiotic resistance is an ancient feature of gut microbial communities and that sharing habitats with humans may have important effects on the structure and function of the primate microbiota

Destination shapes antibiotic resistance gene acquisitions, abundance increases, and diversity changes in Dutch travelers

BACKGROUND: Antimicrobial-resistant bacteria and their antimicrobial resistance (AMR) genes can spread by hitchhiking in human guts. International travel can exacerbate this public health threat when travelers acquire AMR genes endemic to their destinations and bring them back to their home countries. Prior studies have demonstrated travel-related acquisition of specific opportunistic pathogens and AMR genes, but the extent and magnitude of travel\u27s effects on the gut resistome remain largely unknown. METHODS: Using whole metagenomic shotgun sequencing, functional metagenomics, and Dirichlet multinomial mixture models, we investigated the abundance, diversity, function, resistome architecture, and context of AMR genes in the fecal microbiomes of 190 Dutch individuals, before and after travel to diverse international locations. RESULTS: Travel markedly increased the abundance and α-diversity of AMR genes in the travelers\u27 gut resistome, and we determined that 56 unique AMR genes showed significant acquisition following international travel. These acquisition events were biased towards AMR genes with efflux, inactivation, and target replacement resistance mechanisms. Travel-induced shaping of the gut resistome had distinct correlations with geographical destination, so individuals returning to The Netherlands from the same destination country were more likely to have similar resistome features. Finally, we identified and detailed specific acquisition events of high-risk, mobile genetic element-associated AMR genes including qnr fluoroquinolone resistance genes, bla CONCLUSIONS: Our results show that travel shapes the architecture of the human gut resistome and results in AMR gene acquisition against a variety of antimicrobial drug classes. These broad acquisitions highlight the putative risks that international travel poses to public health by gut resistome perturbation and the global spread of locally endemic AMR genes

Impact of international travel and diarrhea on gut microbiome and resistome dynamics

International travel contributes to the global spread of antimicrobial resistance. Travelers\u27 diarrhea exacerbates the risk of acquiring multidrug-resistant organisms and can lead to persistent gastrointestinal disturbance post-travel. However, little is known about the impact of diarrhea on travelers\u27 gut microbiomes, and the dynamics of these changes throughout travel. Here, we assembled a cohort of 159 international students visiting the Andean city of Cusco, Peru and applied next-generation sequencing techniques to 718 longitudinally-collected stool samples. We find that gut microbiome composition changed significantly throughout travel, but taxonomic diversity remained stable. However, diarrhea disrupted this stability and resulted in an increased abundance of antimicrobial resistance genes that can remain high for weeks. We also identified taxa differentially abundant between diarrheal and non-diarrheal samples, which were used to develop a classification model that distinguishes between these disease states. Additionally, we sequenced the genomes of 212 diarrheagenic Escherichia coli isolates and found those from travelers who experienced diarrhea encoded more antimicrobial resistance genes than those who did not. In this work, we find the gut microbiomes of international travelers\u27 are resilient to dysbiosis; however, they are also susceptible to colonization by multidrug-resistant bacteria, a risk that is more pronounced in travelers with diarrhea

Manure microbial communities and resistance profiles reconfigure after transition to manure pits and differ from those in fertilized field soil

In agricultural settings, microbes and antimicrobial resistance genes (ARGs) have the potential to be transferred across diverse environments and ecosystems. The consequences of these microbial transfers are unclear and understudied. On dairy farms, the storage of cow manure in manure pits and subsequent application to field soil as a fertilizer may facilitate the spread of the mammalian gut microbiome and its associated ARGs to the environment. To determine the extent of both taxonomic and resistance similarity during these transitions, we collected fresh manure, manure from pits, and field soil across 15 different dairy farms for three consecutive seasons. We used a combination of shotgun metagenomic sequencing and functional metagenomics to quantitatively interrogate taxonomic and ARG compositional variation on farms. We found that as the microbiome transitions from fresh dairy cow manure to manure pits, microbial taxonomic compositions and resistance profiles experience distinct restructuring, including decreases in alpha diversity and shifts in specific ARG abundances that potentially correspond to fresh manure going from a gut-structured community to an environment-structured community. Further, we did not find evidence of shared microbial community or a transfer of ARGs between manure and field soil microbiomes. Our results suggest that fresh manure experiences a compositional change in manure pits during storage and that the storage of manure in manure pits does not result in a depletion of ARGs. We did not find evidence of taxonomic or ARG restructuring of soil microbiota with the application of manure to field soils, as soil communities remained resilient to manure-induced perturbation

Understand and predict microbiome and resistome dynamics in response to perturbations across diverse populations and environments

Complex microbial communities are at the interface of human, animal and environment interconnected ecosystem, where they can move within and between these entities. These microbial communities are mostly beneficial, maintaining the host health and homeostatic state. However, these communities can also serve as reservoirs of antibiotic resistance (AR) genes that may disseminate to pathogen bacteria, compromising the treatment options. Like other microbial communities, human gut microbiome is highly dynamic and can get acutely perturbed with the changes in the habitat, diet, lifestyle and disease. A perturbed gut community structure has profound impact on the host health and physiology. Use of antibiotics in medical and animal sector, international travel to high infectious burden regions and occupational exposure at workplace like dairy farm or swine farm can significantly alter the microbiome structure and function. Thus, in this thesis, I aim to understand the ecological principles governing the microbiome structure in different habitats and their response to such perturbations. To accomplish this goal, I first developed and optimized the computational pipelines (PARFuMS, Resfams (v2), and resAnnotator) for high-throughput characterization of antibiotic resistance genes in diverse habitats. I then employed these methods along with other bioinformatics suites to understand the dynamics of human gut microbiota and the antibiotic resistant genes that it collectively encodes (the ‘resistome’) in response to perturbations such as international travel and swine farm exposures. Additionally, I studied wild and captive baboon populations to understand the impact of captivity and lifestyle changes associated with human contact on the changes in baboon’s microbiome and antibiotic resistome. To determine the impact of international travel and enteric infections on the gut microbial ecosystem, I have specifically focused on two international travel scenarios. In the first scenario, travelers from different countries, mainly the US and European nations, travel to one location i.e. Cusco, Peru, and in the second scenario, travelers from one country, Netherlands, travel to four different destinations viz. North Africa, East Africa, Southeastern Asia and Southern Asia. In the first scenario, I investigated the impact of travelers’ diarrhea (TD) on travelers’ gut microbiome and resistome, and the dynamics of these changes throughout travel and during specific diarrheal episodes. To this end, we assembled a cohort of 159 travelers visiting the Andean city of Cusco, Peru and applied next-generation sequencing techniques to 718 longitudinally-collected stool samples. I found that the gut microbiome composition of all travelers changed significantly during their stay, but the taxonomic diversity was stable. However, diarrhea disrupts this stability and results in an increased abundance of antibiotic resistance genes which remains high weeks after the diarrheal episode. I also identified several taxa that were differentially abundant between diarrheal and non-diarrheal samples, which were used to develop a classification model that can distinguish between the two sample types. In addition, we sequenced the genomes of 212 diarrheagenic Escherichia coli isolates, and found that isolates from travelers with diarrhea encoded more AR genes than those from healthy subjects. In summary, the gut microbiomes of international travelers’ was found to be surprisingly resilient against dysbiosis; however, they are susceptible to colonization by antibiotic resistant and multidrug-resistant bacteria, a risk that becomes more pronounced if they have travelers’ diarrhea. In the second scenario, I specifically studied the resistome dynamics among travelers to different high infectious burden regions by comparing their pre- and post-travel samples. The study showed that the destination shapes the travelers’ resistome, where travelers to a common destination share similar resistome post-travel compared to their pre-travel resistome. I also found that Southeastern Asia travelers acquired most AMR gene families compared to other travelers, and several high-risk AR genes (e.g. mcr-1, blaCTX-M-1) were borne on mobile genetic elements, highlighting the potential risk of global spread of the locally endemic AR genes. Indiscriminate use of antimicrobials brings agricultural workers at risk for potential long-term health effects from occupational exposure to AR microbes. To understand how exposure to such workplaces impact the gut resistome dynamics, I studied metagenomic samples from fourteen healthy students who visited the confined and controlled swine farms for three consecutive months and were sampled before-, during and after their visit in China. Longitudinal investigation showed extensive sharing of AR genes and microorganisms after exposure to the swine farm environment, along with several evidences of plasmid-associated AR genes that were borne on mobile genetic elements. The study also showed partial reversal of the microbiome and resistome shift within four to six-month period post-visit to the swine farms, attributing to the resiliency of the human gut microbiome. Although the antibiotics and the defense mechanisms adopted by bacteria to combat them are ancient, the spread of AR is steadily rising since the introduction of antibiotics in agriculture and medicine. To understand the impact of human exposure and how primate microbiota have changed in the antibiotic era, I investigated the resistome of wild and captive baboon population. I found expansion of resistome among captive baboons compared to the wild-type counterparts, and the captivity and lifestyle changes associated with human contact can lead to marked changes in the ecology of primate gut communities. In this thesis, I demonstrated the impact of different anthropogenic activities, like international travel and use of antibiotics in livestock farming on the microbiome and resistome of the host, and how these activities perturb the gut microbial ecosystem. Although each project demonstrates the resiliency of the microbiome and its ability to recover from the perturbed state, there are also long-term, indirect impact on the host physiology especially with the acquisition of AR genes upon exposure or enteric infections. I believe these studies lay ground with potential hypothesis that can be further tested with clinical intervention studies

Destination shapes antibiotic resistance gene acquisitions, abundance increases, and diversity changes in Dutch travelers

Abstract Background Antimicrobial-resistant bacteria and their antimicrobial resistance (AMR) genes can spread by hitchhiking in human guts. International travel can exacerbate this public health threat when travelers acquire AMR genes endemic to their destinations and bring them back to their home countries. Prior studies have demonstrated travel-related acquisition of specific opportunistic pathogens and AMR genes, but the extent and magnitude of travel’s effects on the gut resistome remain largely unknown. Methods Using whole metagenomic shotgun sequencing, functional metagenomics, and Dirichlet multinomial mixture models, we investigated the abundance, diversity, function, resistome architecture, and context of AMR genes in the fecal microbiomes of 190 Dutch individuals, before and after travel to diverse international locations. Results Travel markedly increased the abundance and α-diversity of AMR genes in the travelers’ gut resistome, and we determined that 56 unique AMR genes showed significant acquisition following international travel. These acquisition events were biased towards AMR genes with efflux, inactivation, and target replacement resistance mechanisms. Travel-induced shaping of the gut resistome had distinct correlations with geographical destination, so individuals returning to The Netherlands from the same destination country were more likely to have similar resistome features. Finally, we identified and detailed specific acquisition events of high-risk, mobile genetic element-associated AMR genes including qnr fluoroquinolone resistance genes, bla CTX-M family extended-spectrum β-lactamases, and the plasmid-borne mcr-1 colistin resistance gene. Conclusions Our results show that travel shapes the architecture of the human gut resistome and results in AMR gene acquisition against a variety of antimicrobial drug classes. These broad acquisitions highlight the putative risks that international travel poses to public health by gut resistome perturbation and the global spread of locally endemic AMR genes

Destination shapes antibiotic resistance gene acquisitions, abundance increases, and diversity changes in Dutch travelers

BACKGROUND: Antimicrobial-resistant bacteria and their antimicrobial resistance (AMR) genes can spread by hitchhiking in human guts. International travel can exacerbate this public health threat when travelers acquire AMR genes endemic to their destinations and bring them back to their home countries. Prior studies have demonstrated travel-related acquisition of specific opportunistic pathogens and AMR genes, but the extent and magnitude of travel's effects on the gut resistome remain largely unknown. METHODS: Using whole metagenomic shotgun sequencing, functional metagenomics, and Dirichlet multinomial mixture models, we investigated the abundance, diversity, function, resistome architecture, and context of AMR genes in the fecal microbiomes of 190 Dutch individuals, before and after travel to diverse international locations. RESULTS: Travel markedly increased the abundance and α-diversity of AMR genes in the travelers' gut resistome, and we determined that 56 unique AMR genes showed significant acquisition following international travel. These acquisition events were biased towards AMR genes with efflux, inactivation, and target replacement resistance mechanisms. Travel-induced shaping of the gut resistome had distinct correlations with geographical destination, so individuals returning to The Netherlands from the same destination country were more likely to have similar resistome features. Finally, we identified and detailed specific acquisition events of high-risk, mobile genetic element-associated AMR genes including qnr fluoroquinolone resistance genes, blaCTX-M family extended-spectrum β-lactamases, and the plasmid-borne mcr-1 colistin resistance gene. CONCLUSIONS: Our results show that travel shapes the architecture of the human gut resistome and results in AMR gene acquisition against a variety of antimicrobial drug classes. These broad acquisitions highlight the putative risks that international travel poses to public health by gut resistome perturbation and the global spread of locally endemic AMR genes

Destination shapes antibiotic resistance gene acquisitions, abundance increases, and diversity changes in Dutch travelers

Background: Antimicrobial-resistant bacteria and their antimicrobial resistance (AMR) genes can spread by hitchhiking in human guts. International travel can exacerbate this public health threat when travelers acquire AMR genes endemic to their destinations and bring them back to their home countries. Prior studies have demonstrated travel-related acquisition of specific opportunistic pathogens and AMR genes, but the extent and magnitude of travel’s effects on the gut resistome remain largely unknown. Methods: Using whole metagenomic shotgun sequencing, functional metagenomics, and Dirichlet multinomial mixture models, we investigated the abundance, diversity, function, resistome architecture, and context of AMR genes in the fecal microbiomes of 190 Dutch individuals, before and after travel to diverse international locations. Results: Travel markedly increased the abundance and α-diversity of AMR genes in the travelers’ gut resistome, and we determined that 56 unique AMR genes showed significant acquisition following international travel. These acquisition events were biased towards AMR genes with efflux, inactivation, and target replacement resistance mechanisms. Travel-induced shaping of the gut resistome had distinct correlations with geographical destination, so individuals returning to The Netherlands from the same destination country were more likely to have similar resistome features. Finally, we identified and detailed specific acquisition events of high-risk, mobile genetic element-associated AMR genes including qnr fluoroquinolone resistance genes, blaCTX-M family extended-spectrum β-lactamases, and the plasmid-borne mcr-1 colistin resistance gene. Conclusions: Our results show that travel shapes the architecture of the human gut resistome and results in AMR gene acquisition against a variety of antimicrobial drug classes. These broad acquisitions highlight the putative risks that international travel poses to public health by gut resistome perturbation and the global spread of locally endemic AMR genes.</p

Environmental remodeling of human gut microbiota and antibiotic resistome in livestock farms

© The Author(s) 2020.Anthropogenic environments have been implicated in enrichment and exchange of antibiotic resistance genes and bacteria. Here we study the impact of confined and controlled swine farm environments on temporal changes in the gut microbiome and resistome of veterinary students with occupational exposure for 3 months. By analyzing 16S rRNA and whole metagenome shotgun sequencing data in tandem with culture-based methods, we show that farm exposure shapes the gut microbiome of students, resulting in enrichment of potentially pathogenic taxa and antimicrobial resistance genes. Comparison of students’ gut microbiomes and resistomes to farm workers’ and environmental samples revealed extensive sharing of resistance genes and bacteria following exposure and after three months of their visit. Notably, antibiotic resistance genes were found in similar genetic contexts in student samples and farm environmental samples. Dynamic Bayesian network modeling predicted that the observed changes partially reverse over a 4-6 month period. Our results indicate that acute changes in a human’s living environment can persistently shape their gut microbiota and antibiotic resistome.This work was jointly supported by the National Key Research and Development Program of China (2016YFD0501300 to Y.-H.L.), the Program for Changjiang Scholars and Innovative Research Team in University of Ministry of Education of China (IRT_17R39 to Y.-H.L.), the Foundation for Innovation and Strengthening School Project of Guangdong (2016KCXTD010 to Y.-H.L.), the National Natural Science Foundation of China (31730097 to Y.-H.L.), the 111 Project (D20008 to J.S., X.-P.L., and Y.-H.L.), the Institutional Program Unifying Population and Laboratory-Based Sciences Burroughs Wellcome Fund grant to Washington University (supporting A.W.D.), and the National Institutes of Health (NIH) Director’s New Innovator Award (to G.D.)