scholarly journals Linking the Resistome and Plasmidome to the Microbiome

2018 ◽  
Author(s):  
Thibault Stalder ◽  
Max Press ◽  
Shawn Sullivan ◽  
Ivan Liachko ◽  
Eva M. Top
Keyword(s):  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Wastewater Sample ◽  
Proximity Ligation ◽  
Rapid Spread ◽  
Ligation Method ◽  
Novel Associations

AbstractThe rapid spread of antibiotic resistance is a serious human health threat. A range of environments have been identified as reservoirs of the antibiotic resistance genes (ARGs) found in pathogens. However, we lack understanding of the origins of these ARGs and their spread from environment to clinic. This is partly due to our inability to identify the bacterial hosts of ARGs and the mobile genetic elements that mediate this spread, such as plasmids and integrons. Here we demonstrated that the in vivo proximity ligation method Hi-C can determine the in situ host range of ARGs, plasmids, and integrons in a wastewater sample by physically linking them to their host chromosomes. Hi-C detected both previously known and novel associations between ARGs, mobile elements and host genomes, mostly validating this method. A better identification of the natural carriers of ARGs will aid the development of strategies to limit resistance spread to pathogens.

scholarly journals Intra- and Interspecies Conjugal Transfer of Tn916-Like Elements from Lactococcus lactis In Vitro and In Vivo

2009 ◽  
Vol 75 (19) ◽  
pp. 6352-6360 ◽  
Author(s):  
Joanna Boguslawska ◽  
Joanna Zycka-Krzesinska ◽  
Andrea Wilcks ◽  
Jacek Bardowski
Keyword(s):  
Antibiotic Resistance ◽  
Lactococcus Lactis ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Tetracycline Resistance ◽  
Raw Milk ◽  
M Gene ◽  
Transfer Resistance

ABSTRACT Tetracycline-resistant Lactococcus lactis strains originally isolated from Polish raw milk were analyzed for the ability to transfer their antibiotic resistance genes in vitro, using filter mating experiments, and in vivo, using germfree rats. Four of six analyzed L. lactis isolates were able to transfer tetracycline resistance determinants in vitro to L. lactis Bu2-60, at frequencies ranging from 10−5 to 10−7 transconjugants per recipient. Three of these four strains could also transfer resistance in vitro to Enterococcus faecalis JH2-2, whereas no transfer to Bacillus subtilis YBE01, Pseudomonas putida KT2442, Agrobacterium tumefaciens UBAPF2, or Escherichia coli JE2571 was observed. Rats were initially inoculated with the recipient E. faecalis strain JH2-2, and after a week, the L. lactis IBB477 and IBB487 donor strains were introduced. The first transconjugants were detected in fecal samples 3 days after introduction of the donors. A subtherapeutic concentration of tetracycline did not have any significant effect on the number of transconjugants, but transconjugants were observed earlier in animals dosed with this antibiotic. Molecular analysis of in vivo transconjugants containing the tet(M) gene showed that this gene was identical to tet(M) localized on the conjugative transposon Tn916. Primer-specific PCR confirmed that the Tn916 transposon was complete in all analyzed transconjugants and donors. This is the first study showing in vivo transfer of a Tn916-like antibiotic resistance transposon from L. lactis to E. faecalis. These data suggest that in certain cases food lactococci might be involved in the spread of antibiotic resistance genes to other lactic acid bacteria.

scholarly journals Enterococcus faecalisCRISPR-Cas is a robust barrier to conjugative antibiotic resistance dissemination in the murine intestine

2018 ◽  
Author(s):  
Valerie J. Price ◽  
Sara W. McBride ◽  
Karthik Hullahalli ◽  
Anushila Chatterjee ◽  
Breck A. Duerkop ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Enterococcus Faecalis ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Conjugative Plasmids ◽  
Resistance Plasmids ◽  
The Impact ◽  
Mammalian Intestine

AbstractCRISPR-Cas systems are barriers to horizontal gene transfer (HGT) in bacteria. Little is known about CRISPR-Cas interactions with conjugative plasmids, and studies investigating CRISPR-Cas/plasmid interactions inin vivomodels relevant to infectious disease are lacking. These are significant gaps in knowledge because conjugative plasmids disseminate antibiotic resistance genes among pathogensin vivo, and it is essential to identify strategies to reduce the spread of these elements. We use enterococci as models to understand the interactions of CRISPR-Cas with conjugative plasmids.Enterococcus faecalisis a native colonizer of the mammalian intestine and harbors pheromone-responsive plasmids (PRPs). PRPs mediate inter- and intraspecies transfer of antibiotic resistance genes. We assessedE. faecalisCRISPR-Cas anti-PRP activity in the mouse intestine and under varyingin vitroconditions. We observed striking differences in CRISPR-Cas efficiencyin vitroversusin vivo. With few exceptions, CRISPR-Cas blocked intestinal PRP dissemination, whilein vitro, the PRP frequently escaped CRISPR-Cas defense. Our results further the understanding of CRISPR-Cas biology by demonstrating that standardin vitroexperiments do not adequately model thein vivoanti-plasmid activity of CRISPR-Cas. Additionally, our work identifies several variables that impact the apparentin vitroanti-plasmid activity of CRISPR-Cas, including planktonic versus biofilm settings, different donor/recipient ratios, production of a plasmid-encoded bacteriocin, and the time point at which matings are sampled. Our results are clinically significant because they demonstrate that barriers to HGT encoded by normal human microbiota can have significant impacts onin vivoantibiotic resistance dissemination.ImportanceCRISPR-Cas is a type of immune system encoded by bacteria that is hypothesized to be a natural impediment to the spread of antibiotic resistance genes. In this study, we directly assessed the impact of CRISPR-Cas on antibiotic resistance dissemination in the mammalian intestine and under varyingin vitroconditions. We observed a robust effect of CRISPR-Cas onin vivobut notin vitrodissemination of antibiotic resistance plasmids in the native mammalian intestinal colonizerEnterococcus faecalis. We conclude that standard laboratory experiments currently do not appropriately model thein vivoconditions where antibiotic resistance dissemination occurs betweenE. faecalisstrains. Moreover, our results demonstrate that CRISPR-Cas encoded by native members of the mammalian intestinal microbiota can block the spread of antibiotic resistance plasmids.

scholarly journals GeneHunter, a Transposon Tool for Identification and Isolation of Cryptic Antibiotic Resistance Genes

2003 ◽  
Vol 47 (12) ◽  
pp. 3840-3845 ◽  
Author(s):  
Stephen J. Salipante ◽  
Miriam Barlow ◽  
Barry G. Hall
Keyword(s):  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Test Organism ◽  
Antibiotic Resistance Genes ◽  
Typhimurium Strain ◽  
Serovar Typhimurium ◽  
Lactamase Gene ◽  
Novel Antibiotics

ABSTRACT GeneHunter is a transposon tool designed for the experimental activation and identification of silent antibiotic resistance genes. The method permits the identification of novel resistance genes that lack previously identified homologues. Using Salmonella enterica serovar Typhimurium strain LT2 as a test organism for the in vivo version of the GeneHunter method, we were able to activate, clone, and identify two cryptic antibiotic resistance genes, the aminoglycoside acetyltransferase aac(6′)-Iaa and the probable Mar-A regulon activator rma. Because the method requires being able to electroporate the host with an efficiency of at least 1010 transformants per microgram, the in vivo method is not applicable to most microorganisms. We therefore developed an in vitro transposition method, showed that it can also recover the cryptic rma gene from S. enterica serovar Typhimurium strain LT2, and showed that it is generally applicable to a variety of microorganisms by using it to recover a cryptic metallo-β-lactamase gene from the gram-positive organism Bacillus cereus. It is anticipated that the GeneHunter method will be used to identify potential resistance genes during the development and testing of novel antibiotics, new variants of existing antibiotics, and drug inhibitor combinations.

scholarly journals Enterococcus faecalis CRISPR-Cas Is a Robust Barrier to Conjugative Antibiotic Resistance Dissemination in the Murine Intestine

mSphere ◽  
2019 ◽  
Vol 4 (4) ◽  
Author(s):  
Valerie J. Price ◽  
Sara W. McBride ◽  
Karthik Hullahalli ◽  
Anushila Chatterjee ◽  
Breck A. Duerkop ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Enterococcus Faecalis ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Content Type ◽  
Conjugative Plasmids ◽  
Resistance Plasmids ◽  
Mammalian Intestine

ABSTRACT CRISPR-Cas systems are barriers to horizontal gene transfer (HGT) in bacteria. Little is known about CRISPR-Cas interactions with conjugative plasmids, and studies investigating CRISPR-Cas/plasmid interactions in in vivo models relevant to infectious disease are lacking. These are significant gaps in knowledge because conjugative plasmids disseminate antibiotic resistance genes among pathogens in vivo, and it is essential to identify strategies to reduce the spread of these elements. We use enterococci as models to understand the interactions of CRISPR-Cas with conjugative plasmids. Enterococcus faecalis is a native colonizer of the mammalian intestine and harbors pheromone-responsive plasmids (PRPs). PRPs mediate inter- and intraspecies transfer of antibiotic resistance genes. We assessed E. faecalis CRISPR-Cas anti-PRP activity in the mouse intestine and under different in vitro conditions. We observed striking differences in CRISPR-Cas efficiency in vitro versus in vivo. With few exceptions, CRISPR-Cas blocked intestinal PRP dissemination, while in vitro, the PRP frequently escaped CRISPR-Cas defense. Our results further the understanding of CRISPR-Cas biology by demonstrating that standard in vitro experiments do not adequately model the in vivo antiplasmid activity of CRISPR-Cas. Additionally, our work identifies several variables that impact the apparent in vitro antiplasmid activity of CRISPR-Cas, including planktonic versus biofilm settings, different donor-to-recipient ratios, production of a plasmid-encoded bacteriocin, and the time point at which matings are sampled. Our results are clinically significant because they demonstrate that barriers to HGT encoded by normal (healthy) human microbiota can have significant impacts on in vivo antibiotic resistance dissemination. IMPORTANCE CRISPR-Cas is a type of immune system in bacteria that is hypothesized to be a natural impediment to the spread of antibiotic resistance genes. In this study, we directly assessed the impact of CRISPR-Cas on antibiotic resistance dissemination in the mammalian intestine and under different in vitro conditions. We observed a robust effect of CRISPR-Cas on in vivo but not in vitro dissemination of antibiotic resistance plasmids in the native mammalian intestinal colonizer Enterococcus faecalis. We conclude that standard in vitro experiments currently do not appropriately model the in vivo conditions where antibiotic resistance dissemination occurs between E. faecalis strains in the intestine. Moreover, our results demonstrate that CRISPR-Cas present in native members of the mammalian intestinal microbiota can block the spread of antibiotic resistance plasmids.

scholarly journals Pollution Characteristics of Typical Antibiotic Resistance Genes in Sludge Treatment Wetlands

2020 ◽  
Vol 194 ◽  
pp. 04010
Author(s):  
Zhuma Luosang ◽  
Wanjun Zhang ◽  
Junwen Ma ◽  
fengying Huang ◽  
Yubo Cui
Keyword(s):  
Antibiotic Resistance ◽  
Removal Efficiency ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Bottom Layer ◽  
Treatment Wetlands ◽  
Sludge Treatment ◽  
Pollution Characteristics ◽  
Box Method

The in-situ static box method was used to conduct a comparative experiment on sludge treatment wetlands (STWs). STW1 had ventilation structure, without reeds and STW2 had ventilation and was planted with reeds. The absolute abundance of sulfonamide, tetracycline and macrolide antibiotic resistance genes (ARGs) in the two STWs were analyzed, and the paper discussed pollution characteristics of typical antibiotic resistance genes in the two STWs. The results showed that three ARGs, sul1 (sulfonamides), tetC (tetracyclines), ermf (macrolides), were detected in STW1 and STW2. The concentration of arginine was sul1 > tetC > ermf. The concentration level of AGRS in STW1 and STW2 was lower in bottom layer than that of surface layer sludge. The removal efficiency of ARGs in the same system was tetC > sul1 > ermf, and the removal efficiency of surface sludge and bottom sludge in different systems was STW2 > STW1. Planting wetland plants in the STW can promote the removal of ARGs.

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