Exploration of the Neisseria resistome reveals resistance mechanisms in commensals that may be acquired by N. gonorrhoeae through horizontal gene transfer

Non-pathogenic Neisseria have repeatedly been demonstrated to transfer antibiotic resistance genes to their pathogenic relative, Neisseria gonorrhoeae. However, the resistance genotypes and subsequent phenotypes of non-pathogens within the genus have been studied and described less frequently. Here, we use Etests to characterize the minimum inhibitory concentrations (MICs) of a panel of Neisseria (n=26) - including several commensal species - acquired from the CDC & FDA's Antibiotic Resistance (AR) Isolate Bank to a suite of diverse antibiotics. We furthermore use whole genome sequencing and the Comprehensive Antibiotic Resistance Database (CARD) Resistance Gene Identifier (RGI) platform to predict possible causal resistance-encoding mutations. Within this panel, resistant isolates to all tested antimicrobials including penicillin (n=5/26), ceftriaxone (n=2/26), cefixime (n=3/26), tetracycline (n=10/26), azithromycin (n=11/26), and ciprofloxacin (n=4/26) were found. In total we identify 63 distinct mutations predicted by RGI to be involved in resistance. The presence of several of these mutations had clear associations with increases in MIC such as: DNA gyrase subunit A (gyrA) (S91F) and ciprofloxacin, tetracycline resistance protein (tetM) and 30S ribosomal protein S10 (rpsJ) (V57M) and tetracycline, and TEM-type beta-lactamases and penicillin. However, mutations with strong associations to macrolide and cephalosporin resistance were not conclusive. This work serves as an initial exploration into the resistance-encoding mutations harbored by non-pathogenic Neisseria, which will ultimately aid in prospective surveillance for novel resistance mechanisms that may be rapidly acquired by N. gonorrhoeae.

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Exploration of the Neisseria Resistome Reveals Resistance Mechanisms in Commensals That May Be Acquired by N. gonorrhoeae through Horizontal Gene Transfer

Antibiotics ◽

10.3390/antibiotics9100656 ◽

2020 ◽

Vol 9 (10) ◽

pp. 656

Author(s):

Michael A. Fiore ◽

Jordan C. Raisman ◽

Narayan H. Wong ◽

André O. Hudson ◽

Crista B. Wadsworth

Keyword(s):

Antibiotic Resistance ◽

Tetracycline Resistance ◽

Dna Gyrase ◽

Resistance Mechanisms ◽

Gene Identifier ◽

Prospective Surveillance ◽

Commensal Species ◽

Cephalosporin Resistance ◽

Resistance Protein ◽

Initial Exploration

Nonpathogenic Neisseria transfer mutations encoding antibiotic resistance to their pathogenic relative Neisseria gonorrhoeae. However, the resistance genotypes and subsequent phenotypes of nonpathogens within the genus have been described infrequently. Here, we characterize the minimum inhibitory concentrations (MICs) of a panel of Neisseria (n = 26)—including several commensal species—to a suite of diverse antibiotics. We furthermore use whole genome sequencing and the Comprehensive Antibiotic Resistance Database Resistance Gene Identifier (RGI) platform to predict putative resistance-encoding mutations. Resistant isolates to all tested antimicrobials including penicillin (n = 5/26), ceftriaxone (n = 2/26), cefixime (n = 3/26), tetracycline (n = 10/26), azithromycin (n = 11/26), and ciprofloxacin (n = 4/26) were found. In total, 63 distinct mutations were predicted by RGI to be involved in resistance. The presence of several mutations had clear associations with increased MIC such as DNA gyrase subunit A (gyrA) (S91F) and ciprofloxacin, tetracycline resistance protein (tetM) and 30S ribosomal protein S10 (rpsJ) (V57M) and tetracycline, and TEM-type β-lactamases and penicillin. However, mutations with strong associations to macrolide and cephalosporin resistance were not conclusive. This work serves as an initial exploration into the resistance-encoding mutations harbored by nonpathogenic Neisseria, which will ultimately aid in prospective surveillance for novel resistance mechanisms that may be rapidly acquired by N. gonorrhoeae.

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Metagenomics-Based Analysis of the Age-Related Cumulative Effect of Antibiotic Resistance Genes in Gut Microbiota

Antibiotics ◽

10.3390/antibiotics10081006 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1006

Author(s):

Lei Wu ◽

Xinqiang Xie ◽

Ying Li ◽

Tingting Liang ◽

Haojie Zhong ◽

...

Keyword(s):

Antibiotic Resistance ◽

Gut Microbiota ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Tetracycline Resistance ◽

Cumulative Effect ◽

Resistance Mechanisms ◽

Human Gut ◽

Human Gut Microbiota ◽

Age Related

Antibiotic resistance in bacteria has become a major global health problem. One of the main reservoirs of antibiotic resistance genes is the human gut microbiota. To characterise these genes, a metagenomic approach was used. In this study, a comprehensive antibiotic resistome catalog was established using fecal samples from 246 healthy individuals from world’s longevity township in Jiaoling, China. In total, 606 antibiotic resistance genes were detected. Our results indicated that antibiotic resistance genes in the human gut microbiota accumulate and become more complex with age as older groups harbour the highest abundance of these genes. Tetracycline resistance gene type tetQ was the most abundant group of antibiotic resistance genes in gut microbiota, and the main carrier of antibiotic resistance genes was Bacteroides. Antibiotic efflux, inactivation, and target alteration were found to be the dominant antimicrobial resistance mechanisms. This research may help to establish a comprehensive antibiotic resistance catalog that includes extremely long-lived healthy people such as centenarians, and may provide potential recommendations for controlling the use of antibiotics.

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Nitric oxide (NO) elicits aminoglycoside tolerance in Escherichia coli but antibiotic resistance gene carriage and NO sensitivity have not co-evolved

Archives of Microbiology ◽

10.1007/s00203-021-02245-2 ◽

2021 ◽

Author(s):

Cláudia A. Ribeiro ◽

Luke A. Rahman ◽

Louis G. Holmes ◽

Ayrianna M. Woody ◽

Calum M. Webster ◽

...

Keyword(s):

Nitric Oxide ◽

Antibiotic Resistance ◽

Antibiotic Susceptibility ◽

Bacterial Pathogens ◽

Antibiotic Resistance Gene ◽

Antibiotic Resistance Genes ◽

Resistance Mechanisms ◽

Current Work ◽

E Coli ◽

Respiratory Inhibitor

AbstractThe spread of multidrug-resistance in Gram-negative bacterial pathogens presents a major clinical challenge, and new approaches are required to combat these organisms. Nitric oxide (NO) is a well-known antimicrobial that is produced by the immune system in response to infection, and numerous studies have demonstrated that NO is a respiratory inhibitor with both bacteriostatic and bactericidal properties. However, given that loss of aerobic respiratory complexes is known to diminish antibiotic efficacy, it was hypothesised that the potent respiratory inhibitor NO would elicit similar effects. Indeed, the current work demonstrates that pre-exposure to NO-releasers elicits a > tenfold increase in IC50 for gentamicin against pathogenic E. coli (i.e. a huge decrease in lethality). It was therefore hypothesised that hyper-sensitivity to NO may have arisen in bacterial pathogens and that this trait could promote the acquisition of antibiotic-resistance mechanisms through enabling cells to persist in the presence of toxic levels of antibiotic. To test this hypothesis, genomics and microbiological approaches were used to screen a collection of E. coli clinical isolates for antibiotic susceptibility and NO tolerance, although the data did not support a correlation between increased carriage of antibiotic resistance genes and NO tolerance. However, the current work has important implications for how antibiotic susceptibility might be measured in future (i.e. ± NO) and underlines the evolutionary advantage for bacterial pathogens to maintain tolerance to toxic levels of NO.

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Identification and antibiotic resistance profiling of bacterial isolates from septicaemic soft-shelled turtles (Pelodiscus sinensis)

Veterinární Medicína ◽

10.17221/65/2016-vetmed ◽

2017 ◽

Vol 62 (No. 3) ◽

pp. 169-177 ◽

Cited By ~ 13

Author(s):

TH Chung ◽

SW Yi ◽

BS Kim ◽

WI Kim ◽

GW Shin

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Present Report ◽

Antibiotic Resistance Genes ◽

Tetracycline Resistance ◽

Edwardsiella Tarda ◽

Pelodiscus Sinensis ◽

Class 1 Integron ◽

Class 1 ◽

M 14

The present study sought to identify pathogens associated with septicaemia in the Chinese soft-shelled turtle (Pelodiscus sinensis) and to characterise antibiotic resistance in these pathogens. Twenty-three isolates recovered from the livers of diseased soft-shelled turtles were genetically identified as Aeromonas hydrophila (n = 8), A. veronii (n = 3), Citrobacter freundii (n = 4), Morganella morganii (n = 3), Edwardsiella tarda (n = 2), Wohlfahrtiimonas chitiniclastica (n = 1), Chryseobacterium sp. (n = 1), and Comamonas sp. (n = 1). Most isolates (n = 21) were resistant to ampicillin whereas a low percentage of isolates was susceptible to aminoglycosides (amikacin, gentamicin, and tobramycin). PCR assays and sequence analysis revealed the presence of the qnrS2 and blaTEM antibiotic resistance genes in all isolates. The blaDHA-1, blaCTX-M-14 and blaCMY-2 genes were harboured by 17.4% (n = 4), 13.5% (n = 3) and 8.7% (n = 2) of the strains, respectively. One or more tetracycline resistance genes were detected in 60.9% (n = 14) of the isolates. Four isolates (17.4%) harboured single or multiple class 1 integron cassettes. Collectively, a variety of bacterial pathogens were involved in the occurrence of septicaemia in Chinese soft-shelled turtles and most of the isolates had multi-antibiotic resistant phenotypes. To our knowledge, the present report is the first to identify W. chitiniclastica and Comamonas sp. as causes of septicaemia in soft-shelled turtles and the first to identify Aeromonas spp. with blaCTX-M-14 and blaDHA-1 resistance genes.

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Comparative evaluation of Assays for Broad Detection of Molecular Resistance Mechanisms in Enterobacterales Isolates

Journal of Clinical Microbiology ◽

10.1128/jcm.01033-21 ◽

2021 ◽

Author(s):

J. N. Brazelton de Cardenas ◽

C. D. Garner ◽

Y Su ◽

L Tang ◽

R. T. Hayden

Keyword(s):

Antibiotic Resistance ◽

Rapid Evolution ◽

Antibiotic Resistance Genes ◽

Resistance Mechanisms ◽

Beta Lactam ◽

Accurate Identification ◽

Gram Negative ◽

Diagnostic Role ◽

Clinically Significant ◽

Third Generation Cephalosporins

Rapid detection of antimicrobial resistance in both surveillance and diagnostic settings is still a major challenge for the clinical lab, compounded by the rapid evolution of antibiotic resistance mechanisms. This study compares four methods for the broad detection of antibiotic resistance genes in Enterobacterales isolates: two multiplex PCR assays, (the Streck ARM-D® beta-lactamase kit and the OpGen Acuitas AMR Gene Panel u5.47 (research use only (RUO)), and one microarray assay (the Check-MDR CT103XL assay), with whole genome sequencing as a reference standard. A total of 65 Gram-negative bacterial isolates, from 56 patients, classified by phenotypic AST as showing resistance to beta-lactam antimicrobials (ESBL positive, resistance to third generation cephalosporins or carbapenems), were included in the study. Overall concordance between the molecular assays and sequencing was high. While all three assays had similar performance, the OpGen Acuitas AMR assay had the highest overall percent concordance with sequencing results. The primary differences between the assays tested were the number and diversity of targets, ranging from 9 for Streck to 34 for OpGen. This study shows that commercially available PCR-based assays can provide accurate identification of antimicrobial resistant loci in clinically significant Gram-negative bacteria. Further studies are needed to determine the clinical diagnostic role and potential benefit of such methods.

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Survey and Sequence Characterization of Bovine Mastitis-Associated Escherichia coli in Dairy Herds

Frontiers in Veterinary Science ◽

10.3389/fvets.2020.582297 ◽

2020 ◽

Vol 7 ◽

Author(s):

John I. Alawneh ◽

Ben Vezina ◽

Hena R. Ramay ◽

Hulayyil Al-Harbi ◽

Ameh S. James ◽

...

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Bovine Mastitis ◽

Antibiotic Resistance Genes ◽

Tetracycline Resistance ◽

Sequence Type ◽

Genotypic Differences ◽

Beta Lactam ◽

Sequence Characterization ◽

E Coli

Escherichia coli is frequently associated with mastitis in cattle. “Pathogenic” and “commensal” isolates appear to be genetically similar. With a few exceptions, no notable genotypic differences have been found between commensal and mastitis-associated E. coli. In this study, 24 E. coli strains were isolated from dairy cows with clinical mastitis in three geographic regions of Australia (North Queensland, South Queensland, and Victoria), sequenced, then genomically surveyed. There was no observed relationship between sequence type (ST) and region (p = 0.51). The most common Multi Locus Sequence Type was ST10 (38%), then ST4429 (13%). Pangenomic analysis revealed a soft-core genome of 3,463 genes, including genes associated with antibiotic resistance, chemotaxis, motility, adhesion, biofilm formation, and pili. A total of 36 different plasmids were identified and generally found to have local distributions (p = 0.02). Only 2 plasmids contained antibiotic resistance genes, a p1303_5-like plasmid encoding multidrug-resistance (trimethoprim, quaternary ammonium, beta-lactam, streptomycin, sulfonamide, and kanamycin) from two North Queensland isolates on the same farm, while three Victorian isolates from the same farm contained a pCFSAN004177P_01-like plasmid encoding tetracycline-resistance. This pattern is consistent with a local spread of antibiotic resistance through plasmids of bovine mastitis cases. Notably, co-occurrence of plasmids containing virulence factors/antibiotic resistance with putative mobilization was rare, though the multidrug resistant p1303_5-like plasmid was predicted to be conjugative and is of some concern. This survey has provided greater understanding of antibiotic resistance within E. coli-associated bovine mastitis which will allow greater prediction and improved decision making in disease management.

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Characterization of antibiotic resistance genes in the species of the rumen microbiota

Nature Communications ◽

10.1038/s41467-019-13118-0 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 9

Author(s):

Yasmin Neves Vieira Sabino ◽

Mateus Ferreira Santana ◽

Linda Boniface Oyama ◽

Fernanda Godoy Santos ◽

Ana Júlia Silva Moreira ◽

...

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Animal Health ◽

Antibiotic Resistance Genes ◽

Tetracycline Resistance ◽

Multidrug Resistant ◽

Resistant Bacteria ◽

Ruminal Bacteria ◽

Genes Encoding

AbstractInfections caused by multidrug resistant bacteria represent a therapeutic challenge both in clinical settings and in livestock production, but the prevalence of antibiotic resistance genes among the species of bacteria that colonize the gastrointestinal tract of ruminants is not well characterized. Here, we investigate the resistome of 435 ruminal microbial genomes in silico and confirm representative phenotypes in vitro. We find a high abundance of genes encoding tetracycline resistance and evidence that the tet(W) gene is under positive selective pressure. Our findings reveal that tet(W) is located in a novel integrative and conjugative element in several ruminal bacterial genomes. Analyses of rumen microbial metatranscriptomes confirm the expression of the most abundant antibiotic resistance genes. Our data provide insight into antibiotic resistange gene profiles of the main species of ruminal bacteria and reveal the potential role of mobile genetic elements in shaping the resistome of the rumen microbiome, with implications for human and animal health.

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

Applied and Environmental Microbiology ◽

10.1128/aem.00470-09 ◽

2009 ◽

Vol 75 (19) ◽

pp. 6352-6360 ◽

Cited By ~ 25

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.

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Incidence, Distribution, and Spread of Tetracycline Resistance Determinants and Integron-Associated Antibiotic Resistance Genes among Motile Aeromonads from a Fish Farming Environment

Applied and Environmental Microbiology ◽

10.1128/aem.67.12.5675-5682.2001 ◽

2001 ◽

Vol 67 (12) ◽

pp. 5675-5682 ◽

Cited By ~ 178

Author(s):

Anja S. Schmidt ◽

Morten S. Bruun ◽

Inger Dalsgaard ◽

Jens L. Larsen

Keyword(s):

Antibiotic Resistance ◽

Resistance Gene ◽

Resistance Genes ◽

Antibiotic Resistance Genes ◽

Tetracycline Resistance ◽

Class 1 Integron ◽

Gene Cassettes ◽

Class 1 Integrons ◽

Resistance Determinants ◽

Class 1

ABSTRACT A collection of 313 motile aeromonads isolated at Danish rainbow trout farms was analyzed to identify some of the genes involved in high levels of antimicrobial resistance found in a previous field trial (A. S. Schmidt, M. S. Bruun, I. Dalsgaard, K. Pedersen, and J. L. Larsen, Appl. Environ. Microbiol. 66:4908–4915, 2000), the predominant resistance phenotype (37%) being a combined oxytetracycline (OTC) and sulphadiazine/trimethoprim resistance. Combined sulphonamide/trimethoprim resistance (135 isolates) appeared closely related to the presence of a class 1 integron (141 strains). Among the isolates containing integrons, four different combinations of integrated resistance gene cassettes occurred, in all cases including a dihydrofolate reductase gene and a downstream aminoglycoside resistance insert (87 isolates) and occasionally an additional chloramphenicol resistance gene cassette (31 isolates). In addition, 23 isolates had “empty” integrons without inserted gene cassettes. As far as OTC resistance was concerned, only 66 (30%) out of 216 resistant aeromonads could be assigned to resistance determinant class A (19 isolates), D (n = 6), or E (n = 39); three isolates contained two tetracycline resistance determinants (AD, AE, and DE). Forty OTC-resistant isolates containing large plasmids were selected as donors in a conjugation assay, 27 of which also contained a class 1 integron. Out of 17 successful R-plasmid transfers to Escherichia coli recipients, the respective integrons were cotransferred along with the tetracycline resistance determinants in 15 matings. Transconjugants were predominantly tetApositive (10 of 17) and contained class 1 integrons with two or more inserted antibiotic resistance genes. While there appeared to be a positive correlation between conjugative R-plasmids andtetA among the OTC-resistant aeromonads, tetEand the unclassified OTC resistance genes as well as class 1 integrons were equally distributed among isolates with and without plasmids. These findings indicate the implication of other mechanisms of gene transfer besides plasmid transfer in the dissemination of antibiotic resistance among environmental motile aeromonads.

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Antibiotic Resistance Characteristics of Environmental Bacteria from an Oxytetracycline Production Wastewater Treatment Plant and the Receiving River

Applied and Environmental Microbiology ◽

10.1128/aem.02964-09 ◽

2010 ◽

Vol 76 (11) ◽

pp. 3444-3451 ◽

Cited By ~ 174

Author(s):

Dong Li ◽

Tao Yu ◽

Yu Zhang ◽

Min Yang ◽

Zhen Li ◽

...

Keyword(s):

Antibiotic Resistance ◽

River Water ◽

Resistance Genes ◽

Treatment Plant ◽

Antibiotic Resistance Genes ◽

Tetracycline Resistance ◽

Production Plant ◽

Bacterial Strains ◽

High Concentration ◽

Receiving River

ABSTRACT We characterized the bacterial populations in surface water receiving effluent from an oxytetracycline (OTC) production plant. Additional sampling sites included the receiving river water 5 km upstream and 20 km downstream from the discharge point. High levels of OTC were found in the wastewater (WW), and the antibiotic was still detectable in river water downstream (RWD), with undetectable levels in river water upstream (RWU). A total of 341 bacterial strains were isolated using nonselective media, with the majority being identified as Gammaproteobacteria. The MICs were determined for 10 antibiotics representing seven different classes of antibiotics, and the corresponding values were significantly higher for the WW and RWD isolates than for the RWU isolates. Almost all bacteria (97%) from the WW and RWD samples demonstrated multidrug-resistant (MDR) phenotypes, while in RWU samples, these were less frequent (28%). The WW and RWD isolates were analyzed for the presence of 23 tetracycline (tet) resistance genes. The majority of isolates (94.2% and 95.4% in WW and RWD, respectively) harbored the corresponding genes, with tet(A) being the most common (67.0%), followed by tet(W), tet(C), tet(J), tet(L), tet(D), tet(Y), and tet(K) (in the range between 21.0% and 40.6%). Class I integrons were detected in the majority of WW and RWD isolates (97.4% and 86.2%, respectively) but were not associated with the tet genes. We hypothesize that the strong selective pressure imposed by a high concentration of OTC contributes to the wide dissemination of tetracycline resistance genes and other antibiotic resistance genes, possibly through mobile genetic elements.

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