Whole-Genome Sequencing of Neisseria gonorrhoeae Isolates with Antimicrobial Resistance Plasmids from a Homeless Community in Colombia.
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ABSTRACT:
SUBMITTER: Gomez OM
PROVIDER: S-EPMC9670893 | biostudies-literature | 2022 Nov
REPOSITORIES: biostudies-literature
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Whole-Genome Sequencing of Neisseria gonorrhoeae Isolates with Antimicrobial Resistance Plasmids from a Homeless Community in Colombia.
Gómez Oscar Mauricio OM Velez Diego E DE McEwen Juan G JG Villegas Aracelly A
Microbiology resource announcements 20221017 11
Neisseria gonorrhoeae is a pathogenic bacterium causing sexually transmitted infections, and it is associated with high antibiotic resistance rates. Here, we describe the genome sequences of four isolates from a homeless community in Colombia. ...[more]
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Project description:BackgroundNeisseria gonorrhoeae is a major public health problem due to increasing incidence and antimicrobial resistance. Genetic markers of reduced susceptibility have been identified; the extent to which those are representative of global antimicrobial resistance is unknown. We evaluated the performance of whole-genome sequencing (WGS) used to predict susceptibility to ciprofloxacin and other antimicrobials using a global collection of N. gonorrhoeae isolates.MethodsSusceptibility testing of common antimicrobials and the recently developed zolifodacin was performed using agar dilution to determine minimum inhibitory concentrations (MICs). We identified resistance alleles at loci known to contribute to antimicrobial resistance in N. gonorrhoeae from WGS data. We tested the ability of each locus to predict antimicrobial susceptibility.ResultsA total of 481 N. gonorrhoeae isolates, collected between 2004 and 2019 and making up 457 unique genomes, were sourced from 5 countries. All isolates with demonstrated susceptibility to ciprofloxacin (MIC ≤0.06 μg/mL) had a wild-type gyrA codon 91. Multilocus approaches were needed to predict susceptibility to other antimicrobials. All isolates were susceptible to zoliflodacin, defined by an MIC ≤0.25 μg/mL.ConclusionsSingle marker prediction can be used to inform ciprofloxacin treatment of N. gonorrhoeae infection. A combination of molecular markers may be needed to determine susceptibility for other antimicrobials.
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Project description:The whole-genome sequencing data of three N. gonorrhoeae strains isolated in the Russian Federation in 2015 are presented. According to the NG-MAST protocol, these strains are related to the globally spread ST 1407 genogroup. The analysis of their resistomes showed the absence of ermA/B/C/F genes and the presence of wild-type alleles of rpsE, rrs, rrl, rplD, rplV, macAB, and mefA genes, and these patterns explain the susceptibility of the sequenced strains to aminocyclitols (spectinomycin) and macrolides (azithromycin). Conjugative resistance determinants (blaTEM, tetM) were absent in the genomes, and the penC/ pilQ, parE, and norM alleles were shown to be wild-type, whereas single or multiple nucleotide substitutions were identified in the genes encoding targets for β-lactams (ponA, penA), tetracyclines (rpsJ), and fluoroquinolones (gyrA, parC). The additional mutations were found in porB gene and the promoter of mtrR gene, which nonspecifically reduced the susceptibility to antimicrobials due to the membrane permeability decrease and efflux pump overexpression. The diversity of mutations observed in the analyzed genomes prompted a revision of the phylogenetic relationships between the strains by comparing more than 790 groups of housekeeping genes. A high homology between the N. gonorrhoeae ST 1407 and N. gonorrhoeae ST 12556 genomes was confirmed; the latter had probably diverged from a common ancestor as a result of single mutation events. On the other hand, N. gonorrhoeae ST 12450 was an example of phenotypic convergence which appeared in the emergence of new drug resistance determinants that partially coincide with those of the ST 1407 genogroup.
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Project description:Many clinical isolates of the human pathogen Neisseria gonorrhoeae contain conjugative plasmids. The host range of these plasmids is limited to Neisseria species, but presence of a tetracycline (tetM) determinant inserted in several of these plasmids is an important cause of the rapid spread of tetracycline resistance. Previously plasmids with different backbones (Dutch and American type backbones) and with and without different tetM determinants (Dutch and American type tetM determinants) have been identified. Within the isolates tested, all plasmids with American or Dutch type tetM determinants contained a Dutch type plasmid backbone. This demonstrated that tetM determinants should not be used to differentiate between conjugal plasmid backbones. The nucleotide sequences of conjugative plasmids with Dutch type plasmid backbones either not containing the tetM determinant (pEP5233) or containing Dutch (pEP5289) or American (pEP5050) type tetM determinants were determined. Analysis of the backbone sequences showed that they belong to a novel IncP1 subfamily divergent from the IncP1alpha, beta, gamma, delta and epsilon subfamilies. The tetM determinants were inserted in a genetic load region found in all these plasmids. Insertion was accompanied by the insertion of a gene with an unknown function, and rearrangement of a toxin/antitoxin gene cluster. The genetic load region contains two toxin/antitoxins of the Zeta/Epsilon toxin/antitoxin family previously only found in Gram positive organisms and the virulence associated protein D of the VapD/VapX toxin/antitoxin family. Remarkably, presence of VapX of pJD1, a small cryptic neisserial plasmid, in the acceptor strain strongly increased the conjugation efficiency, suggesting that it functions as an antitoxin for the conjugative plasmid. The presence of the toxin and antitoxin on different plasmids might explain why the host range of this IncP1 plasmid is limited to Neisseria species. The isolated plasmids conjugated efficiently between N. gonorrhoeae strains, but did not enhance transfer of a genetic marker.
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Project description:Two extended-spectrum cephalosporin-resistant Neisseria gonorrhoeae isolates were discovered among 6,340 (0.03%) French isolates between 2010 and 2014. One isolate corresponded to the F89 multidrug-resistant N. gonorrhoeae isolate harboring a penA mosaic; whole-genome sequencing highlighted an additional R251H substitution in the ftsX gene recently involved in cephalosporin resistance. The other, ceftriaxone-resistant isolate (MIC, 0.25 mg/liter) harbored the PBP2 pattern XXXVI plus a P551S substitution and belonged to sequence type ST1579 (multilocus sequence typing [MLST]).
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Project description:BackgroundTracking the spread of the Neisseria gonorrhoeae strains with decreased susceptibility or resistance to cephalosporins is a major priority for global surveillance programmes. Whole-genome sequencing (WGS) has been widely used by increasing countries in North America, Europe, and Pacific to determine the decreased susceptible or resistance determinants of Neisseria gonorrhoeae, track the spread of these determinants throughout the gonococcal population at national or regional level. However, no studies to date have examined the genomic epidemiology of gonorrhea in Asia where the antimicrobial resistant strains of Neisseria gonorrhoeae appears to have emerged before disseminating the strains globally.MethodsWe obtained clinical isolates and data from the China Gonococcal Resistance Surveillance Programme (China-GRSP) from 2012 to 2013. We sequenced the genomes of 435 clinical isolates of Neisseria gonorrhoeae, including 112 (25.6%) isolates with decreased susceptibility to ceftriaxone (Cfx-DS). We assessed the association between antimicrobial resistance genotype and phenotype. We also compared our data with the whole genome data of the isolates from the USA and the UK in the GenBank.FindingsThe most prevalent MLST STs in our gonococcal population were MLST ST7827 (n = 74), followed by ST7365 (n = 58), ST1600 (n = 38), ST7367 (n = 35), and ST7363 (n = 29). MLST ST1901 which was reported as the predominant ST in the US was not found in our population. A total of 2512 strains, including additional 2077 published NG strains, were further included for phylogenetic analysis. It generated two distinct lineages - lineage 1 and lineage 2. Analysis of MLST ST1901 in the database indicate that most of MLST ST1901 isolates in the lineage2.6 were Cfx-DS isolates while all isolates in the lineage 2.1 were sensitive to ceftriaxone (77/110 vs. 0/13; p < 0.001). ST1901/lineage 2.6 is a ceftriaxone resistant clone which cannot distinguished by MLST genotyping. In the isolates from our study, the MICs of ceftriaxone for ST7363/lineage 2.6 isolates ranged from 0.008-0.125 mg/L (mean ± SD; 0.054 ± 0.043 mg/L) while those for ST7363/lineage 2.8 isolates ranged from 0.032-0.250 mg/L (0.134 ± 0.085 mg/L). All ST7363/lineage 2.8 isolates contained penA mosaic alleles.InterpretationTo our knowledge, current study is the first WGS-based analysis of gonococcal population at national level in Asia. China harbors the different predominant clones associated with decreased susceptibility to ceftriaxone from those clones circulated in other regions. The findings from the study can be not only used as baseline data for future studies in China but also contributable to our understanding on spread of N. gonorrhoeae and its resistant strains at regional and global levels.FundingThe Chinese Academy Medical Sciences (CAMS) Initiative for Innovative Medicine.
| S-EPMC6537553 | biostudies-literature
Project description:A multiplex assay based on a low-density hydrogel microarray was developed to identify genomic substitutions in N. gonorrhoeae that determine resistance to the currently recommended treatment agents ceftriaxone and azithromycin and the previously used drugs penicillin, tetracycline, and ciprofloxacin. The microarray identifies 74 drug resistance determinants in the N. gonorrhoeae penA, ponA, porB, gyrA, parC, rpsJ, mtrR, blaTEM, tetM, and 23S rRNA genes. The hydrogel elements were formed by automated dispensing of nanoliter-volume droplets followed by UV-induced copolymerization of NH2-containing oligonucleotides with gel-forming monomers. Polybutylene terephthalate plates without special modifications were used as microarray substrates. Sequences and concentrations of immobilized oligonucleotides, gel composition, and hybridization conditions were carefully selected, and the median discrimination ratio ranged from 2.8 to 29.4, allowing unambiguous identification of single-nucleotide substitutions. The mutation identification results in a control sample of 180 N. gonorrhoeae isolates were completely consistent with the Sanger sequencing results. In total, 648 clinical N. gonorrhoeae isolates obtained in Russia during the last 5 years were analyzed and genotyped using these microarrays. The results allowed us to draw conclusions about the present situation with antimicrobial susceptibility of N. gonorrhoeae in Russia and demonstrated the possibility of using hydrogel microarrays to control the spread of antibiotic resistance.
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Project description:BackgroundAntimicrobial resistance (AMR) and genetic determinants of resistance of N. gonorrhoeae isolates from Hefei, China, were characterized adding a breadth of information to the molecular epidemiology of gonococcal resistance in China.Methods126 N. gonorrhoeae isolates from a hospital clinic in Hefei, were collected between January, 2014, and November, 2015. The minimum inhibitory concentration (MIC) of N. gonorrhoeae isolates for seven antimicrobials were determined by the agar dilution method. Isolates were tested for mutations in penA and mtrR genes and 23S rRNA, and also genotyped using N. gonorrhoeae multi-antigen sequence typing (NG-MAST).ResultsAll N. gonorrhoeae isolates were resistant to ciprofloxacin; 81.7% (103/126) to tetracycline and 73.8% (93/126) to penicillin. 39.7% (50/126) of isolates were penicillinase producing N. gonorrhoeae (PPNG), 31.7% (40/126) were tetracycline resistant N. gonorrhoeae (TRNG) and 28.6% (36/126) were resistant to azithromycin. While not fully resistant to extended spectrum cephalosporins (ESCs), a total of 14 isolates (11.1%) displayed decreased susceptibility to ceftriaxone (MIC ≥ 0.125 mg/L, n = 10), cefixime (MIC ≥ 0. 25 mg/L, n = 1) or to both ESCs (n = 3). penA mosaic alleles XXXV were found in all isolates that harbored decreased susceptibility to cefixime, except for one. Four mutations were found in mtrR genes and mutations A2143G and C2599T were identified in 23S rRNA. No isolates were resistant to spectinomycin. Gonococcal isolates were distributed into diverse NG-MAST sequence types (STs); 86 separate STs were identified.ConclusionsN. gonorrhoeae isolates from Hefei during 2014-2015, displayed high levels of resistance to antimicrobials that had been recommended previously for treatment of gonorrhea, e.g., penicillin, tetracycline and ciprofloxacin. The prevalence of resistance to azithromycin was also high (28.6%). No isolates were found to be fully resistant to spectinomycin, ceftriaxone or cefixime; however, 11.1% isolates, overall, had decreased susceptibility to ESCs.
| S-EPMC5445337 | biostudies-literature
Project description:Surveillance of gonococcal antimicrobial resistance and the molecular characterization of the mechanisms underlying these resistance phenotypes are essential in order to establish correct empirical therapies, as well as to describe the emergence of new mechanisms in local bacterial populations. To address these goals, 149 isolates were collected over a 1-month period (October-November 2008) at the Ontario Public Health Laboratory, Toronto, Canada, and susceptibility profiles (8 antibiotics) were examined. Mutations in previously identified targets or the presence of some enzymes related to resistance (r), nonsusceptibility (ns) (resistant plus intermediate categories), or reduced susceptibility (rs) to the antibiotics tested were also studied. A significant proportion of nonsusceptibility to penicillin (PEN) (89.2%), tetracycline (TET) (72.3%), ciprofloxacin (CIP) (29%), and macrolides (erythromycin [ERY] and azithromycin; 22.3%) was found in these strains. Multidrug resistance was observed in 18.8% of the collection. Although all the strains were susceptible to spectinomycin and extended-spectrum cephalosporins (ESC) (ceftriaxone and cefixime), 9.4% of them displayed reduced susceptibility to extended-spectrum cephalosporins. PBP 2 mosaic structures were found in all of these ESC(rs) isolates. Alterations in the mtrR promoter, MtrR repressor (TET(r), PEN(ns), ESC(rs), and ERY(ns)), porin PIB (TET(r) and PEN(ns)), and ribosomal protein S10 (TET(r)) and double mutations in gyrA and parC quinolone resistance-determining regions (QRDRs) (CIP(r)) were associated with and presumably responsible for the resistance phenotypes observed. This is the first description of ESC(rs) in Canada. The detection of this phenotype indicates a change in the epidemiology of this resistance and highlights the importance of continued surveillance to preserve the last antimicrobial options available.
| S-EPMC3028768 | biostudies-literature
Project description:BackgroundThe increasing rate of antimicrobial-resistant Neisseria gonorrhoeae poses a considerable public health threat due to the difficulty in treating gonococcal infections. This study examined antimicrobial resistance (AMR) to drugs recommended for gonorrhea treatment between 2015 and 2017, and the AMR determinants and genetic compositions of plasmids in 3 gonococcal strains with high-level penicillin resistance.MethodsWe collected 117 N. gonorrhoeae isolates from patients with gonococcal infections who attended Siriraj Hospital, Bangkok, Thailand, between 2015 and 2017. Minimum inhibitory concentrations (MICs) of penicillin, tetracycline, ciprofloxacin, azithromycin, spectinomycin, cefixime, and ceftriaxone were determined by the agar dilution method. PCR amplification and sequencing of 23S rRNA and mtrR (a negative regulator of MtrCDE efflux pump) were performed. Whole genomes of 3 PPNG strains with high-level penicillin resistance (MIC ≥ 128 μg/ml) were sequenced using Illumina and Nanopore sequencing platforms.ResultsThe proportions of N. gonorrhoeae isolates with resistance were 84.6% for penicillin, 91.5% for tetracycline, and 96.6% for ciprofloxacin. All isolates were susceptible to spectinomycin, azithromycin, cefixime, and ceftriaxone. An adenine deletion within a 13 bp inverted repeat sequence in the mtrR promoter and an H105Y mutation in the mtrR coding region were found in the N. gonorrhoeae isolate with the highest azithromycin MIC value (1 μg/ml). Three high-level penicillin-resistant isolates contained nonmosaic type II penA and had mutations in penB and the mtrR coding region. All isolates with high-level penicillin resistance carried the conjugative plasmids with or without the Dutch type tetM determinant, the beta-lactamase plasmid (Rio/Toronto), and the cryptic plasmid.ConclusionsThe gonococcal population in Thailand showed high susceptibility to ceftriaxone and azithromycin, current dual therapy recommended for gonorrhea treatment. As elevated MIC of azithromycin has been observed in 1 strain of N. gonorrhoeae, expanded and enhanced surveillance of antimicrobial susceptibility and study of genetic resistance determinants are essential to improve treatment guidelines.
| S-EPMC9337635 | biostudies-literature
Project description:Whole genome sequencing of Neisseria gonorrhoeae isolates
| PRJEB40855 | ENA