Comparative genomics of Flavobacterium columnare unveils novel insights in virulence and antimicrobial resistance mechanisms

AbstractThis study reports the comparative analyses of four Flavobacterium columnare isolates that have different virulence and antimicrobial resistance patterns. The main research goal was to reveal new insights into possible virulence genes by comparing the genomes of bacterial isolates that could induce tissue damage and mortality versus the genome of a non-virulent isolate. The results indicated that only the genomes of the virulent isolates possessed unique genes encoding amongst others a methyl-accepting chemotaxis protein possibly involved in the initial colonization of tissue, and several VgrG proteins engaged in interbacterial competition. Furthermore, comparisons of genes unique for the genomes of the highly virulent (HV) carp and trout isolates versus the, respectively, low and non-virulent carp and trout isolates were performed. An important part of the identified unique virulence genes of the HV-trout isolate was located in one particular gene region identified as a genomic island. This region contained araC and nodT genes, both linked to pathogenic and multidrug-resistance, and a luxR-gene, functional in bacterial cell-to-cell communication. Furthermore, the genome of the HV-trout isolate possessed unique sugar-transferases possibly important in bacterial adhesion. The second research goal was to obtain insights into the genetic basis of acquired antimicrobial resistance. Several point-mutations were discovered in gyrase-genes of an isolate showing phenotypic resistance towards first and second-generation quinolones, which were absent in isolates susceptible to quinolones. Tetracycline-resistance gene tetA was found in an isolate displaying acquired phenotypic resistance towards oxytetracycline. Although not localized on a prophage, several flanking genes were indicative of the gene’s mobile character.

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AMRFinderPlus and the Reference Gene Catalog facilitate examination of the genomic links among antimicrobial resistance, stress response, and virulence

Scientific Reports ◽

10.1038/s41598-021-91456-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Michael Feldgarden ◽

Vyacheslav Brover ◽

Narjol Gonzalez-Escalona ◽

Jonathan G. Frye ◽

Julie Haendiges ◽

...

Keyword(s):

Stress Response ◽

Antimicrobial Resistance ◽

Reference Gene ◽

Virulence Genes ◽

Stop Codon ◽

Point Mutations ◽

Resistance Mechanisms ◽

Gene Content ◽

Gene Catalog ◽

Gene Database

AbstractAntimicrobial resistance (AMR) is a significant public health threat. With the rise of affordable whole genome sequencing, in silico approaches to assessing AMR gene content can be used to detect known resistance mechanisms and potentially identify novel mechanisms. To enable accurate assessment of AMR gene content, as part of a multi-agency collaboration, NCBI developed a comprehensive AMR gene database, the Bacterial Antimicrobial Resistance Reference Gene Database and the AMR gene detection tool AMRFinder. Here, we describe the expansion of the Reference Gene Database, now called the Reference Gene Catalog, to include putative acid, biocide, metal, stress resistance genes, in addition to virulence genes and species-specific point mutations. Genes and point mutations are classified by broad functions, as well as more detailed functions. As we have expanded both the functional repertoire of identified genes and functionality, NCBI released a new version of AMRFinder, known as AMRFinderPlus. This new tool allows users the option to utilize only the core set of AMR elements, or include stress response and virulence genes, too. AMRFinderPlus can detect acquired genes and point mutations in both protein and nucleotide sequence. In addition, the evidence used to identify the gene has been expanded to include whether nucleotide or protein sequence was used, its location in the contig, and presence of an internal stop codon. These database improvements and functional expansions will enable increased precision in identifying AMR genes, linking AMR genotypes and phenotypes, and determining possible relationships between AMR, virulence, and stress response.

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Diversity and Antimicrobial Resistance in the Streptococcus bovis/Streptococcus equinus Complex (SBSEC) Isolated from Korean Domestic Ruminants

Microorganisms ◽

10.3390/microorganisms9010098 ◽

2021 ◽

Vol 9 (1) ◽

pp. 98

Author(s):

Seon Young Park ◽

Mingyung Lee ◽

Se Ra Lim ◽

Hyemin Kwon ◽

Ye Seul Lee ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Phylogenetic Analyses ◽

Tetracycline Resistance ◽

Current Status ◽

Streptococcus Bovis ◽

Limited Information ◽

Phenotypic Resistance ◽

Domestic Ruminants ◽

Antimicrobial Resistance Genes ◽

Acid Producing Bacteria

S. bovis/S. equinus complex (SBSEC) includes lactic acid-producing bacteria considered as the causative agent associated with acute rumen lactic acidosis in intensive ruminants. Considering the limited information on the detailed characteristics and diversity of SBSEC in Korea and the emergence of antimicrobial resistance (AMR), we investigated the diversity of SBSEC from domestic ruminants and verified the presence of antimicrobial resistance genes (ARGs) against several antimicrobials with their phenotypic resistance. Among 51 SBSEC isolates collected, two SBSEC members (S. equinus and S. lutetiensis) were identified; sodA-based phylogenetic analyses and comparisons of overall genome relatedness revealed potential plasticity and diversity. The AMR rates of these SBSEC against erythromycin, clindamycin, and tetracycline were relatively lower than those of other SBSEC isolates of a clinical origin. An investigation of the ARGs against those antimicrobials indicated that tetracycline resistance of SBSECs generally correlated with the presence of tet(M)-possessing Tn916-like transposon. However, no correlation between the presence of ARGs and phenotypic resistance to erythromycin and clindamycin was observed. Although a limited number of animals and their SBSEC isolates were examined, this study provides insights into the potential intraspecies biodiversity of ruminant-origin SBSEC and the current status on antimicrobial resistance of the bacteria in the Korean livestock industry.

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Genomic Analysis of Antimicrobial Resistance Genotype-to-Phenotype Agreement in Helicobacter pylori

Microorganisms ◽

10.3390/microorganisms9010002 ◽

2020 ◽

Vol 9 (1) ◽

pp. 2

Author(s):

Tal Domanovich-Asor ◽

Yair Motro ◽

Boris Khalfin ◽

Hillary A. Craddock ◽

Avi Peretz ◽

...

Keyword(s):

Helicobacter Pylori ◽

Antimicrobial Resistance ◽

Point Mutations ◽

Genomic Analysis ◽

23S Rrna ◽

Phenotype Correlation ◽

Bioinformatics Pipeline ◽

Phenotypic Resistance ◽

Commercial Kits ◽

H Pylori

Antimicrobial resistance (AMR) in Helicobacter pylori is increasing and can result in treatment failure and inappropriate antibiotic usage. This study used whole genome sequencing (WGS) to comprehensively analyze the H. pylori resistome and phylogeny in order to characterize Israeli H. pylori. Israeli H. pylori isolates (n = 48) underwent antimicrobial susceptibility testing (AST) against five antimicrobials and WGS analysis. Literature review identified 111 mutations reported to correlate with phenotypic resistance to these antimicrobials. Analysis was conducted via our in-house bioinformatics pipeline targeting point mutations in the relevant genes (pbp1A, 23S rRNA, gyrA, rdxA, frxA, and rpoB) in order to assess genotype-to-phenotype correlation. Resistance rates of study isolates were as follows: clarithromycin 54%, metronidazole 31%, amoxicillin 10%, rifampicin 4%, and levofloxacin 2%. Genotype-to-phenotype correlation was inconsistent; for every analyzed gene at least one phenotypically susceptible isolate was found to have a mutation previously associated with resistance. This was also observed regarding mutations commonly used in commercial kits to diagnose AMR in H. pylori cases. Furthermore, 11 novel point mutations associated with a resistant phenotype were detected. Analysis of a unique set of H. pylori isolates demonstrates that inferring resistance phenotypes from WGS in H. pylori remains challenging and should be optimized further.

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Antimicrobial resistance mechanisms, MLST and NG-STAR sequence types of diverse Neisseria gonorrhoeae isolates in KwaZulu-Natal, South Africa

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00759-21 ◽

2021 ◽

Author(s):

Nireshni Mitchev ◽

Ravesh Singh ◽

Mushal Allam ◽

Stanford Kwenda ◽

Arshad Ismail ◽

...

Keyword(s):

South Africa ◽

Antimicrobial Resistance ◽

Neisseria Gonorrhoeae ◽

Point Mutations ◽

Resistance Mechanisms ◽

Public Healthcare ◽

Multiple Sequence ◽

Genotypic Analysis ◽

Kwazulu Natal ◽

Sequence Types

Objective: Antimicrobial resistance (AMR) is a major challenge to managing infectious diseases. Africa has the highest incidence of gonorrhoea but there is a lack of comprehensive data from sparse surveillance programs. This study investigated the molecular epidemiology and AMR profiles of Neisseria gonorrhoeae isolates in KwaZulu-Natal province (KZN), South Africa. Methods: Repository isolates, from patients attending public healthcare clinics for STI care, were used for phenotypic and genotypic analysis. Etest® was performed to determine antimicrobial susceptibility. Whole-genome sequencing (WGS) was used to determine epidemiology and to predict susceptibility by detecting resistance-associated genes and mutations. Results: Among the 61 isolates, multiple sequence types were identified. Six isolates were novel as determined by multilocus sequence typing. N.gonorrhoeae Sequence Typing for Antimicrobial Resistance (NG-STAR) determined 48 sequence types, of which 35 isolates had novel antimicrobial profiles. Two novel penA alleles and eight novel mtrR alleles were identified. Point mutations were detected in gyrA , parC , mtrR , penA , ponA and porB1 . This study revealed a high prevalence of AMR (penicillin 67%, tetracycline 89% and ciprofloxacin 52%). However, spectinomycin, cefixime, ceftriaxone and azithromycin remained 100% effective. Conclusion: This study is one of the first to comprehensively describe the epidemiology and AMR of N. gonorrhoeae in KZN, South Africa and Africa, using WGS. KZN has a wide strain diversity and most of these sequence types have been detected in multiple countries, however more than half of our isolates have novel antimicrobial profiles. Continued surveillance is crucial to monitor the emergence of resistance to cefixime, ceftriaxone and azithromycin.

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Resistance Patterns, mcr-4 and OXA-48 Genes, and Virulence Factors of Escherichia coli from Apennine Chamois Living in Sympatry with Domestic Species, Italy

Animals ◽

10.3390/ani12020129 ◽

2022 ◽

Vol 12 (2) ◽

pp. 129

Author(s):

Camilla Smoglica ◽

Alberto Vergara ◽

Simone Angelucci ◽

Anna Rita Festino ◽

Antonio Antonucci ◽

...

Keyword(s):

Escherichia Coli ◽

Red Deer ◽

Virulence Genes ◽

Central Italy ◽

Resistance Mechanisms ◽

Phenotypic Resistance ◽

Domestic Species ◽

Resistance Patterns ◽

Biomolecular Assays ◽

Potential Pathogenicity

The aim of this study was to determine and characterize potential resistance mechanisms against selected Critically Important Antibiotics in Escherichia coli isolates collected from wild and domestic ruminants living in the Maiella National Park, in Central Italy. A total of 38 isolates were obtained from red deer, Apennine chamois, cattle, sheep, and goats grazing in lands with different levels of anthropic pressure. Antimicrobial susceptibility was determined by Minimal Inhibitory Concentration testing, showing phenotypic resistance to colistin, meropenem, or ceftazidime in 9 isolates along with one bacterial strain being resistant to three of the tested antibiotics. In addition, the biomolecular assays allowed the amplification of the genes conferring the colistin (mcr-4), the carbapenems (OXA-48), penicillins and cephalosporins (TEM, SHV, CMY-1, CMY-2) resistance. In order to describe the potential pathogenicity of isolates under study, virulence genes related to Shiga toxin-producing (STEC) and enteropathogenic (EPEC) pathovars were identified. This study is the first report of mcr-4 and OXA-48 genes in resistant E. coli harboring virulence genes in Italian wildlife, with special regard to Apennine chamois and red deer species. The multidisciplinary approach used in this study can improve the early detection of emerging antibiotic resistance determinants in human-animal-environment interfaces by means of wildlife monitoring.

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Antimicrobial Susceptibility and Characterization of Resistance Mechanisms of Corynebacterium urealyticum Clinical Isolates

Antibiotics ◽

10.3390/antibiotics9070404 ◽

2020 ◽

Vol 9 (7) ◽

pp. 404

Author(s):

Itziar Chapartegui-González ◽

Marta Fernández-Martínez ◽

Ana Rodríguez-Fernández ◽

Danilo J. P. Rocha ◽

Eric R. G. R. Aguiar ◽

...

Keyword(s):

Antimicrobial Susceptibility ◽

Antimicrobial Agents ◽

Point Mutations ◽

Tetracycline Resistance ◽

Multidrug Resistant ◽

Resistance Mechanisms ◽

Resistance Rate ◽

Clinical Isolates ◽

Tract Infections ◽

Corynebacterium Urealyticum

Corynebacterium urealyticum is a non-diphtherial urease-producing clinically relevant corynebacterial, most frequently involved in urinary tract infections. Most of the C. urealyticum clinical isolates are frequently resistant to several antibiotics. We investigated the susceptibility of 40 C. urealyticum isolated in our institution during the period 2005–2017 to eight compounds representative of the main clinically relevant classes of antimicrobial agents. Antimicrobial susceptibility was determined by the Epsilometer test. Resistance genes were searched by PCR. All strains were susceptible to vancomycin whereas linezolid and rifampicin also showed good activity (MICs90 = 1 and 0.4 mg/L, respectively). Almost all isolates (39/40, 97.5%) were multidrug resistant. The highest resistance rate was observed for ampicillin (100%), followed by erythromycin (95%) and levofloxacin (95%). Ampicillin resistance was associated with the presence of the blaA gene, encoding a class A β-lactamase. The two rifampicin-resistant strains showed point mutations driving amino acid replacements in conserved residues of RNA polymerase subunit β (RpoB). Tetracycline resistance was due to an efflux-mediated mechanism. Thirty-nine PFGE patterns were identified among the 40 C. urealyticum, indicating that they were not clonally related, but producing sporadic infections. These findings raise the need of maintaining surveillance strategies among this multidrug resistant pathogen.

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Dynamics of antimicrobial resistance and virulence genes in Enterococcus faecalis during swine manure storage

Canadian Journal of Microbiology ◽

10.1139/w10-055 ◽

2010 ◽

Vol 56 (8) ◽

pp. 683-691 ◽

Cited By ~ 8

Author(s):

Anil Mangalappalli-Illathu ◽

Patrick Duriez ◽

Luke Masson ◽

Moussa Sory Diarra ◽

Andrew Scott ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Enterococcus Faecalis ◽

Virulence Genes ◽

Swine Manure ◽

Tetracycline Resistance ◽

Storage Facility ◽

Low Frequencies ◽

Pure Cultures ◽

Manure Storage ◽

The Stability

In this study we used 2 experimental approaches to evaluate the stability of antimicrobial resistance (AMR) phenotypes, selected AMR genes, and selected virulence genes in Enterococcus faecalis during manure storage on a commercial swine farm. Isolates of E. faecalis were obtained directly from fresh fecal material (n = 120) and from the manure storage facility (n = 85) and compared. Tetracycline resistance and the virulence genes cob, esp, eep, and ccf were detected at lower frequency in manure isolates than in fecal isolates. A second approach consisted of immersing in diffusion chambers pure cultures of E. faecalis that varied in their AMR phenotypes and virulence genotypes in the swine manure storage facility for 8 weeks, sampling periodically, and evaluating the recovered strains for changes in their genotypic or phenotypic characteristics. Enterococcus faecalis populations declined exponentially, with rate constants ranging from 0.011 to 0.022 h–1. Among the AMR and virulence genes examined, 1 AMR gene (sat4) and 7 virulence genes (agrBfs, cob, cpd, cylB, efaAfs, enlA, and esp) were lost at low frequencies in the recovered strains. The AMR phenotypes were stable during the incubation, with minimal loss (P > 0.05) of the streptomycin-resistance phenotype. Overall, these results suggest that some attributes of public health significance in populations of E. faecalis decrease in frequency during manure storage.

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Genomic Analysis of Antimicrobial Resistance and Resistance Plasmids in Salmonella Serovars from Poultry in Nigeria

Antibiotics ◽

10.3390/antibiotics10020099 ◽

2021 ◽

Vol 10 (2) ◽

pp. 99

Author(s):

Abdurrahman Hassan Jibril ◽

Iruka N. Okeke ◽

Anders Dalsgaard ◽

Vanesa García Menéndez ◽

John Elmerdahl Olsen

Keyword(s):

Antimicrobial Resistance ◽

Resistance Genes ◽

Draft Genome ◽

Point Mutations ◽

Antibiotic Resistance Genes ◽

Genomic Analysis ◽

Health Concern ◽

Global Public Health ◽

Phenotypic Resistance ◽

Virulence Potential

Antimicrobial resistance is a global public health concern, and resistance genes in Salmonella, especially those located on mobile genetic elements, are part of the problem. This study used phenotypic and genomic methods to identify antimicrobial resistance and resistance genes, as well as the plasmids that bear them, in Salmonella isolates obtained from poultry in Nigeria. Seventy-four isolates were tested for susceptibility to eleven commonly used antimicrobials. Plasmid reconstruction and identification of resistance and virulence genes were performed with a draft genome using in silico approaches in parallel with plasmid extraction. Phenotypic resistance to ciprofloxacin (50.0%), gentamicin (48.6%), nalidixic acid (79.7%), sulphonamides (71.6%) and tetracycline (59.5%) was the most observed. Antibiotic resistance genes (ARGs) detected in genomes corresponded well with these observations. Commonly observed ARGs included sul1, sul2, sul3, tet (A), tet (M), qnrS1, qnrB19 and a variety of aminoglycoside-modifying genes, in addition to point mutations in the gyrA and parC genes. Multiple ARGs were predicted to be located on IncN and IncQ1 plasmids of S. Schwarzengrund and S. Muenster, and most qnrB19 genes were carried by Col (pHAD28) plasmids. Seventy-two percent (19/24) of S. Kentucky strains carried multidrug ARGs located in two distinct variants of Salmonella genomic island I. The majority of strains carried full SPI-1 and SPI-2 islands, suggesting full virulence potential.

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Associations between Antimicrobial Resistance Phenotypes, Antimicrobial Resistance Genes, and Virulence Genes of Fecal Escherichia coli Isolates from Healthy Grow-Finish Pigs

Applied and Environmental Microbiology ◽

10.1128/aem.01253-08 ◽

2009 ◽

Vol 75 (5) ◽

pp. 1373-1380 ◽

Cited By ~ 51

Author(s):

Leigh B. Rosengren ◽

Cheryl L. Waldner ◽

Richard J. Reid-Smith

Keyword(s):

Escherichia Coli ◽

Antimicrobial Resistance ◽

Resistance Genes ◽

Large Scale ◽

Virulence Genes ◽

Antimicrobial Use ◽

Phenotypic Resistance ◽

E Coli ◽

Antimicrobial Resistance Genes ◽

Antimicrobial Resistance Gene

ABSTRACT Escherichia coli often carries linked antimicrobial resistance genes on transmissible genetic elements. Through coselection, antimicrobial use may select for unrelated but linked resistance or virulence genes. This study used unconditional statistical associations to investigate the relationships between antimicrobial resistance phenotypes and antimicrobial resistance genes in 151 E. coli isolates from healthy pigs. Phenotypic resistance to each drug was significantly associated with phenotypic resistance to at least one other drug, and every association found that the probability of observing the outcome resistance was increased by the presence of the predictor resistance. With one exception, each statistical association that was identified between a pair of resistance genes had a corresponding significant association identified between the phenotypes mediated by those genes. This suggests that associations between resistance phenotypes might predict coselection. If this hypothesis is confirmed, evaluation of the associations between resistance phenotypes could improve our knowledge of coselection dynamics and provide a cost-effective way to evaluate existing data until large-scale genotypic data collection becomes feasible. This could enable policy makers and users of antimicrobials to consider coselection in antimicrobial use decisions. This study also considered the unconditional relationships between resistance and virulence genes in E. coli from healthy pigs (aidA-1, eae, elt, estA, estB, fedA1, stx1, and stx2). Positive statistical associations would suggest that antimicrobial use may select for virulence in bacteria that may contaminate food or cause diarrhea in pigs. Fortunately, the odds of detecting a virulence gene were rarely increased by the presence of an antimicrobial resistance gene. This suggests that on-farm antimicrobial use did not select for the examined virulence factors in E. coli carried by this population of healthy pigs.

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Establishing Genotypic Cutoff Values To Measure Antimicrobial Resistance in Salmonella

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02140-16 ◽

2016 ◽

Vol 61 (3) ◽

Cited By ~ 22

Author(s):

Gregory H. Tyson ◽

Shaohua Zhao ◽

Cong Li ◽

Sherry Ayers ◽

Jonathan L. Sabo ◽

...

Keyword(s):

Antimicrobial Resistance ◽

Large Scale ◽

Acquired Resistance ◽

Resistance Mechanisms ◽

Wild Type ◽

Content Type ◽

Phenotypic Resistance ◽

Large Scale Assessment ◽

Resistance Determinants ◽

Cutoff Values

ABSTRACT Whole-genome sequencing (WGS) has transformed our understanding of antimicrobial resistance, helping us to better identify and track the genetic mechanisms underlying phenotypic resistance. Previous studies have demonstrated high correlations between phenotypic resistance and the presence of known resistance determinants. However, there has never been a large-scale assessment of how well resistance genotypes correspond to specific MICs. We performed antimicrobial susceptibility testing and WGS of 1,738 nontyphoidal Salmonella strains to correlate over 20,000 MICs with resistance determinants. Using these data, we established what we term genotypic cutoff values (GCVs) for 13 antimicrobials against Salmonella. For the drugs we tested, we define a GCV as the highest MIC of isolates in a population devoid of known acquired resistance mechanisms. This definition of GCV is distinct from epidemiological cutoff values (ECVs or ECOFFs), which currently differentiate wild-type from non-wild-type strains based on MIC distributions alone without regard to genetic information. Due to the large number of isolates involved, we observed distinct MIC distributions for isolates with different resistance gene alleles, including for ciprofloxacin and tetracycline, suggesting the potential to predict MICs based on WGS data alone.

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