Presence Of Antimicrobial Resistance Genes Research Articles

Whole genome sequencing and de novo assembly of Staphylococcus pseudintermedius: a pangenome approach to unravelling pathogenesis of canine pyoderma.

Staphylococcus pseudintermedius is the main aetiological agent of canine pyoderma. Whole genome sequencing is the most comprehensive way of obtaining relevant genomic information about micro-organisms. Oxford Nanopore technology enables quality sequencing and de novo assembly of the whole genome of S. pseudintermedius. Whole genome analysis of S. pseudintermedius may help to better understand the pathogenesis of canine pyodermas. Twenty-two strains of S. pseudintermedius isolated from the skin of five healthy dogs and 33 strains isolated from skin of 33 dogs with pyoderma were analysed. DNA was extracted and sequenced using Oxford Nanopore MinION, a new technology that delivers longer reads in a hand-held device. The pangenome was analysed and visualised with Anvi'o 6.1. Nanopore technology allowed the sequencing and de novo assembly of the genomes of 55 S. pseudintermedius strains isolated from healthy dogs and from dogs with pyoderma. The average genome size of S. pseudintermedius was 2.62 Mbp, with 48% being core genome. Pyoderma isolates contained a higher number of antimicrobial resistance genes, yet the total number of virulence factors genes did not change between isolates from healthy dogs and from dogs with pyoderma. Genomes of meticillin-resistant S. pseudintermedius (MRSP) strains were larger than those of meticillin-susceptible (MSSP) strains (2.80 Mbp versus 2.59 Mbp), as a consequence of a greater presence of antimicrobial resistance genes, phages and prophages. This technique allows much more precise and easier characterisation of canine S. pseudintermedius populations and may lead to a better understanding of the pathogenesis of canine pyodermas.

Draft genome sequence of a carbapenem-resistant clinical Acinetobacter baumannii revealing co-existence of four classes of β-lactamases

Nosocomial carbapenem-resistant Acinetobacter baumannii is a challenge in the treatment of intensive care unit (ICU) patients. The presence of antimicrobial resistance genes (ARGs) and mobile genetic elements can further complicate effects to combat antibiotic resistance in this post-antibiotic era. The aim of this study was to analyse the molecular basis of carbapenem resistance in A. baumannii strain DMC-32a. Strain DMC-32a, isolated from a wound swab of an ICU patient, was screened phenotypically and genotypically for carbapenem resistance. The isolate was subjected whole-genome sequencing (WGS) to investigate the resistance mechanisms. Strain DMC-32a was resistant to all tested antibiotics belonging to seven classes, except for the polymyxins. MICs determined against imipenem and meropenem were 512 mg/L and >512 mg/L, respectively. Gene-specific PCR confirmed the presence of blaNDM-1 and intI1. WGS confirmed the isolate as sequence type ST1053, the presence of four classes of β-lactamases [A (blaPER-7, blaTEM-116), B (blaNDM-1), C (blaADC-26) and D (blaOXA-23, blaOXA-51)], and four genes encoding aminoglycoside-modifying enzymes [aph(6)-Id, aph(3'')-Ib, aph(3')-VI and ant(3'')-IIc] conferring resistance to streptomycin, amikacin, kanamycin and spectinomycin. The analysis also confirmed the presence of a class 1 integron gene cassette harbouring ARGs to rifamycin (arr-2) and phenicols (cmlA5). No plasmid replicon was found from the sequence data. The co-existence of four β-lactamase classes in A. baumannii DMC-32a has not been reported previously from Bangladesh and not in this ST elsewhere. The emergence of such a nosocomial pathogen creates the need for effective therapeutics for critically-ill patients and for controlling hospital-acquired infections.

Prevalence, Risk Factors, and Antimicrobial Resistance Profile of Respiratory Pathogens Isolated From Suckling Beef Calves to Reprocessing at the Feedlot: A Longitudinal Study.

Here, we investigated the prevalence and risk factors for the presence of Histophilus somni, Mannheimia haemolytica, Mycoplasma bovis, and Pasteurella multocida in the respiratory tract of calves from the spring processing to the reprocessing at feedlots. Additionally, we characterized, phenotypically and genotypically, the antimicrobial resistance (AMR) profile of the four species. Calves from 22 cow–calf operations were enrolled in the study (n = 30 calves per operation) and sampled by deep nasopharyngeal swabs at three time points: spring processing, weaning, or induction into feedlots, and at reprocessing at the feedlot. Isolates were tested for susceptibility using the minimum inhibitory concentration (MIC) test against commonly administered antimicrobials. Additionally, a subset of isolates underwent whole-genome sequencing to infer presence of AMR genes and resistance determinants. Among studied pathogens, P. multocida was the most prevalent species, regardless of time point, followed by M. haemolytica, M. bovis, and H. somni. For M. bovis, a sharp increase in prevalence was detected at the reprocessing sampling, whereas for P. multocida, an increase in prevalence was observed at the weaning/induction sampling. Comingling and co-location of feedlots were not associated with prevalence of any respiratory pathogen. In terms of AMR, resistance against macrolides was prevalent in M. bovis, with most isolates resistant against tildipirosin, tilmicosin, and tylosin. In general, there was limited evidence to support an increase in resistance rates of respiratory bacteria from the spring processing to reprocessing at feedlots, with the exception of florfenicol resistance in M. bovis, which increased at reprocessing. Metaphylactic administration of tetracyclines at feedlot induction was not associated with the MIC of tetracyclines in any respiratory bacteria. Conversely, there were clear associations between the parenteral use of macrolides as metaphylaxis at the feedlot induction, and increased MIC against macrolides in P. multocida, M. haemolytica, and H. somni. Overall, the AMR phenotypes were corroborated by presence of AMR genes. We hypothesize that the administration of macrolides such as tulathromycin at feedlot induction contributes to historical changes in macrolides MIC data of respiratory bacteria of beef cattle.

Distribution of Salmonella spp. serotypes in poultry flocks in Abruzzo and Molise, Italy (2015-2020)

Abstract Issue/problem The Italian National Health System has implemented a National Control Program for Salmonella in poultry populations in order to reduce the prevalence of infection. Starting from 2019, the control program has introduced health measures against Salmonella breeding flocks of Gallus gallus, positive to Salmonella Infantis, an emerging serotype at European level, and widespread along the entire poultry primary production chain. Description of the problem Recent data from the European Union One Health 2019 Zoonoses Report published by EFSA, highlighted that most of the Salmonella isolates (70%) originated from broilers. Interestingly, Salmonella Infantis accounted for 29.7% of the total. Results In the framework of the Italian Salmonella National Control Program the isolation of Salmonella spp. is carried out according to ISO 6579-1: 2017 from faeces, dust and boot socks. During the 2015-2020 period, testing in the Italian Regions Abruzzo and Molise was performed on 2,791 samples and 5,039 sample units. The percentage of samples positive to Salmonella spp. in Abruzzo and Molise was 16,9% and 10,1%, respectively. Out of 395 Salmonella spp. strains isolated, Salmonella Infantis was the most prevalent, representing the 54,9% and 61,9% for Abruzzo and Molise, respectively, followed by Salmonella Livingstone (18,4%) for Abruzzo and (14,4%) for Molise. Lessons The record of Salmonella Infantis in the areas under study reflects the European trend highlighted by the EFSA 2019 Report and justifies the decision to increase the biosecurity measures in farms where this serotype is isolated and to proceed with destruction of positive animals according to the Regulation 1069/2009/EC. Key messages Salmonella Infantis represents a risk for human health due to the presence of antimicrobial resistance genes that confer resistance to relevant antibiotics used in the treatment of human infections. Biosafety guidelines and disinfection practices on poultry farms should be improved in order to reduce the spread of this serotype, also considering its capability to form biofilm.

Virulence and Antibiotic Resistance Characteristics of Vibrio Isolates From Rustic Environmental Freshwaters.

The study investigated the occurrence of antimicrobial resistance genes and virulence determinants in Vibrio species recovered from different freshwater sheds in rustic milieu. A total of 118 Vibrio isolates comprising Vibrio fluvialis (n=41), Vibrio mimicus (n=40) and V. vulnificus (n=37) was identified by amplification of ToxR, vmh and hsp60 genes. The amplification of virulence genes indicated that V. mimicus (toxR, zot, ctx, VPI, and ompU) genes were detected in 12.5%, 32.5%, 45%, 37.5% and 10% respectively. V. fluvialis genes (stn, hupO and vfh) were harboured in 48.8%, 14.6% and 19.5% isolates congruently. The other virulence genes that include vcgC and vcgE were observed in 63.1% and 29% of isolates belonging to V. vulnificus. With the exceptions of imipenem, meropenem and ciprofloxacin, most isolates exhibited more than 50% resistance to antibiotics. The antimicrobial resistance was more prevalent for polymyxin B (100%), azithromycin (100%) and least in ciprofloxacin (16.1%). Multiple antibiotic resistance index range was 0.3 and 0.8 with most isolates showing MARI of 0.8. The blaTEM, AmpC, blaGES, blaIMP, blaOXA-48 and blaKPC genes were detected in 53.3%, 42%, 29.6%, 16.6%, 15%, 11.3% and 5.6% of the isolates. Non-beta lactamases such as streptomycin resistance (aadA and strA), gentamicin resistance (aphA1) and quinolone resistance gene (qnrVC) were found in 5.2%, 44.3%, 26% and 2.8%. Chloramphenicol resistance genes (cmlA1 and catII) were found in 5.2% and 44.3% among the isolates. Our findings reveal the presence of antimicrobial resistance genes and virulent Vibrio species in aquatic environment which can have potential risk to human and animal’s health.

Prevalence and Characteristics of Multidrug-Resistant Livestock-Associated Methicillin-Resistant Staphylococcus aureus (LA-MRSA) CC398 Isolated from Quails (Coturnix Coturnix Japonica) Slaughtered for Human Consumption

Simple SummaryMethicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in both humans and animals worldwide. MRSA associated with livestock is a zoonotic pathogen that has been reported in several animals and, although its infections in humans are rare, this strain is recognized as an occupational hazard for people working in direct contact with livestock. Thus, we aimed to isolate MRSA from quails and to characterize their antimicrobial resistance and genetic lineages. One hundred swab samples were recovered from quails at the slaughterhouse. To investigate the prevalence and antimicrobial resistance of MRSA in poultry, we conducted this study on 100 quails slaughtered for human consumption. The antimicrobial resistance was investigated in all isolates as well as virulence genes and genetic lineages. Twenty-nine MRSA were isolated. The results showed that all MRSA isolates had resistance to multiple antibiotics. All strains were classified as livestock-associated. Most strains belonged to a well-known livestock-associated lineage: CC398.Livestock-associated MRSA (LA-MRSA) is a zoonotic pathogen that has been reported in several animals, and it is often associated with clonal complex (CC) 398. We aimed to isolate MRSA from quails and to characterize their antimicrobial resistance and genetic lineages. One hundred swab samples were recovered from quails at the slaughterhouse. The swabs were inoculated onto CHROMagar™ MRSA agar plates for MRSA isolation. The presence of antimicrobial-resistant genes and virulence factors was investigated by PCR. All strains were typed by MLST, SCCmec-, spa- and agr-typing. From the 100 samples, 29 MRSA were isolated. All strains were resistant to penicillin, cefoxitin, ciprofloxacin, erythromycin and clindamycin and carried the blaZ, mecA, ermB and ermC genes. All strains, except one, showed resistance to tetracycline and harbored the tetM, tetK and tetL genes in different combinations. Twenty strains belonged to ST398 and SCCmec type V, and nine strains belonged to the new ST6831. Twenty-eight out of twenty-nine strains were ascribed to t011 and one to t108. As far as we know, this is the first report of MRSA from quails slaughtered for human consumption. Most strains belonged to ST398-t011, which is the most common LA-MRSA clone found in livestock in Europe.

Virulence determinants and antimicrobial resistance of E. coli isolated from bovine clinical mastitis in some selected dairy farms of Bangladesh.

E. coli is one of the major significant pathogens causing mastitis, the most complex and costly diseases in the dairy industry worldwide. Present study was undertaken to isolate, detect the virulence factors, phylogroup, antimicrobial susceptibility and antimicrobial resistance genes in E. coli from cows with clinical mastitis. A total of 68 milk samples comprising 53 from clinical mastitis and 15 from apparently healthy cattle were collected from four different established dairy farms in Bangladesh. E. coli was isolated from the milk samples and identified by PCR targeting malB gene and sequencing of 16S rRNA gene. E. coli isolates were screened by PCR for the detection of major virulence genes (stx, eae and cdt) of diarrheagenic E. coli followed by phylogenetic grouping. Antimicrobial susceptibility of the E. coli isolates was determined by disk diffusion test and E. coli showing resistance was further screened for the presence of antimicrobial resistance genes. E. coli was isolated from 35.8% of the mastitis milk samples but none from the apparently healthy cattle milk. All the E. coli isolates were negative for stx, eae and cdt genes and belonged to the phylogenetic groups A and B1 which comprising of commensal E. coli. Antibiotic sensitivity testing revealed 84.2% (16/19) of the isolates as multidrug resistant. Highest resistance was observed against amoxicillin (94.5%) followed by ampicillin (89.5%) and tetracycline (89.5%). E. coli were found resistant against all the classes of antimicrobials used at the farm level. Tetracycline resistance gene (tetA) was detected in 100% of the tetracycline resistant E. coli and blaTEM-1 was present in 38.9% of the E. coli isolates. Findings of this study indicate a potential threat of developing antimicrobial resistance in commensal E. coli and their association with clinical mastitis. Occurrence of multidrug resistant E. coli might be responsible for the failure of antibiotic therapies in clinical mastitis as well as pose potential threat of transmitting and development of antibiotic resistance in human.

Molecular Characterization and Antimicrobial Susceptibility of Methicillin-Resistant Staphylococcus Aureus Isolates from Clinical Samples and Asymptomatic Nasal Carriers in Istanbul (Turkey)

Methicillin-resistant Staphylococcus aureus (MRSA) has been a widespread problem in Turkish hospitals. The aim of this study was to investigate the staphylococcal toxin genes of the clinical and nasal MRSA isolates, and their antibiotic resistance profiles. Isolation of nasal and clinical bacteria was done following standard microbiological methods. The presence of antimicrobial resistance genes (mec A, pvl, tsst-1, and SEs genes) was determined using the real-time polymerase chain reaction (PCR) assay. Among nasal MRSA isolates, 66.7% were toxigenic. The distribution of genes was as follows: pvl 26.7%, tsst-1 3.3%, and SEs 36.7%. Therefore, the nasal MRSA isolates had a rate of 23.3% multidrug resistance (MDR) pattern to the non-beta-lactams antibiotics. All (100%) clinical MRSA isolates were found to be toxigenic. The distribution of genes was as follows; pvl 10%, tsst-1 6.7%, and SEs 100%. The clinical MRSA isolates had a rate of 60% MDR. Following detection of pvl, tsst-1, and SEs among nasal and clinical MRSA isolates, and the presence of high antimicrobial resistance, the spread of these strains may be an additional factor contributing to the emergence of community-acquired (CA)-MRSA and hospital-acquired (HA)-MRSA. This study is the first to determine the resistance to linezolid and tigecycline in both nasal and clinical MRSA isolates, for the first time in Turkey. All nasal and clinical MRSA isolates were uniformly susceptible to vancomycin and quinupristin-dalfopristin. Our findings show that MRSA infections in Turkey can be empirically treated with vancomycin and quinupristin-dalfopristin based on the lack of demonstrable resistance to these drugs.

Phenotypic and genotypic characterization of mcr-1-positive multidrug-resistant Escherichia coli ST93, ST117, ST156, ST10, and ST744 isolated from poultry in Poland

BackgroundA plasmid-mediated mechanism of bacterial resistance to polymyxin is a serious threat to public health worldwide. The present study aimed to determine the occurrence of plasmid-mediated colistin resistance genes and to conduct the molecular characterization of mcr-positive Escherichia coli strains isolated from Polish poultry.MethodsIn this study, 318 E. coli strains were characterized by the prevalence of mcr1–mcr5 genes, antimicrobial susceptibility testing by minimal inhibitory concentration method, the presence of antimicrobial resistance genes was screened by PCR, and the biofilm formation ability was tested using the crystal violet staining method. Genetic relatedness of mcr-1-positive E. coli strains was evaluated by multilocus sequence typing method.ResultsAmong the 318 E. coli isolates, 17 (5.35%) harbored the mcr-1 gene. High antimicrobial resistance rates were observed for ampicillin (100%), tetracycline (88.24%), and chloramphenicol (82.35%). All mcr-1-positive E. coli strains were multidrug-resistant, and as many as 88.24% of the isolates contained the blaTEM gene, tetracycline (tetA and tetB), and sulfonamide (sul1, sul2, and sul3) resistance genes. Additionally, 41.18% of multidrug-resistant, mcr-1-positive E. coli isolates were moderate biofilm producers, while the rest of the strains showed weak biofilm production. Nine different sequence types were identified, and the dominant ST was ST93 (29.41%), followed by ST117 (17.65%), ST156 (11.76%), ST 8979 (11.76%), ST744 (5.88%), and ST10 (5.88%). Moreover, the new ST was identified in this study.ConclusionsOur results showed a low occurrence of mcr-1-positive E. coli strains isolated from Polish poultry; however, all the isolated strains were resistant to multiple antimicrobial agents and were able to form biofilms at low or medium level.

Molecular Analysis of Antimicrobial Resistance among Enterobacteriaceae Isolated from Diarrhoeic Calves in Egypt.

Simple SummaryBacterial antimicrobial resistance is a serious global health challenge. This study investigated the occurrence of major antimicrobial resistance genes, including integrons, ß-lactamases, and florfenicol in Enterobacteriaceae that were isolated from diarrhoeic calves in Egypt. From 120 calves, 149 isolates of bacteria were recovered, identified, and screened phenotypically against 12 antimicrobials, and molecularly for the presence of the resistance determinants of integrons, ß-lactamases and florfenicol. The findings revealed that 24.8% of the isolates exhibited multidrug resistance. Escherichia coli was found to be the most prevalent multidrug resistant species. Class 1 integrons, blaTEM, and floR genes were detected at incidence rates of 18.8%, 24.8%, and 1.3%, respectively, whereas class 2 integrons and blaCTX-M were not detected in any isolates. The higher incidence of the antimicrobial resistance genes indicate the importance of regular monitoring of the antibiotic susceptibilities of isolated bacteria to minimise the risk of human exposure to pathogens that are resistant to antimicrobials.The present study was designed to investigate the presence of genes that conferred resistance to antimicrobials among Enterobacteriaceae that were isolated from diarrhoeic calves. A total of 120 faecal samples were collected from diarrhoeic calves that were raised in Kafr El-Sheikh governorate, Egypt. The samples were screened for Enterobacteriaceae. A total of 149 isolates of bacteria were recovered and identified; Escherichia coli was found to be the most overwhelming species, followed by Citrobacter diversus, Shigella spp., Serratia spp., Providencia spp., Enterobacter spp., Klebsiella pneumoniae, Proteus spp., Klebsiella oxytoca, and Morganella morganii. All isolates were tested for susceptibility to 12 antimicrobials; resistant and intermediately resistant strains were screened by conventional polymerase chain reaction for the presence of antimicrobial resistance genes. Of the 149 isolates, 37 (24.8%) exhibited multidrug resistant phenotypes. The most prevalent multidrug resistant species were E. coli, C. diversus, Serratia spp., K. pneumoniae, Shigella spp., Providencia spp., and K. oxytoca. Class 1 integrons were detected in 28 (18.8%) isolates. All isolates were negative for class 2 integrons. The blaTEM gene was identified in 37 (24.8%) isolates, whereas no isolates carried the blaCTX-M gene. The florfenicol gene (floR) was detected in two bacterial isolates (1.3%). The findings of this study reveal that calves may act as potential reservoirs of multidrug resistant bacteria that can be easily transmitted to humans.

Next generation sequencing for pathogen detection in periprosthetic joint infections.

Periprosthetic joint infections (PJI) represent one of the most catastrophic complications following total joint arthroplasty (TJA). The lack of standardized diagnostic tests and protocols for PJI is a challenge for arthroplasty surgeons.Next generation sequencing (NGS) is an innovative diagnostic tool that can sequence microbial deoxyribonucleic acids (DNA) from a synovial fluid sample: all DNA present in a specimen is sequenced in parallel, generating millions of reads. It has been shown to be extremely useful in a culture-negative PJI setting.Metagenomic NGS (mNGS) allows for universal pathogen detection, regardless of microbe type, in a 24–48-hour timeframe: in its nanopore-base variation, mNGS also allows for antimicrobial resistance characterization.Cell-free DNA (cfDNA) NGS, characterized by lack of the cell lysis step, has a fast run-time (hours) and, together with a high sensitivity and specificity in microorganism isolation, may provide information on the presence of antimicrobial resistance genes.Metagenomics and cfDNA testing have reduced the time needed to detect infecting bacteria and represent very promising technologies for fast PJI diagnosis.NGS technologies are revolutionary methods that could disrupt the diagnostic paradigm of PJI, but a comprehensive collection of clinical evidence is still needed before they become widely used diagnostic tools.Cite this article: EFORT Open Rev 2021;6:236-244. DOI: 10.1302/2058-5241.6.200099

Spatiotemporal persistence of multiple, diverse clades and toxins of Corynebacterium diphtheriae

Diphtheria is a respiratory disease caused by the bacterium Corynebacterium diphtheriae. Although the development of a toxin-based vaccine in the 1930s has allowed a high level of control over the disease, cases have increased in recent years. Here, we describe the genomic variation of 502 C. diphtheriae isolates across 16 countries and territories over 122 years. We generate a core gene phylogeny and determine the presence of antimicrobial resistance genes and variation within the tox gene of 291 tox+ isolates. Numerous, highly diverse clusters of C. diphtheriae are observed across the phylogeny, each containing isolates from multiple countries, regions and time of isolation. The number of antimicrobial resistance genes, as well as the breadth of antibiotic resistance, is substantially greater in the last decade than ever before. We identified and analysed 18 tox gene variants, with mutations estimated to be of medium to high structural impact.

Population Genomic Analysis of Listeria monocytogenes From Food Reveals Substrate-Specific Genome Variation.

Listeria monocytogenes is the major causative agent of the foodborne illness listeriosis. Listeriosis presents as flu-like symptoms in healthy individuals, and can be fatal for children, elderly, pregnant women, and immunocompromised individuals. Estimates suggest that L. monocytogenes results in ∼1,600 illnesses and ∼260 deaths annually in the United States. L. monocytogenes can survive and persist in a variety of harsh environments, including conditions encountered in production of fermented dairy products such as cheese. For instance, microbial growth is often limited in soft cheese fermentation because of harsh pH, water content, and salt concentrations. However, L. monocytogenes has caused a number of deadly listeriosis outbreaks through the contamination of cheese. The purpose of this study was to understand if genetically distinct populations of L. monocytogenes are associated with particular foods, including cheese and dairy. To address this goal, we analyzed the population genetic structure of 504 L. monocytogenes strains isolated from food with publicly available genome assemblies. We identified 10 genetically distinct populations spanning L. monocytogenes lineages 1, II, and III and serotypes 1/2a, 1/2b, 1/2c, 4b, and 4c. We observed an overrepresentation of isolates from specific populations with cheese (population 2), fruit/vegetable (population 2), seafood (populations 5, 8 and 9) and meat (population 10). We used the Large Scale Blast Score Ratio pipeline and Roary to identify genes unique to population 1 and population 2 in comparison with all other populations, and screened for the presence of antimicrobial resistance genes and virulence genes across all isolates. We identified > 40 genes that were present at high frequency in population 1 and population 2 and absent in most other isolates. Many of these genes encoded for transcription factors, and cell surface anchored proteins. Additionally, we found that the virulence genes aut and ami were entirely or partially deleted in population 2. These results indicate that some L. monocytogenes populations may exhibit associations with particular foods, including cheese, and that gene content may contribute to this pattern.

Whole genome sequence of a New Delhi metallo-β-lactamase 1-producing Proteus mirabilis isolate SNYG35 from broiler chicken in China

ObjectivesThe global spread of the New Delhi metallo-β-lactamase 1 (NDM-1) gene poses a significant challenge to worldwide public health. Here, we characterize the whole genome of NDM-1-producing Proteus mirabilis isolate SNYG35 of broiler chicken origin in China. MethodsThe genome of SNYG35 was sequenced using a PacBio RS II sequencing instrument and Illumina HiSeq platform. SMRT cell data were assembled independently using HGAP4 and Canu v1.6, and were further polished with Illumina data using Pilon v1.22. The presence of antimicrobial resistance genes was identified using CGE ResFinder 3.2. A conjugation experiment was performed using the sodium azide-resistant Escherichia coli J53AziR strain as the recipient. ResultsThe chromosome of SNYG35 is 4 014 504 bp in size and consists of one chromosome and one plasmid named pSNYG35. It contains 3646 coding sequences, 82 tRNA genes, 22 rRNAs, and four non-coding RNAs. Besides blaNDM-1, SNYG35 harbours 26 different antimicrobial resistance genes including ESBL gene blaCTX-M-65 as well as fluoroquinolone and aminoglycoside resistance gene aac(6')-Ib-cr. The blaNDM-1-harbouring pSNYG35 is a pPrY2001-like plasmid and shares highest nucleotide identity to pHFK418-NDM. It carries a Tn1696-like multidrug-resistant region harbouring 12 different antimicrobial resistance genes, and could be transferred to E. coli J53. ConclusionsHere, we report for the first time the whole genome sequence of a NDM-1-producing P. mirabilis isolate from broiler chicken in China, which provides valuable information for tracing the potential transmission of NDM-1-producing P. mirabilis from broiler chicken to humans, as well as revealing the spread and evolution of blaNDM-1-harbouring pPrY2001-like plasmids.

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

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.

Characterization of Salmonella spp. Isolates from Swine: Virulence and Antimicrobial Resistance.

Simple SummarySalmonella is a pathogenic bacterium able to infect both humans and animals. It is diffused worldwide and, generally, animals are a source of infection for humans. Among domestic animals, swine represents an important reservoir and a frequent source of human infection, especially in some countries like Italy. To acquire information on Salmonella, in particular about epidemiology, but also virulence, pathogenesis and antimicrobial resistance, is the basis for a cohesive control program. This manuscript describes an investigation conducted on Salmonella isolates from swine, where two important characteristics were evaluated: the pathogenicity and the antimicrobial resistance. A great variability was observed among investigated strains. Salmonella serovar Typhimurium was confirmed as one of the most virulent serovars; indeed, most isolates belonging to this serovar presented many of the searched virulence factors. A high level of antimicrobial resistance was observed for some compounds (sulfonamide, tetracycline, streptomycin and ampicillin), but not for the so-called “last line antibiotics”, such as, for example, ciprofloxacin. The constant monitoring on circulating strains in reservoir animals is important to acquire information and set up adequate prophylaxis measures.Salmonella is one of the most important zoonotic pathogens worldwide. Swine represent typical reservoirs of this bacterium and a frequent source of human infection. Some intrinsic traits make some serovars or strains more virulent than others. Twenty-nine Salmonella spp. isolated from pigs belonging to 16 different serovars were analyzed for gastric acid environment resistance, presence of virulence genes (mgtC, rhuM, pipB, sopB, spvRBC, gipA, sodCI, sopE), antimicrobial resistance and presence of antimicrobial resistance genes (blaTEM, blaPSE-1, aadA1, aadA2, aphA1-lab, strA-strB, tetA, tetB, tetC, tetG, sul1, sul2, sul3). A percentage of 44.83% of strains showed constitutive and inducible gastric acid resistance, whereas 37.93% of strains became resistant only after induction. The genes sopB, pipB and mgtC were the most often detected, with 79.31%, 48.28% and 37.93% of positive strains, respectively. Salmonella virulence plasmid genes were detected in a S. enterica sup. houtenae ser. 40:z4,z23:-strain. Fifteen different virulence profiles were identified: one isolate (ser. Typhimurium) was positive for 6 genes, and 6 isolates (3 ser. Typhimurium, 2 ser. Typhimurium monophasic variant and 1 ser. Choleraesuis) scored positive for 5 genes. None of the isolates resulted resistant to cefotaxime and ciprofloxacin, while all isolates were susceptible to ceftazidime, colistin and gentamycin. Many strains were resistant to sulfonamide (75.86%), tetracycline (51.72%), streptomycin (48.28%) and ampicillin (31.03%). Twenty different resisto-types were identified. Six strains (4 ser. Typhimurium, 1 ser. Derby and 1 ser. Typhimurium monophasic variant) showed the ASSuT profile. Most detected resistance genes sul2 (34.48%), tetA (27.58%) and strA-strB (27.58%). Great variability was observed in analyzed strains. S. ser. Typhimurium was confirmed as one of the most virulent serovars. This study underlines that swine could be a reservoir and source of pathogenic Salmonella strains.

Genomic epidemiology of Corynebacterium striatum from three regions of China: an emerging national nosocomial epidemic

Corynebacteritum straitum has been considered as an emerging multi-drug-resistant (MDR) pathogen. Isolation of MDR C.striatum as the only organism from respiratory samples from hospitalized patients is increasing in China. To elucidate the genomic epidemiology and evolution of C. striatum in China. A total of 260 isolates from 2016 to 2018 were collected from three hospitals in three regions of China. Antibiotic sensitivity testing was performed on all isolates. Whole-genome sequencing was applied to all isolates to assess their genomic diversity and relationships and detect the presence of antimicrobial resistance genes (ARG) and ARG cassettes. Almost all isolates (96.2%, 250/260) showed multi-drug-resistance. Genome sequencing revealed four major lineages with lineage IV emerging as the epidemic lineage. Most of the diversity was developed in the last 6 years. Each hospital has its own predominant clones with potential spread between Hebei and Guangdong hospitals. Genomic analysis further revealed multiple antimicrobial resistance genes. Our results suggested that four lineages of C.striatum have spread in parallel across China, causing persistent and extensive transmissions within hospitals. MDR C.striatum infection has become a national epidemic. Antibiotic-driven selection pressure may have played significant roles in forming persistent and predominant clones. Our data provide the basis for surveillance and prevention strategies to control the epidemic caused by MDR C.striatum.

Prevalence of antimicrobial resistance genes and its association with restricted antimicrobial use in food-producing animals: a systematic review and meta-analysis.

There is ongoing debate regarding potential associations between restrictions of antimicrobial use and prevalence of antimicrobial resistance (AMR) in bacteria. To summarize the effects of interventions reducing antimicrobial use in food-producing animals on the prevalence of AMR genes (ARGs) in bacteria from animals and humans. We published a full systematic review of restrictions of antimicrobials in food-producing animals and their associations with AMR in bacteria. Herein, we focus on studies reporting on the association between restricted antimicrobial use and prevalence of ARGs. We used multilevel mixed-effects models and a semi-quantitative approach based on forest plots to summarize findings from studies. A positive effect of intervention [reduction in prevalence or number of ARGs in group(s) with restricted antimicrobial use] was reported from 29 studies for at least one ARG. We detected significant associations between a ban on avoparcin and diminished presence of the vanA gene in samples from animals and humans, whereas for the mecA gene, studies agreed on a positive effect of intervention in samples only from animals. Comparisons involving mcr-1, blaCTX-M, aadA2, vat(E), sul2, dfrA5, dfrA13, tet(E) and tet(P) indicated a reduced prevalence of genes in intervention groups. Conversely, no effects were detected for β-lactamases other than blaCTX-M and the remaining tet genes. The available body of scientific evidence supported that restricted use of antimicrobials in food animals was associated with an either lower or equal presence of ARGs in bacteria, with effects dependent on ARG, host species and restricted drug.

Faecal microbiota and antimicrobial resistance gene profiles of healthy foals.

The human and domestic animal faecal microbiota can carry various antimicrobial resistance genes (ARGs), especially if they have been exposed to antimicrobials. However, little is known about the ARG profile of the faecal microbiota of healthy foals. A high-throughput qPCR array was used to detect ARGs in the faecal microbiota of healthy foals. To characterise the faecal microbiota and ARG profiles in healthy Australian foals aged less than 1month. Observational study. The faecal microbiota and ARG profiles of 37 Thoroughbred foals with no known gastrointestinal disease or antimicrobial treatment were determined using 16S rRNA gene sequencing and a high-throughput ARG qPCR array. Each foal was sampled on one occasion. Firmicutes and Bacteroidetes were dominant in the faecal microbiota. Foals aged 1-2weeks had significantly lower microbiota richness than older foals. Tetracycline resistance genes were the most common ARGs in the majority of foals, regardless of age. ARGs of high clinical concern were rarely detected in the faeces. The presence of ARGs was associated with the presence of class I integron genes. Samples were collected for a case-control study so foals were not sampled longitudinally, and thus the development of the microbiota as individual foals aged could not be proven. The history of antimicrobial treatment of the dams was not collected and may have affected the microbiota of the foals. The ARGs in foal faeces varied concomitantly with age-related microbiota shifts. The high abundance of tetracycline resistance genes was likely due to the dominance of Bacteroides spp.

Detection of Antimicrobial Resistance Genes in the Milk Production Environment: Impact of Host DNA and Sequencing Depth.

Over the past decades, antimicrobial resistance (AMR) has been recognized as one of the most serious threats to public health. Although originally considered a problem to human health, the emerging crisis of AMR requires a “One Health” approach, considering human, animal, and environmental reservoirs. In this regard, the extensive use of antibiotics in the livestock production systems to treat mastitis and other bacterial diseases can lead to the presence of AMR genes in bacteria that contaminate or naturally occur in milk and dairy products, thereby introducing them into the food chain. The recent development of high-throughput next-generation sequencing (NGS) technologies is improving the fast characterization of microbial communities and their functional capabilities. In this context, whole metagenome sequencing (WMS), also called shotgun metagenomic sequencing, allows the generation of a vast amount of data which can be interrogated to generate the desired evidence, including the resistome. However, the amount of host DNA poses a major challenge to metagenome analysis. Given the current absence of literature concerning the application of WMS on milk to detect the presence of AMR genes, in the present study, we evaluated the effect of different sequencing depths, host DNA depletion methods and matrices to characterize the resistome of a milk production environment. WMS was conducted on three aliquots of bulk tank milk and three aliquots of the in-line milk filter collected from a single dairy farm; a fourth aliquot of milk and milk filter was bioinformatically subsampled. Two commercially available host DNA depletion methods were applied, and metagenomic DNA was sequenced to two different sequencing depth. Milk filters proved to be the most suitable matrices to evaluate the presence of AMR genes; besides, the pre-extraction host DNA depletion method was the most efficient approach to remove host reads. To our knowledge, this is the first study to evaluate the limitations posed by the host DNA in investigating the milk resistome with a WMS approach, confirming the circulation of AMR genes in the milk production environment.