Β-lactamase Resistance Genes Research Articles

Screening of Antimicrobial Resistance Genes and Epidemiological Features in Hospital and Community-Associated Carbapenem-Resistant Pseudomonas aeruginosa Infections.

IntroductionResearching carbapenem-resistant isolates enables the identification of carbapenemase-producing bacteria and prevents their spread.MethodsP. aeruginosa isolates were recovered from Medicine Faculty of Recep Tayyip Erdoğan University and identified by conventional methods and the automated Vitek 2 Compact system. Antimicrobial susceptibility experiments were performed in accordance with CLSI criteria and the automated Vitek 2 Compact system. The PCR method was investigated for the presence of β-lactamase resistance genes. PFGE typing was performed to show clonal relation among samples.ResultsSeventy P. aeruginosa isolates were isolated from seventy patients. Of the patients, 67.1% had contact with the health service in the last 90 days and 75.7% of the patients had received antimicrobial therapy in the previous 90 days. Twenty-four isolates were carbapenem resistant, 2 isolates were multidrug-resistant except colistin, and none of the samples had colistin resistance. The gene encoding β-lactamase or metallo-β-lactamase was found in a total of 36 isolates. The blaVEB and blaPER genes were identified in 1 and 5 isolates alone or 17 and 13 isolates in combination with other resistance genes, respectively. The blaNDM was the most detected metallo-β-lactamase encoding gene (n=18), followed by blaKPC (n=12). blaIMP and blaVIM were detected in 5 and 1 isolates, respectively. Also, the association of blaVEB-blaPER and blaVEB-blaKPC-blaNDM was found to be very high. Much more resistance genes and co-occurrence were detected in hospital-acquired samples than community-acquired samples. No difference was found between the community and hospital-associated isolates according to PFGE results. Simultaneously from 6 patients, other microorganisms were also isolated and 5 of them died.ConclusionThe average length of stay (days) was found to be significantly higher in HAI group than CAI group. The death of 5 patients with fewer or no resistance genes showed that the co-existence of other microorganisms in addition to resistance genes was important on death.

Bacterial Causes of Hemorrhagic Gastroenteritis in Dogs and Cats with Detection of Some Virulence and β-lactamase Resistance Genes in Escherichia coli and Salmonella by Multiplex PCR

Hemorrhagic Gastroenteritis (HGE) is a life-threatening disease caused by bacteria or viruses or endoparasites or irritant drugs or food allergies. Out of 202 sampled animals, the total bacterial isolates were 104. The identified bacterial isolates were 46 (44.23%) E. coli, 9 (8.65%) Klebsiella, 5 (4.80%) C. perfringens, 22 (21.15%) Proteus spp., 3 (2.88%) Salmonella spp., 3 (2.88%) Shigella spp. and 4 (3.84%) Pseudomonas aeruginosa, 4 (3.84%) Enterobacter species, 2 (1.92%) Citrobacter species, 2 (1.92%) Providencia rettgeri, 1 (0.96%) Hafnia species, 1 (0.96%) Serratia liquefaciens, 1 (0.96%) C. bifermentans and 1 (0.96%) C. putrefaciens. EHEC (12/26, 46.15%), EPEC (9/26, 34.61%) and ETEC (4/26, 15.38%) strains were detected by E. coli serotyping. Salmonella isolates were serotyped as Salmonella Typhimurium, S. Heidelberg, and S. Infantis. E. coli isolates from dogs were resistant to amoxicillin/clavulanic acid, cephalexin, ceftriaxone, trimethoprim/sulphamethoxazole, tetracycline, and erythromycin. Feline E. coli isolates had moderate resistance to amoxicillin/clavulanic acid, trimethoprim/ sulphonamides, and tetracycline. Salmonella isolates were highly resistant to amoxicillin/clavulanic acid, cephalexin, trimethoprim/sulphamethoxazole, tetracycline, and erythromycin. stx1 and stx2 E. coli virulence genes were detected in 80% and 60% of tested E. coli isolates, respectively while S. Typhimurium was positive for invA, hilA, and fimH virulence genes and S. Heidelberg was positive for invA and fimH genes. blaTEM and blaCTX-M1 β-lactamase resistance genes were detected in 60% and 20% of tested E. coli isolates, respectively. Salmonella Typhimurium was positive for blaCMY-1 and blaOXA-2 genes, Salmonella Heidelberg was positive for the blaCMY-1 gene. In conclusion, E. coli, Salmonella, C. perfringens, Klebsiella were major bacterial causes of HGE in dogs and cats. Additionally, E. coli and Salmonella isolated from companion animals can carry multidrug resistance genes encoding for extended-spectrum β-lactamases.

Antimicrobial resistance genes of Vibrio parahaemolyticus and Aeromonas hydrophila isolated from Nile tilapia and Mugil fish farms in Kafr-Elsheikh governorate, Egypt

The progressive expansion of aquaculture practices led to the development of bacterial disease outbreaks, otherwise the continuous and extensive use of antibiotics to overcome these diseases. Vibrio and Aeromonas species are responsible for the most severe losses affecting the aquaculture industry in Egypt. The objective of our study to investigate the antibiotic susceptibility of Vibrio parahemolyticus and Aeromonas hydrophila species isolated from Nile tilapia and Mugil fish farms in Kafr El-Sheikh province, Egypt, and molecular detection of antibiotic resistance genes. A total of 305 samples were gathered from apparently path-genomic lesions in different organs from 100 diseased fishes, 65 isolates of Vibrio species were isolated and V. parahemolyticus was isolated with an incidence of 55.4%. Out of 100 examined fish samples 72 Aeromonas species were isolated, A. hydrophila was isolated with an incidence of 99.3 %. Vibrio parahemolyticus showed high resistance for amoxicillin and colistin followed by cefotaxime and streptomycin. Meanwhile, Aeromonas hydrophila were highly resistant to amoxicillin and tetracycline followed by streptomycin, cefotaxime, and colistin. Five isolates of V. Parahemolyticus and A. hydrophila were screened using PCR for detection of 4 antibiotic resistance genes β-lactamase resistance gene (blaTEM); aminoglycosides (aadA1); tetracycline-resistant A tetA (A) and polymyxin resistance (mcr1) which were distinguished in all five V. parahaemolyticus and A. hydrophila isolates. The high detection of V. parahaemolyticus and A. hydrophila antibiotic resistance genes in our study could pose a potential risk to public health so, control methods should be taken to prevent the emergence of antibiotic resistance.

The β-lactamase profile of Escherichia coli isolates from patients with urinary tract infections in Teaching Hospital in Sulaimani, Iraq.

Escherichia coli bearing β-lactamase resistance genes are a leading cause of developing multi-drug resistance. The aim of this work was to study the molecular characterization and genotypic pattern of β-lactamase resistance genes in Escherichia coli. In total, 203 urine samples of patients who have symptoms of urinary tract infections (UTI) were screened to isolate E. coli and characterize resistance genes. Out of 203 patients, 32 (15.7%) cases were infected with E. coli. All E. coli samples showed a complete resistance against many antibiotics, including tetracycline (100%), rifampin (100%), and gentamycin (100%), but recorded the lowest resistance rate against imipenem (12.5%). Based on the existence of one or more gene of the chuA, yjaA and DNA fragment TSPE4.C2, E. coli is classified under three phylogenetic groups, A, B1, B2, and D. The highest rate of pathogenic E. coli was characterized under phylogenetic groups B2 (37.5%), and D (34.3%). Fifty β-lactamase resistance genes were recovered in this study and some isolates harbored more than one resistance genes. Among them, blaCTX recorded the highest rate, 27 (84.3%), while none of the isolates was detected to bear blaSHV resistance gene. Among five blaCMY genes, three different variants were revealed via sequencing and phylogenetic tree. Two mutations were found in one isolate at position 65 and 566, and three mutations were detected in another isolate at position number 413, 574 and 584, in comparison to a wild type variant. In conclusion, it was revealed that 15.7% cases of urinary tract infections were caused by E. coli. E. coli isolates were completely resistant to many antibiotics, but they were more sensitive to imipenem. Among the fifty β-lactamase resistance genes recovered in this study, blaCTX was the most common gene. There were three variants among blaCMY genes in a single area of study.

Detection of carbapenem-resistance genes in bacteria isolated from wastewater in Ontario

Bacterial carbapenem resistance is a major public health concern since these antimicrobials are often the last resort to treat serious human infections. To evaluate methodologies for detection of carbapenem resistance, carbapenem-tolerant bacteria were isolated from wastewater treatment plants in Toronto, Ottawa, and Arnprior, Ontario. A total of 135 carbapenem-tolerant bacteria were recovered. Polymerase chain reaction (PCR) indicated the presence of carbapenem hydrolysing enzymes KPC ( n = 10), GES ( n = 5), VIM ( n = 7), and IMP ( n = 1), and β-lactamases TEM ( n = 7), PER ( n = 1), and OXA-variants ( n = 16). A subset of 46 isolates were sequenced and analysed using ResFinder and CARD-RGI. Both programs detected carbapenem resistance genes in 35 sequenced isolates and antimicrobial resistance genes (ARGs) conferring resistance to multiple class of other antibiotics. Where β-lactamase resistance genes were not initially identified, lowering the thresholds for ARG detection enabled identification of closely related β-lactamases. However, no known carbapenem resistance genes were found in seven sequenced Pseudomonas spp. isolates. Also of note was a multi-drug-resistant Klebsiella pneumoniae isolate from Ottawa, which harboured resistance to seven antimicrobial classes including β-lactams. These results highlight the diversity of genes encoding carbapenem resistance in Ontario and the utility of whole genome sequencing over PCR for ARG detection where resistance may result from an assortment of genes.

An Acinetobacter non-baumannii Population Study: Antimicrobial Resistance Genes (ARGs).

Acinetobacter non-baumannii species are becoming common etiologic agents of nosocomial infections. Furthermore, clinical isolates belonging to this group of bacteria are usually resistant to one or more antibiotics. The current information about antibiotic resistance genes in the different A. non-baumannii species has not yet been studied as a whole. Therefore, we did a comparative study of the resistomes of A. non-baumannii pathogens based on information available in published articles and genome sequences. We searched the available literature and sequences deposited in GenBank to identify the resistance gene content of A. calcoaceticus, A. lwoffii, A. junii, A. soli, A. ursingii, A. bereziniae, A. nosocomialis, A. portensis, A. guerrae, A. baylyi, A. calcoaceticus, A. disperses, A. johnsonii, A. junii, A. lwoffii, A. nosocomialis, A. oleivorans, A. oryzae, A. pittii, A. radioresistens, and A. venetianus. The most common genes were those coding for different β-lactamases, including the carbapenemase genes blaNDM-1 and blaOXA-58. A. pittii was the species with the most β-lactamase resistance genes reported. Other genes that were commonly found include those encoding some aminoglycoside modifying enzymes, the most common being aph(6)-Id, ant(3″)-IIa, and aph(3″)-Ib, and efflux pumps. All or part of the genes coding for the AdeABC, AdeFGH, and AdeIJK efflux pumps were the most commonly found. This article incorporates all the current information about A. non-baumannii resistance genes. The comparison of the different resistomes shows that there are similarities in the genes present, but there are also significant differences that could impact the efficiency of treatments depending on the etiologic agent. This article is a comprehensive resource about A. non-baumannii resistomes.

Comparison of the elimination effectiveness of tetracycline and AmpC β-lactamase resistance genes in a municipal wastewater treatment plant using four parallel processes.

Municipal wastewater treatment plants (mWWTPs), considered reservoirs of antibiotic resistance genes (ARGs), are selected to compare the contributions of technology and process to ARG removal. Fifteen ARGs (tetA, tetB, tetC, tetE, tetG, tetL, tetM, tetO, tetQ, tetS, tetX, MOX, CIT, EBC, and FOX) and two integron genes (intI1, intI2) were tracked and detected in wastewater samples from a large-scale mWWTP with four parallel processes, including three biological technologies of AAO (anaerobic-anoxic-oxic), AB (adsorption-biodegradation), and UNITANK, two different disinfection technologies, and two primary sedimentation steps. The results showed that ARGs were widely detected, among which tetA and tetM had the highest detection rate at 100%. AAO was the most effective process in removing ARGs, followed by the AB and UNITANK processes, where the separation step was critical: 37.5% AmpC β-lactamase genes were reduced by the secondary clarifier. UV disinfection was more efficient than chlorination disinfection by 47.0% in ARG removal. Both disinfection and primary sedimentation processes could effectively remove integrons, and the swirling flow grit chamber was a more effective primary settling facility in total ARG removal than the aerated grit chamber. The tet genes and AmpC β-lactamase genes were significantly correlated with the water quality indexes of BOD5, CODCr, SS, TP, TOC, pH and NH4+-N (p < 0.05). In addition, the correlation between efflux pump genes and AmpC β-lactamase genes was strongly significant (r2 = 0.717, p < 0.01). This study provides a more powerful guide for selecting and designing treatment processes in mWWTPs with additional consideration of ARG removal.

Interpenetration of metal cations into polyelectrolyte-multilayer-films via layer-by-layer assembly: Selective antibacterial functionality of cationic guar gum/ polyacrylic acid- Ag+ nanofilm against resistant E. coli

There is an urging need for the development of effective coating systems to avoid the rapidly growing resistant bacterial strains on medical devices surfaces that causes nosocomial infections. To this end, we studied the interpenetration of metal ions into polysaccharide thin films. Using several types of polyelectrolyte multilayer (PEM) films constructed from polycations; Quaternized cellulose (QC) and Cationic guar gum (CGG), and poly anions; Carboxymethyl cellulose (CMC) and Polyacrylic acid (PAA), we found that doping of Cu2+, Fe2+ and Ag+ metal ions into PEM matrix occurred through the interactions with polyacids groups. The fabricated thin films were characterized before and after metal interpenetration by FTIR, XRD, SEM, EDX, UV–vis, and optical reflectometry (OR). In comparison to QC/CMC and CGG/CMC thin films, CGG/PAA system exhibited maximum metal loading capacity confirmed by EDX and UV–vis, and 75 % reduction in thickness (215−290 nm) confirmed by OR. Ag+ loaded CGG/PAA nanofilm exhibited a robust antibacterial activity as 50-folds than Fe2+ and 150-folds than Cu2+, against resistant E. coli bacteria via down regulation of erm and β-Lactamase resistance gene expression. Which highlights the efficacy of CGG/PAA-Ag+ as an attractive antibacterial coating material for medical devices.

Swine Manure Composting With Compound Microbial Inoculants: Removal of Antibiotic Resistance Genes and Their Associations With Microbial Community.

In this study, compound microbial inoculants, including three Bacillus strains and one Yeast strain, were inoculated into swine manure composting to explore the effects on antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs), microbial community structure, and pathogenic bacteria. The results indicated that the abundances of the detected ARGs ranged from 3.6 × 103 to 1.13 × 108 copies/g. The ARGs with the highest abundance was sul2, and the lowest was blaCTX. Composting removes most of the ARGs and MGEs by 22.8–99.7%. These ARGs were significantly reduced during the thermophilic phase of compost. The removal rate of ARGs at the different layers of compost pile was different as follows: middle layer > upper layer > lower layer. But some ARGs proliferated significantly in the maturation phase of compost, especially the sulfonamide resistance genes. Compound microbial inoculants increased the temperature of compost, accelerated water loss, nitrogen fixation, and increased the removal rate of β-lactamase resistance genes, the transposon gene tn916 and part of tetracycline resistance genes by 3.7–23.8% in compost. Compound microbial inoculants changed the community structure and increased the Bacillus abundance in the thermophilic phase of compost. And it was helpful for removing pathogens during composting. The addition of compound microbial inoculants causes the decrease of Firmicutes and the increase of Bacteroidetes, which may be related to the removal and proliferation of ARGs.

Phenotypic and genotypic characterization of Antibiotic resistant strains of Flavobacterium columnare isolated from Oreochromis niloticus (Nile tilapia)

Flavobacterium columnare (F.columnare) is an important pathogen affecting gills and skin of freshwater fishes causing columnaris disease. The objective of the study to isolate F.columnare from Nile tilapia and study the prevalence of antibiotics resistance strains with genotypic identification of β-lactamase resistance genes (blaTEM, bla SHV) and tetracycline resistance gene (tetA) among isolated strains of F.columnare. Two hundred diseased fish were randomly collected from different localities throughout year (2018-2019). F. columnare showed seasonal prevalence 0.00%, 10%, 40.00%, and 20.00% in winter, spring, summer and autumn respectively with total prevalence 35/200 (17.5%). F.columnare was isolated in high percent from skin and gills and rare in internal organs. The invitro antibacterial sensitivity test was applied on 20 F.columnare isolated strains and revealed resistant to pencillin, cephalosporin, aminoglycoside, nitrofurans, polymixinB and tetracycline. The isolated strains have three antibacterial resistance genes, bla TEM with amplicon size 516bp, bla SHV with amplicon size 392bp and tetA with amplicon size 576bp. Conclusion: According to this study tested strains were resistant to multiple antibiotics and their genes were widely distributed, so phenotypic and genotypic characterization of antibiotic resistant strains of isolated F. columnare are similar.

Antimicrobial Resistance Profile and Detection of Extended Spectrum and Amp C β-Lactamase Resistance Genes in Escherichia coli Isolated from Diarrheic Children in Lafia, Nasarawa State, Nigeria

Aims: This study evaluated the presence of extended spectrum β-lactamase (ESBL) and AmpC β-lactamase resistance genes in E. coli from stool of diarrheic children in some hospitals in Lafia metropolis, Nigeria.&#x0D; Methodology: A total of 70 stool samples of children were obtained from Dalhatu Araf Specialist Hospital, Lafia, M &amp; D Hospital, Olivet Medical Centre and Sandaji Medical Centre, Lafia. Escherichia coli were isolated and identified using standard microbiological methods. Antimicrobial susceptibility of the isolates was tested using Clinical and Laboratory Standards Institute (CLSI) method. The phenotypic detection of ESBL and AmpC β-lactamase production in some antibiotic resistant isolates were carried out using disc method. The molecular detections of ESBL and AmpC resistance genes were carried out using Polymerase Chain reaction (PCR) method.&#x0D; Results: Of the 70 samples, the occurrence of E. coli was 100%. The isolates were highly resistant to ampicillin (97.14%), ciprofloxacin (90.00%), sulfamethoxazole/trimethoprim (84.29%), streptomycin (78.57%), amoxicillin/clavulanic acid (70.00%); moderate to gentamicin (38.57%), ceftazidime (37.14%) and cefotaxime (30.00%); and less resistant to cefoxitin (15.71%) and imipenem (8.57%). Twenty-one (30.00%) isolates were jointly resistant to both cefotaxime and ceftazidine. Of this number, 66.67% (14/21) were phenotypically confirmed ESBL producers; and the occurrences of ESBL resistance genes were: 7.14% (SHV), 42.86% (CTX-M) and 50.00% (TEM). Out of 11isolates resistant to cefoxitin, 4(36.36%) were phenotypically confirmed as AmpC β-lactamase producers; and the occurrence of AmpC genes were: 50.00% (CIT), 25.00% (FOX) and 25.00% (MOX).&#x0D; Conclusion: The isolates were least resistant to imipenem and cefoxitin and highly resistant to ampicillin, ciprofloxacin and sulfamethoxazole/trimethoprim. TEM and CTX-M ESBL genes were more frequent than SHV. CIT AmpC gene was more frequent than FOX and MOX.

Gut carriage of antimicrobial resistance genes in women exposed to small-scale poultry farms in rural Uganda: A feasibility study.

Antibiotic use for livestock is presumed to be a contributor to the acquisition of antimicrobial resistance (AMR) genes in humans, yet studies do not capture AMR data before and after livestock introduction. We performed a feasibility study by recruiting a subset of women in a delayed-start randomized controlled trial of small-scale chicken farming to examine the prevalence of clinically-relevant AMR genes. Stool samples were obtained at baseline and one year post-randomization from five intervention women who received chickens at the start of the study, six control women who did not receive chickens until the end of the study, and from chickens provided to the control group at the end of the study. Stool was screened for 87 clinically significant AMR genes using a commercially available qPCR array (Qiagen). Chickens harbored 23 AMR genes from classes found in humans as well as additional vancomycin and β-lactamase resistance genes. AMR patterns between intervention and control women appeared more similar at baseline than one year post randomization (PERMANOVA R2 = 0.081, p = 0.61 at baseline, R2 = 0.186, p = 0.09 at 12 months) Women in the control group who had direct contact with the chickens sampled in the study had greater similarities in AMR gene patterns to chickens than those in the intervention group who did not have direct contact with chickens sampled (p = 0.01). However, at one year there was a trend towards increased similarity in AMR patterns between humans in both groups and the chickens sampled (p = 0.06). Studies designed to evaluate human AMR genes in the setting of animal exposure should account for high baseline AMR rates. Concomitant collection of animal, human, and environmental samples over time is recommended to determine the directionality and source of AMR genes. ClinicalTrials.gov Identifier NCT02619227.

Antimicrobial Resistance and Molecular Characterization of Gene Cassettes from Class 1 Integrons in Pseudomonas aeruginosa Strains.

We investigated the antibiotic-resistance phenotypes and molecularly characterized class 1 integron gene cassettes from 113 Pseudomonas aeruginosa isolates from patients. Primers specific for the class 1 integron integrase (intI1) gene were used to screen for these integrons using polymerase chain reactions (PCRs). The variable regions of the integrons were PCR-amplified and sequenced. Sputum was the most common specimen (69.9%; 79/113) followed by aseptic sites (21.2%; 24/113). Of the 113 isolates with phenotypic resistance to the tested antimicrobials, the highest resistances were to ciprofloxacin (CIP) (26.55%), imipenem (IPM) (23.89%), and meropenem (MEM) (23%). Carbapenem-sensitive P. aeruginosa (CS-PA) isolates displayed 23 patterns, and the predominant multidrug resistance phenotype was CIP-levofloxacin (7.23%, 6/83). Carbapenem-resistant P. aeruginosa (CR-PA) isolates displayed 12 patterns, and the predominant multidrug resistance phenotype was IPM-MEM (23.33%, 7/30). Class 1 integrons were detected in 14 (12.4%, 14/113) isolates, 7.22% (6/83) in CS-PA isolates, and 26.67% (8/30) in CR-PA isolates. Six gene cassette arrays were detected, the most prevalent being aacA4-blaOXA101-aadA5 in five isolates (4.4%, 5/113). Seventeen gene cassettes were detected. The most prevalent antibiotic-resistance gene cassettes were aacA4 (6.2%, 7/113), blaOXA-1, and blaOXA-101. Extended-spectrum β-lactamase resistance genes were detected. Some of the genes carried were similar to those in other species, but some had shared characteristics among the P. aeruginosa isolates. Long-standing drug resistance genes appeared to be under elimination in P. aeruginosa, whereas integrons conferring resistance to commonly used clinical drugs such as β-lactamases, fluoroquinolones, and even carbapenems, as well as some other gene elements, were found to be newly integrated.

Antibiotic susceptibility and molecular analyses of clinical Enterobacter cloacae isolates in Eastern Heilongjiang Province, China.

Enterobacter cloacae is an emerging opportunistic pathogen. We retrospectively conducted a study to assess antimicrobial susceptibility and investigated the Molecular characteristics of carbapenem-resistant Enterobacter cloacae (CREL) isolates. Three hundred forty-two isolates of Enterobacter cloacae were collected from January 2014 to December 2018. Ten strains of CREL were collected for further research. The species identifications and minimum inhibitory concentrations (MICs) of all antibiotics tested were analyzed using the Vitek 2 Compact system (BioMerieux, France) and supplemented by the disk diffusion method. Polymerase chain reaction (PCR) was performed to detect extended-spectrum β-lactamase (ESBL) and carbapenemase resistance genes. The results showed that most of the isolates remained susceptible to tested antibiotics; however, the resistance rate of Cefepime has been increasing in recent years. One strain co-producing New Delhi Metallo-β-lactamase NDM-1 and Imipenem hydrolase IMP-4. NDM-1 and IMP-4-producing isolates highlight that active surveillance is necessary to prevent the further spread of the bacteria. Multilocus sequence typing (MLST) showed that two KPC-producing isolates assigned to ST93, two isolates carrying NDM-1 assigned to ST1120. Moreover, the MEGA analysis showed that ST93, ST256, and ST1120 have homology, showing that CREL in our area has a potential spread risk. These findings indicating that CREL clonal dissemination may occurred in this region and should be taken seriously concern. Our study highlights an urgent need to monitor these isolates to prevent their further spread.

Evolution and distribution of resistance genes and bacterial community in water and biofilm of a simulated fish-duck integrated pond with stress.

Integrated fish-duck pond is a common circular farming model in South China, besides, it is also hot-spot for the co-selection of antibiotic resistance genes (ARGs). The aim of this study was to investigate the effects of duck manure, As and cefotaxime on the bacterial community, and the evolution and distribution of ARGs and metal(loid) resistance genes (MRGs) in water and biofilm. Five groups of fish tanks included a control and four test groups. The experimental period lasted for 100 days. Six ARGs (CIT, DHA, EBC, FOX, MOX, TEM), two MRGs (arsB, arsC), and two integron genes (intI1, intI2) were tracked and detected in water and biofilm. The results showed that duck manure brought ARGs and MRGs into fish tanks. Stress factors (cefotaxime, As) increased the relative abundance of resistance genes, and this was positively correlated with stress concentrations. The biofilm was visible significantly at the end of stage 3, and the total relative abundance of resistance genes in biofilm was higher than water from stage 5 onwards. Evolution of AmpC β-lactamase resistance genes was more obvious than MRGs, especially for MOX, which increased by 3 orders of magnitude. The abundance of Flavobacterium was higher in biofilm than in water. Moreover, correlation analysis showed that both arsB and MOX were significantly correlated with intI1 (p<0.05), which suggested a potentially dissemination risk of resistance genes. This study provides a reference for health risk assessment in integrated aquaculture environment contaminated with duck manure, antibiotics and metalloids.

Occurrence of ESBL-Producing Escherichia coli ST131, Including the H30-Rx and C1-M27 Subclones, Among Urban Seagulls from the United Kingdom.

Antimicrobial resistance is a public health concern. Understanding any role that urban seagulls may have as a reservoir of resistant bacteria could be important for reducing transmission. This study investigated fecal Escherichia coli isolates from seagulls (herring gulls and lesser black-backed gulls) to determine the prevalence of extended-spectrum cephalosporin-resistant (ESC-R) and fluoroquinolone-resistant E. coli among gull species from two cities (Taunton and Birmingham) in the United Kingdom (UK). We characterized the genetic background and carriage of plasmid-mediated resistance genes in extended-spectrum β-lactamase (ESBL)-producing E. coli obtained from these birds. Sixty ESC-R E. coli isolates were obtained from 39 seagulls (39/78, 50%), of which 28 (28/60, 46.7%) were positive for plasmid-mediated CTX-M and/or AmpC β-lactamase resistance genes. Among these, blaCTX-M-15, blaCTX-M-14, and blaCMY-2 predominated. Three isolates belonging to the B2-ST131 clone were detected, of which two harbored blaCTX-M-15 (typed to C2/H30Rx) and one harbored blaCTX-M-27 and was typed to C1/H30-R (recently described as the C1-M27 sublineage). The plasmid-mediated quinolone resistance (PMQR) gene carriage prevalence (11.7%) consisted of aac(6')-Ib-cr and qnrB genes. No carbapenem or colistin resistance genes were detected. Urban seagulls in the UK are colonized and can spread major antimicrobial-resistant E. coli isolates harboring ESBL and PMQR determinants, including clinically important strains such as the pandemic clone B2-ST131 and the C1-M27 subclade. This is the first report of ST131-C1-M27 subclade in wildlife in the UK and in seagulls worldwide.

Distribution of fosfomycin and AmpC β-lactamase resistance genes in urinary Escherichia coli isolates obtained from patients admitted to an educational hospital in Ahvaz, southwest Iran

BackgroundEscherichia coli is the most important etiologic agent of urinary tract infections (UTIs). Today, the emergence of antibiotic resistance in UTI-causing E. coli isolates is increasing. This study aimed to investigate the extended-spectrum β-lactamases (ESBLs) production, AmpC enzymes, and fosfomycin resistance genes in E. coli clinical isolates obtained from patients with UTIs. MethodsFrom December 2018 to June 2019, the E. coli isolates were collected from patients with UTI who were referred to Golestan teaching hospital in Ahvaz, Iran. Then, the antimicrobial susceptibility testing, ESBL and AmpC β-lactamase production were performed by disk diffusion method. Finally, the polymerase chain reaction was done to investigate the presence of fosfomycin resistance (fosC2, fosA3) and AmpC (blaCMY-2, blaACC) genes. ResultsIn total, 90 E. coli isolates were collected. The most effective antibiotics were fosfomycin, nitrofurantoin and imipenem with 86 (95.5%), 82 (91.1%), and 78 (86.7%) susceptibility, respectively. AmpC beta-lactamase was confirmed in 18 (20%) isolates. Moreover, 63 (90%) isolates were found as ESBL producers and 6 (6.6%) had both AmpC and ESBL β-lactamases. The frequency rates of blaCMY-2 and blaACC resistance genes in AmpC-producing isolates was 12 (13.3%) and 15 (83.3%), respectively. The fosC2 and fosA3 genes were not detected in E. coli isolates. ConclusionCurrent study revealed the emergence of E. coli isolates harboring ESBL and AmpC enzymes in southwest region of Iran. This study highlighted the need of our laboratories to adopt an appropriate method for continuous screening of ESBL and AmpC in clinical isolates on a daily basis.

Genetic Features of mcr-1 Mediated Colistin Resistance in CMY-2-Producing Escherichia coli From Romanian Poultry.

Colistin is a last resort antibiotic used for the treatment of human infections associated with carbapenemase-producing Enterobacteriales. Here, we evaluated the occurrence of mcr-1 and -2 plasmid-mediated colistin resistance in colistin and/or carbapenem resistant human clinical Enterobacteriales and other gram-negative bacteria (n = 543) as well as third generation cephalosporin-resistant (3GCR) Escherichia coli isolates from poultry abattoir workers (n = 15) and poultry fecal samples (n = 92) collected from two geographically separate abattoirs in Romania. which revealed that mcr-1 was present within four sequence types (STs): ST744 (n = 7), ST57 (n = 7), ST156 (n = 2), and ST10 (n = 1). Within STs, serotypes were conserved and, notably, all except one of the mcr-1-positive isolates were found to exhibit fluoroquinolone-resistance (FQR) associated SNPs in both gyrA and parC. While there were variations in genotypes, all isolates belonging to ST744, ST57, and ST156 were rich in resistance determinants, carrying aminoglycoside-modifying enzymes genes, sulfonamide resistance gene blaTEM–1 as well as blaCMY–2 AmpC β-lactamase resistance genes. They also exhibited high similarity in carriage of virulence genes; ST10, however, only carried the mcr-1 gene. Whole genome sequencing (WGS) analysis also revealed that although the mcr-1 gene was identified in a diverse population of E. coli, two STs (ST57 and ST744) predominated and interestingly, were found in isolates across both abattoirs providing evidence for clonal transmission. Also, two main genomic contexts of mcr-1 isolates were revealed with all ST57 isolates harboring the mcr-1 gene between two copies of ISApl1 (or the Tn6330 transposon) whilst a common mcr-1 containing scaffold, highly similar to IncX type mcr-1-bearing plasmids (pWI2-mcr, Accession number: LT838201), was present among mcr-1 isolates of varying phylogenetic backgrounds (ST10, ST744 and ST156). The high prevalence of the mcr-1 gene in poultry E. coli isolates with co-resistance to cephalosporins and quinolones, in a country where antimicrobial use in food production species is poorly regulated, is concerning and the findings from this study should lead to better surveillance of antimicrobial resistance (AMR) in food-production animals in Romania.

Multiple β-Lactam Resistance Gene-Carrying Plasmid Harbored by Klebsiella quasipneumoniae Isolated from Urban Sewage in Japan.

The continuous emergence of carbapenemase-producing Enterobacteriaceae (CPE) presents a great public health challenge. Mitigation of CPE spread in the environment is crucial, particularly from a One Health perspective. Here we describe the isolation of CPE strain SNI47 from influent water of a sewage treatment plant in Japan. SNI47 was identified as Klebsiella quasipneumoniae subsp. quasipneumoniae by phylogenetic analysis and was resistant to β-lactams, including carbapenems. Of four plasmids detected from SNI47, the 185,311-bp IncA/C2 plasmid (pTMSNI47-1), which carried 10 drug resistance genes, including genes for four β-lactamases (blaCTX-M-2, blaDHA-1, blaKHM-1, and blaOXA-10), was transferred to Escherichia coli J53 via conjugation. The MICs of all tested β-lactams for the transconjugant were higher than for the recipient. We constructed recombinant plasmids, into which each β-lactamase gene was inserted, and used them to transform E. coli DH5α cells, demonstrating that KHM-1 enhanced carbapenem resistance. In addition, these β-lactamases were responsible for a wide-spectrum β-lactam resistance acquisition with mutual compensation. KHM-1, recognized as a rare type of metallo-β-lactamase, was detected in a transferable plasmid, from a sewage treatment plant, involved in horizontal gene transfer. The detection of such plasmids raises a health risk alarm for CPE dissemination.IMPORTANCE In our investigation of urban wastewater in Japan, carbapenem-resistant Klebsiella quasipneumoniae subsp. quasipneumoniae was isolated that carried the pTMSNI47-1 plasmid, which carries four β-lactamase genes and has transferability among Enterobacteriaceae pTMSNI47-1 was found to encode a rarely reported carbapenemase, KHM-1. Cooperative effects of β-lactamases encoded by pTMSNI47-1 appeared to have broad-spectrum resistance to β-lactams. The detection of the KHM-1 gene in urban wastewater suggests that such a rare antimicrobial resistance (AMR) gene can be pooled in the environment, potentially emerging as an AMR determinant in a pathogen. When the number of β-lactamase resistance genes is increased in one plasmid, the transfer of this plasmid can confer broad-spectrum resistance to β-lactams, even if the individual gene confers narrow-spectrum resistance. The present study adds important information about the potential risk of sewage treatment plants as reservoirs and environmental suppliers of AMR genes, contributing to the public health from a One Health perspective.

Non-typhoidal Salmonella contamination in egg shells and contents from retail in Western Australia: Serovar diversity, multilocus sequence types, and phenotypic and genomic characterizations of antimicrobial resistance

In recent years, the number of human salmonellosis cases in Western Australia (WA) has increased more dramatically than in any other Australian state. In 2017, the number of cases in WA was more than double the five-year average, and eggs had emerged as the key culprit for several Salmonella foodborne disease outbreaks. To better understand such an epidemiologically intriguing situation, our research goal was to investigate the prevalence, serovar diversity, multilocus sequence types, and antimicrobial resistance of non-typhoidal Salmonella contamination in retail eggs produced and sold in WA. A total of 200 visually clean and intact retail egg samples (each containing a dozen eggs) were purchased for one year (2017–2018) from supermarkets in metropolitan Perth, the capital of WA. For each sample, the contents and shells of the 12 eggs were separately pooled and cultured according to standard methods. Overall, Salmonella was detected in 11.5% (23/200) of the tested egg samples. Salmonella was isolated from 4.5% (9/200) and 3% (6/200) of eggshells and egg contents, respectively. In 4% (8/200) of the samples, Salmonella was recovered from both eggshell and egg contents. Isolates from positive retail egg samples were serotyped as either S. Typhimurium (52.2% [12/23]) or S. Infantis (39.1% [9/23]). Both serotypes were concurrently recovered from two different retail egg samples. We retained a set of both S. Typhimurium (n = 29) and S. Infantis (n = 12) isolates from all Salmonella-positive retail packs (n = 23) for further characterization. Only two (S. Typhimurium) isolates showed resistance to ampicillin, of which one carried β-lactamase resistance gene blaTEM-1b. The remaining isolates (39/41) were susceptible to all 14 antimicrobials included in the minimum inhibitory concentrations (MICs) testing panel. Multilocus sequence typing and serotyping were perfectly mirrored, as all S. Typhimurium isolates were characterized as sequence type (ST)-19, and all S. Infantis isolates were ST-32. This study points to the noteworthy Salmonella prevalence rate in retail egg samples in WA. Our results illustrate minimal public health risks arising from antimicrobial resistance Salmonella from Australian eggs.