Genetic markers for detection of Escherichia coli K-12 harboring ampicillin-resistance plasmid from an industrial wastewater treatment effluent pond. - Related Documents




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204901.0000Genetic markers for detection of Escherichia coli K-12 harboring ampicillin-resistance plasmid from an industrial wastewater treatment effluent pond. Biotechnology industries that use recombinant DNA technology are potential sources for release of genetically modified organisms to the environment. Antibiotic-resistance marker genes are commonly used for recombinant bacteria selection. One example is the marker gene coding for β-lactamase (bla) in plasmids found in Escherichia coli K-12. The aim of this study was to provide an approach to develop a molecular method for genetic marker detection in E. coli K-12 harboring bla genes from an industrial wastewater treatment effluent pond (IWTEP). For the detection of bla and Achromobacter lyticus protease I (api) genes in samples from IWTEP, we employed multiplex polymerase chain reaction (PCR) using E. coli K-12 genetic marker detection primers, previously described in the literature, and primers designed in our laboratory. The microbiological screening method resulted in 22 bacterial colony-forming units isolated from three different IWTEP harvesting points. The multiplex PCR amplicons showed that five isolates were positive for the bla gene marker and negative for the E. coli K-12 and api genes. The 16S rRNA regions of positive microorganisms carrying the bla gene were genotyped by the MicroSeq®500 system. The bacteria found were Escherichia spp (3/5), Chromobacterium spp (1/5), and Aeromonas spp (1/5). None of the 22 isolated microorganisms presented the molecular pattern of E. coli K-12 harboring the bla gene. The presence of microorganisms positive for the bla gene and negative for E. coli K-12 harboring bla genes at IWTEP suggests that the ampicillin resistance found in the isolated bacteria could be from microorganisms other than the E. coli K-12 strain harboring plasmid.201627323199
273210.9998Biofilms in hospital effluents as a potential crossroads for carbapenemase-encoding strains. Bacterial resistance to carbapenem, which is mainly due to the successful dissemination of carbapenemase-encoding genes, has become a major health problem. Few studies have aimed to characterize the level of resistance in the environment, notably in hospital wastewater, which is a likely hotspot for exchange of antibiotic resistance genes. In this work, we looked for the presence of imipenem-resistant bacteria and imipenem in the effluent of the teaching hospital of Clermont-Ferrand, France. Selective growth of bacteria from 14-day old biofilms formed in the pipe sewer showed that 22.1% of the isolates were imipenem-resistant and identified as Aeromonas (n = 23), Pseudomonas (n = 10), Stenotrophomonas (n = 4) and Acinetobacter (n = 1). Fifteen of these strains harbored acquired carbapenemase-encoding genes bla(VIM) (n = 11), bla(OXA-48) (n = 2), bla(GES) (n = 1), bla(NDM) (n = 1). All isolates also harbored associated resistances to aminoglycosides, fluoroquinolones and/or tetracyclin. S1-nuclease pulsed-field gel electrophoresis analysis of eight selected isolates showed that four of them harbored one to two plasmids of molecular weight between 48.5 Kb and 194 Kb. In vitro transformation assays evidenced the presence of bla(VIM) and bla(NDM) on plasmids with the bla(VIM) harboring 80 Kb plasmid having conjugative capacity. The predicted environmental concentration of imipenem in the hospital effluent was 3.16 μg/L, suggesting that biofilm bacteria are subjected to sub-MICs of imipenem within the effluent. However, no imipenem molecule was detected in the hospital effluent, probably owing to its instability: in vitro assays indicated that imipenem's biological activity was no longer detectable after 45 h of storage. However, the predictive value of the hazard quotient relative to the development of resistance was >1.0 (HQr = 28.9 ± 1.9), which indicates a possible risk. The presence of carbapenemase-encoding genes in hospital effluent biofilm strains and their ability to transfer are therefore a potential hazard that should not be neglected and points to the need for monitoring antibiotic resistance in hospital wastewater.201930530220
205020.9998Identification of a novel fosfomycin resistance gene (fosA2) in Enterobacter cloacae from the Salmon River, Canada. AIMS: To investigate the occurrence of fosfomycin-resistant (fos(R) ) bacteria in aquatic environments. METHODS AND RESULTS: A fos(R) strain of Enterobacter cloacae was isolated from a water sample collected at a site (50°41'33·44″N, 119°19'49·50″W) near the mouth of the Salmon River at Salmon Arm, in south-central British Columbia, Canada. The strain was identified by PCR screening for plasmid-borne, fosA-family amplicons, followed by selective plating. Sequencing of the resistance gene cloned using PCR primers to conserved flanking DNA revealed a new allele (95% amino acid identity to fosA), and I-Ceu I PFGE showed that it was chromosomally located. In Escherichia coli, the cloned DNA conferred a greater resistance to fosfomycin than its fosA counterpart. CONCLUSIONS: Gene fosA2 conferred fosfomycin resistance in an environmental isolate of Ent. cloacae. SIGNIFICANCE AND IMPACT OF THE STUDY: The repurposing of older antibiotics should be considered in the light of existing reservoirs of resistance genes in the environment.201121392044
170930.9998High prevalence of bla(VIM-1) gene in bacteria from Brazilian soil. This study investigated bacteria from soil samples to (i) determine the main bacterial genera and species having resistance to carbapenem and other β-lactams and (ii) establish if the mechanism of resistance was due to the production of metallo-β-lactamases. The isolates were characterized by PCR for metallo-β-lactamases and integrons, by antimicrobial susceptibility testing, and by sequencing. The antimicrobial profile of 40 imipenem-resistant Gram-positive soil isolates from all Brazilian regions demonstrated that 31 (77.5%) of them were multidrug resistant. Among the 40 isolates, 19 presented the bla(VIM) gene and class 1 integrons by PCR. Six of the 19 isolates were identified as Paenibacillus sp., 12 as Bacillus sp., and just 1 was classified as Staphylococcus sp., by sequencing of the 16S rRNA gene. These results suggest that bacteria from soil can act as a source of bla(VIM-1) genes, representing a threat to public health.201627392282
298840.9998Establishment of a Multiplex Loop-Mediated Isothermal Amplification Method for Rapid Detection of Sulfonamide Resistance Genes (sul1, sul2, sul3) in Clinical Enterobacteriaceae Isolates from Poultry. Antimicrobial resistance genes play an important role in mediating resistance to sulfonamide in Gram-negative bacteria. While PCR is the current method to detect sulfonamide resistance genes (sul1, sul2, sul3), it is time-consuming and costly and there is an urgent need to develop a more convenient, simpler and rapid test for the sul. In this study, we describe a multiplex loop-mediated isothermal amplification (m-LAMP) assay we developed for the rapid and simultaneous detection of three sul. This m-LAMP assay successfully detected seven reference strains with different sul genotypes, but was negative for nine sul-negative reference strains. The m-LAMP products were verified by HinfI restriction enzyme digestion and the detection limit of the test was 0.5 pg genomic DNA per reaction. Testing 307 sulfonamide-resistant Enterobacteriaceae clinical isolates with the m-LAMP revealed all were positive for the sul with sul2 (79.5%) and sul1 (64.5%) being most prevalent, and sul3 the least (12.1%). Of the Enterobacteriaceae isolates tested, the Salmonella Indiana, a newly emerging serovar resistant to numerous antimicrobials, were most commonly positive with 33% having sul3.201829708802
225550.9998Diversity and metallo-β-lactamase-producing genes in Pseudomonas aeruginosa strains isolated from filters of household water treatment systems. The microbiological quality of drinking water has long been a critical element in public health. Considering the high clinical relevance of Pseudomonas aeruginosa, we examined the filters of household water treatment systems for its presence and characteristics to determine the systems' efficiency in eliminating the bacteria. In total, filters of 50 household water treatment systems were examined. Microbiological and molecular methods were used for the detection and confirmation of P. aeruginosa isolates. Random Amplification of Polymorphic DNA-polymerase chain reaction (RAPD-PCR) was performed to detect similarities and differences among P. aeruginosa isolates. Combined disk (CD) method and double disk synergy test (DDST) were performed to detect metallo-beta-lactamase (MBL)-producing P. aeruginosa isolates. Finally, PCR was performed to detect MBL genes in MBL-producing strains. From the 50 analyzed systems, 76 colonies of P. aeruginosa were identified. In some systems, isolated bacteria from different filters harbored similar genetic profiles, indicating that these isolates may be able to pass through the filter and reach higher filters of the system. Phenotypic tests revealed 7 (9.2%) MBL-producing strains. Two isolates were positive for bla(VIM-1), whereas one isolate was positive for bla(NDM) and bla(IMP-1). The wide distribution of resistant phenotypes and genetic plasticity of these bacteria in household water treatment systems indicate that resistance mechanisms circulate among P. aeruginosa isolates in the environment of the filtration systems. The presence of MBL-producing genes in these systems and P. aeruginosa as a potential reservoir of these resistance genes can be a major concern for public health.201930368151
159860.9998A method to detect Escherichia coli carrying the colistin-resistance genes mcr-1 and mcr-2 using a single real-time polymerase chain reaction and its application to chicken cecal and porcine fecal samples. Colistin is one of the last-resort antibiotics for the treatment of multidrug-resistant infections in humans, but transmissible colistin-resistance genes have emerged in bacteria from animals. The rapid and sensitive detection among animals of colonization with bacteria carrying these genes is critical in helping to control further spread. Here we describe a method for broth enrichment of colistin-resistant Escherichia coli from animal fecal and cecal samples followed by real-time polymerase chain reaction (PCR) for the simultaneous detection of two of the main colistin-resistance genes, mcr-1 and mcr-2. The PCR uses a single set of nondegenerative primers, and mcr variants can be differentiated by melt-curve analysis. Overnight culture enrichment was effective for amplifying colistin-resistant E. coli, even when initially present in numbers as low as 10 bacteria per gram of sample. The mcr-1 and mcr-2 genes were not found in any of the Ontario swine and poultry samples investigated.201830363381
204870.9998The Role of Plasmids in the Multiple Antibiotic Resistance Transfer in ESBLs-Producing Escherichia coli Isolated From Wastewater Treatment Plants. We compared the diversity of extended-spectrum β-lactamases (ESBLs) producing Escherichia coli (E. coli) in wastewater of a municipal wastewater treatment plant. This was done by analyzing multiple antibiotic resistant phenotypes and genotypes. Also, we investigated the antibiotic resistance transfer mechanism of the plasmid by comparing the antibiotic resistance gene linked transfer using a conjugative test, and by analyzing the full-length DNA sequence of one plasmid. The results showed that 50 ESBLs-producing E. coli isolates were isolated from 80 wastewater samples at the rate of 62.5% (50/80), out of which 35 transconjugants were obtained with the multiple antibiotic resistant transfer rate as high as 70.0% (35/50). Multiple antibiotic resistance was shown in all transconjugants and donor bacteria, which were capable of resistance to 11 out of 15 kinds of antibiotics. Both transconjugants and donors were capable of resistance to the Ampicillin and Cefalotin at a rate of 100.00% (35/35), while the total antibiotic resistant spectrum of transconjugants narrowed at the rate of 94.29% (33/35) and broadened at the rate of 5.71% (2/35) after conjugate to the donor bacteria. PCR showed that the resistant genotypes decreased or remained unchanged when compared to donor bacteria with transconjugants while the bla(TEM) and bla(CTX-M) genes were transferred and linked at a rate of 100.00% (35/35) and the bla(SHV) gene was at the rate as high as 94.29% (33/35). However, the qnrS gene was transferred at a low rate of 4.17% (1/24). In addition, the major resistance gene subtypes were bla(TEM-) (1), bla(SHV -11) , and bla(CTX-M-15) according to sequencing and Blast comparison. Plasmid wwA8 is a closed-loop DNA molecule with 83157 bp, and contains 45 predicted genes, including three antibiotic resistant resistance genes, bla(CTX-M-15) , bla(TEM-1) and qnrS1, which can be transferred with E. coli in vitro. This study shows that E. coli isolated from wastewater was capable of transferring resistance genes and producing antibiotic resistant phenotypes. The plasmids containing different resistance genes in E. coli play an important role in the multiple antibiotic resistant transfer. Most importantly, antibiotic resistant resistance genes have different transfer efficiencies, the bla(TEM) and bla(CTX-M) genes transferred at a rate of 100.00% and linked transfer in all 35 transconjugants.201931001218
194780.9998Search for carbapenem-resistant bacteria and carbapenem resistance genes along swine food chains in Central Italy. The presence of carbapenem-resistant bacteria and carbapenem resistance genes (CRGs) in livestock is increasing. To evaluate the presence of carbapenemase-producing Enterobacteriaceae (CPE) and the main CRGs along swine food chains of the Marche Region (Central Italy), samples of faeces, feed, and animal-food derived products were collected from seven small/medium, medium, and large-scale pig farms. A total of 191 samples were analysed using a culture-dependent method, with the aim of isolating CPE. Isolates were analysed for their resistance to carbapenems using a modified Hodge test and the microdilution method for the minimum inhibitory concentration (MIC) determination. Moreover, the extraction of microbial DNA from each sample was performed to directly detect selected CRGs via qPCR. Among the 164 presumptive resistant isolates, only one strain from a liver sample, identified as Aeromonas veronii, had an ertapenem MIC of 256 μg/mL and carried a carbapenemase- (cphA) and a β-lactamase- (blaOXA-12) encoding genes. A low incidence of CRGs was found; only nine and four faecal samples tested positive for blaNDM-1 and blaOXA-48, respectively. Overall, the importance of monitoring CPE and CRGs in livestock and their food chains should be stressed to control all potential non-human CPE and CRGs reservoirs and to determine safety levels for human health.202438181018
205390.9998Replicon typing of plasmids in environmental Achromobacter sp. producing quinolone-resistant determinants. This study aimed to investigate the antimicrobial resistance profile to quinolones, the presence of quinolone-resistant determinants and the plasmid replicon typing in environmental Achromobacter sp. isolated from Brazil. Soil and water samples were used for bacterial isolation. The antimicrobial susceptibility testing was performed by minimum inhibitory concentration method. The detection of mutations in the quinolone resistance-determining regions (QRDR) genes, the presence of plasmid-mediated quinolone resistance (PMQR) genes, and plasmid replicons were performed by PCR. A total of 16 isolates was obtained from different cultures, cities, and states of Brazil. All isolates were non-susceptible to ciprofloxacin, norfloxacin, and levofloxacin. Some mutations in QRDR genes were found, including Gln-83-Leu and Asp-87-Asn in the gyrA and Gln-80-Ile and Asp-84-Ala in the parC. Different PMQR genes were detected, such as qnrA, qnrB, qnrS, oqxA, and oqxB. Three different plasmid families were detected, being most presented the ColE-like, followed by IncFIB and IncA/C. The presence of different PMQR genes and plasmids in the isolates of the present study shows that environmental bacteria can act as reservoir of important genes of resistance to fluoroquinolones, which is of great concern, due to the potential of horizontal dissemination of these genes. Besides that, there are no studies reporting these results in Achromobacter sp. isolates.201830357960
1619100.9998Evidence of colistin resistance genes (mcr-1 and mcr-2) in wild birds and its public health implication in Egypt. BACKGROUND: Antimicrobial resistance has become one of the most severe global threats to human and veterinary Medicine. colistin is an effective therapeutic agent against multi-drug-resistant pathogens. However, the discovery of transferable plasmids that confer resistance to colistin (mcr-1) has led to challenges in medical science. This study describes the role of wild birds in the harbouring and environmental spread of colistin-resistant bacteria, which could pose a potential hazard to human and animal health. METHODS: In total, 140 faecal samples from wild birds (migratory and resident birds) were tested. Twenty surface water samples were collected from the area in which wild bird trapping was conducted, and 50 human stool samples were collected from individuals residing near the surface water sources and farm buildings. Isolation and identification of Enterobacteriaceae and Pseudomonas aeruginosa from the different samples were performed using conventional culture techniques and biochemical identification. PCR amplification of the mcr genes was performed in all positive isolates. Sequencing of mcr-1 genes from three randomly selected E. coli carrying mcr-1 isolates; wild birds, water and humans was performed. RESULT: The bacteriological examination of the samples showing isolates of Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca and P. aeruginosa. The results of multiplex PCR of the mcr genes revealed that E. coli was the most prevalent gram-negative bacterium harbouring the mcr genes, whereas a low prevalence was observed for K. pneumoniae. The prevalence of mcr-1 in resident birds, migratory birds, water sources and humans were 10.4, 20,16.6 and 9.6% while the prevalence of mcr-2 were 1.4, 3.6, 11.1 and 9.6%, respectively. Sequencing of the mcr-1 gene from the three E. coli carrying mcr-1 isolates indicated a possible correlation between the wild bird and surface water isolates. CONCLUSION: The detection of mcr-1-positive bacteria in wild birds in Egypt indicates the possible environmental dissemination of this gene through bird activity. The impact of the interaction between domestic and wild animals on public health cannot be overlooked.201931827778
2922110.9997Tetracycline-resistance genes in gram-negative isolates from estuarine waters. AIMS: To investigate the diversity and dissemination of tetracycline resistance genes in isolates from estuarine waters. METHODS AND RESULTS: Forty-two out of 164 multi-resistant isolates previously obtained were resistant or less-susceptible to tetracycline, as evaluated by the disc diffusion method. Minimal inhibitory concentration for resistant bacteria ranged from 16 to 256 mg l(-1). Screening of tet genes by polymerase chain reaction showed that 88% of the isolates carried at least one of the genes tested, namely tet(A) (present in 13 isolates), tet(B) (present in 13 isolates), tet(C) (present in 3 isolates), tet(D) (present in 1 isolate), tet(E) (present in 6 isolates) and tet(M) (present in 1 isolate). One isolate carried tet(A) and tet(M). To our knowledge, this study presents the first description of a tet(D) gene in Morganella morganii. Hybridization revealed that tet genes were plasmid-located in 31% of the isolates. Those isolates were included as donors in conjugation experiments and 38% transferred tetracycline resistance. CONCLUSIONS: A considerable diversity of tet genes was detected in the estuary. Frequently, these genes were associated with plasmids and could be transferred to Escherichia coli. SIGNIFICANCE AND IMPACT OF THE STUDY: The results presented provide further evidence of the role played by estuarine reservoirs in antibiotic resistance maintenance and dissemination.200819120920
2731120.9997Prevalence of tetracycline resistance genes among multi-drug resistant bacteria from selected water distribution systems in southwestern Nigeria. BACKGROUND: Antibiotic resistance genes [ARGs] in aquatic systems have drawn increasing attention they could be transferred horizontally to pathogenic bacteria. Water treatment plants (WTPs) are intended to provide quality and widely available water to the local populace they serve. However, WTPs in developing countries may not be dependable for clean water and they could serve as points of dissemination for antibiotic resistant bacteria. Only a few studies have investigated the occurrence of ARGs among these bacteria including tetracycline resistance genes in water distribution systems in Nigeria. METHODOLOGY: Multi-drug resistant (MDR) bacteria, including resistance to tetracycline, were isolated from treated and untreated water distribution systems in southwest Nigeria. MDR bacteria were resistant to >3 classes of antibiotics based on break-point assays. Isolates were characterized using partial 16S rDNA sequencing and PCR assays for six tetracycline-resistance genes. Plasmid conjugation was evaluated using E. coli strain DH5α as the recipient strain. RESULTS: Out of the 105 bacteria, 85 (81 %) and 20 (19 %) were Gram- negative or Gram- positive, respectively. Twenty-nine isolates carried at least one of the targeted tetracycline resistance genes including strains of Aeromonas, Alcaligenes, Bacillus, Klebsiella, Leucobacter, Morganella, Proteus and a sequence matching a previously uncultured bacteria. Tet(A) was the most prevalent (16/29) followed by tet(E) (4/29) and tet30 (2/29). Tet(O) was not detected in any of the isolates. Tet(A) was mostly found with Alcaligenes strains (9/10) and a combination of more than one resistance gene was observed only amongst Alcaligenes strains [tet(A) + tet30 (2/10), tet(A) + tet(E) (3/10), tet(E) + tet(M) (1/10), tet(E) + tet30 (1/10)]. Tet(A) was transferred by conjugation for five Alcaligenes and two E. coli isolates. CONCLUSIONS: This study found a high prevalence of plasmid-encoded tet(A) among Alcaligenes isolates, raising the possibility that this strain could shuttle resistance plasmids to pathogenic bacteria.201526108344
1032130.9997Molecular investigation of antibiotic resistant bacterial strains isolated from wastewater streams in Pakistan. Antibiotic resistance is a global public health issue and it is even more daunting in developing countries. The main objective of present study was to investigate molecular responses of antibiotic-resistant bacteria. The 48 bacterial strains, which were previously isolated and identified were subjected to disc diffusion and MIC (minimum inhibitory concentration) determination, followed by investigating the production of the three beta-lactamases (ESBLs (Extended-spectrum Beta-lactamases), MBLs (Metallo Beta-lactamases), AmpCs) and exploring prevalence of the two antibiotic-resistant genes (ARGs); blaTEM and qnrS. Higher MIC values were observed for penicillin(s) than that for fluoroquinolones (ampicillin > amoxicillin > ofloxacin > ciprofloxacin > levofloxacin). Resistance rates were high (58-89%) for all of the tested beta-lactams. Among the tested strains, 5 were ESBL producers (4 Aeromonas spp. and 1 Escherichia sp.), 2 were MBL producers (1 Stenotrophomonas sp. and 1 Citrobacter sp.) and 3 were AmpC producers (2 Pseudomonas spp. and 1 Morganella sp.). The ARGs qnrS2 and blaTEM were detected in Aeromonas spp. and Escherichia sp. The results highlighted the role of Aeromonas as a vector. The study reports bacteria of multidrug resistance nature in the wastewater environment of Pakistan, which harbor ARGs of clinical relevance and could present a public health concern.202032802720
2082140.9997Rapid screening technique for class 1 integrons in Enterobacteriaceae and nonfermenting gram-negative bacteria and its use in molecular epidemiology. A screening technique for integrons in members of the family Enterobacteriaceae and nonfermenting gram-negative bacteria by real-time PCR is reported. A total of 226 isolates of gram-negative bacteria obtained from a variety of clinical specimens were screened for class 1 integrons by real-time PCR performed on a LightCycler instrument. This technique used a primer pair specific for a 300-bp conserved region at the 5' ends of class 1 integrons. The screening assay was evaluated by comparison with results obtained by the conventional, thermal-block PCR (long PCR) by using established conditions and primers for the detection of class 1 integrons, and the real-time PCR technique was thus shown to be both sensitive and specific. DNA from 50 of 226 (22%) isolates screened was identified as containing an integron by the screening PCR, and sequence data were obtained across the integron for 34 of 50 (68%) of these isolates. In an attempt to study the molecular epidemiology of antimicrobial resistance genes carried within integrons, a comparison of the types of gene cassettes carried by isolates from different patients was made. Adenyltransferase genes conferring resistance to streptomycin and spectinomycin were the predominant gene cassettes amplified in the study. Resistance to trimethoprim was also frequently found to be encoded within integrons. Furthermore, multiple bacterial isolates obtained from one patient over a 5-month period were all shown to carry an integron containing the same single adenyltransferase gene cassette, suggesting that these elements were relatively stable in this case.200111257011
2730150.9997Multidrug Resistance in Quinolone-Resistant Gram-Negative Bacteria Isolated from Hospital Effluent and the Municipal Wastewater Treatment Plant. This study is aimed to assess if hospital effluents represent an important supplier of multidrug-resistant (MDR) Gram-negative bacteria that, being discharged in the municipal collector, may be disseminated in the environment and bypassed in water quality control systems. From a set of 101 non-Escherichia coli Gram-negative bacteria with reduced susceptibility to quinolones, was selected a group of isolates comprised by those with the highest indices of MDR (defined as nonsusceptibility to at least one agent in six or more antimicrobial categories, MDR ≥6) or resistance to meropenem or ceftazidime (n = 25). The isolates were identified and characterized for antibiotic resistance phenotype, plasmid-mediated quinolone resistance (PMQR) genes, and other genetic elements and conjugative capacity. The isolates with highest MDR indices were mainly from hospital effluent and comprised ubiquitous bacterial groups of the class Gammaproteobacteria, of the genera Aeromonas, Acinetobacter, Citrobacter, Enterobacter, Klebsiella, and Pseudomonas, and of the class Flavobacteriia, of the genera Chryseobacterium and Myroides. In this group of 25 strains, 19 identified as Gammaproteobacteria harbored at least one PMQR gene (aac(6')-Ib-cr, qnrB, qnrS, or oqxAB) or a class 1 integron gene cassette encoding aminoglycoside, sulfonamide, or carbapenem resistance. Most of the E. coli J53 transconjugants with acquired antibiotic resistance resulted from conjugation with Enterobacteriaceae. These transconjugants demonstrated acquired resistance to a maximum of five classes of antibiotics, one or more PMQR genes and/or a class 1 integron gene cassette. This study shows that ubiquitous bacteria, other than those monitored in water quality controls, are important vectors of antibiotic resistance and can be disseminated from hospital effluent to aquatic environments. This information is relevant to support management options aiming at the control of this public health problem.201626469134
5088160.9997A Multiplex SYBR Green Real-Time PCR Assay for the Detection of Three Colistin Resistance Genes from Cultured Bacteria, Feces, and Environment Samples. The aim of the study was to develop a multiplex assay for rapid detection of mcr-1, mcr-2, and mcr-3, a group of genes of conferring resistance to colistin mediated by plasmid in Enterobacteriaceae. A SYBR Green based real-time PCR assay has been designed to detect the mcr genes, and applied to cultured bacteria, feces and soil samples. All three mcr genes could be detected with a lower limit of 10(2) cultured bacteria. This test was highly specific and sensitive, and generated no false-positive results. The assay was also conclusive when applied to feces and soil samples containing mcr-1-positive Escherichia coli, which could facilitate the screening of mcr genes not only in the bacteria, but also directly from the environment. This simple, rapid, sensitive, and specific multiplex assay will be useful for rapid screening of the colistin resistance in both clinical medicine and animal husbandry.201729163387
1692170.9997Phenotypic and genotypic detection of antibiotic-resistant bacteria in fresh fruit juices from a public hospital in Rio de Janeiro. Gram-negative bacteria are worrisome because they are becoming resistant to many antibiotic available options, mainly in hospital environment. Several studies have noted the presence of bacteria producing extended-spectrum beta-lactamase, with the presence of antibiotic-resistance genes in fresh vegetables and fruits. This study aimed to detect the presence of phenotypic and genotypic resistance in eight samples of fresh fruit juices served to patients admitted to a hospital in Rio de Janeiro. The growth of microorganisms on MacConkey and XLD agar was carried out to obtain a "pool" of Gram-negative bacteria. The disk diffusion test and the polymerase chain reaction were performed to detect the phenotypic and genotypic resistance of Gram-negative bacteria to the tested antibiotics. The multidrug resistance was detected in all samples and the shv, tem, ctx, tetA, tetB and oxa- 48 genes were found in the samples, including the presence of class 2 and 3 integrons. We can conclude that the selection methodology allows the detection of a greater number of genes and this found warns about the risk of making these foods available to patients in hospitals.202133398401
1616180.9997Characterization of Extended-Spectrum β-Lactamase-Producing Escherichia coli Isolated from Fresh Produce and Agricultural Environments in Korea. ABSTRACT: This study was conducted to characterize Escherichia coli strains and evaluate the spread of antimicrobial resistance among these strains from fresh produce and farm environments in Korea. We then conducted phenotypic and genetic studies on antimicrobial-resistant isolates. We determined the genetic epidemiological characteristics of isolates that produced extended-spectrum β-lactamase (ESBL) and confirmed plasmid transfer in isolates that carried blaCTX-M-type genes. E. coli strains were isolated from 8 samples of fresh produce and 152 samples from the farm environment collected from May 2014 to June 2016. Cephalosporin resistance was the most prevalent (61.8%) type of resistance among the isolates. Five ESBL-producing strains with high genetic homology with E. coli of human or livestock origin were identified. Lateral transfer of plasmids harboring blaCTX-M-type genes to transconjugants was successful. Two isolates from Chinese cabbage and from water samples collected from a nearby stream harbored the ISEcp1-blaCTX-M-55-orf477 operon and were confirmed as sequence type 1196 and the same type of plasmid replicon, suggesting that cross-contamination was highly likely. A high-risk clone of sequence type 69 (clonal complex 69) isolates was also recovered from the farm environment. This study provides genetic evidence that antimicrobial resistance factors in E. coli from farm environments originate in the clinic or in livestock, highlighting the fact that good agricultural practices in farming are important to inhibit the spread of antimicrobial resistance to bacteria on fresh produce.202032083678
2690190.9997Characterization of Cefotaxime- and Ciprofloxacin-Resistant Commensal Escherichia coli Originating from Belgian Farm Animals Indicates High Antibiotic Resistance Transfer Rates. Food-producing animals represent one of the sources of antibiotic resistant commensal bacteria. There is an increasing awareness that these bacteria might have the potential to transfer their resistance genes to other (pathogenic) bacteria. In this study, 50 commensal Escherichia coli strains originating from food-producing animals and resistant to the "highest priority, critically important antibiotics" cefotaxime and/or ciprofloxacin, were selected for further characterization. For each strain (i) an antibiogram, (ii) the phylogenetic group, (iii) plasmid replicon type, (iv) presence and identification of integrons, and (v) antibiotic resistance transfer ratios were determined. Forty-five of these strains were resistant to 5 or more antibiotics, and 6 strains were resistant to 10 or more antibiotics. Resistance was most common to ampicillin (100%), sulfamethoxazole, ciprofloxacin (82%), trimethoprim, tetracycline (74%), cefotaxime, (70%) and ceftazidime (62%). Phylogenetic groups A (62%) and B1 (26%) were most common, followed by C (8%) and E (4%). In 43 strains, more than 1 replicon type was detected, with FII (88%), FIB (70%), and I1 (48%) being the most encountered types. Forty strains, positive for integrons, all harbored a class I integron and seven of them contained an additional class II integron. No class III integrons were detected. The antibiotic resistance transfer was assessed by liquid mating experiments. The transfer ratio, expressed as the number of transconjugants per recipient, was between 10(-5) and 10(0) for cefotaxime resistance and between 10(-7) and 10(-1) for ciprofloxacin resistance. The results of the current study prove that commensal E. coli in food-production animals can be a source of multiple resistance genes and that these bacteria can easily spread their ciprofloxacin and cefotaxime resistance.201829148895