Bla CTX-M Research Articles

Origin, maintenance and spread of antibiotic resistance genes within plasmids and chromosomes of bloodstream isolates of Escherichia coli.

Blood stream invasion by Escherichia coli is the commonest cause of bacteremia in the UK and elsewhere with an attributable mortality of about 15–20 %; antibiotic resistance to multiple agents is common in this microbe and is associated with worse outcomes. Genes conferring antimicrobial resistance, and their frequent location on horizontally transferred genetic elements is well-recognised, but the origin of these determinants, and their ability to be maintained and spread within clinically-relevant bacterial populations is unclear. Here, we set out to examine the distribution of antimicrobial resistance genes in chromosomes and plasmids of 16 bloodstream isolates of E. coli from patients within Scotland, and how these genes are maintained and spread. Using a combination of short and long-read whole genome sequencing methods, we were able to assemble complete sequences of 44 plasmids, with 16 Inc group F and 20 col plasmids; antibiotic resistance genes located almost exclusively within the F group. bla CTX-M15 genes had re-arranged in some strains into the chromosome alone (five strains), while others contained plasmid copies alone (two strains). Integrons containing multiple antibiotic genes were widespread in plasmids, notably many with a dfrA7 gene encoding resistance to trimethoprim, thus linking trimethoprim resistance to the other antibiotic resistance genes within the plasmids. This will allow even narrow spectrum antibiotics such as trimethoprim to act as a selective agent for plasmids containing antibiotic resistance genes mediating much broader resistance, including blaCTX-M15. To our knowledge, this is the first analysis to provide complete sequence data of chromosomes and plasmids in a collection of pathogenic human bloodstream isolates of E. coli . Our findings reveal the interplay between plasmids and integrative and conjugative elements in the maintenance and spread of antibiotic resistance genes within pathogenic E. coli .

Untreated HWWs Emerged as Hotpots for ARGs.

Hospital wastewaters (HWWs) are reported to be hotspots for antibiotics and antibiotic-resistant bacteria. However, limited information involves the impact of these effluents on dissemination of antibiotic-resistance genes (ARGs). In this study, therefore, seasonally collected HWWs were monitored for overall bacterial load and seven ARGs aadA, tetA, cmlA, sul1, qnrS, ermBandblaCTX-Mby using quantitative polymerase chain reaction method. Overall bacterial 16S rRNA copy number was found to be the lowest in winter with 103copy number/mL, while the highest copy number, with 105copy number/mL, was observed in both summer and spring. All hospitals tested displayed similar seasonal ARG copy number profile ofaadA > tetA > cmlA≈sul1 > ermB≈qnrS > blaCTX-M. The results indicated that untreated HWWs were hotspots for ARGs and required attention before discharging into public sewer.

Multidrug-Resistance Genes in Pseudomonas aeruginosa from Wound Infections in a Tertiary Health Institution in Osogbo, Nigeria.

Multidrug Resistant Pseudomonas aeruginosa (MDRPA) is a ubiquitous opportunistic organism that poses threat to the management of infections globally. The objectives of the current research were to assess the antibiotic resistance profiles as well as Multiple Antibiotic Resistance (MAR) Index of clinical isolates of P. aeruginosa associated with wound infections. Presence of Extended Spectrum Beta Lactamase genes (bla CTX-M, bla SHV and bla TEM) and Carbapenemase genes (bla KPC and blaNDM) were also determined among the isolates. Swab samples were collected from 255 patients with wound infections. Bacterial identification was done by standard diagnostic tests. The identity of isolates was confirmed by the detection of the exoA gene using the PCR technique. Antibiotic susceptibility testing and resistance profile were determined using the disc diffusion method. Resistance genes were amplified by the PCR method. A total of 235 (92.2%) bacterial isolates were recovered from the wounds of the 255 patients, of these, 124 (52.8%) were Gram-negative bacilli while the remaining 111 (47.2%) were Gram-positive cocci. A total of 69 Pseudomonas aeruginosa strains were recovered from the wound specimens. Imipenem was the most effective antibiotic against these isolates (92.8% isolates were susceptible) while all isolates were resistant to Meropenem, Cefepime, Ticarcillin, Amoxicillin-clavulanic acid, Cefotaxime, Ampicillin and Cefpodoxime. All 69 Pseudomonas aeruginosa isolates were multidrug resistant (MDR). Of the isolates selected for PCR, all were positive for TEM, CTX-M and SHV genes while one-third were blaKPC and blaNDM producers. This study demonstrated high prevalence of carbapenem-resistant strains of P. aeruginosa, suggesting that there is an urgent need in Nigeria for the enactment and enforcement of policies and necessary laws restricting the availability and indiscriminate use of antibiotics.

A High Resolution DNA Melting Curve Analysis for the Rapid and Efficient Molecular Diagnostics of Extended Spectrum β-Lactamase Determinants from Foodborne Escherichia coli.

The accurate identification of Extended-Spectrum β-Lactamase (ESBL) genes in Gram-negative bacteria is necessary for surveillance and epidemiological studies of transmission through foods. We report a novel rapid, cheap, and accurate closed tube molecular diagnostic tool based on two multiplex HRM protocols for analysis of the predominant ESBL families encountered in foods. The first multiplex PCR assay targeted blaCTX-M including phylogenetic groups 1 (CTX-M-1-15, including CTX-M-1, CTX-M-3 and CTX-M-15), 2 (CTX-M-2), and 9 (CTX-M-9-14, including CTX-M-9 and CTX-M-14). The second assay involved blaTEM /bla CTX-M /blaSHV, including TEM variants (TEM-1 and TEM-2), SHV-1-56 (SHV-1, SHV-2 and SHV-56), and CTX-M-8-41 (CTX-M-8, CTX-M-25, CTX-M-26 and CTX-M-39 to CTX-M-41). The individual melting curves were differentiated by a temperature shift according to the type of ESBL gene. The specificity and sensitivity of the first assay were 100% and 98%, respectively. For the second assay, the specificity and sensitivity were 87% and 89%, respectively. The detection of ESBL variants or mutations in existing genes was also demonstrated by the subtyping of a variant of the CTXM-1-15. The HRM is a potential tool for the rapid detection of present β-lactamase genes and their characterization in a highly sensitive, closed-tube, inexpensive method that is applicable in high throughput studies.

Characteristics of virulence, resistance and genetic diversity of strains of Salmonella Infantis isolated from broiler chicken in Brazil

ABSTRACT: Salmonella Infantis is frequently associated with human infections worldwide and is transmitted by consumption of contaminated foods, particularly those of animal origin, especially the chicken meat. We aimed to evaluate virulence characteristics, antimicrobial resistance and the genetic similarity of 51 strains of S. Infantis isolated from samples of poultry origin. The strains were isolated from 2009 to 2010 in a company with full cycle of broiler’s production in the state of São Paulo, Brazil. The antimicrobial susceptibility test was performed and, by PCR, we evaluated the presence of the genes lpfA (hem-adhesion), agfA (hem-biofilm) and sefA (hem-adhesion) and resistance genes to beta-lactams (blaTEM, blaSHV, bla CTX-M and blaAmpC ). The phylogenetic relationship was determined by RAPD-PCR method. Among the drugs tested, the highest percentages of resistance were to amoxicillin (35.3%) and to sulfonamide (15.7%). Eleven antimicrobial resistance patterns were identified (A1 to A11), none of them presented a multiresistance profile (> 3 antimicrobials classes). There was 100% of positivity for the agfA gene, 92.2% for the lpfA gene, and no strain presented the sefA gene. Most of the isolates showed similarities in virulence potential, since they were simultaneously positive for two studied genes, agfA and lpfA (92.2%, 47/51). Of the 18 (35.3%) strains resistant to antimicrobials of the β-lactam class, 10 (55.5%) were positive to blaAmpC gene, five (27.8%) for blaCTX-M , two (11.1%) to blaSHV and no strain presented the blaTEM gene. The phylogenetic evaluation has shown the presence of five clusters (A, B, C, D and E) with similarity greater than 80%, and three distinct strains which were not grouped in any cluster. Cluster B grouped 33 strains, all positive for lpfA and agfA genes, from both, the broiler farming facility and the slaughterhouse, persistent throughout all the study period. This cluster also grouped 18 strains clones with genetic similarity greater than 99%, all isolated in the slaughterhouse. The presence of virulence genes associated with persistent strains clones for a long period, warns to the possibility of S. Infantis to form biofilm, and should be constantly monitored in broilers’ production chain, in order to know the profile of the strains that may contaminate the final product and evaluate the hazards that represents to public health.

English

This present study was undertaken for detection of extended spectrum β-lactamases (ESBLS) enzyme genes among clinical isolates of Pseudomonas aeruginosa using phenotypic and molecular techniques. Thirty-four P. aeruginosa isolates from different hospitals in Nsukka and University of Nigeria Teaching Hospital (UNTH), Enugu were screened for the presence of ESBL-encoding genes. Phenotypic screening for ESBL producers was carried out using double disk synergy test and combined disk test. Genomic DNA was extracted from the isolates by modified boiling method. Extracted DNA was amplified by polymerase chain reaction (PCR) using ESBL specific primers namely Bla GES, PER, OXA-50, SHV, CTX-M and TEM. The results revealed that a total of 15 isolates of P. aeruginosa were identified as ESBL producer by phenotypic approaches which exhibited varying degrees of resistance to an array of antibiotics tested. While, the PCR screening revealed that 53.33% (n=8) of the isolates that were phenotypically ESBL positive harboured bla OXA-50 gene. However, the genes that encode PER, GES, SHV, TEM and CTX-M were not found in any of the P. aeruginosa isolates. This study highlights the need to establish antimicrobial resistance surveillance network to determine the appropriate empirical treatment regimen for Pseudomonas infections.   Key words: Pseudomonas aeruginosa, antibiotic resistance extended spectrum β-lactamase (ESBL), polymerase chain reaction (PCR), Nsukka.

Bacteriological and Molecular Study Characterization of Klebsiella pneumoniae Isolated from Different Clinical Specimens

Klebsiella pneumoniae is one of the foremost imperative opportunistic pathogens. Urinary characteristic disease is the common infectious bacterial contamination caused by K. pneumonia, that are rising around the world comprising a danger to community and clinic settings. K. pneumonia Isolates were evaluated for their antimicrobial susceptibility. Samples were taken from 80 patients with diverse diseases infection. Genomic DNA of K. pneumonia Confines were extricated and detection of ESBL Genes was 53.75% of the isolates were predominance for ESBL Genes blaTEM, blaSHV and bla CTX-M 82.5, 92.5 and 70 %, respectively. Out of 100 obtained clinical isolates of K. pneumonia from diverse healing centers and therapeutic research facilities in Duhok/ Iraq as it were (80%) isolates had a place to the genus K. pneumonia. Ampicillin and Aztreonam 100 % anti-microbial resistance where was Imipenem Ertapenem Meropenem 100% sensitive. Conveyance of ESBLs creating K. pneumoniae among different clinical tests because it was 71.42% in urine, 40. 90 % in wound swabs, 42. 10 % in sputum and 50 % in blood culture. The recurrence of the ESBL production can easily be thought little of within the clinical isolates of K. pneumoniae with the utilize of the current CLSI suggested strategies, an ideal recognizable proof of the ESBL creating isolates is basic to define approaches for an experimental antimicrobial treatment

Clonal dissemination of KPC-2-producing Klebsiella pneumoniae ST11 and ST48 clone among multiple departments in a tertiary teaching hospital in Jiangsu Province, China.

The world-wide prevalence of carbapenem-resistant Klebsiella pneumoniae (CRKP) poses a threat to the public health. The objective of this study was to determine the epidemiological and molecular patterns of KPC-producing Klebsiella pneumoniae (K. pneumoniae) clinical isolates. In this study, a total of 82 non-duplicated CRKP isolates were analyzed for the prevalence of resistant determinants including carbapenemase, extended spectrum β-lactamase (ESBLs), and AmpC as well as integrons and cassette regions by polymerase chain reaction (PCR) and DNA sequencing. The genetic relatedness was investigated by pulsed field gel electrophoresis (PFGE) and multi-locus sequencing typing (MLST). Overall, bla KPC-2 (n=75) was the predominant carbapenemase gene, followed by high prevalence of bla SHV (92.7%) and bla CTX-M (90.2%). PFGE and MLST analysis revealed that 65 out of 68 KPC-2-producing CRKP belonged to the ST11 clone and were distributed mainly in the department of neurology ICU. Moreover, first report on clonal dissemination of KPC-2-producing CRKP ST48 clone and NDM-5-producing CRKP ST337 clone was also identified. Class I integron were detected in 17 (20.7%) of 82 isolates with aadA2 being the most common cassette. And a novel cassette array of integron, aac(6')-II-bla CARB/PSE-1 was identified. All in all, KPC-2-producing CRKP ST11 and ST48 clone were widely disseminated in multiple departments of our hospital, which triggers the need for active surveillance and implementation of infection control measures.

PREVALENCE OF CTX-M-PRODUCING GRAM-NEGATIVE UROPATHOGENS IN SOKOTO, NORTH-WESTERN NIGERIA

Objective: Infections of the urinary tract remains one of the most common bacterial infections with many implicated organisms being Gram-negative, which are increasingly resistant to antimicrobial agents. The aim of the study was to evaluate the resistance of ESBL producing Gram-negative enterobacteriaceae to commonly prescribed antibiotics and the prevalence of CTX-M genes from these isolates using polymerase chain reaction (PCR).
 Methods: The isolates were collected from urine over a period of 4 mo and studied, and were identified using Microgen Identification Kit (GN-ID). Susceptibility testing was performed by the modified Kirby Bauer disc diffusion method, and results were interpreted according to Clinical and Laboratory Standard Institute (CLSI). Extended-Spectrum Beta-Lactamase (ESBL) production was detected by the double-disc synergy test (DDST). Molecular characterization was based on the isolates that were positive for the phenotypic detection of ESBL.
 Results: Sixty one (61) isolates of Gram-negative uropathogens were identified. Of these, 19 (31.2%) were E. coli, 15 (24.6%) were Salmonella arizonae, Klebsiella pneumoniae were 7 (11.5%), Klebsiella oxytoca were 3 (4.9%), Enterobacter gergoviae were 6 (9.8%), 4 (6.6%) were Citrobacter freundii, 4 (6.6%) were Serratia marscence, and 1 (1.6%) were Enterobacter aerogenes, Proteus mirabilis, and Edwardsiella tarda each. Analysis of the bacterial susceptibility to antibiotics revealed most of them to be generally resistant to cotrimoxazole (73.3%), nalidixic acid (66.7%), norfloxacin (53.5%), ciprofloxacin (50.5%), gentamicin (48.6%), amoxicillin/clavulanate (45%), and the least resistant was displayed in nitrofurantoin (30%). Of the 15 ESBL producers, 11 (73.3%) were harbouring bla CTX-M genes.
 Conclusion: The study revealed a high susceptibility to nitrofurantoin, whereas susceptibility to cotrimoxazole was lowest. It further portrays a high prevalence of enterobacteriaceae isolates harbouring bla CTX-M genes in Sokoto metropolis.

شناسایی و توالی‌یابی ژن‌های blaCTX-M در ایزوله های بالینی کلبسیلا پنومونیه جدا شده از بیمارستان میلاد

شناسایی و توالی‌یابی ژن‌های blaCTX-M در ایزوله های بالینی کلبسیلا پنومونیه جدا شده از بیمارستان میلاد

Sequence-based epidemiology of an OXA-48 plasmid during a hospital outbreak.

Objectives. A large OXA-48 outbreak in the Netherlands involved the spread of OXA-48producing Enterobacteriaceae among at least 118 patients, suggesting horizontal transfer of this resistance gene through one or more plasmids. Elucidating transmission dynamics of resistance plasmids is hampered by the low resolution of classic typing methods. This study aimed to investigate the molecular epidemiology of plasmids carrying OXA-48 carbapenemase using a next-generation sequencing approach.Methods. A total of 68 OXA-48-producing Enterobacteriaceae isolated from the hospital outbreak, as well as 22 non-outbreak related OXA-48-producing Enterobacteriaceae from the Netherlands, Libya and Turkey were selected. Plasmids were sequenced using the Illumina Miseq platform, and read sets were assembled and analysed.Results. In all plasmids bla OXA-48 was embedded in transposon Tn1999.2 and located on a ca. 62 kb IncL/M conjugative plasmid in 14 different species. There were a maximum of 2 SNPs (single nucleotide polymorphisms) between the core sequence alignment of all plasmids. Closely related sequence variants of this plasmid were detected in non-outbreak isolates from the Netherlands and other countries. Thirty-one of 89 OXA-48-producing isolates also harboured bla CTX-M-15, which was not located on the bla OXA-48-carrying plasmid. Sequencing of four plasmids harbouring bla CTX-M15 revealed extensive plasmid heterogeneity.Conclusions. A ca 62 kb plasmid was responsible for the OXA-48 outbreak in a Dutch hospital. Our findings provide strong evidence for both within-host inter-species and between host dissemination of plasmid-based OXA-48 during a nosocomial outbreak. These findings exemplify the complex epidemiology of carbapenemase producing Enterobacteriaceae (CPE).

Determination of the frequency of β-lactamase genes (bla SHV, bla TEM, bla CTX-M) and phylogenetic groups among ESBL-producing uropathogenic Escherichia coli isolated from outpatients

Abstract Background Escherichia coli accounts for 70–95% of community-acquired urinary tract infections (UTIs). Recently, there has been an increase in the prevalence of extended-spectrum β-lactamase (ESBL) in the community which required an accurate identification for better management. Therefore, the current study was performed to determine the antimicrobial resistance pattern, investigate ESBL phenotypes and genotypes (blaCTX-M, bla TEM and bla SHV genes) and determine the phylogenetic groups among ESBL-positive isolates from outpatients. Methods One hundred and eighty-three positive urine samples were collected from 4450 outpatient clinic attendees. Antibiotic susceptibility was determined and ESBL phenotype screening was carried out using disk diffusion agar and combination disk techniques, respectively. The assessment of the presence of the blaCTX-M, bla TEM and blaSHV genes and phylogenetic grouping were performed using the polymerase chain reaction (PCR) method. Results Out of 183 E. coli isolates, 59 (32.2%) showed a positive ESBL phenotype. The prevalence of ESBL-producing E. coli was higher in males (57.4%). Fifty-seven of the ESBL-producing strains carried at least one of the β-lactamase genes (bla CTX-M, bla TEM, bla SHV). Phylotyping of multi-drug resistant isolates indicated that the isolates belonged to B2, A and D phylogroups. Analysis of resistance patterns among these phylogroups revealed that 74.4%, 55.3% and 29.7% of the isolates in the B2 group were resistant to trimethoprim-sulfamethoxazole, ciprofloxacin and gentamicin, respectively. Most of the strains in the phylogroup B2 carried the bla CTX-M gene. Conclusions All the ESBL-producing isolates were placed in one of the four phylogenetic groups. The presence of CTX-M and resistance to quinolones were more frequent in B2 strains than in non-B2 strains.

Geographical Distribution of β-Lactam Resistance among Klebsiella spp. from Selected Health Facilities in Ghana.

β-Lactam-resistant Klebsiella isolates continue to cause multidrug resistance infections worldwide. This study aimed to describe the geographical distribution of extended spectrum β-lactamase (ESBL), AmpC β-lactamase (AmpC), and carbapenemase production among 139 Klebsiella isolates recovered from patients at major referral health facilities in Ghana. The phenotypic methods of combined disc diffusion test, modified three-dimensional test, modified Hodge test (MHT), and combined disc test were performed for each isolate to detect ESBL, AmpC, carbapenemase, and metallo-β-lactamase (MBL) producers, respectively. Except for MBL, all other β-lactam resistance mechanisms were highest in the healthcare facilities situated in the northern belt of Ghana. Significant regional difference of ESBL producers was observed between the northern and middle belts as well as the northern and southern belts. Genotypic detection with polymerase chain reaction (PCR) revealed the presence of bla TEM 36/139 (25.9%), bla SHV 40/139 (28.8%), bla CTX-M 37/139 (26.6%), bla OXA-48 3/139 (2.16%), and bla NDM 1/139 (0.72%) genotypes. In conclusion, there were variations in β-lactam resistance among Klebsiella spp. from health facilities situated in the northern, middle, and southern belts of Ghana. The study provides preliminary evidence that emphasizes the need to direct more attention to antimicrobial resistance control, especially in the northern belt of Ghana. Findings from this study may be critical for creating and fine-tuning effective antimicrobial resistance control strategies and for informing accurate antibiotic prescription by practitioners.

Urban brown rats (Rattus norvegicus) as possible source of multidrug-resistant Enterobacteriaceae and meticillin-resistant Staphylococcus spp., Vienna, Austria, 2016 and 2017.

BackgroundBrown rats (Rattus norvegicus) are an important wildlife species in cities, where they live in close proximity to humans. However, few studies have investigated their role as reservoir of antimicrobial-resistant bacteria.AimWe intended to determine whether urban rats at two highly frequented sites in Vienna, Austria, carry extended-spectrum β-lactamase-producing Enterobacteriaceae, fluoroquinolone-resistant Enterobacteriaceae and meticillin-resistant (MR) Staphylococcus spp. (MRS).MethodsWe surveyed the presence of antimicrobial resistance in 62 urban brown rats captured in 2016 and 2017 in Vienna, Austria. Intestinal and nasopharyngeal samples were cultured on selective media. We characterised the isolates and their antimicrobial properties using microbiological and genetic methods including disk diffusion, microarray analysis, sequencing, and detection and characterisation of plasmids.ResultsEight multidrug-resistant Escherichia coli and two extensively drug-resistant New Delhi metallo-β-lactamases-1 (NDM-1)-producing Enterobacter xiangfangensis ST114 (En. cloacae complex) were isolated from nine of 62 rats. Nine Enterobacteriaceae isolates harboured the bla CTX-M gene and one carried a plasmid-encoded ampC gene (bla CMY-2). Forty-four MRS were isolated from 37 rats; they belonged to seven different staphylococcal species: S. fleurettii, S. sciuri, S. aureus, S. pseudintermedius, S. epidermidis, S. haemolyticus (all mecA-positive) and mecC-positive S. xylosus.ConclusionOur findings suggest that brown rats in cities are a potential source of multidrug-resistant bacteria, including carbapenem-resistant En. xiangfangensis ST114. Considering the increasing worldwide urbanisation, rodent control remains an important priority for health in modern cities.

Detection and genetic characterization of extended-spectrum beta-lactamases producers in a tertiary care hospital.

BACKGROUND:Extended-spectrum beta-lactamase (ESBL)-producing organisms inactivate extended beta-lactam antibiotics and monobactams and also exhibit coresistance to many other classes of antibiotics. The present study was carried out to assess the prevalence of the ESBLs and to determine the most prevalent genotype in our hospital.MATERIALS AND METHODS:All clinically significant Gram-negative isolates were identified, and their antimicrobial susceptibility testing was done by Kirby–Bauers' disc diffusion method. ESBL detection was confirmed by minimal inhibitory concentration method using agar dilution technique for those who screened positive by ceftazidime (30 μg) disc. Further, the established ESBL-positive isolates were subjected to genotyping for bla TEM, bla CTX-M, and bla SHV genes by using conventional polymerase chain reaction.RESULTS:Escherichia coli was the most common (28.84%) Gram-negative bacillus followed by Klebsiella pneumoniae (18.07%), while Pseudomonas spp. (9.61%) was the most commonly identified nonfermenter. ESBL production was detected in 160 (30.8%) isolates. Klebsiella oxytoca (46.7%) followed by E. coli (44%) were the common ESBL producers. Most predominant ESBL gene was bla TEM, found in 122 (76.25%) isolates. Combinations of two genes were seen in 109 (68.1%) isolates, the most common (43.12%) combination being blaTEM and blaCTX-M. In this study, 16 (10%) strains had all the three types of genes. Most of the isolated Gram-negative bacilli (GNB) were sensitive to amikacin, imipenem, and colistin.CONCLUSION:In our study, the 30.8% of GNB were ESBL producers. This is the only study that shows that TEM is the most prevalent ESBL genotypes in our area. Of concern is a good number of isolates showing all three patterns of genes (TEM, SHV, and CTX-M). Amikacin, imipenem, and colistin were the most useful antibiotics in our setup.

Occurrence of antibiotic resistance among Gram negative bacteria isolated from effluents of fish processing plants in and around Mangalore

ABSTRACT The presence of antibiotic-resistant bacteria in seafood not only poses a serious health risk for the consumers but also contributes to the spread of these antibiotic-resistant bacteria in the natural environments through the effluents discharged from the fish processing plants. The aims of this study were to isolate Gram-negative bacteria from the effluents of fish processing plants in and around Mangalore, India and to profile their antibiotic resistance pattern. Maximum resistance was seen for ampicillin (40.78%) followed by tetracycline (40.22%) and nitrofurantoin (29.05%). Further, the detection of genes that contribute to antibiotic resistance revealed the presence of sulfonamide resistance genes (sul1 and sul2) and extended spectrum β-lactamase genes (bla CTX-M, bla TEM) in a few isolates. The presence of such bacteria in fish processing effluents is a matter of great concern because they can contribute significantly to the antibiotic resistance in the natural environment. It is imperative that seafood processing plants follow the safe disposal of effluents in order to reduce or eliminate the antibiotic resistance menace.

CTX-M Β-LACTAMASE–PRODUCING ESCHERICHIA COLI IN SUDAN TERTIARY HOSPITALS: DETECTION GENOTYPES VARIANTS AND BIOINFORMATICS ANALYSIS

Background: Extended spectrum β-lactamase (ESBL)-producing Escherichia coli (ESBL-EC) constitute an emerging public-health concern. Consider that ESBL genes type CTX-M types has been increased significantly in most parts of the world. Few data are available on the CTX-M variants circulating in Sudan. Objective: This study used polymerase chain reaction (PCR) and bioinformatics tools to identify blaCTX-M and its variants for Extended-spectrum β-lactamase (ESBLs) producing clinical isolates of Escherichia coli (E. coli) obtained from hospitals in Khartoum-Sudan. Methods: A total of 216 non-repetitive isolates were selected during 2007-2018. The phenotypic identification of ESBL production was confirmed according to CLSI guidelines. CTX-M genotype was analyzed by uniplex PCRs reactions subsequently sequences performed later sequences were analyzed using bioinformatics tools with nBLAST program, multiple alignments to determine CTX-M genotype variants . Nucleotide sequences were submitted to Gen Bank and accession numbers were obtained. Result: ESBL phenotype among 212 confirmed E.coli isolates was (34.9% of 212,n=74). (62.2% of 74, n=46) strains carried bla CTX-M genes, CTX-M genotype variants identified in this study as followed: blaCTX-M-15 gene was the most prevalent one (78.6%) followed by blaCTX 90 (14.3%) and CTX-M55 (7.1%),
 Conclusion: This study reveled high ESBL occurrence among E.coli isolates, with CTX-M 15 the predominant variants and highlights the incidence of CTX-M-55 for the first time from Sudan.
 Keywords: E. coli, PCR, ESBLs, CTX-M, Sudan, bioinformatics

Prevalence of extended spectrum beta-lactamase producing Enterobacteriaceae in urine samples from Thumbay hospitals, U.A.E

Beta Lactamases is proven to be one of the leading cause of resistance to β-lactam antibiotics among gram-negative bacteria. Many up to date researches have shown increase in the incidence and prevalence of ESBL worldwide. This study aimed to determine the prevalence of ESBL strains of Klebsiella spp. and Escherichia coli species in urinary isolates from the patients admitted in Thumbay hospitals around United Arab Emirates. Furthermore, drug resistant genes (SHV and CTX-M) in the ESBL positive samples were detected. 237 urine samples were collected from November 2017 to January 2018. Based on the lactose utilization, colony morphology, and biochemical utilization of the gram negative bacilli were identified as E. coli (53), Klebsiella pneumoniae (10) and Citrobacter species (2). Antibiotic sensitivity test, double disc diffusion test and combination disc tests all confirmed that the 65 (27.4 %) out of 237 isolates were ESBL producing bacteria. There was high prevalence of bacteria in females than male and the number of E. coli strains is higher than Klebsiella spp. DNA isolation was performed on the 65 samples, out of which 50 samples were selected for PCR based on their concentration. The selected DNA samples were used to detect the presence of bla CTX-M and bla SHV genes. Only 24 DNA samples (48 %) contains blaCTX-M genes, bla SHV or both the genes. 14 samples had bla CTX-M gene, 2 bla SHV genes, and 8 with both bla SHV and bla CTX-M. At the rate at which ESBL is spreading, further research, close observation and cautious use of antibiotics is important.

Molecular characteristics and antibiotic resistance profiles of Escherichia coli strains isolated from urinary tract infections in children admitted to children's referral hospital of Qom, Iran.

Urinary tract infections (UTIs) are a highly prevalent infection among children and Escherichia coli is one of the most important pathogens causing pediatric UTIs. Production of extended spectrum b-lactamase (ESBL) enzymes is an important factor in the emergence of antibiotic resistance among these bacteria. This study aimed to determine the resistance patterns, the frequency of ESBL-encoding genes and the genetic diversity of E. coli strains isolated from children with UTIs who were admitted to children's referral hospital of Hazrat Masoumeh, Qom, Iran. A total of 102 consecutive non-duplicative strains of E.coli that were isolated from children with UTIs were included into the study. Antibiotic susceptibility of the isolates was determined by disk diffusion method according to the CLSI guidelines. The ability of the isolates to produce ESBLs was phenotypically determined by both combined disk test and double disk synergy test. The ESBL encoding genes (bla CTX-M, bla SHV, and bla TEM) in phenotypically confirmed ESBL-positive isolates was detected by PCR method. The genetic relatedness of the isolates was designated by enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR). Eighty-three percent (n=85) of the children were female. Most of the infected boys (88%, n=15) were less than 1 year of age and most of the infected girls (48%, n=41) aged 1 to 6 years old. The highest sensitivity was observed to nitrofurantoin (8%, n=8), followed by amikacin (12%, n=12) and piperacillin-tazobactam (17%, n=17). In contrast, the highest resistance rate was seen to ampicillin (94%, n=96) and cefazolin (93%, n=95). Eighty-eight percent (90 out of 102) of the strains were multidrug-resistant (MDR). Fifty-eight percent (n=59) of the strains were ESBL-positive and results of the combined disk test was in concordance with PCR. The blaCTX-M was the most frequent ESBL encoding gene (88%, n=52), followed by blaTEM (54%, n=32), and blaSHV (15%, n=9). Based on the ERIC-PCR technique, isolates were clustered in 13 different types. There was no relationship between different ERIC types and origin of the isolates (i.e. hospitalized or outpatients), ESBL-producing ability, and antibiotic resistance patterns. High prevalence of ESBL-positive isolates of E. coli (58%) was found in our study and all of them were MDR. In addition, there were statistically significant differences in the resistance rates of ESBL-producers, and non-producers to some antibiotics, which result in limiting their therapeutic options. Continuous surveillance of the emergence of ESBL-producing isolates and their antibiotic resistance profiles as well as using appropriate typing methods is needed for reducing their spread, selecting appropriate treatment regimens and finding hospital outbreaks.

Distribution of ExPEC Virulence Factors, bla CTX-M, fosA3, and mcr-1 in Escherichia coli Isolated From Commercialized Chicken Carcasses.

Pathogenic Escherichia coli found in humans and poultry carcasses harbor similar virulence and resistance genes. The present study aimed to analyze the distribution of extraintestinal pathogenic E. coli (ExPEC) virulence factors (VF), blaCTX−M groups, fosA3, and mcr-1 genes in E. coli isolated from commercialized chicken carcasses in southern Brazil and to evaluate their pathogenic risk. A total of 409 E. coli strains were isolated and characterized for genes encoding virulence factors described in ExPEC. Results of antimicrobial susceptibility testing confirmed that the strains were resistant to β-lactams, fosfomycin, colistin, and others resistance groups. The highest prevalence of VFs was observed in isolates belonging to the CTX-M groups, especially the CTX-M-2 group, when compared to those in other susceptible strains or strains with different mechanisms of resistance. Furthermore, ESBL strains were found to be 1.40 times more likely to contain three to five ExPEC virulence genes than non-ESBL strains. Our findings revealed the successful conjugation between ESBL-producing E. coli isolated from chicken carcass and the E. coli recipient strain J53, which suggested that genetic determinants encoding CTX-M enzymes may have originated from animals and could be transmitted to humans via food chain. In summary, chicken meat is a potential reservoir of MDR E. coli strains harboring resistance and virulence genes that could pose serious risks to human public health.