blaTEM Genes Research Articles

Tracking the Phylogenetic Origins of Proteus mirabilis and P. vulgaris in Iraq

Abstract Background: Common gram-negative rod-shaped bacteria called Proteus spp. are widely known for their swarming motility and urease activity. It is the main pathogen that causes complicated urethral tract infections. Objectives: Proteus mirabilis and P. vulgaris isolates were isolated, and the positions of the bla TEM, bla CTX-M, and bla SHV genes on the P. mirabilis isolates’ genomes were identified. The MEGA X10.2.4 software tool was employed to achieve this, and the genetic trees of P. mirabilis were created. Additionally, the isolates that represent the prevalence and degree of pathogenicity of these bacteria in Iraq were used to produce biofilms. Materials and Methods: A total of 210 urine samples from various patients who were hospitalized in general hospitals in Iraq between September 2021 and July 2022 were collected and yielded forty isolates of P. mirabilis and P. vulgaris. Traditional methods and the VITEK 2 compact were used to identify the isolates, which were given the names PM1 to PV40. Results: The results showed that all isolates were biofilm-forming, with the following ratios: 5 (15%) were weak, 31 (5.77%) were medium, and 4 (5.7%) were strong. The matching of this study isolates with NCBI-Gen bank global strains found that four isolates (PM1, PM2, PM3, and PM4) were originated in India, two isolates (PM5 and PM8) were originated in Iraq, and one (PM6) isolate was originated in Egypt. Conclusion: According to the results of this investigation, P. mirabilis and P. vulgaris isolates from various geographical areas show varying degrees of virulence factors. Therefore, additional researches are required to create a genetic diversity map of P. mirabilis and P. vulgaris due to variable frequencies in the sequencing of bla TEM, bla CTX-M, and bla SHV genes of the isolates.

Molecular analysis of ESBL- and AmpC-producing Enterobacteriaceae in fecal samples from broiler and backyard chickens

ObjectiveTo analyse the molecular profile of antibiotic resistance in Enterobacteriaceae isolates from fecal samples of healthy broilers and backyard chickens in Tamil Nadu, India. MethodsA total of 115 and 57 bacterial strains were isolated from fecal samples of broiler and backyard chickens, respectively using conventional bacterial culture tests. Bacterial isolates were screened for antibiotic susceptibility by Kirby-Bauer’s disk diffusion method. Colistin susceptibility was tested using the broth microdilution method. Extended-spectrum β-lactamase (ESBL), AmpC β-lactamase and metallo-β-lactamase (MBL) producers were screened by combination disk method. The drug-resistant genes’ positivity was analysed by polymerase chain reaction (PCR) method. ResultsEscherichia coli was the predominant bacterium isolated from both broiler (53%) and backyard (64.9%) chickens. Bacterial isolates from broilers and backyard chickens showed 100% resistance to ampicillin, piperacillin, and piperacillin-tazobactam. Five bacterial isolates from broiler chickens were resistant to colistin. ESBL producers were 34.7% in broilers and 19.1% in backyard chickens. AmpC production was detected in 44.3% of the isolates from broilers and 26.2% of those from backyard chickens. Molecular analysis revealed that bacterial isolates from broiler chickens were positive for drug-resistant genes such as blaTEM (25%), blaSHV (5%), blaCTX-M (4%), blaAmpC (56%), integron (39%), sul1 (30%), and sul2 (23%). Bacterial isolates from backyard chickens were positive for the blaTEM (22%), blaSHV (14%), blaAmpC (54.3%), and integron (20%) genes. None of the isolates from either broiler or backyard chickens harboured New Delhi metallo-β-lactamase (blaNDM) or colistin-resistant genes (mcr-1 to −3). ConclusionsBacterial isolates from broiler chickens harboured a higher number of drug resistance genes, especially blaTEM, blaCTX-M, sul1, sul2, and integron, than those from backyard chickens.

Prevalence and antimicrobial resistance profiles of mastitis causing bacteria isolated from dairy goats in Mukurweini Sub-County, Nyeri County, Kenya.

Ruminant mastitis continues to be a cause of economic losses in the dairy industry and remains a major public health hazard globally. This cross-sectional study was carried out in Mukurweini Sub-County of Nyeri County, Kenya, to investigate the prevalence of bacteria causing mastitis, risk factors associated with goat mastitis and the antibiotic resistance profiles of bacteria isolated from the goat milk. Farm level data on risk factors for mastitis was obtained from 56 farmers using a semi structured questionnaire. A total of 189 goat milk samples were collected. The goat's udder was observed for signs of clinical mastitis and the California Mastitis Test (CMT) used to test the milk for sub-clinical mastitis. All samples were then cultured for morphological identification of bacteria and strain typing by Matrix Assisted Laser Desorption/Ionization (MALDI)-Time of Flight (ToF) technique. Antimicrobial susceptibility of the isolated Staphylococcus aureus, coagulase-negative Staphylococcus (CoNS), Escherichia coli, Klebsiella oxytoca, Pseudomonas spp., Enterobacter spp., Proteus vulgaris and Escherichia vulneris to eight commonly used antibiotics was done by the disc diffusion method and validated by determining the presence of antibiotic resistance genes (mecA and blaTEM) using polymerase chain reaction method. The prevalence of clinical mastitis was 1.1% (2/189) while that of sub-clinical mastitis was 84.7% (160/189). Higher (p<0.05) prevalence of mastitis was observed in goats whose houses were cleaned fortnightly and in cases where farmers used same towel to dry different does' udders during the milking process. Thirteen different bacterial species were isolated from the milk samples and identified by MALDI-ToF, and these included S. aureus (22.0%), CoNS (20.3%), E. coli (18.1%), Pseudomonas spp. (14.3%), Enterobacter spp. (10.4%), K. oxytoca (6.0%), E. vulneris (1.7%), P. vulgaris (1.7%), Raoutella ornithinolytica (1.7%), Stenotrophomonas maltophilia (1.1%), Pantoea agglomerans (1.1%), Serratia marcescens (1.1%) and Cedeceas spp. (0.6%). One hundred pathogenic bacterial isolates were randomly selected and tested for antibiotic sensitivity to eight antibiotics out of which S. aureus were 97.5% resistant to Oxacillin and 100% sensitive to Ciprofloxacin. The CoNSs were 100% resistant to Oxacillin and 100% sensitive to Ciprofloxacin. E. coli were 93.9% resistant to Oxacillin, 69.7% sensitive to Ciprofloxacin and 87.9% sensitive to both Amoxicillin/Clavulanic acid and Meropenem. The antimicrobial resistant genes detected in S. aureus and E. coli were mecA [66.7%, 0%], and blaTEM [20% and 78.3%], respectively. In conclusion, the study showed that most of the does were affected by subclinical mastitis with the main causative bacteria being Staphylococci spp. and coliforms. Farmers need to be trained on improved control of mastitis by adoption of good milking practices and use of CMT kit for early detection of mastitis. Occurrence of multidrug resistance by key mastitis causing pathogens was shown to be prevalent and therefore there is need for development of intervention strategies.

Antibiotic Resistance Profile of Salmonella sp. Isolates from Commercial Laying Hen Farms in Central-Western Brazil.

Microbial resistance to antibiotics poses a significant threat to both human and animal health, necessitating international efforts to mitigate this issue. This study aimed to assess the resistance profiles of Salmonella sp. isolates and identify the presence of intl1, sul1, and blaTEM resistance genes within antigenically characterized isolates, including Agona, Livingstone, Cerro, Schwarzengrund, Salmonella enterica subsp. enterica serotype O:4.5, Anatum, Enteritidis, Johannesburg, Corvallis, and Senftenberg. These isolates underwent susceptibility testing against 14 antibiotics. The highest resistance percentages were noted for sulfamethoxazole (91%), sulfonamides (51%), and ceftiofur (28.9%), while no resistance was observed for ciprofloxacin. Salmonella Johannesburg and Salmonella Corvallis showed resistance to one antibiotic, whereas other serovars were resistant to at least two. Salmonella Schwarzengrund exhibited resistance to 13 antibiotics. The intl1 gene was detected in six out of the ten serovars, and the sul1 gene in three, always co-occurring with intl1. The blaTEM gene was not identified. Our findings highlight the risk posed by the detected multiple resistances and genes to animal, human, and environmental health. The multidrug resistance, especially to third-generation cephalosporins and fluoroquinolones, highlights the need for stringent monitoring of Salmonella in laying hens. The potential of the environment, humans, eggs, and their products to act as vectors for antibiotic resistance represents a significant concern for One Health.

Study of Antibiotic Resistance in ESKAPE Bacteria Using β-lactamase and ESBL Genes

Background: Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species, collectively known as ESKAPE bacteria, pose a significant threat in healthcare settings due to their increasing antibiotic resistance. Understanding the mechanisms underlying their resistance is crucial for developing effective therapeutic strategies. Methods: This study aimed to investigate the phenotypic and genotypic properties of β-lactamase enzymes in ESKAPE bacteria isolated from ulcer infections. Clinical specimens were collected from patients with various ulcer diseases, and bacterial isolates were identified using standard bacteriological methods and the Vitek-2 automated system. DNA extraction, polymerase chain reaction (PCR), and antibiotic susceptibility testing using the Kirby-Bauer disk diffusion method were performed. The presence of specific β-lactamase genes (blaKPC, blaTEM, blaCTX-M, and blaAMPC) was examined through molecular techniques. Results: Among the 104 clinical specimens collected, 88% yielded positive cultures, with Gram-negative bacteria predominating. Antibiotic susceptibility testing revealed high resistance rates, particularly to β-lactam antibiotics, among ESKAPE isolates. Molecular analysis identified the presence of extended-spectrum β-lactamases (ESBLs) in all isolates, with blaKPC and blaTEM being the most prevalent β-lactamase genes. Notably, blaKPC was detected in 72% of E. cloacae, 13.3% of S. aureus, 33.33% of Klebsiella pneumoniae, 50% of Pseudomonas aeruginosa, 0% of E. faecium, and 100% of A. baumannii isolates. Similarly, blaCTX-M and blaAMPC genes showed distinct distribution patterns across the different species. Conclusion: The study highlights the widespread presence of ESBL-producing ESKAPE bacteria in ulcer infections and underscores the importance of molecular techniques for accurate detection of β-lactamase production.

Identification of Extended Spectrum Beta Lactamases Producing Pseudomonas aeruginosa from Subclinical Mastitis Milk Samples in an Organized Dairy Farm of SVVU, Andhra Pradesh

The present study was conducted during August 2021 at Buffalo Research Station, Venkatarammanagudem of West Godavari District, Andhra Pradesh, India and was aimed to detect the presence of extended-spectrum beta-lactamases (ESBL) producing P. aeruginosa from sub-clinical mastitis that are apparently healthy Murrah buffaloes. Inflammation of the udder by microbial infection is one of the leading economic disease in the dairy sector. Most of the pathogens especially Pseudomonas aeruginosa are refractory to the antibiotic theraphy. Nowadays, the antimicrobial resistance owed by the bacteria is at an alarming rate. Pseudomonas aeruginosa, a multi drug resistant pathogen carry the extended spectrum beta lactamase genes in its plasmids and thus become resistant to antibiotic therapy. A total of 276 milk samples were collected from 69 milch animals in the month of August, 2021 comprising from all the four quarters of each buffalo. Nine (n=9) (3.26%) P. aeruginosa isolates were identified from the sub-clinical mastitis milk samples from the total 276 samples collected in the study. The isolates on morphological analysis were typical to P. aeruginosa. Seven isolates were positive for phenotypical beta-lactamases production. A total of 1 and 2 isolates were found reactive of the blaSHV and blaOXA genes, and two isolates harboured both blaSHV and blaOXA genes, respectively on PCR assay. No blaTEM gene was found in the isolates. Antibiogram of the ESBL producing P. aeruginosa isolates (n=7) possessed 100% resistance against most of the commonly used antimicrobials like ampicillin, amoxycillin, clindamycin, co-trimoxazole oxytetracycline, streptomycin and ceftriaxone (85%). The P. aeruginosa isolates were sensitive to amikacin (100%), ciprofloxacin (87.4%), gentamicin (84%), norfloxacin (84.8%) and enrofloxacin (82%).

Antimicrobial resistance, β-lactamase genotypes, and plasmid replicon types of Shiga toxin-producing Escherichia coli isolated from different animal hosts.

The primary objective of this study was to analyze antimicrobial resistance (AMR), with a particular focus on β-lactamase genotypes and plasmid replicon types of Shiga toxin-producing Escherichia coli (STEC) strains originating from various animal hosts. A total of 84 STEC strains were isolated from cattle (n=32), sheep/goats (n=26), pigeons (n=20), and wild animals (n=6) between 2010 and 2018 in various regions of Iran. The Kirby-Bauer susceptibility test and multiple polymerase chain reaction (PCR) panels were employed to elucidate the correlation between AMR and plasmid replicon types in STEC isolates. The predominant replicon types were IncFIC and IncFIB in cattle (46.8%), IncFIC in sheep/goats (46.1%), IncA/C in pigeons (90%), and IncP in wild animals (50%). STEC of serogroups O113, O26, and O111 harbored the IncFIB (100%), IncI1 (80%), and IncFIC+IncA/C (100%) plasmids, respectively. A remarkable AMR association was found between ciprofloxacin (100%), neomycin (68.7%), and tetracycline (61.7%) resistance with IncFIC; amoxicillin+clavulanic acid (88.8%) and tetracycline (61.7%) with IncA/C; ciprofloxacin (100%) with IncFIB; fosfomycin (85.7%) and sulfamethoxazole+trimethoprim (80%) with IncI1. IncI1 appeared in 83.3%, 50%, and 100% of the isolates harboring blaCTX-M, blaTEM, and blaOXA β-lactamase genes, respectively. The emergence of O26/IncI1/blaCTX-M STEC in cattle farms poses a potential risk to public health.

Genetic insights of antibiotic resistance, pathogenicity (virulence) and phylogenetic relationship of Escherichia coli strains isolated from livestock, poultry and their handlers - a one health snapshot.

Pathogenic and non-pathogenic strains of Escherichia coli harbouring antibiotic resistance genes (ARGs) from any source (clinical samples, animal settings, or environment) might be transmitted and contribute to the spread and increase of antibiotic resistance in the biosphere. The goal of this study was to investigate the genome to decipher the repertoire of ARGs, virulence genes carried by E. coli strains isolated from livestock, poultry, and their handlers (humans), and then unveil the genetic relatedness between the strains. Whole genome sequencing was done to investigate the genetic makeup of E. coli isolates (n = 20) [swine (n = 2), cattle (n = 2), sheep (n = 4), poultry (n = 7), and animal handlers (n = 5)] from southern India. The detection of resistome, virulome, biofilm forming genes, mobile genetic elements (MGE), followed by multilocus sequence typing (MLST)and phylogenetic analyses, were performed. E. coli strains were found to be multi drug resistant, with a resistome encompassing > 20 ARGs, the virulome-17-22 genes, and > 20 key biofilm genes. MGE analysis showed four E. coli isolates (host: poultry, swine and cattle) harbouring composite transposons with ARGs/virulence genes (blaTEM, dfr, qnr/nleB, tir, eae,and esp) with the potential for horizontal transfer. MLST analyses revealed the presence of ST937 and ST3107 in both livestock/poultry and their handlers. Phylogenomic analyses with global E. coli isolates (human/livestock/poultry hosts) showed close relatedness with strains originating from different parts of the world (the United States, China, etc.). The current study emphasizes the circulation of strains of pathogenic sequence types of clinical importance, carrying a diverse repertoire of genes associated with antibiotic resistance, biofilm formation and virulence properties in animal settings, necessitating immediate mitigation measures to reduce the risk of spread across the biosphere.

ANTIBIOTIC SUSCEPTIBILITY AND DETECTION OF EXTENDED-SPECTRUM B-LACTAMASE GENES IN GRAM-NEGATIVE BACTERIA ISOLATED FROM AUTOMATED TELLER MACHINES

Antibiotic-resistant (AR) bacteria especially from commonly shared surfaces in the environment, pose a serious threat to global public health. The increasing demand for electronic banking and the associated risk of the spread of pathogenic bacteria makes it crucial to assess the antibiotic susceptibility and the presence of extended-spectrum beta-lactamase genes in bacteria isolated from automated teller machines (ATMs). Following standard procedures, a total of 22 samples were collected randomly from different ATMs using sterile cotton swabs soaked in physiologic saline and cultured on selective media. Isolates were characterized biochemically. Antibiotics susceptibility test was carried out on isolates using the Kirby-Bauer disc diffusion method. All Gram-negative isolates were subjected to polymerase chain reaction (PCR) to evaluate the common extended-spectrum beta-lactamase –encoding (ESBL) genes. Bacterial isolates were characterized as Klebsiella pneumoniae (33.33%), Pseudomonas aeruginosa (19.61%), Escherichia coli (15.69%), Staphylococcus aureus (9.80%), Acinetobacter spp. (7.84%), Enterobacter spp. and Bacillus subtilis (5.88%), and Enterococcus faecalis (1.96%). The highest antibiotic resistance pattern was displayed against ceftazidime (45.1%), followed by tetracycline (43.1%), cefixime (41.2%), ciprofloxacin and chloramphenicol (29.4%) and meropenem (9.8%). The TEM gene (54.8%) was recovered most, followed by CTX-M (28.6%) and SHV (19.0%). This study reveals that ATMs could harbour pathogenic bacteria with antibiotic resistance (AR) genes especially ESBL genes (blaTEM) which could be responsible for the widespread resistance to antibiotics. Therefore, adequate personal hygiene by users, proper cleaning regimen to sanitize these facilities regularly and public enlightenment are recommended to reduce the accompanying risks of spreading AR genes within the environment

Molecular characterization of virulence and resistance genes in Salmonella strains isolated from chickens sold at the informal chicken market in Gauteng Province, South Africa

AbstractThis cross‐sectional study determined the occurrence of virulence and antimicrobial resistance genes in Salmonella strains recovered from chicken obtained from informal markets in Gauteng province, South Africa. The study also assessed the relationship between these characteristics, the source, the type of samples, and the serotypes of Salmonella isolates. A total of 151 samples (cloacal swabs, chicken carcasses, and carcass drips) were randomly collected from 47 informal market outlets in six townships in Gauteng province. Salmonella spp. was isolated and identified based on ISO 6579:2002 methods and confirmed by polymerase chain reaction (PCR) targeting invA gene fragment. Conventional PCR was used to detect 12 virulence and 18 antimicrobial resistance (AMR) genes in Salmonella spp. The most frequently detected virulence genes were invA (100%), shdA (91%), mgtB (87.7%), and sopE (81%), but considerably low for spvC (2.2%), sefC (1.5%), and pefC (0.4%). The differences in detection frequency were statistically significant (p &lt; 0.05). Tetracycline‐resistant genes tetA (34.7%) and tetB (16%) were the most frequently detected, while Beta‐lactam‐resistant genes blaTEM (0.4%), blaCMY‐2 (0.4%) and quinolones resistant gene qnrS (0.4%) were detected in low frequency (p &lt; 0.05). The locations of the outlets and the types of samples were significantly associated with detecting some virulence and AMR genes. Significant but moderately to substantial positive correlations were observed for qnrS, sul2; shdA, and mgtB genes. The pipA and spiC were, however, substantially negatively correlated. Our findings show that detecting these virulence and AMR genes in Salmonella isolates serves as a potential health hazard to the public, environment, and poultry farming in Gauteng, South Africa.

Antimicrobial Resistance and Molecular Characterization of Salmonella Rissen Isolated in China During 2008-2019.

This study aimed to provide epidemiological features of Salmonella enterica serovar Rissen, determine antimicrobial susceptibility, virulence gene profiles, and describe the potential association of S. Rissen from different sources in China. During 2008-2019, a total of non-repetitive 228 S. Rissen isolates were collected from human, animals and environment in China. The antimicrobial susceptibility test, screening of antimicrobial and virulence genes by PCR, and pulsed-field gel electrophoresis (PFGE) were performed. Among the 154 isolates from human, the majority of the cases (80.5%) occurred in summer, and S. Rissen was mainly detected in people aged 21-40 (37.7%) and 41-60 (28.6%) years old, and 74 non-human source S. Rissen strains were identified, with pork being the most common source. About 93.4% isolates were resistant to at least one of the 12 tested antimicrobial agents, and high frequencies of resistance were observed for tetracyclines (91.2%), trimethoprim-sulfamethoxazole (74.1%) and ampicillin (67.5%). A total of 171 (75%) isolates were resistant to at least three categories of antimicrobials, and the most common resistance profile was Tetracycline(s)-β-Lactams-Sulfonamides. The resistance rates to chloramphenicol, quinolones and sulfafurazole were significantly higher in strains isolated from human compared to non-human source strains. Among these isolates, the β-Lactams resistance was mainly associated with gene blaTEM (54.7%), sulfonamide resistance with sul2 (45.7%) and sul3 (54.3%), tetracycline resistance with tetA (81.3%). All the isolates harbored virulence genes hilA, sopB, sciN, stn and ssrB, and most of them harbored ssaQ (98.7%), mgtC (98.7%) and invA (98.2%). The majority (91.7%) of S. Rissen isolates showed high similarity (>80%) with each other in PFGE patterns and came from human, animals and environment. The high frequencies of multidrug resistance and probable clonal dissemination in this serovar call for the necessity of systematic surveillance on S. Rissen in China.

Quantification of β-lactamase producing bacteria in German surface waters with subsequent MALDI-TOF MS-based identification and β-lactamase activity assay

Environmental oligotrophic bacteria are suspected to be highly relevant carriers of antimicrobial resistance (AMR). However, there is a lack of validated methods for monitoring in the aquatic environment. Since extended-spectrum β-lactamases (ESBLs) play a particularly important role in the clinical sector, a culturing method based on R2A-medium spiked with different combinations of β-lactams was applied to quantify β-lactamase-producing environmental bacteria from surface waters. In German surface water samples (n = 28), oligotrophic bacteria ranging from 4.0 × 103 to 1.7 × 104 CFU per 100 mL were detected on the nutrient-poor medium spiked with 3rd generation cephalosporins and carbapenems. These numbers were 3 log10 higher compared to ESBL-producing Enterobacteriales of clinical relevance from the same water samples. A MALDI-TOF MS identification of the isolates demonstrated, that the method leads to the isolation of environmentally relevant strains with Pseudomonas, Flavobacterium, and Janthinobacterium being predominant β-lactam resistant genera. Subsequent micro-dilution antibiotic susceptibility tests (Micronaut-S test) confirmed the expression of β-lactamases. The qPCR analysis of surface waters DNA extracts showed the presence of β-lactamase genes (blaTEM, blaCMY-2, blaOXA-48, blaVIM-2, blaSHV, and blaNDM-1) at concentrations of 3.7 (±1.2) to 1.0 (±1.9) log10 gene copies per 100 mL. Overall, the results demonstrate a widespread distribution of cephalosporinase and carbapenemase enzymes in oligotrophic environmental bacteria that have to be considered as a reservoir of ARGs and contribute to the spread of antibiotic resistance.

Antibiotic Resistance in Seawater Samples from East Coast of Spain

Seawater has been proposed as a reservoir for antibiotic-resistant bacteria and antibiotic resistance genes, thus representing a risk to public health. In this study, we evaluated the presence of antibiotic resistance determinants (bacteria and genes) in 77 seawater samples collected at different points along the coast of the Gulf of Valencia (Spain). Specifically, indicators of fecal contamination bacteria, Escherichia coli and Enterococcus sp., were isolated, and their antibiotic resistance profiles were analyzed through the use of the Sensititre® system, followed by the detection of the main antibiotic resistance genes (blaTEM, qnrS, tetW, sulI, and ermB). The highest frequencies of resistance in the E. coli isolates were detected for ampicillin (35.1%) and ciprofloxacin (17.5%), followed by sulfamethoxazole and trimethoprim (15.7%), while 23% of enterococci isolates showed resistance to a single antibiotic, 20% against tetracycline and 3% against daptomycin. Through PCR analysis, 93% of the E. coli strains showed the blaTEM and sulI resistance genes. Among the enterococci, the presence of the blaTEM gene was detected in 40% of the isolates, while the rest of the genes were present at very low rates. Among the water samples, 57% were positive for at least one of the tested genes, italic format with blaTEM being the most commonly found gene (47%), followed by the qnrS (33%) and sulI (23%) genes. These results show that seawater, in addition to being subjected to a high rate of fecal contamination, can contribute to the spread of antibiotic resistance.

Prevalence and molecular characterization of multi-resistant Escherichia coli isolates from clinical bovine mastitis in China

Different antibiotics are used to treat mastitis in dairy cows that is caused by Escherichia coli (E. coli). Antimicrobial resistance in food-producing animals in China has been monitored since 2000. Surveillance data have shown that the prevalence of multiresistant E. coli in animals has increased significantly. This study aimed to investigate the occurrence and molecular characteristics of resistance determinants in E. coli strains (n = 105) obtained from lactating cows with clinical bovine mastitis (CBM) in China. A total of 220 cows with clinical mastitis, which has swollen mammary udder with reduced and red or gangrenous milk, were selected from 5000 cows. The results showed 94.3% of the isolates were recognized as multidrug resistant. The isolates (30.5%) were positive for the class I integrase gene along with seven gene cassettes that were accountable for resistance to trimethoprim resistance (dfrA17, dfr2d and dfrA1), aminoglycosides resistance (aadA1 and aadA5) and chloramphenicol resistance (catB3 and catB2), respectively. The bla TEM gene was present in all the isolates, and these carried the bla CTX gene. A double mutation in gyrA (i.e., Ser83Leu and Asp87Asn) was observed in all fluoroquinolone-resistant isolates. In total, nine fluoroquinolone-resistant E. coli isolates were identified with five different types of mutations in parC. In four (44.4%) isolates, Ser458Ala was present in parE, and in all nine (9/9) fluoroquinolone-resistant isolates, Pro385Ala was present in gyrB. Meanwhile, fluoroquinolone was observed as highly resistant, especially in isolates with gyrA and parC mutations. In summary, the findings of this research recognize the fluoroquinolone resistance mechanism and disclose integron prevalence and ESBLs in E. coli isolates from lactating cattle with CBM.

Prevalence of extended-spectrum beta-lactamase and molecular detection of blaTEM, blaSHV, and blaCTX-M genotypes among gram-negative Bacilli isolates from hospital acquired infections in pediatrics, one institutional study

BackgroundGram-negative bacilli represents an important pathogen in hospital-acquired infections (HAIs) worldwide. The emergence of antibiotic resistance in these pathogens warrants attention for the proper management of infections. Extended-spectrum beta-lactamase (ESBL) resistance represents a major therapeutic problem in infections due to Gram-negative bacilli.The present study aimed to study the extended-spectrum beta-lactamase genes blaTEM, blaSHV, and blaCTX-M by multiplex polymerase reaction in isolated Gram-negative bacilli from HAIs in pediatric patients.MethodsThe study included one hundred-five isolates of Gram-negative bacilli from pediatric patients with different types of HAIs. The isolates were subjected to full microbiological identification, antibiotics susceptibility by disc diffusion method, the phenotypic study of ESBL, and the genetic study of ESBL genes by multiplex PCR.ResultsFifty isolates of Gram-Negative bacilli showed ESBL activity by a phenotypic study by double disc diffusion method (50/105). All ESBL producers’ isolates were positive by PCR for ESBL genes. The most frequent gene was blaTEM (64%), followed by blaSHV (30%) and CTX-M (22%). Mixed genes were found in 4 isolates (8%) for blaTEM and blaSHV, blaTEM and CTX-M. There was a significant association between PCR for ESBL genes and phenotypic ESBL detection (P = 0.001). There was significant detection of ESBL genes in E. coli (28%), followed by Enterobacter spp. (26%), Klebsiella spp. (24%), Serratia (14%), Pseudomonas spp. (6%) and Proteus (2%), P = 0.01. There Seventy percent of isolates positive for ESBL production had an insignificant association between MDR and PCR for ESBL genes (P = 0.23).ConclusionThe present study highlights the prevalence of ESBL activity among clinical isolates of Gram-negative bacilli isolated from hospital-acquired infections in pediatric patients. The most common gene responsible for this activity was blaTEM gee followed by blaSHV and blaCTX-M. There was a high prevalence of multiple antibiotic resistance among isolates with ESBL activity. The finding of the present study denotes the importance of screening extended beta-lactamase among Gram-negative bacilli associated with HAIs in pediatric patients.

Typhoidal salmonella disease in Mukuru informal settlement, Nairobi Kenya; carriage, diversity, and antimicrobial resistant genes.

Multiple studies have shown that typhoid fever is endemic in developing countries characterized by poor hygiene. A unique way of Salmonella Typhi (S.Typhi) pathogenicity is establishing a persistent, usually asymptomatic carrier state in some infected individuals who excrete large numbers of bacteria in faeces. This study aimed to determine the isolation rate of S.Typhi from blood and stool samples among cases and asymptomatic individuals in the Mukuru informal settlement and identify antibiotic resistance patterns within the same population. We recruited 1014 outpatient participants presenting with typhoid-like symptoms in selected health centres in Nairobi, Kenya. Bacterial isolation was done on Xylose Lysine Deoxycholate agar (XLD) and Mac Conkey agar (Oxoid), followed by standard biochemical tests. Identification was done using API20E, and S.Typhi was confirmed by serotyping using polyvalent antisera 0-9 and monovalent antisera d. The Kirby-Bauer disc diffusion method was used to test the antimicrobial susceptibility of S.Typhi isolates, while Multi-Drug Resistant (MDR) strains were characterized using conventional PCR. Of 1014 participants, 54 (5%) tested positive for S.Typhi. Thirty-eight (70%) of the S.Typhi isolated were from stool samples, while sixteen (30%) were from blood. Three (0.2%) of the isolates were from asymptomatic carriers. Of the 54 S.Typhi isolates, 20 (37%) were MDR. Resistance to ciprofloxacin and nalidixic acid was 43% and 52%, respectively. Resistance to amoxicillin-clavulanic acid (a beta-lactam inhibitor) was 2%. The BlaTEM-1 gene was present in 19/20 (95%) MDR isolates. MDR S.Typhi is prevalent in Mukuru Informal settlement. The sharp increase in nalidixic acid resistance is an indication of reduced susceptibility to fluoroquinolones, which are currently the recommended drugs for the treatment of typhoid fever. This study highlights the need for effective antimicrobial stewardship and routine surveillance of antimicrobial resistance (AMR) to inform policy on the prevention and control of MDR Typhoid disease.

Outbreak of colistin and carbapenem-resistant Klebsiella pneumoniae ST16 co-producing NDM-1 and OXA-48 isolates in an Iranian hospital

BackgroundColistin and carbapenem-resistant Klebsiella pneumoniae (Col-CRKP) represent a significant and constantly growing threat to global public health. We report here an outbreak of Col-CRKP infections during the fifth wave of COVID-19 pandemic.MethodsThe outbreak occurred in an intensive care unit with 22 beds at a teaching university hospital, Isfahan, Iran. We collected eight Col-CRKP strains from seven patients and characterized these strains for their antimicrobial susceptibility, determination of hypermucoviscous phenotype, capsular serotyping, molecular detection of virulence and resistance genes. Clonal relatedness of the isolates was performed using MLST.ResultsThe COVID-19 patients were aged 24–75 years with at least 50% pulmonary involvement and were admitted to the intensive care unit. They all had superinfection caused by Col-CRKP, and poor responses to antibiotic treatment and died. With the exception of one isolate that belonged to the ST11, all seven representative Col-CRKP strains belonged to the ST16. Of these eight isolates, one ST16 isolate carried the iucA and ybtS genes was identified as serotype K20 hypervirulent Col-CRKP. The blaSHV and blaNDM-1 genes were the most prevalent resistance genes, followed by blaOXA-48 and blaCTX-M-15 and blaTEM genes. Mobilized colistin-resistance genes were not detected in the isolates.ConclusionsThe continual emergence of ST16 Col-CRKP strains is a major threat to public health worldwide due to multidrug-resistant and highly transmissible characteristics. It seems that the potential dissemination of these clones highlights the importance of appropriate monitoring and strict infection control measures to prevent the spread of resistant bacteria in hospitals.

The presence of antibiotic-resistant bacteria at four Norwegian wastewater treatment plants: seasonal and wastewater-source effects

Wastewater treatment plants receive low concentrations of antibiotics. Residual concentrations of antibiotics in the effluent may accelerate the development of antibiotic resistance in the receiving environments. Monitoring of antimicrobial resistance genes (ARGs) in countries with strict regulation of antibiotic use is important in gaining knowledge of how effective these policies are in preventing the emergence of ARGs or whether other strategies are required, for example, at-source treatment of hospital effluents. This study evaluates the presence of certain common resistance genes (blaSHV-1, blaTEM-1, msrA, ermA, ermC, tetM, tetL, tetA, vanA, and vanC) in the influent, sludge, and effluent of four wastewater treatment plants (WWTPs) in the North Jæren region of Norway at two different sampling times (January and May). These WWTPs vary in drainage area and wastewater composition and were selected based on their differing wastewater characteristics. Randomly selected colonies from the activated sludge samples were used to determine the minimum inhibitory concentration (MIC) for ampicillin, vancomycin, and tetracycline. In addition, variations in the bacterial composition of the wastewater were characterized via 16S rRNA sequencing and were analyzed in terms of bacterial host taxa that explain the presence of the ARGs in wastewater. The MIC tests revealed MIC90 values of &amp;gt;128 µg/mL for ampicillin, ≥128 µg/mL for vancomycin, and 32 µg/mL for tetracycline. In addition, the three resistance genes, ermB, tetA, and tetM, that were present in the influent and activated sludge were still present in the effluent. These results indicate that WWTPs represent a direct route into the environment for resistance genes and do not significantly reduce their abundance. Hence, the development of treatment methods for the removal of these genes from WWTPs in the future is of utmost importance.

Occurrence characteristics and influencing factors of antibiotic resistance genes in rural groundwater in Henan Province.

This study determined the antibiotic-resistant gene (ARG) contents of 34 groundwater samples in Henan Province collected from September to October 2022, then assessed the roles of both water quality parameters and intI1 in ARG propagation in groundwater. The results show that there existed universal ARG pollution in groundwater, and sulfonamides-, β-lactem-, and tetracycline-resistance genes were the most prevalent gene types during the time. Sul1 contributed the majority proportion of the total resistance genes (TARGs). The prevalence of ESBLs gene blaTEM and the occurrence of Carbapenems resistant gene blaOXA-1 suggests the pollution of high-risk ARGs in groundwater demands more attention. IntI1 is prevalent and had a significantly positive correlation with almost 50% ARGs, indicating its contribution to ARG propagation in groundwater. Well types contribute little to ARG propagation in rural groundwater of Henan, which means the protective facilities established by the local government for public wells can effectively prevent contamination from exogenous ARGs. However, the economic level has no impact on the abundance of ARGs in rural groundwater, which suggests the local government should pay greater attention to investment in controlling ARG pollution in Henan rural areas.

Antimicrobial resistance of avian pathogenic Escherichia coli isolated from broiler, layer, and breeder chickens.

Antimicrobials are extensively used in poultry production for growth promotion as well as for the treatment and control of diseases, including avian pathogenic Escherichia coli (APEC). Poor selection, overuse, and misuse of antimicrobial agents may promote the emergence and dissemination of antimicrobial resistance (AMR) in APEC. This study aimed to assess antimicrobial susceptibility patterns and detect antibiotic resistance genes (ARGs) in APEC isolated from clinical cases of colibacillosis in commercial broiler, layer, and breeder chickens. A total of 487 APEC were isolated from 539 across 300 poultry farms in various regions of Nepal. Antimicrobial susceptibility patterns was determined using the Kirby-Bauer disk diffusion and broth microdilution methods. The index of AMR, such as multiple antibiotic resistance (MAR) index, resistance score (R-score), and multidrug resistance (MDR) profile, were determined. Polymerase chain reaction was employed to detect multiple ARGs and correlations between phenotypic and genotypic resistance were analyzed. The prevalence of APEC was 91% (487/539). All of these isolates were found resistant to at least one antimicrobial agent, and 41.7% of the isolates were resistant against 8-9 different antimicrobials. The antibiogram of APEC isolates overall showed the highest resistance against ampicillin (99.4%), whereas the highest intermediate resistance was observed in enrofloxacin (92%). The MAR index and R-score showed significant differences between broiler and layers, as well as between broiler breeder and layers. The number of isolates that were resistant to at least one agent in three or more antimicrobial categories tested was 446 (91.6%) and were classified as MDR-positive isolates. The ARGs were identified in 439 (90.1%) APEC isolates, including the most detected mobilized colistin resistance (mcr1) which was detected in the highest (52.6%) isolates. Overall, resistance gene of beta-lactam (blaTEM), mcr1, resistance gene of sulphonamide (sul1) and resistance gene of tetracycline (tetB) (in broiler), were detected in significantly higher than other tested genes (p < 0.001). When examining the pair-wise correlations, a significant phenotype-phenotype correlation (p < 0.001) was observed between levofloxacin and ciprofloxacin, chloramphenicol and tetracycline with doxycycline. Similarly, a significant phenotype-genotype correlation (p < 0.001) was observed between chloramphenicol and the tetB, and colistin with blaTEM and resistance gene of quinolone (qnrA). In this study, the current state of APEC AMR in commercial chickens is revealed for the first time in Nepal. We deciphered the complex nature of AMR in APEC populations. This information of molecular surveillance is useful to combat AMR in APEC and to contribute to manage APEC associated diseases and develop policies and guidelines to enhance the commercial chicken production.