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The rapid emergence of multidrug-resistant (MDR) uropathogenic Escherichia coli (UPEC) strains pose a critical challenge in urinary tract infection (UTI) treatments. However, little work elucidated the resistance mechanisms of the MDR UPEC clinical strains in Malaysia. Therefore, this study aimed to determine the antimicrobial susceptibility profiles and the prevalence of antimicrobial resistance genes among the UPEC strains. Polymerase chain reactions were conducted to detect the presence of 6 antimicrobial resistance genes among 60 UPEC strains. Meanwhile, the antimicrobial resistance profiles against 9 antimicrobials were examined through the Kirby-Bauer disk diffusion method. In this study, the MDR isolates accounted for 40.0% (24/60), with the highest prevalence of resistance towards ampicillin (43/60; 71.7%), followed by tetracycline (31/60; 51.7%), nalidixic acid (30/60; 50.0%), co-trimoxazole (20/60, 33.3%), ciprofloxacin (19/60, 31.7%), levofloxacin (16/60, 21.6%) and chloramphenicol (10/60, 16.7%). In contrast, low resistance rates were observed among minocycline (1/60; 1.7%) and imipenem (0/60; 0.0%). bla TEM was the most prevalent gene (36/60; 60.0%), followed by tetA (27/60; 45.0%), sul2 (25/60; 41.7%), sul1 (13/60; 21.7%) and tetB (8/60; 13.3%). Surprisingly, bla SHV was not detected among the UPEC isolates. The MDR, ampicillin and tetracycline-resistant isolates were significantly associated with a higher prevalence of tetA, sul1, sul2 and bla TEM. In contrast, tetB displayed no significant relationship with any of the antimicrobials tested. The patient's age and gender were not the risk factors for the carriage of the resistance genes. Our findings identified the common resistance genes carried by the antimicrobial resistant UPEC isolates and provide valuable insights into developing the best antibiotic prescription regime to treat UTIs in our local scene.

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Background and Aim: The excessive use of antimicrobials in livestock farming leads to the emergence and dissemination of antimicrobial-resistant organisms. This study aimed to detect extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli genes in integrated poultry-fish farms in Bogor, Indonesia. Materials and Methods: A total of 256 samples were collected from six poultry-fish farms. One hundred and seventy-five chicken cloaca swabs, 60 fish skin swabs, six pond water samples, and 15 farmer's hand swabs. ESBL-producing E. coli was confirmed through double-disk diffusion. The specific primers and probe genes for quantitative polymerase chain reaction detection of ESBL-producing E. coli targeted blaTEM, blaCTX-M, blaSHV, and blaOXA-48 genes. Results: Among the 256 samples tested, 145 (56.6%) were positive for E. coli, and 67.6% (98/145) were identified as ESBL-producing E. coli. The most ESBL-producing E. coli isolates were obtained from chicken cloaca (78.3%, 72/92), followed by pond water (66.7%, 4/6), fish skin (47.6%, 20/42), and farmer's hand swabs (40%, 2/5). About 100% of the isolates carried the genes blaTEM and blaCTX-M, whereas 17.3% and 24.5% carried blaSHV and blaOXA-48, respectively. Conclusion: ESBL-producing E. coli genes were investigated in chicken cloaca, fish, pond water, and farmers' hands within an interconnected poultry-fish farming operation. The ESBL-producing E. coli in chickens can transfer resistant genes to aquatic environments. The transfer could harm other aquatic species and food chains, potentially threatening human health.

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Urban wastewater is a significant by-product of human activities. Conventional urban wastewater treatment plants have limitations in their treatment, mainly concerning the low removal efficiency of conventional and emerging contaminants. Discharged wastewater also contains harmful microorganisms, posing risks to public health, especially by spreading antibiotic-resistant bacteria and genes. Therefore, this study assesses the potential of a native microalgae-bacteria system (MBS) for urban wastewater bioremediation and disinfection, targeting NH4+-N and PO43--P removal, coliform reduction, and antibiotic resistance gene mitigation. The MBS showed promising results, including a high specific growth rate (0.651 ± 0.155 d-1) and a significant average removal rate of NH4+-N and PO43--P (9.05 ± 1.24 mg L-1 d-1 and 0.79 ± 0.06 mg L-1 d-1, respectively). Microalgae-induced pH increase rapidly reduces coliforms (r > 0.9), including Escherichia coli, within 3 to 6 days. Notably, the prevalence of intI1 and the antibiotic resistance genes sul1 and blaTEM are significantly diminished, presenting the MBS as a sustainable approach for tertiary wastewater treatment to combat eutrophication and reduce waterborne disease risks and antibiotic resistance spread.

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Salmonella enterica serovar Kentucky ST198 is a major health threat due to its resistance to ciprofloxacin and several other drugs, including third-generation cephalosporins. Many drug-resistant genes have been identified in the Salmonella genomic island 1 variant K (SGI1-K). In this study, we investigated the antimicrobial resistance (AMR) profile and genotypic relatedness of two isolates of ciprofloxacin-resistant (CIPR) S. Kentucky ST198 from poultry in Northeastern Thailand. We successfully assembled the complete genomes of both isolates, namely SSSE-01 and SSSE-03, using hybrid de novo assembly of both short- and long-read sequence data. The complete genomes revealed their highly similar genomic structures and a novel variant of SGI1-K underlying multidrug-resistant (MDR) patterns, including the presence of blaTEM-1b, which confers resistance to beta-lactams, including cephalosporins and lnu(F) which confers resistance to lincomycin and other lincosamides. In addition, the chromosomal mutations in the quinolone resistance-determining region (QRDR) were found at positions 83 (Ser83Phe) and 87 (Asp87Asn) of GyrA and at positions 57 (Thr57Ser) and 80 (Ser80Ile) of ParC suggesting high resistance to ciprofloxacin. We also compared SSSE-01 and SSSE-03 with publicly available complete genome data and revealed significant variations in SGI1-K genetic structures and variable relationships to antibiotic resistance. In comparison to the other isolates, SGI1-K of SSSE-01 and SSSE-03 had a relatively large deletion in the backbone, spanning from S011 (traG∆) to S027 (resG), and the inversion of the IS26-S044∆-yidY segment. Their MDR region was characterized by the inversion of a large segment, including the mer operon and the relocation of IntI1 and several resistance genes downstream of the IS26-S044∆-yidY segment. These structural changes were likely mediated by the recombination of IS26. The findings broaden our understanding of the possible evolution pathway of SGI1-K in fostering drug resistance, which may provide opportunities to control these MDR strains.IMPORTANCEThe emergence of ciprofloxacin-resistant (CIPR) Salmonella Kentucky ST198 globally has raised significant concerns. This study focuses on two poultry isolates from Thailand, revealing a distinct Salmonella genomic island 1 variant K (SGI1-K) genetic structure. Remarkably, multiple antibiotic resistance genes (ARGs) were identified within the SGI1-K as well as other locations in the chromosome, but not in plasmids. Comparing the SGI1-K genetic structures among global and even within-country isolates unveiled substantial variations. Intriguingly, certain isolates lacked ARGs within the SGI1-K, while others had ARGs relocated outside. The presence of chromosomal extended-spectrum ß-lactamase (ESBL) genes and lincosamide resistance, lnu(F), gene, could potentially inform the choices of the treatment of CIPRS. Kentucky ST198 infections in humans. This study highlights the importance of understanding the diverse genetic structures of SGI1-K and emphasizes the role of animals and humans in the emergence of antimicrobial resistance.

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This study examines the genetic contexts and evolutionary steps responsible for the formation of the widely spread transposon Tn6925 carrying blaTEM and aacC2e, which confers resistance to beta-lactam and aminoglycoside antibiotics in Gram-negative bacteria. The blaTEM-1 and aacC2e genes were found in several transposons. They were first observed within an IS26 bounded 3.7 kb transposon (Tn6925) on several Acinetobacter baumannii plasmids located within a 4.7 kb dif module. Truncated and expanded variations of Tn6925 were found across other A. baumannii plasmids, as well as in other Gram-negative bacteria (including Vibrio cholerae). Moreover, blaTEM-1 and aacC2e were in much larger resistance-heavy transposons including the ISAba1-bounded 24.6 kb (here called Tn6927), found in an A. baumannii chromosome. A novel ISKpn12-bounded transposon was also observed to contain blaTEM and aacC2e which was found interrupting Tn5393 along with an IS26 pseudo-compound transposon to form a 24.9 kb resistance island in an Acinetobacter pittii plasmid. Multiple mobile genetic elements are involved in the formation of transposon structures that circulate blaTEM and aacC2e. Among these, IS26 and ISAba1 appear to have played a major role in the formation and spread of these elements in the Acinetobacter species.

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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.

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Background: The discovery of antibiotic-resistant Enterobacteriaceae bacteria in wild animals is an indication of their potential for wildlife as a reservoir. Bats are natural reservoir hosts and a source of infection for several microorganisms and have the potential to become vectors for the spread of zoonotic diseases. Aim: A study was conducted based on these characteristics to identify and detect the blaTEM gene in Eschericia coli isolated from bat excrements in Tanjung Ringgit Cave, East Lombok. Methods: Bat fecal samples were firstly inoculated onto eosin methylene blue agar media. Recovered bacterial isolates were further characterized using standard microbiological techniques. Antimicrobial susceptibility testing was done using the Kirby-Bauer disc diffusion method. blaTEM gene detection was carried out using polymerase chain reaction (PCR). Results: Out of the 150 bat fecal samples obtained from Tanjung Ringgit cave, Lombok Island, Indonesia, 56 (37%) were positive for E. coli. Eight (8) out of the 56 E. coli isolates that underwent antimicrobial susceptibility testing using the disc diffusion method were confirmed to be multidrug-resistant as they exhibited resistance to at least three different classes of antibiotics. Out of the eight (8) multidrug resistance E. coli isolates recovered from fecal samples of bats, 2 (two) harbored the blaTEM gene. Conclusion: The discovery of the blaTEM gene in bat fecal samples indicates the potential for wild animals, especially bats, to spread ESBL resistance genes to the environment and to humans.

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Poultry products in Zambia form an integral part of the human diet in many households, as they are cheap and easy to produce. The burden of poultry diseases has, however, remained a major challenge. Growing consumer demand for poultry products in Zambia has resulted in non-prudent antimicrobial use on farms, intending to prevent and treat poultry diseases for growth optimisation and maximising profits. This cross-sectional study aimed to identify the different types of bacteria causing diseases in chickens in Lusaka and to detect the extended-spectrum lactamase (ESBL)-encoding genes. We collected 215 samples from 91 diseased chickens at three post-mortem facilities and screened them for Gram-negative bacteria. Of these samples, 103 tested positive for various clinically relevant Enterobacteriaceae, including Enterobacter (43/103, 41.7%), Escherichia coli (20/103, 19.4%), Salmonella (10/103, 9.7%), and Shigella (8/103, 7.8%). Other isolated bacteria included Yersinia, Morganella, Proteus, and Klebsiella, which accounted for 21.4%. E. coli, Enterobacter, Salmonella, and Shigella were subjected to antimicrobial susceptibility testing. The results revealed that E. coli, Enterobacter, and Shigella were highly resistant to tetracycline, ampicillin, amoxicillin, and trimethoprim-sulfamethoxazole, while Salmonella showed complete susceptibility to all tested antibiotics. The observed resistance patterns correlated with antimicrobial usage estimated from sales data from a large-scale wholesale and retail company. Six (6/14, 42.9%) E. coli isolates tested positive for blaCTX-M, whilst eight (8/14, 57.1%) Enterobacter samples tested positive for blaTEM. Interestingly, four (4/6, 66.7%) of the E. coli isolates carrying blaCTX-M-positive strains were also positive for blaTEM. Sanger sequencing of the PCR products revealed that five (5/6, 83.3%) of the abovementioned isolates possessed the blaCTX-M-15 allele. The results suggest the presence of potentially pathogenic ESBL-producing Enterobacteriaceae in poultry, threatening public health.

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Nontyphoid salmonella can cause severe infections in newborns and is therefore declared a pathogen of major health significance at this age. The aim of the study was molecular and antimicrobial characterization of ß-lactamase-producing Salmonella Mikawasima outbreak clone on a Neonatal ward, University Hospital of Split (UHS), Croatia during the COVID-19 pandemic. From April 2020, until April 2023, 75 nonrepetitive strains of Salmonella Mikawasima were isolated from stool specimens and tested for antimicrobial resistance. All 75 isolates were resistant to ampicillin and gentamicin, while 98% of isolates were resistant to amoxicillin/clavulanic acid. A high level of resistance was observed to third-generation cephalosporins (36% to ceftriaxone and 47% to ceftazidime). Extended-spectrum ß-lactamase production was phenotypically detected by double-disk synergy test in 40% of isolates. Moderate resistance to quinolones was detected; 7% of isolates were resistant to pefloxacin and ciprofloxacin. All isolates were susceptible to carbapenems, chloramphenicol, and co-trimoxazole. Fourteen representative isolates, from 2020, 2021, 2022, and 2023, were analyzed with PFGE and all of them belong to the same clone. Whole-genome sequencing (WGS) analysis of three outbreak-related strains (SM1 and SM2 from 2020 and SM3 from 2023) confirmed that these strains share the same serotype (Mikawasima), multilocus sequence typing profile (ST2030), resistance genes [blaTEM-1B, aac(6')-Iaa, aac(6')-Im, and aph(2'')-Ib)] and carry incompatibility group C (IncC) plasmid. Furthermore, the gene blaSHV-2 was detected in SM1 and SM2. In summary, WGS analysis of three representative strains clearly demonstrates the persistence of ß-lactamase-producing Salmonella Mikawasima in UHS during the 4-year period.

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In the 2019-2020 summer, wildfires decimated the Australian bush environment and impacted wildlife species, including koalas (Phascolarctos cinereus) and grey headed flying fox pups (Pteropid bats, Pteropus poliocephalus). Consequently, hundreds of koalas and thousands of bat pups entered wildlife hospitals with fire-related injuries/illness, where some individuals received antimicrobial therapy. This study investigated the dynamics of antimicrobial resistance (AMR) in pre-fire, fire-affected and post-fire koalas and Pteropid bat pups. PCR and DNA sequencing were used to screen DNA samples extracted from faeces (koalas and bats) and cloacal swabs (koalas) for class 1 integrons, a genetic determinant of AMR, and to identify integron-associated antibiotic resistance genes. Class 1 integrons were detected in 25.5% of koalas (68 of 267) and 59.4% of bats (92 of 155). Integrons contained genes conferring resistance to aminoglycosides, trimethoprim and beta-lactams. Samples were also screened for blaTEM (beta-lactam) resistance genes, which were detected in 2.6% of koalas (7 of 267) and 25.2% of bats (39 of 155). Integron occurrence was significantly higher in fire-affected koalas in-care compared to wild pre-fire koalas (P < 0.0001). Integron and blaTEM occurrence were not significantly different in fire-affected bats compared to pre-fire bats (P > 0.05), however, their occurrence was significantly higher in fire-affected bats in-care compared to wild fire-affected bats (P < 0.0001 and P = 0.0488 respectively). The observed shifts of AMR dynamics in wildfire-impacted species flags the need for judicious antibiotic use when treating fire-affected wildlife to minimise unwanted selective pressure and negative treatment outcomes associated with carriage of resistance genes and antibiotic resistant bacteria.

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BACKGROUND: Gram-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. METHODS: The 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. RESULTS: Fifty 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). CONCLUSION: The 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.

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OBJECTIVES: This study reports the whole-genome sequences of two strains of extended-spectrum beta-lactamase (ESBL)-producing and multidrug-resistant (MDR) K. pneumoniae ST268 and explores their acquired antibiotic resistance genes (ARGs) and the mobile genetic elements (MGEs). METHODS: Two strains of K. pneumoniae ST268 were isolated from different samples of one patient. Assessment of antimicrobial susceptibility was performed, and then whole-genome sequencing was conducted. Acquired ARGs, insertion sequences, and transposons harboured by the two strains of K. pneumoniae ST268 were identified, and then the genetic contexts associated with the ARGs were analysed systematically. RESULTS: Two strains of K. pneumoniae ST268 were found to carry the 118.6-kb hybrid IncFIIK:IncQ1:repBR1701 plasmid. All the acquired ARGs carried by the IncF plasmid were found to be situated on the 25.3-kb MDR region bracketed by ISKpn19 and IS26, which was widely present in the plasmids in 14 STs of strains in K. pneumoniae but also in IncF plasmids from Shigella flexneri and Klebsiella quasipneumoniae. Notably, the IncF plasmids harbouring the 25.3-kb MDR region were geographically distributed mainly in China, and the pKP161637-1/pKP160802-1 in our study was the first report on the IncF plasmid carrying the 25.3-kb MDR region bracketed in K. pneumoniae ST268. CONCLUSIONS: Two strains of ESBL-producing K. pneumoniae ST268 with a MDR IncF plasmid were identified in a hospital in China. The ARGs were identified on the 25.3-kb MDR region, bracketed by ISKpn19 and IS26, of the IncF plasmids, which were present not only in the K. pneumoniae but also in the S. flexneri and K. quasipneumoniae.

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BACKGROUND: The increasing cases of bloodstream infections among children at neonatal intensive care units (NICUs) led this work to investigate biofilm production, antibiotics and the presence of ESßL genes in Serratia marcescens (S. marcescens) strains isolated from blood. METHODS: Twenty S. marcescens strains were isolated and identified by the VITEK-2 system over 7 months from late 2022 to mid-2023 from Ibn Al-Balady Hospital in Baghdad. Kirby-Bauer test was used to measure antibiotic susceptibility. RESULTS: The results revealed that 95% of twenty S. marcescens isolates were non-susceptible to Ampicillin and Amoxicillin-clavulanic. Furthermore, S. marcescens isolates showed a high sensitivity rate 70% toward Imipenem. All S. marcescens strains 100% were produced biofilm. This work clarifies that, out of 20 S. marcescens strains, 80% were harbored ESßL genes. The coexistence of blaTEM, blaCTX and blaSHV genes was shown in 43.75% of strains, while 56.25% of S. marcescens strains harbored single ES[Formula: see text]L genes. The biofilm values increase with the accuracy of EsßL genes. Phylogenetic analyses based on the sequence of blaCTX-M and blaTEM were done with closely related genes in the GenBank using MEGA6 software. CONCLUSIONS: The distribution of blaTEM, blaCTX and blaSHV genes among local S. marcescens strains may be attributed to the indiscriminate use of antibiotics. The results confirmed the spread of ESßL genes in S. marcescens from blood infections among newborn infants.

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The Enterobacteriaceae family exhibits resistance to antibiotics by producing ß-Lactamase. Mutations in ß-Lactamase, have led to the generation of inhibitor resistant variants known as IR-TEM. In the present study, IR-TEM ß-Lactamase of Enterobacter hormaechei and Enterobacter asburiae was compared. To investigate the mechanism behind the conferred mutation, we studied its interaction with Clavulanic acid, (ß-Lactamase inhibitor) with different lineages of TEM and IR-TEM. We found that Clavulanic acid quickly left the binding pockets of both variants using molecular dynamics (MD) simulations. Interestingly, mutations at the V84I and A184V positions were found to drastically influence the protein dynamics. Mutating the residues at V84I and A184V positions by computational mutagenesis in Enterobacter hormaechei, it was observed that the residues on the Ω-loop as well as a few downstream residues were primarily involved in generating resistance towards inhibitors by conferring increased flexibility to the loop. This further strongly supports the notion that inhibitor resistance in ß-Lactamase is conferred through allosteric regulation, wherein mutations in positions 84 and 184 may play an important role in regulating the movement of the Ω-loop. These two positions determine the lineage pattern between two clusters in TEM-1 and TEM-116. To date, no reports have been made regarding the importance of these mutations and their dynamics in Ω-loop.Communicated by Ramaswamy H. Sarma.

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Introduction: Infections of humans and animals by multidrug resistant bacteria are increasing because of the inappropriate use of antibiotics. Disease management may be more challenging if Escherichia coli produce extended-spectrum beta-lactamase (ESBL), which could cause resistance to aztreonam and third-generation cephalosporins. This study was aimed at determining the prevalence of the blaCTX-M and blaTEM genes among ESBL-producing E. coli isolated from broiler chickens in Indonesia. Material and Methods: A total of 115 broiler cloacal swab samples were obtained from 22 farms and studied for the presence of E. coli. The isolates were identified using approved standard methods and were purified on eosin methylene blue agar media. The E. coli isolates were subjected to sensitivity testing using beta-lactam antibiotics, and ESBL production was confirmed by a double-disc synergy test. The presence of the blaCTX-M and blaTEM genes was identified using a PCR. Results: It was found that 99/115 (86.1%) of the isolated E. coli were resistant to beta-lactam antibiotics and 34/115 (29.6%) of them were phenotypically detected to be ESBL producers. Of the 34 isolates that were confirmed ESBL producers, 32/34 (94.1%) of them harboured the blaCTX-M and 13/34 (38.2%) the blaTEM genes. The blaCTX-M and blaTEM genes were detected together in 12/34 (35.3%) isolates. Conclusion: This study discovered that broiler chickens are possible reservoirs of ESBL-producing E. coli that may infect humans. Thus, a committed public health education campaign is recommended in order to mitigate the potential threat to human health.

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Background The oral microbiome consists of various bacterial genera, with Neisseria spp. being a prominent part of this niche. While Neisseria gonorrhoeae and Neisseria meningitidis are human-restricted pathogens, non-pathogenic Neisseria species like Neisseria sicca, Neisseria perflava, etc., are primarily commensals that can also behave as opportunistic pathogens. With increasing penicillin resistance in commensal Neisseria, there is a concern that these bacteria might harbor resistance genes that can be transferred to other pathogens. This study aimed to characterize the blaTEM gene (encodes for the plasmid-mediated ß-lactamase enzyme that hydrolyzes the ß-lactam ring) of commensal Neisseria spp. isolated from respiratory samples. Methodology The research was conducted in the Department of Clinical Microbiology at Sri Ramachandra University, Chennai. The specimens used were sputum and throat swabs, which were subjected to a series of phenotypic methods and matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) for speciation. The antibiogram was determined using the Kirby-Bauer disk diffusion method, and a PCR assay was utilized to identify the blaTEM gene responsible for ß-lactamase production. Results Out of 274 processed samples, 65 unique commensal Neisseria spp. were identified. The study highlighted the presence of the blaTEM gene in 93.9% (61) of the isolates, which is responsible for ß-lactamase production. All isolates exhibited resistance to penicillin. Most blaTEM-positive commensal Neisseria spp. were susceptible to cefuroxime (83.6%), ceftriaxone (85.2%), and cefotaxime (85.2%). The high prevalence of the blaTEM gene in commensal Neisseria is alarming. The gene, found on plasmids, could potentially transfer to other related species like Neisseria gonorrhoeae and Neisseria meningitidis, as well as other Gram-negative bacilli. Conclusion The presence of resistance genes in commensal bacteria is of concern, as they might be reservoirs for resistance transfer to pathogenic strains. The study emphasizes the importance of continuous monitoring and deeper investigations into commensal bacteria, emphasizing the need for a broader community screening approach to understand resistance mechanisms in the normal microbiome.

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Introduction: Escherichia coli is an opportunistic bacteria that can grow easily, produce toxins, and resist antibiotics. The phenomenon of E. coli developing multidrug resistance is currently the subject of extensive research. The objective of this study was to molecularly identify blaTEM and blaCTX-M genes in multidrug-resistant E. coli found in milk samples from dairy cattle farms in Tulungagung, Indonesia. Material and Methods: One hundred and ten milk samples were collected from 45 dairy cattle farms in Tulungagung, Indonesia. Indole, methyl red, Voges-Proskauer and in citrate tests and triple iron sugar agar tests were used to identify E. coli. Multidrug resistance was determined in isolates through antibiotic sensitivity tests using tetracycline, streptomycin, trimethoprim, chloramphenicol and aztreonam. Extended-spectrum beta lactamase enzyme production was confirmed by double-disc synergy test (DDST). Molecular identification was performed to confirm the blaTEM and blaCTX-M genes. Results: One hundred and one (91.82%) E. coli strains were isolated from the samples. The antibiotic sensitivity test showed four (3.96%) multidrug-resistant (MDR) and one (0.99%) ESBL-positive E. coli by DDST confirmation. There were three (77.78%) blaTEM genes and one (0.99%) blaCTX-M gene discovered in the MDR E. coli isolates using PCR for molecular identification. Conclusion: The findings of the blaTEM and blaCTX-M genes encoding ESBL E. coli in dairy cattle milk in Tulungagung, Indonesia is concerning and argues for prompt action to stop the emergence of antibiotic resistance which has an impact on public health.

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In this study, we sought to profile the abundances and drivers of antibiotic resistance genes in an urban river impacted by combined sewage overflow (CSO) events. Water samples were collected weekly during the summer for two years; then, quantitative PCR was applied to determine the abundance of resistance genes associated with tetracycline, quinolones, and ß-lactam antibiotics. In addition to sampling a CSO-impacted site near the city center, we also sampled a less urban site ~12 km upstream with no proximal sewage inputs. The tetracycline genes tetO and tetW were rarely found upstream, but were common at the CSO-impacted site, suggesting that the primary source was untreated sewage. In contrast, ampC was detected in all samples indicating a more consistent and diffuse source. The two other genes, qnrA and blaTEM, were present in only 40-50% of samples and showed more nuanced spatiotemporal patterns consistent with upstream agricultural inputs. The results of this study highlight the complex sources of ARGs in urban riverine ecosystems, and that interdisciplinary collaborations across diverse groups of stakeholders are necessary to combat the emerging threat of antibiotic resistance through anthropogenic pollution.

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Horizontal gene transfer (HGT) in food matrices has been investigated under conditions that favor gene exchange. However, the major challenge lies in determining the specific conditions pertaining to the adapted microbial pairs associated with the food matrix. HGT is primarily responsible for enhancing the microbial repertoire for the evolution and spread of antimicrobial resistance and is a major target for controlling pathogens of public health concern in food ecosystems. In this study, we investigated Salmonella Heidelberg (SH) and Escherichia coli (EC) regarding gene exchange under conditions mimicking the industrial environment, with the coproducts whey (SL) and chicken juice (CJ). The S. Heidelberg strain was characterized by antibiotic susceptibility standards and PCR to detect the blaTEM gene. A concentration of 0.39 mg/mL was determined to evaluate the anti-conjugation activity of nanostructured lipid nanocarriers (NLCs) of essential oils to mitigate ß-lactam resistance gene transfer. The results showed that the addition of these coproducts promoted an increase of more than 3.5 (whey) and 2.5 (chicken juice) orders of magnitude in the conjugation process (p < 0.01), and NLCs of sage essential oil significantly reduced the conjugation frequency (CF) by 74.90, 90.6, and 124.4 times when compared to the transfers in the absence of coproducts and the presence of SL and CJ, respectively. For NLCs from olibanum essential oil, the decrease was 4.46-fold for conjugations without inhibitors and 3.12- and 11.3-fold in the presence of SL and CJ. NLCs associated with sage and olibanum essential oils effectively control the transfer of antibiotic resistance genes and are a promising alternative for use at industrial levels.

RESUMEN

This preliminary study aimed to detect biological and chemical contaminants in vegetables sold in Sicily for human consumption, assess the spread of antimicrobial-resistant (AMR) strains in these foods, and characterize their antimicrobial-resistance genes. A total of 29 fresh and ready-to-eat samples were analyzed. Microbiological analyses were performed for the detection of Salmonella spp. and the enumeration of Enterococci, Enterobacteriaceae, and Escherichia coli. Antimicrobial resistance was assessed by the Kirby-Bauer method, according to the Clinical and Laboratory Standards Institute guidelines. Pesticides were detected by high-performance liquid chromatography and gas chromatography coupled with mass spectrometry. No samples were contaminated by Salmonella spp., E. coli was detected in 1 sample of fresh lettuce at a low bacterial count (2 log cfu/g). 17.24% of vegetables were contaminated by Enterococci and 65.5% by Enterobacteriaceae (bacterial counts between 1.56 log cfu/g and 5.93 log cfu/g and between 1.6 log cfu/g and 5.48 log cfu/g respectively). From 86.2% of vegetables, 53 AMR strains were isolated, and 10/53 isolates were multidrug resistant. Molecular analysis showed that the blaTEM gene was detected in 12/38 ß-lactam-resistant/intermediate-resistant isolates. Genes conferring tetracycline resistance (tetA, tetB, tetC, tetD, tetW) were detected in 7/10 isolates. The qnrS gene was detected in 1/5 quinolone-resistant isolates, the sulI gene was detected in 1/4 sulfonamide- resistant/intermediate-resistant isolates and the sulIII gene was never detected. Pesticides were detected in 27.3% of samples, all of which were leafy vegetables. Despite the satisfactory hygienic status of samples, the high percentage of AMR bacteria detected stresses the need for an effective monitoring of these foods as well as adequate strategies to counteract the spread of AMR bacteria along the agricultural chain. Also, the chemical contamination of vegetables should not be underestimated, especially considering that leafy vegetables are commonly consumed raw and that no official guidelines about maximum residue limits of pesticides in ready-to-eat vegetables are available.