Linking active rectal mucosa-attached microbiota to host immunity reveals its role in host-pathogenic STEC O157 interactions.

The rectal-anal junction (RAJ) is the major colonization site of Shiga toxin-producing Escherichia coli (STEC) O157 in beef cattle, leading to transmission of this foodborne pathogen from farms to food chains. To date, there is limited understanding regarding whether the mucosa-attached microbiome has a profound impact on host-STEC interactions. In this study, the active RAJ mucosa-attached microbiota and its potential role in host immunity-STEC commensal interactions were investigated using RAJ mucosal biopsies collected from calves orally challenged with two STEC O157 strains with or without functional stx2a (stx2a+ or stx2a-). The results revealed that shifts of microbial diversity, topology, and assembly patterns were subjected to stx2a production post-challenge and Paeniclostridium and Gallibacterium were the keystone taxa for both microbial interactions and assembly. Additional mucosal transcriptome profiling showed stx2a-dependent host immune responses (i.e. B- and T-cell signaling and antigen processing and presentation) post-challenge. Further integrated analysis revealed that mucosa-attached beneficial microbes (i.e. Provotella, Faecalibacterium, and Dorea) interacted with host immune genes pre-challenge to maintain host homeostasis; however, opportunistic pathogenic microbes (i.e. Paeniclostridium) could interact with host immune genes after the STEC O157 colonization and interactions were stx2a-dependent. Furthermore, predicted bacterial functions involved in pathogen (O157 and Paeniclostridium) colonization and metabolism were related to host immunity. These findings suggest that during pathogen colonization, host-microbe interactions could shift from beneficial to opportunistic pathogenic bacteria driven and be dependent on the production of particular virulence factors, highlighting the potential regulatory role of mucosa-attached microbiota in affecting pathogen-commensal host interactions in calves with STEC O157 infection.

Antimicrobial resistance among indicator Enterococcus faecium and Escherichia coli in Swedish pig farms.

Monitoring the use of antimicrobials and the emergence of resistance in animals and people is important for the control of antimicrobial resistance, and for establishing sustainable and effective disease management practices. In this study, we used Enterococcus spp. and Escherichia coli as indicator species to investigate antimicrobial susceptibility patterns and how these change over time, on ten Swedish pig farms. Indoor environmental sock sampling was performed once a month during the entire production cycle of one batch of pigs on each farm, resulting in 60 samples collected in total. Selective culture for E. coli and Enterococcus spp. resulted in 122 isolates of E. coli, 74 isolates of E. faecium, but no isolates of E. faecalis. Microdilution was used to determine minimum inhibitory concentrations for twelve antimicrobial substances in E. coli and fifteen substances in E. faecium. The overall prevalence of resistance was low. Among the E. coli isolates, the proportions non-wild type (resistant, NWT) isolates were as follows: azithromycin and amikacin 1% (n = 1), trimethoprim and sulfamethoxazole 2% (n = 3), ampicillin 6% (n = 7) and tetracycline 9% (n = 11). Among the E. faecium isolates, the NWT proportions were: teicoplanin, linezolid and gentamicin 1% (n = 1), daptomycin 3% (n = 2), erythromycin 26% (n = 19), tetracycline 27% (n = 20), quinupristin/dalfopristin 58% (n = 42). The resistance patterns differed between the farms, likely due to different antimicrobial use, biosecurity measures and source of the animals. The NWT prevalence among E. coli decreased over time, whereas no similar trend could be observed in E. faecium. The results of the current study illustrate the complex factors affecting the antimicrobial resistance patterns observed on each farm, indicating that specific practices and risk factors have an impact on the prevalence and type of antimicrobial resistance. Further studies of the farm environments in combination with antimicrobial use and other risk factor data are needed to elucidate the multifaceted drivers of antimicrobial resistance development on livestock farms.

Characterization of antimicrobial resistance profiles in Escherichia coli isolated from captive mammals in Ecuador.

BACKGROUND: This study focuses on the AMR profiles in E. coli isolated from captive mammals at EcoZoo San Martín, Baños de Agua Santa, Ecuador, highlighting the role of wildlife as reservoirs of resistant bacteria. AIMS: The aim of this research is to investigate the antimicrobial resistance profiles of E. coli strains isolated from various species of captive mammals, emphasizing the potential zoonotic risks and the necessity for integrated AMR management strategies. MATERIALS & METHODS: A total of 189 fecal samples were collected from 70 mammals across 27 species. These samples were screened for E. coli, resulting in 90 identified strains. The resistance profiles of these strains to 16 antibiotics, including 10 ß-lactams and 6 non-ß-lactams, were determined using the disk diffusion method. Additionally, the presence of Extended-Spectrum Beta-Lactamase (ESBL) genes and other resistance genes was analyzed using PCR. RESULTS: Significant resistance was observed, with 52.22% of isolates resistant to ampicillin, 42.22% to ceftriaxone and cefuroxime, and 27.78% identified as ESBL-producing E. coli. Multiresistance (resistance to more than three antibiotic groups) was found in 35.56% of isolates. Carnivorous and omnivorous animals, particularly those with prior antibiotic treatments, were more likely to harbor resistant strains. DISCUSSION: These findings underscore the role of captive mammals as indicators of environmental AMR. The high prevalence of resistant E. coli in these animals suggests that zoos could be significant reservoirs for the spread of antibiotic-resistant bacteria. The results align with other studies showing that diet and antibiotic treatment history influence resistance profiles. CONCLUSION: The study highlights the need for an integrated approach involving veterinary care, habitat management, and public awareness to prevent captive wildlife from becoming reservoirs of antibiotic-resistant bacteria. Improved waste management practices and responsible antibiotic use are crucial to mitigate the risks of AMR in zoo environments and reduce zoonotic threats.

Comparative genomics of quinolone-resistant Escherichia coli from broilers and humans in Norway.

BACKGROUND: The usage of fluoroquinolones in Norwegian livestock production is very low, including in broiler production. Historically, quinolone-resistant Escherichia coli (QREC) isolated from Norwegian production animals rarely occur. However, with the introduction of a selective screening method for QREC in the Norwegian monitoring programme for antimicrobial resistance in the veterinary sector in 2014; 89.5% of broiler caecal samples and 70.7% of broiler meat samples were positive. This triggered the concern if there could be possible links between broiler and human reservoirs of QREC. We are addressing this by characterizing genomes of QREC from humans (healthy carriers and patients) and broiler isolates (meat and caecum). RESULTS: The most frequent mechanism for quinolone resistance in both broiler and human E. coli isolates were mutations in the chromosomally located gyrA and parC genes, although plasmid mediated quinolone resistance (PMQR) was also identified. There was some relatedness of the isolates within human and broiler groups, but little between these two groups. Further, some overlap was seen for isolates with the same sequence type isolated from broiler and humans, but overall, the SNP distance was high. CONCLUSION: Based on data from this study, QREC from broiler makes a limited contribution to the incidence of QREC in humans in Norway.

Population structure and antibiotic resistance of swine extraintestinal pathogenic Escherichia coli from China.

Extraintestinal Pathogenic Escherichia coli (ExPEC) pose a significant threat to human and animal health. However, the diversity and antibiotic resistance of animal ExPEC, and their connection to human infections, remain largely unexplored. The study performs large-scale genome sequencing and antibiotic resistance testing of 499 swine-derived ExPEC isolates from China. Results show swine ExPEC are phylogenetically diverse, with over 80% belonging to phylogroups B1 and A. Importantly, 15 swine ExPEC isolates exhibit genetic relatedness to human-origin E. coli strains. Additionally, 49 strains harbor toxins typical of enteric E. coli pathotypes, implying hybrid pathotypes. Notably, 97% of the total strains are multidrug resistant, including resistance to critical human drugs like third- and fourth-generation cephalosporins. Correspondingly, genomic analysis unveils prevalent antibiotic resistance genes (ARGs), often associated with co-transfer mechanisms. Furthermore, analysis of 20 complete genomes illuminates the transmission pathways of ARGs within swine ExPEC and to human pathogens. For example, the transmission of plasmids co-harboring fosA3, blaCTX-M-14, and mcr-1 genes between swine ExPEC and human-origin Salmonella enterica is observed. These findings underscore the importance of monitoring and controlling ExPEC infections in animals, as they can serve as a reservoir of ARGs with the potential to affect human health or even be the origin of pathogens infecting humans.

Isolation of multi-drug-resistant strains of Escherichia coli from faecal samples of dogs and cats from Harare, Zimbabwe.

BACKGROUND: The escalation of antimicrobial resistance (AMR) in recent years has been of major public health concern globally. Escherichia coli are amongst the bacteria that have been targeted for AMR surveillance due to their ability to cause infection in both animals and humans. Their propensity to produce extended spectrum beta-lactamases further complicates the choices of treatment regimens. OBJECTIVES: To investigate the prevalence of antimicrobial-resistance in E. coli strains isolated from faecal samples of dogs and cats from selected veterinary surgeries and animal shelters from Harare, Zimbabwe. MATERIALS AND METHODS: A cross-sectional study was carried out to select animals by a systematic random procedure. Faecal samples were collected for culture and isolation of E. coli. Their susceptibility to antimicrobial drugs was assessed using the disc diffusion method. RESULTS: A total of 95% (133/140) of the samples from cats (n = 40) and dogs (n = 93) yielded E. coli. Resistance was recorded for ampicillin (45.9%), trimethoprim-sulphamethoxazole (44.4%), nalidixic acid (29.3%), ceftazidime (15.8%) and azithromycin (12.8%), but not for gentamicin and imipenem. A total of 18% of the isolates were multi-drug-resistant where resistance to nalidixic acid, ampicillin and trimethoprim-sulphamethoxazole predominated. CONCLUSION: We observed relatively high AMR of E. coli strains against ampicillin. The isolation of multi-drug-resistant strains of E. coli may signal the dissemination of resistance genes in the ecosystem of these bacteria which may have a public health impact.

Evaluation of the efficacy of a new inactivated vaccine against Staphylococcus aureus, Echerchia coli and Mycoplasma bovis mastitis in cows.

Staphylococcus aureus, Escherichia coli and Mycoplasma bovis are the most commonly isolated mastitis pathogens. The aim of this study was to evaluate the efficacy of a new mixed vaccine against mastitis caused by  Staphylococcus aureus, Escherichia coli, and Mycoplasma bovis. For this purpose, a mixed inactivated vaccine was administered subcutaneously to 24 heifers as one dose (2 mL) on the 45th day before birth and the second dose 21 days later. In 9 heifers, 2 mL of PBS was administered as placebo instead of vaccine. Then, heifers were divided into 3 groups as 7 vaccinated and 3 unvaccinated animals. Staphylococcus aureus, Escherichia coli, and Mycoplasma bovis were administered to the groups through intramammary route. Three vaccinated heifers were considered the common control without bacteria in all groups. The parameters considered to assess the effect of vaccination were clinical findings, bacterial count in milk, somatic cell count, and antibody titers. Clinical signs were observed only in the unvaccinated placebo group. Bacteria count and somatic cell count in milk increased in vaccinated and unvaccinated heifers. However, this increase was less in vaccinated animals and gradually returned to the normal level. In the unvaccinated heifers, it was ever high. Serum antibody titers were measured before and after vaccination. Antibody titers were high in vaccinated heifers after vaccination and were negative in unvaccinated heifers. In conclusion, the mixed vaccine had beneficial effect against Staphylococcus aureus, Escherichia coli, and Mycoplasma bovis mastitis and stimulated the immune response of vaccinated heifers.

Correlation analysis of whole genome sequencing of a pathogenic Escherichia coli strain of Inner Mongolian origin.

Anal swabs of 1-month-old Holstein calves with diarrhea were collected from an intensive cattle farm, and a highly pathogenic Escherichia coli strain was obtained by isolation and purification. To study the virulence and resistance genes of pathogenic E. coli that cause diarrhea in calves, a strain of E. coli E12 isolated from calf diarrhea samples was used as experimental material in this experiment, and the virulence of the E12 strain were identified by the mouse infection test, and the whole genome map of the E12 strain were obtained by whole-genome sequencing and analyzed for genome characterization. The results showed that the lethality of strain E12 was 100%, the total length of E12-encoded genes was 4,294,530 bp, Cluster of Orthologous Groups of proteins (COG) annotated to 4,194 functional genes, and the virulence genes of sequenced strain E12 were compared with the virulence genes of sequenced strain E12 from the Virulence Factors of Pathogenic Bacteria (VFDB), which contained a total of 366 virulence genes in sequenced strain E12. The analysis of virulence genes of E12 revealed a total of 52 virulence genes in the iron transferrin system, 56 virulence genes in the secretory system, 41 virulence genes in bacterial toxins, and a total of 217 virulence genes in the Adhesin and Invasins group. The antibiotic resistance genes of sequenced strain E12 were identified through the Antibiotic Resistance Genes Database (ARDB) and Comprehensive Antibiotic Research Database, and it was found that its chromosome and plasmid included a total of 127 antibiotic resistance genes in four classes, and that E12 carried 71 genes related to the antibiotic efflux pumps, 36 genes related to antibiotic inactivation, and 14 antibiotic target alteration and reduced penetration into antibiotics, and 6 antibiotic resistance genes, and the resistance phenotypes were consistent with the genotypes. The pathogenic E. coli that causes diarrhea in calves on this ranch contains a large number of virulence and resistance genes. The results provide a theoretical basis for the prevention and treatment of diarrhea and other diseases caused by E. coli disease.

The phylogroups and antibiotic susceptibilities of Escherichia coli isolates from the feces of Anatolian Ground Squirrels (Spermophilus xanthoprymnus).

The current study was conducted to determine the phylogroups and antibiotic susceptibilities of Escherichia coli isolates recovered from fecal samples of Anatolian Ground Squirrels (Spermophilus xanthoprymnus) and to examine the relationship between them. Eighty-two E. coli isolates obtained from 150 fecal samples were investigated. The quadruplex polymerase chain reaction (PCR), phylogroup C-, and E-specific mPCR were subjected to phylogenetic typing of the isolates. The susceptibilities to fifteen antibiotics of the isolates were detected by the disk diffusion method. In the result of phylogenetic typing, phylogroup B2 was most predominant (58.6 %), followed by B1 (25.6 %), E (8.5 %), C (4.9 %), and D (2.4 %). The phylogroup A, F, and Escherichia clades were not detected. The antibiotic susceptibility test revealed that 59.8 % (49/82) and 19.5 % (16/82) of E. coli isolates were resistant to at least one antibiotic and multidrug-resistant (MDR), respectively. Twenty-six (31.7 %), 19 (23.2 %), 11 (13.4 %), and 10 (12.2 %) of the isolates were found to be resistant to gentamicin, tetracycline, amoxicillin-clavulanic acid, and cefoxitin. Of the 49 E. coli isolates that were found to be resistant to any antibiotic analyzed, 30, 13, 4, and 2 were located in phylogroup B2, B1, E, and D, respectively. MDR isolates were mostly located in both phylogroup B1 (31.3 %) and B2 (31.3 %). In conclusion, data from the current study suggest that the isolates may potentially have pathogenic properties, since the majority (69.5 %) of E. coli isolates from fecal samples of Spermophilus xanthoprymnus were located in the pathogenic phylogroup and resistance to various antibiotics was detected.

Designing a multiepitope mRNA-based vaccine against enterotoxigenic Escherichia coli infection in calves: Immunoinformatics and molecular modeling approach.

Background: Escherichia coli is one of the serious pathogens causing various infections in the animal field, such as neonatal calf diarrhea, which is responsible for mortality associated with diarrhea during the first days of life. Aim: Current work is aimed at designing an effective and safe multiepitope vaccine candidate against E. coli infection in calves based on the fimbrial protein K99 of Enterotoxigenic E. coli (ETEC) and Immuno-informatics. Methods: A conserved sequence of K99 protein was generated, and then highly antigenic, nonallergic, and overlapped epitopes were used to construct a multiepitope vaccine. Five THL, six MHC II, and four beta cell epitopes were targeted to create the candidate. The candidate vaccine was produced utilizing 15 epitopes and three types of linkers, two types of untranslated region (UTR) human hemoglobin subunit beta (HBB), UTR beta-globin (Rabb), and RpfE protein as an immunomodulation adjuvant. Results: Immuno-informatics analysis of the constructed protein showed that the protein was antigenic (antigenic score of 0.8841), stable, nonallergen, and soluble. Furthermore, the Immuno-informatics and physiochemical analysis of the constructed protein showed a stable, nonallergic, soluble, hydrophilic, and acidic PI (isoelectric point). of 9.34. Docking of the candidate vaccine with the toll-like receptor TLR3 was performed, and results showed a strong interaction between the immune receptor and the vaccine. Finally, the expression efficiency of the construct in E. coli was estimated via computational cloning of the vaccine sequence into Pet28a. Conclusion: Results of immunoinformatics and in silico approaches reveal that the designed vaccine is antigenic, stable, and able to bind to the immune cell receptors. Our results interpret the proposed multiepitope mRNA vaccine as a good preventive option against E. coli infection in calves.

The roles of Braun Lipoprotein in inducing tolerance of bovine endometrium infected by Escherichia coli.

Escherichia coli (E. coli), a Gram-negative bacterium, is the primary pathogen responsible for endometritis in dairy cattle. The outer membrane components of E. coli, namely lipopolysaccharide (LPS) and bacterial lipoprotein, have the capacity to trigger the host's innate immune response through pattern recognition receptors (PRRs). Tolerance to bacterial cell wall components, including LPS, may play a crucial role as an essential regulatory mechanism during bacterial infection. However, the precise role of Braun lipoprotein (BLP) tolerance in E. coli-induced endometritis in dairy cattle remains unclear. In this study, we aimed to investigate the impact of BLP on the regulation of E. coli infection-induced endometritis in dairy cattle. The presence of BLP was found to diminish the expression and release of proinflammatory cytokines (IL-8 and IL-6), while concurrently promoting the expression and release of the anti-inflammatory cytokine IL-10 in endometrial epithelial cells (EECs). Furthermore, BLP demonstrated the ability to impede the activation of MAPK (ERK and p38) and NF-κB (p65) signaling pathways, while simultaneously enhancing signaling through the STAT3 pathway in EECs. Notably, BLP exhibited a dual role, acting both as an activator of TLR2 and as a regulator of TLR2 activation in LPS- and E. coli-treated EECs. In E. coli-infected endometrial explants, the presence of BLP was noted to decrease the release of proinflammatory cytokines and the expression of HMGB1, while simultaneously enhancing the release of anti-inflammatory cytokines. Collectively, our findings provide evidence that the bacterial component BLP plays a protective role in E. coli-induced endometritis in dairy cattle.

Multidrug resistance of Escherichia coli isolated from the urinary bladder of dogs and cats with suspected urinary tract infections.

INTRODUCTION AND OBJECTIVE: Escherichia coli is one of the most common bacteria isolated from urine samples collected from dogs and cats with urinary tract infection (UTI). Uncomplicated UTIs in dogs and cats can be treated with short courses of first-line antimicrobial drugs, e.g. amoxicillin, amoxicillin with clavulanic acid, or trimethoprim/sulfonamide. Recurrent or complicated UTIs often require long-term treatment with broad-spectrum antibiotics. However, the choice of drug should be based on antimicrobial susceptibility. MATERIAL AND METHODS: Between March - September 2022, E. coli isolates cultured from the urine of 66 dogs and 41 cats with UTI symptoms were tested for antimicrobial resistance by using Minimum Inhibitory Concentration (MIC). Antimicrobial susceptibility was tested for ampicillin, ampicillin/sulbactam, cefazolin, cefuroxime, aztreonam, gentamycin, amikacin, colistin, trimethoprim/sulfamethoxazole, ciprofloxacin, chloramphenicol and tetracycline. RESULTS: The highest prevalence of resistance was documented for ampicillin (68% in dogs, 100% in cats) and ampicillin with sulbactam (59% in dogs, 54% in cats). The most common antimicrobial resistance patterns of E. coli were ampicillin alone (12 isolates, 29.3% in cats) and beta-lactams, including aztreonam (14 isolates, 21.2% in dogs). CONCLUSIONS: High resistance to aztreonam (61% and 32% of isolates from dogs and cats, respectively), other beta-lactams, and fluoroquinolones should cause be alarm due to zoonotic potential and cross-transmission of antimicrobial-resistant microorganisms between animals and humans.

Virulent-MDR-ESBL E. coli and Klebsiella pneumoniae report from North Sinai calves diarrhea and in vitro antimicrobial by Moringa oleifera.

The health of calves has a significant impact on the production of cows and livestock. Some desert plants have pharmacological importance, as they can be used to reduce antibiotic resistance. Our hypothesis is designed to detect Virulent- Multidrug-Resistant and Extended- spectrum Beta- lactamase Enterobacteriaceae (Virulent-MDR-ESBL Enterobacteriaceae and to determine whether Moringa oleifera has antibacterial activity against the detected isolates. A total of 39 Enterobacteriaceae isolates from 28 diarrheic samples were collected from calves aged between 20 days and 20 months from 3 different flocks in North Sinai, Sahl-Eltina region, Egypt. E.coli 46% (18/39), O157 13% (5/39), Klebsiella pneumoniae 41% (16/39). MDR members accounted for 87%, while ESBL isolates accounted for 43%. The antibacterial activity is represented by microdilution. Minimum inhibition concentration (MIC) for the methanol extract of Moringa oleifera ranged from 2.5,5,10, and 25mg/ ml among E.coli isolates, and O157 was susceptible to (2.5mg/ ml), Klebsiella pneumoniae isolates were susceptible to (5-50mg/ ml). Analysis of the methanol extract revealed that ferulic acid was the dominant phenolic compound with a concentration of 29,832 parts per million (ppm). In silico docking study expected the active site of ferulic acid to act on the tyrosine bacterial enzyme through Pi-alkyl, Pi-anion, Carbon hydrogen bonds, and extra ionic attractive interactions with copper ions which can stabilize ferulic acid inside the targeted pocket Diverse virulent gene profiles were observed in E. coli. The Shiga toxin-producing Escherichia coli (STEC) was reported in 83% of the isolated E. coli, while the DNA gyrase (gyrA) was harbored in 100% of Klebsiella pneumoniae isolates. Various profiles of antibiotic resistance genes for both E. coli and Klebsiella pneumoniae isolates were distinguished. blaTEM genes were detected in 99% of E. coli and 100% of Klebsiella pneumoniae. Sequence analysis for E. coli strain DRC-North Sinai-Eg was placed in accession numbers (OP955786) for the Shiga toxin 2 gene (Stx2A), (OP997748) and (OP997749) for the Adhesion to host cell gene (Eae). For the hemolysine gene (hylA), the accession number was (OP946183). Klebsiella pneumoniae strain DRC-North Sinai-Eg was placed in (OP946180) for (gyrA). This study has proven the broad range of Moringa oliefera's antibacterial effects in vitro against the virulent-MDR- ESBL E. coli and Klebsiella pneumoniae isolated from North Sinai calves diarrhea. These are congruent with the disability effect on bacterial tyrosinase enzyme through docking study therefore, we recommend the usage of this desert plant as a prospective feed additive, we endorse this as an antibacterial new insight natural source and for the medication of considered pathogens with zoonotic impacts.

Reversal of gentamicin sulfate resistance in avian pathogenic Escherichia coli by matrine combined with berberine hydrochloride.

In recent years, the evolution of antibiotic resistance has led to the inefficacy of several antibiotics, and the reverse of resistance was a novel method to solve this problem. We previously demonstrated that matrine (Mat) and berberine hydrochloride (Ber) had a synergistic effect against multidrug-resistant Escherichia coli (MDREC). This study aimed to demonstrate the effect of Mat combined with Ber in reversing the resistance of MDREC. The MDREC was sequenced passaged in the presence of Mat, Ber, and a combination of Mat and Ber, which did not affect its growth. The reverse rate was up to 39.67% after MDREC exposed to Mat + Ber for 15 days. The strain that reversed resistance was named drug resistance reversed E. coli (DRREC) and its resistance to ampicillin, streptomycin, gentamicin, and tetracycline was reversed. The MIC of Gentamicin Sulfate (GS) against DRREC decreased 128-fold to 0.63 µg/mL, and it was stable within 20 generations. Furthermore, the susceptible phenotype of DRREC remained stable within 20 generations, as well. The LD50 of DRREC for chickens was 8.69 × 109 CFU/mL. qRT-PCR assays revealed that the transcript levels of antibiotic-resistant genes and virulence genes in the DRREC strain were significantly lower than that in the MDREC strain (P < 0.05). In addition, GS decreased the death, decreased the bacterial loading in organs, alleviated the injury of the spleen and liver, and decreased the cytokine levels in the chickens infected by the DRREC strain. In contrast, the therapeutic effect of GS in chickens infected with MDREC was not as evident. These findings suggest that the combination of Mat and Ber has potential for reversing resistance to MDREC.

Whole genome sequencing analysis on antibiotic-resistant Escherichia coli isolated from pig farms in Banten Province, Indonesia.

IMPORTANCE: The emergence and rapid increase in the incidence of multidrug-resistant (MDR) bacteria in pig farms has become a serious concern and reduced the choice of effective antibiotics. OBJECTIVE: This study analyzed the phylogenetics and diversity of antibiotic resistance genes (ARGs) and molecularly identified the source of ARGs in antibiotic-resistant Escherichia coli isolated from pig farms in Banten Province, Indonesia. METHODS: Forty-four antibiotic-resistant E. coli isolates from fecal samples from 44 pig farms in Banten Province, Indonesia, were used as samples. The samples were categorized into 14 clusters. Sequencing was performed using the Oxford Nanopore Technologies MinION platform, with barcoding before sequencing with Nanopore Rapid sequencing gDNA-barcoding (SQK-RBK110.96) according to manufacturing procedures. ARG detection was conducted using ResFinder, and the plasmid replicon was determined using PlasmidFinder. RESULTS: Three phylogenetic leaves of E. coli were identified in the pig farming cluster in Banten Province. The E. coli isolates exhibited potential resistance to nine classes of antibiotics. Fifty-one ARGs were identified across all isolates, with each cluster carrying a minimum of 10 ARGs. The ant(3'')-Ia and qnrS1 genes were present in all isolates. ARGs in the E. coli pig farming cluster originated mainly from plasmids, accounting for an average of 89.4%. CONCLUSIONS AND RELEVANCE: The elevated potential for MDR events, coupled with the dominance of ARGs originating from plasmids, increases the risk of ARG spread among bacterial populations in animals, humans, and the environment.

Widespread prevalence of plasmid-mediated blaCTX-M type extended-spectrum beta-lactamase Escherichia coli in backyard broiler production systems in the United States.

Extended-spectrum beta-lactamase (ESBL) Escherichia coli (E. coli) is an emerging pathogen of high concern given its resistance to extended-spectrum cephalosporins. Broiler chicken, which is the number one consumed meat in the United States and worldwide, can be a reservoir of ESBL E. coli. Backyard poultry ownership is on the rise in the United States, yet there is little research investigating prevalence of ESBL E. coli in this setting. This study aims to identify the prevalence and antimicrobial resistance profiles (phenotypically and genotypically) of ESBL E. coli in some backyard and commercial broiler farms in the U.S. For this study ten backyard and ten commercial farms were visited at three time-points across flock production. Fecal (n = 10), litter/compost (n = 5), soil (n = 5), and swabs of feeders and waterers (n = 6) were collected at each visit and processed for E. coli. Assessment of ESBL phenotype was determined through using disk diffusion with 3rd generation cephalosporins, cefotaxime and ceftazidime, and that with clavulanic acid. Broth microdilution and whole genome sequencing were used to investigate both phenotypic and genotypic resistance profiles, respectively. ESBL E. coli was more prevalent in backyard farms with 12.95% of samples testing positive whereas 0.77% of commercial farm samples were positive. All isolates contained a blaCTX-M gene, the dominant variant being blaCTX-M-1, and its presence was entirely due to plasmids. Our study confirms concerns of growing resistance to fourth generation cephalosporin, cefepime, as roughly half (51.4%) of all isolates were found to be susceptible dose-dependent and few were resistant. Resistance to non-beta lactams, gentamicin and ciprofloxacin, was also detected in our samples. Our study identifies prevalence of blaCTX-M type ESBL E. coli in U.S. backyard broiler farms, emphasizing the need for interventions for food and production safety.

VirBR, a transcription regulator, promotes IncX3 plasmid transmission, and persistence of blaNDM-5 in zoonotic bacteria.

IncX3 plasmids carrying the New Delhi metallo-ß-lactamase-encoding gene, blaNDM-5, are rapidly spreading globally in both humans and animals. Given that carbapenems are listed on the WHO AWaRe watch group and are prohibited for use in animals, the drivers for the successful dissemination of Carbapenem-Resistant Enterobacterales (CRE) carrying blaNDM-5-IncX3 plasmids still remain unknown. We observe that E. coli carrying blaNDM-5-IncX3 can persist in chicken intestines either under the administration of amoxicillin, one of the largest veterinary ß-lactams used in livestock, or without any antibiotic pressure. We therefore characterise the blaNDM-5-IncX3 plasmid and identify a transcription regulator, VirBR, that binds to the promoter of the regulator gene actX enhancing the transcription of Type IV secretion systems (T4SS); thereby, promoting conjugation of IncX3 plasmids, increasing pili adhesion capacity and enhancing the colonisation of blaNDM-5-IncX3 transconjugants in animal digestive tracts. Our mechanistic and in-vivo studies identify VirBR as a major factor in the successful spread of blaNDM-5-IncX3 across one-health AMR sectors. Furthermore, VirBR enhances the plasmid conjugation and T4SS expression by the presence of copper and zinc ions, thereby having profound ramifications on the use of universal animal feeds.

Paeoniflorin protects chicken against APEC-induced acute lung injury by affecting the endocannabinoid system and inhibiting the PI3K/AKT and NF-κB signaling pathways.

Avian pathogenic Escherichia coli (APEC) is the causative agent of chicken colibacillosis. Paeoniflorin, a natural ingredient extracted from Paeonia lactiflora, has a variety of pharmacological effects including anti-inflammatory and immunomodulatory. However, its effects and mechanism in APEC-induced acute lung injury (ALI) in chicken is not clear. The aim of this study was to investigate the protective effect of paeoniflorin on APEC-induced ALI and its possible mechanism. Paeoniflorin (25, 50, and 100 mg/kg) was administered by gavage for 5 d starting at 9 d of age and the chicken were infected with APEC by intraperitoneal injection at 12 d of age. The tissues were collected after APEC infection for 36 h for analysis. The results showed that paeoniflorin significantly alleviated the symptoms, increased the survival rate and body weight gain of APEC-infected chicken, and improved the histopathological damages, and reduced APEC loads in lung tissues. In addition, paeoniflorin restored the gene expression of ZO-1, Occludin and Claudin-3 during APEC infection. Moreover, paeoniflorin pretreatment significantly affected the endocannabinoid system (ECs) by increasing DAGL, decreasing MAGL, increasing secretion of 2-AG. Then, paeoniflorin significantly decreased the secretion of IL-1ß, IL-6 and TNF-α in lung tissues, and decreased the mRNA expression of CXCL8, CXCL12, CCL1, CCL5, and CCL17. In addition, paeoniflorin significantly reduced the phosphorylation levels of PI3K, AKT, P65, and IκB. In summary, we found that paeoniflorin inhibited APEC-induced ALI, and its mechanism may be through affecting ECs and inhibiting the activation of PI3K/AKT and NF-κB signaling pathways, which provides a new idea for the prevention and treatment of chicken colibacillosis.

Pathogenic profile and antimicrobial resistance of Escherichia coli, Escherichia marmotae and Escherichia ruysiae detected from hunted wild boars in Sardinia (Italy).

The objective of this study was to evaluate the occurrence of E. coli in hunted wild boars in Sardinia (Italy) and to further characterize the isolates with Whole Genome Sequencing to assess the genetic relatedness and the presence of virulence and antimicrobial resistance (AMR) genes. Samples were taken from 66 wild boars between 2020 and 2022 slaughtered in five hunting houses. A total of 181 samples were tested, including 66 samples from mesenteric lymph nodes, 66 samples from colon content and 49 samples from carcass surface. Isolates referable to Escherichia species were detected in all of the wild boars sampled. On a selection of 61 isolates, sequencing was conducted and antimicrobial susceptibility was tested. Among these, three isolates were confirmed to be two Escherichia marmotae (cryptic clade V) and one Escherichia ruysiae (cryptic clade III). E. coli pathotypes identified were UPEC (13 %), ExPEC-UPEC (5.6 %) and ETEC (3.7 %). Moreover, 3/6 E. marmotae isolates had typical ExPEC genes. Genetic similarity was observed in isolates collected from animals slaughtered in the same hunting house; this suggests epidemiological links deriving from the presence of animals infected with closely related strains or the result of cross-contamination. Antimicrobial resistance genes were detected in three non-pathogenic E. coli isolates: one isolate had sul2, tet(B), aph(6)-ld and aph(3″)-lb resistance genes and two had the fosA7 gene. This study confirmed that wild boars can act as reservoirs and spreaders of pathogenic Escherichia species and it provides information for future comparative genomic analysis in wildlife. Although isolates showed a limited resistome, the detection of resistance in non-pathogenic isolates underlines the need to monitor antimicrobial resistance in the wild boar population. To the best of our knowledge, this is the first detection of E. mamotae and E. ruysiae isolates in wild boars in Italy and the presence of this pathogen in wildlife and livestock need to be investigated further.

Pediatric urinary tract infections caused by poultry-associated Escherichia coli.

Escherichia coli is the leading cause of urinary tract infections (UTIs) in children and adults. The gastrointestinal tract is the primary reservoir of uropathogenic E. coli, which can be acquired from a variety of environmental exposures, including retail meat. In the current study, we used a novel statistical-genomic approach to estimate the proportion of pediatric UTIs caused by foodborne zoonotic E. coli strains. E. coli urine isolates were collected from DC residents aged 2 months to 17 years from the Children's National Medical Center Laboratory, 2013-2014. During the same period, E. coli isolates were collected from retail poultry products purchased from 15 sites throughout DC. A total of 52 urine and 56 poultry isolates underwent whole-genome sequencing, core genome phylogenetic analysis, and host-origin prediction by a Bayesian latent class model that incorporated data on the presence of mobile genetic elements (MGEs) among E. coli isolates from multiple vertebrate hosts. A total of 56 multilocus sequence types were identified among the isolates. Five sequence types-ST10, ST38, ST69, ST117, and ST131-were observed among both urine and poultry isolates. Using the Bayesian latent class model, we estimated that 19% (10/52) of the clinical E. coli isolates in our population were foodborne zoonotic strains. These data suggest that a substantial portion of pediatric UTIs in the Washington DC region may be caused by E. coli strains originating in food animals and likely transmitted via contaminated poultry meat.IMPORTANCEEscherichia coli UTIs are a heavy public health burden and can have long-term negative health consequences for pediatric patients. E. coli has an extremely broad host range, including humans, chickens, turkeys, pigs, and cattle. E. coli derived from food animals is a frequent contaminant of retail meat products, but little is known about the risk these strains pose to pediatric populations. Quantifying the proportion of pediatric UTIs caused by food-animal-derived E. coli, characterizing the highest-risk strains, and identifying their primary reservoir species could inform novel intervention strategies to reduce UTI burden in this vulnerable population. Our results suggest that retail poultry meat may be an important vehicle for pediatric exposure to zoonotic E. coli strains capable of causing UTIs. Vaccinating poultry against the highest-risk strains could potentially reduce poultry colonization, poultry meat contamination, and downstream pediatric infections.