Extraintestinal Pathogenic Escherichia Coli Research Articles

Public health concern of antimicrobial resistance and virulence determinants in E. coli isolates from oysters in Egypt

The emergence of critical-priority E. coli, carrying a wide array of resistance and virulence factors through food sources, poses a significant challenge to public health. This study aimed to investigate the potential role of oysters sold in Egypt as a source for E. coli, identify their resistance and virulence-associated gene profiles, and assess associated zoonotic risks. A total of 33 pooled fresh oyster samples were obtained from various retail fish markets in Egypt and examined bacteriologically for the presence of E. coli. Antimicrobial resistance was performed by the disk-diffusion method, and the multiple antibiotic resistance index (MAR) was calculated. All isolates were screened for extended-spectrum beta-lactamase (ESBL) (blaTEM, blaSHV, blaCTX−M, and blaOXA−1), plasmid-mediated AmpC blaCMY−2, and carbapenemases (blaKPC, blaNDM, blaVIM, and blaOXA−48) genes by Polymerase chain reaction. Moreover, the presence of virulence-encoding genes was investigated. The virulent MDR strains were clustered using R with the pheatmap package. The prevalence of E. coli was 72.7% (24 out of 33), with 66.7% of the isolates classified as multi-drug resistant, and 75% exhibited MAR values exceeding the 0.2 threshold. Different antimicrobial sensitivity phenotypes and genotype profiles were identified in E. coli isolates. The most prevalent gene detected among all isolates was blaTEM (22/24, 91.7%). Notably, all non-ESBL producers were positive for blaCMY2. Carbapenem-resistant and carbapenem-intermediate strains were carbapenemase producers, with the predominance of the blaKPC gene (11/24, 45.8%). Remarkably, twelve out of sixteen virulence genes were identified, with papC (21/24, 87.5%) and sfa (16/24, 66.7%) genes being the most prevalent. Most isolates carry virulence genes primarily associated with extra-intestinal pathogenic E. coli (ExPEC) (87.5%) and enteropathogenic (EPEC) (70.8%) pathotypes. Four E. coli isolates exhibit cluster patterns. This study provides the first insight into the emergence of virulent MDR E. coli among oysters in Egypt. It underscores the potential role of oysters as a source for disseminating these strains within aquatic ecosystems, presenting a possible threat to public health.

Selective colonization of microplastics, wood and glass by antimicrobial-resistant and pathogenic bacteria.

The Plastisphere is a novel niche whereby microbial communities attach to plastic debris, including microplastics. These communities can be distinct from those found in the surrounding environment or those attached to natural substrates and may serve as a reservoir of both pathogenic and antimicrobial-resistant (AMR) bacteria. Owing to the frequent omission of appropriate comparator particles (e.g. natural substrates) in previous studies, there is a lack of empirical evidence supporting the unique risks posed by microplastics in terms of enrichment and spread of AMR pathogens. This study investigated selective colonization by a sewage community on environmentally sampled microplastics with three different polymers, sources and morphologies, alongside natural substrate (wood), inert substrate (glass) and free-living/planktonic community controls. Culture and molecular methods (quantitative polymerase chain reaction (qPCR)) were used to ascertain phenotypic and genotypic AMR prevalence, respectively, and multiplex colony PCR was used to identify extra-intestinal pathogenic Escherichia coli (ExPECs). From this, polystyrene and wood particles were found to significantly enrich AMR bacteria, whereas sewage-sourced bio-beads significantly enriched ExPECs. Polystyrene and wood were the least smooth particles, and so the importance of particle roughness on AMR prevalence was then directly investigated by comparing the colonization of virgin vs artificially weathered polyethylene particles. Surface weathering did not have a significant effect on the AMR prevalence of colonized particles. Our results suggest that the colonization of plastic and non-plastic particles by AMR and pathogenic bacteria may be enhanced by substrate-specific traits.

Characterization of Extraintestinal Pathogenic Escherichia coli Strains Causing Canine Pneumonia in China: Antibiotic Resistance, Virulence Genes, and Sequence Typing.

To determine the etiological agents responsible for acute pneumonia in puppies in China, this study utilized bronchoalveolar lavage (BAL) fluid extraction to enable the isolation, culture, biochemical identification, and 16S rRNA PCR amplification of the pathogens. Following preliminary identification, the pathogens underwent analysis for antibiotic resistance phenotypes and resistance genes. Additionally, the study examined the presence of virulence genes, conducted multilocus sequence typing (MLST), and performed whole-genome sequencing (WGS). The findings revealed that all four isolated pathogens were characterized as extraintestinal pathogenic Escherichia coli (ExPEC). The examined ExPEC strains demonstrated resistance to cephalosporins, tetracyclines, and penicillins, while remaining susceptible to aminoglycosides, beta-lactamase inhibitors, carbapenems, chloramphenicols, and sulfonamides. An analysis of virulence genes identified the presence of eight genes, namely CNF-I, fyuA, fimC, papC, ompA, fimH, irp2, and iroN, which are implicated in their invasiveness and potential to inflict tissue damage. The MLST analysis revealed that all ExPEC strains were classified under either sequence type ST131 (Achtman database) or ST43 (Pasteur database). The study further determined that these strains were absent in the kennel's drinking water source, thereby ruling out water contamination as a potential factor in the emergence of ST131-type ExPEC. This study offers a theoretical framework and empirical evidence for elucidating the potential pathogenic mechanisms and clinical therapeutic strategies of ExPEC in the etiology of acute pneumonia in puppies.

Efflux of TolC protein to different antimicrobials can be replaced by other outer membrane proteins with similar β-barrel structures in extraintestinal pathogenic Escherichia coli.

As a major efflux pump system in Gram-negative bacteria, AcrAB-TolC plays a key role in the transport of multiple drug substrates and is considered a potential target for the development of novel antimicrobials. Our previous study found that TolC inactivation compromised the resistance to different antimicrobials in porcine extraintestinal pathogenic Escherichia coli (ExPEC) strain PPECC042 (WT). This study was designed to investigate the functional substitution of TolC by other outer membrane proteins (OMPs) with similar β-barrel structures in pumping out different antimicrobials. In this study, we found that over-expression of several OMPs with similar β-barrel structures, OmpX, OmpC, OmpN, OmpW, and PhoE, in the ΔtolC strain restored the resistance to macrolides, quinolones, or tetracyclines to the level of WT strain. However, the introduction of any one of the five OMPs did not affect the resistance of the strains ΔacrA, ΔacrB, and ΔacrAΔtolC. Further study revealed that the efflux activity was significantly reduced in the ΔtolC strain, but not in the WT strain and the ΔtolC strains over-expressing various OMPs. Additionally, Nile red dye test and ciprofloxacin accumulation test confirmed that the lost efflux activity and drug accumulation in bacterial periplasm by TolC inactivation was restored by the over-expression of each OMP, depending on the presence of genes acrA and acrB. All five OMPs can replace the TolC protein to play the efflux role in pumping out the drugs from the periplasm to the extracellular space with the help of proteins AcrA and AcrB.

Enteroaggregative Escherichia coli (EAEC) isolates obtained from non-diarrheic children carry virulence factor-encoding genes from Extraintestinal Pathogenic E. Coli (ExPEC).

Enteroaggregative E. coli (EAEC) is one of the most frequent pathogens isolated from diarrheal patients as well as from healthy individuals in Brazil and has recently also been implicated as an extraintestinal pathogenic E. coli (ExPEC) associated with bloodstream and urinary tract infections. In this study, 37 EAEC isolates, obtained from fecal samples of non-diarrheic children, were molecularly and phenotypically characterized to access the pathogenic features of these isolates. The EAEC isolates were assigned into the phylogroups A (54.1%), D (29.7%), B1 (13.5%) and B2 (2.7%); and harbored genes responsible for encoding the major pilin subunit of the aggregative adherence fimbriae (AAFs) or aggregate-forming pili (AFP) adhesins as follows: aggA (24.3%), agg3A (5.4%), agg4A (27.0%), agg5A (32.4%) and afpA (10.8%). The most frequent O:H serotypes were O15:H2 (8.1%), O38:H25 (5.4%) and O86:H2 (5.4%). Twenty-one isolates (56.8%) produce the aggregative adherence (AA) pattern on HeLa cells, and biofilm formation was more efficient among EAEC isolates harboring the aggA and agg5A genes. PFGE analysis showed that 31 (83.8%) of the isolates were classified into 10 distinct clusters, which reinforces the high diversity found among the isolates studied. Of note, 40.5% (15/37) of the EAEC isolates have a genetic profile compatible with E. coli isolates with intrinsic potential to cause extraintestinal infections in healthy individuals, and therefore, classified as EAEC/ExPEC hybrids. In conclusion, we showed the presence of EAEC/ExPEC hybrids in the intestinal microbiota of non-diarrheic children, possibly representing the source of some endogenous extraintestinal infections.

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 and characterization of cefotaxime resistant Escherichia coli from household floors in rural Bangladesh

Antimicrobial resistance (AMR) is a rising health concern worldwide. As an indicator organism, E. coli, specifically extended-spectrum β-lactamase (ESBL) producing E. coli, can be used to detect AMR in the environment and estimate the risk of transmitting resistance among humans, animals and the environment. This study focused on detecting cefotaxime resistant E. coli in floor swab samples from 49 households in rural villages in Bangladesh. Following isolation of cefotaxime resistant E. coli, DNA extracted from isolates was subjected to molecular characterization for virulence and resistance genes, determination of resistance to multiple classes of antibiotics to define multidrug resistant (MDR) and extensively drug resistant (XDR) strains, and the biofilm forming capacity of the isolates. Among 49 households, floor swabs from 35 (71 %) households tested positive for cefotaxime resistant E. coli. Notably, all of the 91 representative isolates were ESBL producers, with the majority (84.6 %) containing the blaCTX-M gene, followed by the blaTEM and blaSHV genes detected in 22.0 % and 6.6 % of the isolates, respectively. All isolates were MDR, and one isolate was XDR. In terms of pathogenic strains, 8.8 % of the isolates were diarrheagenic and 5.5 % were extraintestinal pathogenic E. coli (ExPEC). At 25 °C, 45 % of the isolates formed strong biofilm, whereas 43 % and 12 % formed moderate and weak biofilm, respectively. On the other hand, at 37 °C, 1.1 %, 4.4 % and 93.4 % of the isolates were strong, moderate and weak biofilm formers, respectively, and 1.1 % showed no biofilm formation. The study emphasizes the importance of screening and characterizing cefotaxime resistant E. coli from household floors in a developing country setting to understand AMR exposure associated with floors.

Uropathogenic E. coli and Hybrid Pathotypes in Mexican Women with Urinary Tract Infections: A Comprehensive Molecular and Phenotypic Overview.

Uropathogenic Escherichia coli (UPEC) is the main cause of urinary tract infections (UTIs) and carries virulence and resistance factors often found in mobilizable genetic elements, such as plasmids or pathogenicity islands (PAIs). UPEC is part of the extraintestinal pathogenic E. coli (ExPEC), but hybrid strains possessing both diarrheagenic E. coli (DEC) and ExPEC traits, termed "hypervirulent", present a significant health threat. This study assessed the prevalence of UPEC PAIs, ExPEC sequence types (ST), DEC genes, carbapenemase and extended-spectrum β-lactamase (ESBL) phenotypes, resistance genotypes, and plasmids in 40 clinical isolates of UPEC. Results showed that 72.5% of isolates had PAIs, mainly PAI IV536 (53%). ESBL phenotypes were found in 65% of β-lactam-resistant isolates, with 100% of carbapenem-resistant isolates producing carbapenemase. The predominant ESBL gene was blaCTX-M-2 (60%), and the most common resistance gene in fluoroquinolone and aminoglycoside-resistant isolates was aac(6')Ib (93%). Plasmids were present in 57% of isolates, and 70% belonged to the ST131 clonal group. Molecular markers for DEC pathotypes were detected in 20 isolates, with 60% classified as hybrid pathotypes. These findings indicate significant pathogenic potential and the presence of hybrid pathotypes in E. coli UTI clinical isolates in the Mexican population.

Multidrug-resistant Escherichia coli causing canine pyometra and urinary tract infections are genetically related but distinct from those causing prostatic abscesses

Despite extensive characterisation of uropathogenic Escherichia coli (UPEC) causing urinary tract infections (UTIs), the genetic background of non-urinary extraintestinal pathogenic E. coli (ExPEC) in companion animals remains inadequately understood. In this study, we characterised virulence traits of 104 E. coli isolated from canine pyometra (n = 61) and prostatic abscesses (PAs) (n = 38), and bloodstream infections (BSIs) in dogs (n = 2), and cats (n = 3). A stronger association with UPEC of pyometra strains in comparison to PA strains was revealed. Notably, 44 isolates exhibited resistance to third-generation cephalosporins and/or fluoroquinolones, 15 were extended-spectrum ß-lactamase-producers. Twelve multidrug-resistant (MDR) strains, isolated from pyometra (n = 4), PAs (n = 5), and BSIs (n = 3), along with 7 previously characterised UPEC strains from dogs and cats, were sequenced. Genomic characteristics revealed that MDR E. coli associated with UTIs, pyometra, and BSIs belonged to international high-risk E. coli clones, including sequence type (ST) 38, ST131, ST617, ST648, and ST1193. However, PA strains belonged to distinct lineages, including ST12, ST44, ST457, ST744, and ST13037. The coreSNPs, cgMLST, and pan-genome illustrated intra-clonal variations within the same ST from different sources. The high-risk ST131 and ST1193 (phylogroup B2) contained high numbers of ExPEC virulence genes on pathogenicity islands, predominating in pyometra and UTI. Hybrid MDR/virulence IncF multi-replicon plasmids, containing aerobactin genes, were commonly found in non-B2 phylogroups from all sources. These findings offer genomic insights into non-urinary ExPEC, highlighting its potential for invasive infections in pets beyond UTIs, particularly with regards to high-risk global clones.

Molecular Markers and Antimicrobial Resistance Patterns of Extraintestinal Pathogenic Escherichia coli from Camel Calves Including Colistin-Resistant and Hypermucoviscuous Strains.

Extraintestinal pathogenic Escherichia coli (ExPEC) strains are capable of causing various systemic infections in both humans and animals. In this study, we isolated and characterized 30 E. coli strains from the parenchymatic organs and brains of young (<3 months of age) camel calves which died in septicemia. Six of the strains showed hypermucoviscous phenotype. Based on minimum inhibitory concentration (MIC) values, seven of the strains were potentially multidrug resistant, with two additional showing colistin resistance. Four strains showed mixed pathotypes, as they carried characteristic virulence genes for intestinal pathotypes of E. coli: three strains carried cnf1, encoding cytotoxic necrotizing factor type 1, the key virulence gene of necrotoxigenic E. coli (NTEC), and one carried eae encoding intimin, the key virulence gene of enteropathogenic E. coli (EPEC). An investigation of the integration sites of pathogenicity islands (PAIs) and the presence of prophage-related sequences showed that the strains carry diverse arrays of mobile genetic elements, which may contribute to their antimicrobial resistance and virulence patterns. Our work is the first to describe ExPEC strains from camels, and points to their veterinary pathogenic as well as zoonotic potential in this important domestic animal.

First Isolation of the Heteropathotype Shiga Toxin-Producing and Extra-Intestinal Pathogenic (STEC-ExPEC) E. coli O80:H2 in French Healthy Cattle: Genomic Characterization and Phylogenetic Position.

The emerging heteropathotype shigatoxigenic (STEC) and extra-intestinal pathogenic Escherichia coli (ExPEC) O80:H2 has been the second leading cause of pediatric HUS in France since the mid-2010s. In contrast with other highly pathogenic STEC serotypes, for which ruminants have clearly been identified as the main human infection source, this heteropathotype's reservoir remains unknown. In this context, we describe for the first time the isolation of seven STEC O80:H2 strains from healthy cattle on a single cattle farm in France. This study aimed at (i) characterizing the genome and (ii) investigating the phylogenetic positions of these O80:H2 STEC strains. The virulomes, resistomes, and phylogenetic positions of the seven bovine isolates were investigated using in silico typing tools, antimicrobial susceptibility testing and cgMLST analysis after short-read whole genome sequencing (WGS). One representative isolate (A13P112V1) was also subjected to long-read sequencing. The seven isolates possessed ExPEC-related virulence genes on a pR444_A-like mosaic plasmid, previously described in strain RDEx444 and known to confer multi-drug resistance. All isolates were clonally related and clustered with human clinical strains from France and Switzerland with a range of locus differences of only one to five. In conclusion, our findings suggest that healthy cattle in France could potentially act as a reservoir of the STEC-ExPEC O80:H2 pathotype.

Progress toward a vaccine for extraintestinal pathogenic E. coli (ExPEC) II: efficacy of a toxin-autotransporter dual antigen approach.

Extraintestinal pathogenic Escherichia coli (ExPEC) is a leading cause of worldwide morbidity and mortality, the top cause of antimicrobial-resistant (AMR) infections, and the most frequent cause of life-threatening sepsis and urinary tract infections (UTI) in adults. The development of an effective and universal vaccine is complicated by this pathogen's pan-genome, its ability to mix and match virulence factors and AMR genes via horizontal gene transfer, an inability to decipher commensal from pathogens, and its intimate association and co-evolution with mammals. Using a pan virulome analysis of >20,000 sequenced E. coli strains, we identified the secreted cytolysin α-hemolysin (HlyA) as a high priority target for vaccine exploration studies. We demonstrate that a catalytically inactive pure form of HlyA, expressed in an autologous host using its own secretion system, is highly immunogenic in a murine host, protects against several forms of ExPEC infection (including lethal bacteremia), and significantly lowers bacterial burdens in multiple organ systems. Interestingly, the combination of a previously reported autotransporter (SinH) with HlyA was notably effective, inducing near complete protection against lethal challenge, including commonly used infection strains ST73 (CFT073) and ST95 (UTI89), as well as a mixture of 10 of the most highly virulent sequence types and strains from our clinical collection. Both HlyA and HlyA-SinH combinations also afforded some protection against UTI89 colonization in a murine UTI model. These findings suggest recombinant, inactive hemolysin and/or its combination with SinH warrant investigation in the development of an E. coli vaccine against invasive disease.

Chromosomal and plasmid-encoded virulence and multidrug resistance of Escherichia coli ST58/24 infecting a 2-year-old sickle cell patient with sepsis in Kampala Uganda, East Africa

Sepsis and drug resistance represent a complex of the most common global causes of mortality in intensive care units (ICUs) especially among patients with comorbidities. Extraintestinal pathogenic Escherichia coli (ExPEC) strains are highly implicated in systemic infections, with multidrug resistance exacerbating the risk of chronic conditions and patient mortality. The diversity of virulence and evolution of multidrug resistance are yet to be fully deciphered. In this work, we aimed at unveiling the pathogens and their genomic determinants of virulence and drug resistance relevant to increased sepsis in a sickle cell child admitted to ICU. From a rectal swab, we isolated a strain of E. coli from the patient and phenotypically tested it against a panel of selected beta lactams, fluoroquinolones, macrolides, aminoglycosides and colistin. We then sequenced the entire genome and integrated multiple bioinformatic pipelines to divulge the virulence and multidrug resistance profiles of the isolate. Our results revealed that the isolate belongs to the sequence type (ST) 58/24, which (ST58), is a known ExPEC. With the use of PathogenFinder, we were able to confirm that this isolate is a human pathogen (p = 0.936). The assembled chromosome and two plasmids encode virulence factors related to capsule (antiphagocytosis), serum survival and resistance, type 6 secretion system (T6SS), multiple siderophores (iron acquisition), and biosynthetic gene clusters for polyketides and nonribosomal peptides exhibiting host cell damaging activity in silico. The genome also harbors multidrug resistance genotypes including extended spectrum beta lactamase (ESBL) genes such as blaTEM-1A/B, sulfonamide resistance genes sul1/2, fluoroquinolone resistance genes dfrA5 and nonsynonymous mutations of the gene pmrB, conferring intrinsic colistin resistance. Conclusively, this pathogen holds the potential to cause systemic infection and might exacerbate sickle cell anemia in the patient. The virulence and multidrug resistance profiles are encoded by both the chromosome and plasmids. Genomic surveillance of pathogens with multidrug resistance among patients with comorbidities is crucial for effective disease management.

Raw milk cheeses from Beira Baixa, Portugal—A contributive study for the microbiological hygiene and safety assessment

Due to specific bacterial microbiota, raw milk cheeses have appreciated sensory properties. However, they may pose a threat to consumer safety due to potential pathogens presence. This study evaluated the microbiological contamination of 98 raw milk cheeses from Beira Baixa, Portugal. Presence and enumeration of Coagulase Positive Staphylococci (CPS), Listeria monocytogenes, Salmonella spp., pathogenic Escherichia coli, and indicator microorganisms (non-pathogenic E. coli and Listeria spp.) was attained. E. coli antimicrobial resistance (AMR) was also evaluated. PCR and/or Whole genome sequencing (WGS) was used to characterize E. coli, Salmonella spp. and L. monocytogenes isolates. Sixteen cheeses (16.3%) were classified as Satisfactory, 59 (60.2%) as Borderline and 23 (23.5%) as Unsatisfactory/Potential Injurious to Health. L. monocytogenes, CPS > 104 cfu g−1, Extraintestinal pathogenic E. coli (ExPEC) and Salmonella spp. were detected in 4.1%, 6.1%, 3.1% and 1.0% of the samples, respectively. Listeria innocua (4.1%) and E. coli > 104 cfu g−1 (16.3%) were also detected. AMR E. coli was detected in 23/98 (23.5%) of the cheese samples, of which two were multidrug resistant. WGS identified genotypes already associated to human disease and Listeria spp. cluster analysis indicated that cheese contamination might be related with noncompliance with Good Hygiene Practices during cheese production.

Baicalin Weakens the Virulence of Porcine Extraintestinal Pathogenic Escherichia coli by Inhibiting the LuxS/AI-2 Quorum-Sensing System.

Porcine extraintestinal pathogenic Escherichia coli (ExPEC) is a pathogenic bacterium that causes huge economic losses to the pig farming industry and considerably threatens human health. The quorum sensing (QS) system plays a crucial role in the survival and pathogenesis of pathogenic bacteria. Hence, it is a viable approach to prevent ExPEC infection by compromising the QS system, particularly the LuxS/AI-2 system. In this study, we investigated the effects of baicalin on the LuxS/AI-2 system of ExPEC. Baicalin at concentrations of 25, 50, and 100 μg/mL significantly diminished the survival ability of ExPEC in hostile environments and could inhibit the biofilm formation and autoagglutination ability in ExPEC. Moreover, baicalin dose-dependently decreased the production of AI-2 and down-regulated the expression level of luxS in PCN033. These results suggest that baicalin can weaken the virulence of PCN033 by inhibiting the LuxS/AI-2 system. After the gene luxS was deleted, AI-2 production in PCN033 was almost completely eliminated, similar to the effect of baicalin on the production of AI-2 in PCN033. This indicates that baicalin reduced the production of AI-2 by inhibiting the expression level of luxS in ExPEC. In addition, the animal experiment further showed the potential of baicalin as a LuxS/AI-2 system inhibitor to prevent ExPEC infection. This study highlights the potential of baicalin as a natural quorum-sensing inhibitor for therapeutic applications in preventing ExPEC infection by targeting the LuxS/AI-2 system.

Antibacterial activity of the antimicrobial peptide PMAP-36 in combination with tetracycline against porcine extraintestinal pathogenic Escherichia coli in vitro and in vivo

The increase in the emergence of antimicrobial resistance has led to great challenges in controlling porcine extraintestinal pathogenic Escherichia coli (ExPEC) infections. Combinations of antimicrobial peptides (AMPs) and antibiotics can synergistically improve antimicrobial efficacy and reduce bacterial resistance. In this study, we investigated the antibacterial activity of porcine myeloid antimicrobial peptide 36 (PMAP-36) in combination with tetracycline against porcine ExPEC PCN033 both in vitro and in vivo. The minimum bactericidal concentrations (MBCs) of AMPs (PMAP-36 and PR-39) against the ExPEC strains PCN033 and RS218 were 10 μM and 5 μM, respectively. Results of the checkerboard assay and the time-kill assay showed that PMAP-36 and antibiotics (tetracycline and gentamicin) had synergistic bactericidal effects against PCN033. PMAP-36 and tetracycline in combination led to PCN033 cell wall shrinkage, as was shown by scanning electron microscopy. Furthermore, PMAP-36 delayed the emergence of PCN033 resistance to tetracycline by inhibiting the expression of the tetracycline resistance gene tetB. In a mouse model of systemic infection of PCN033, treatment with PMAP-36 combined with tetracycline significantly increased the survival rate, reduced the bacterial load and dampened the inflammatory response in mice. In addition, detection of immune cells in the peritoneal lavage fluid using flow cytometry revealed that the combination of PMAP-36 and tetracycline promoted the migration of monocytes/macrophages to the infection site. Our results suggest that AMPs in combination with antibiotics may provide more therapeutic options against multidrug-resistant porcine ExPEC.

Two Novel Bacteriophage Species Against Hybrid Intestinal Pathogenic Escherichia coli/Extraintestinal Pathogenic Escherichia coli Strains.

Colibacillosis caused by Escherichia coli is one of the main problems in the swine industry. In addition, the emergence of antimicrobial resistance and the combination of virulence genes among pathotypes have led to the emergence of more virulent pathogenic E. coli strains. Phage therapy has become a promising approach to address these issues. Virulence genes for intestinal pathogenic E. coli (IPEC) and extraintestinal pathogenic E. coli (ExPEC) were investigated in pathogenic E. coli isolated from pigs. In addition, two potential phages, vB_EcoM-RPN187 and vB_EcoM-RPN226, isolated in our previous study, were further characterized in this study. Both phages were lytic and were highly effective at 20-37°C. Interestingly, they infected the hybrid IPEC/ExPEC strains. vB_EcoM-RPN187 and vB_EcoM-RPN226 possess 167 kbp of linear double-stranded DNA without virulence or antibiotic resistance genes and may be classified as new phage species in the genera Mosigvirus and Tequatrovirus, respectively. Both phages could be promising candidates for phage therapy against pathogenic E. coli.

Antimicrobial resistance and phylogenetic relatedness of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli in peridomestic rats (Rattus norvegicus and Rattus tanezumi) linked to city areas and animal farms in Hong Kong

Extended-spectrum β-lactamase-producing Escherichia (E.) coli (ESBL-EC) in the clinical setting have emerged as a major threat to public and animal health. Wildlife, including Rattus spp. may serve as reservoirs and spreaders of ESBL-EC in the environment. Peridomestic rats are well adapted to living in proximity to humans and animals in a variety of urban and agricultural environments and may serve as sentinels to identify variations of ESBL-EC within their different habitats. In this study, a set of 221 rats (Rattus norvegicus, R. tanezumi, R. andamanensis, and Niviventer huang) consisting of 104 rats from city areas, 44 from chicken farms, 52 from pig farms, and 21 from stables of horse-riding schools were screened for ESBL-EC. Overall, a total of 134 ESBL-EC were isolated from the caecal samples of 130 (59%) rats. The predominant blaESBL genes were blaCTX-M-14, blaCTX-M-15, blaCTX-M-55, and blaCTX-M-65. Phylogenetic analysis revealed a total of 62 sequence types (STs) and 17 SNP clusters. E. coli ST10 and ST155 were common to ESBL-EC from city areas and chicken farms, and ST44 were found among ESBL-EC from city areas and pig farms. Extra-intestinal pathogenic E. coli (ExPEC) ST69, ST131 and ST1193 were found exclusively among rats from city areas, and avian pathogenic E. coli (APEC) ST177 was restricted to ESBL-EC originating from chicken farms. Phylogenetic analysis showed that the populations of rodent ESBL-EC from city areas, chicken farms and pig farms were genetically different, suggesting a certain degree of partitioning between the human and animal locations. This study contributes to current understanding of ESBL-EC occurring in rats in ecologically diverse locations.

Phenotypic and genotypic characterization of Escherichia coli isolated from the chicken liver in relation to slaughterhouse conditions

Avian pathogenic Escherichia coli (APEC) has been identified as a sub-group of extraintestinal pathogenic E. coli (ExPEC). Recent studies indicate APEC as a potential foodborne zoonotic pathogen and a source or reservoir of human extraintestinal infections. The slaughtering and processing of poultry in low-income countries such as Jordan occurs in two distinct ways: in informal facilities known as Natafat and in formal slaughterhouses. This study compared E. coli phenotypes and genotypes according to slaughtering conditions (formal slaughterhouses vs. informal slaughter facilities). Therefore, liver samples (n = 242) were collected from formal (n = 121) and informal slaughter facilities (n = 121). Results revealed a high prevalence (94.2%) of E. coli among all isolates, with 59 (17 formal and 42 informal) isolates considered avian pathogenic E. coli (APEC) based on the virulence-associated genes. The prevalence of resistance among isolates was relatively high, reaching up to 99% against penicillin and 97% against nalidixic acid. However, the prevalence of resistance was the lowest (1.3%) against both meropenem and imipenem. Based on the MIC test findings, colistin resistance was 46.9% (107/228). The mcr −1 gene prevalence was 51.4% (55/107), of which 17.1 % were from formal plants (6/36) and 68.1% from informal facilities (49/72). Interestingly, only one isolate (0.9%) expressed mcr-10. Escherichia coli O157:H7 and associated virulence genes were found more in informal (n = 15 genes) than in formal slaughterhouses (n = 8). Phylogroups B1, C, and A were the most frequent in 228 E. coli isolates, while G, B2, and clade were the least frequent. In conclusion, these findings highlight the importance of implementing biosecurity measures in slaughterhouses to reduce antibiotic-resistant E. coli spread. Furthermore, this study provides valuable insights into the effects of wet market (Natafat) slaughter conditions on increasing bacterial resistance and virulence.

Shiga toxin-producing Escherichia coli isolated from hunted wild boar (Sus scrofa) in Switzerland.

Shiga toxin-producing Escherichia (E.) coli (STEC) are zoonotic foodborne pathogens of significant public health importance. While ruminants are considered the main reservoir, wild animals are increasingly acknowledged as carriers and potential reservoirs of STEC. The aim of this study was to determine the occurrence of STEC in a total of 59 faecal samples of hunted wild boars (Sus scrofa) from two different regions in Switzerland (canton Thurgau in northern Switzerland and canton Ticino in southern Switzerland), and to characterise the isolates using a whole genome sequencing approach. After an enrichment step, Shiga-toxin encoding genes (stx) were detected by real-time PCR in 41 % (95 % confidence interval (95 %CI) 0,29 - 0,53) of the samples, and STEC were subsequently recovered from 22 % (95 %CI 0,13 - 0,34) of the same samples. Seven different serotypes and six different sequence types (STs) were found, with O146:H28 ST738 (n = 4) and O100:H20 ST2514 (n = 4) predominating. Subtyping of stx identified isolates with stx1c/stx2b (n = 1), stx2a (n = 1), stx2b (n = 6), and stx2e (n = 6). No isolate contained the eae gene, but all harboured additional virulence genes, most commonly astA (n = 10), hlyE (n = 9), and hra (n = 9). STEC O11:H5, O21:H21, and O146:H28 harboured virulence factors associated with extra-intestinal pathogenic E. coli (ExPEC), and STEC O100:H20 and O155:H26 possessed sta1 and/or stb and were STEC/enterotoxigenic E. coli (ETEC) hybrid pathotypes. Our results show that wild boars are carriers of STEC which may be distributed in the environment, possibly leading to the contamination of agricultural crops and water sources. The serogroups included STEC O146 which belongs to the most common non-O157 serogroups associated with human illness in Europe, with implications for public health. Since Stx2e-producing STEC have frequently been reported in swine and pork, STEC O100:H20 harbouring stx2e in faeces of wild boars may be relevant to free-range systems of pig farming because of the potential risk of transmission events at thewildlife-livestock interface.