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Enterohemorrhagic Escherichia coli (EHEC) is a foodborne pathogen that infects humans by colonizing the large intestine. Upon reaching the large intestine, EHEC mediates local signal recognition and the transcriptional regulation of virulence genes to promote adherence and colonization in a highly site-specific manner. Two-component systems (TCSs) represent an important strategy used by EHEC to couple external stimuli with the regulation of gene expression, thereby allowing EHEC to rapidly adapt to changing environmental conditions. An increasing number of studies published in recent years have shown that EHEC senses a variety of host- and microbiota-derived signals present in the human intestinal tract and coordinates the expression of virulence genes via multiple TCS-mediated signal transduction pathways to initiate the disease-causing process. Here, we summarize how EHEC detects a wide range of intestinal signals and precisely regulates virulence gene expression through multiple signal transduction pathways during the initial stages of infection, with a particular emphasis on the key roles of TCSs. This review provides valuable insights into the importance of TCSs in EHEC pathogenesis, which has relevant implications for the development of antibacterial therapies against EHEC infection. [ABSTRACT FROM AUTHOR]

Multi-drug resistant-Shiga toxin producing Escherichia coli (MDR-STEC), poses considerable health risks for humans, animals and birds. Migratory and captive wild birds are known carriers of this pathogen. This study aimed to investigate prevalence of MDR-STEC along with its associated virulence genes from migratory and captive wild birds in Bangladesh. A total of 247 fecal specimens were obtained from both migratory (119) and captive wild birds (128) for the isolation and characterization of E. coli. Standard microbiological and biochemical methods were used for identification and Polymerase Chain Reaction (PCR) was employed for confirmation of E. coli isolates followed by disc diffusion method to determine antibiotic susceptibility. The overall E. coli prevalence was 80.97% (200/247; 95% CI: 75.51-85.67), with a prevalence of 77.31% (92/119) in migratory birds and 84.38% (108/128) in captive wild birds. Among the 200 E. coli isolates, 53 (26.5%; 95% CI: 20.5-33.2) were identified as multidrug-resistant (MDR), with 21.7% (20/92) of MDR isolates originating from migratory birds and 30.6% (33/108) from captive wild birds. Only 24 (12%) isolates were positive for virulent gene stx2 whereas 167 (83.50%) isolates were positive for fimC. Among the ß-lactamase resistant genes, blaTEM (91.50%; 183/200) was found significantly (P<0.0001) higher than blaSHV (9.00%; 18/200). Among the antimicrobial resistant genes, 175 (87.50%) isolates were found positive for qnrS resistant gene. E. coli isolates of birds exhibited diverse phenotypic AMR patterns, with complete (100%) resistance to several antibiotics (ampicillin, ceftazidime, cefuroxime, and tetracycline) while being entirely sensitive to others (ceftriaxone, amikacin and aztreonam). This research underscores the concerning prevalence of E. coli strains having various virulent genes and resistant to multiple drugs among the wild birds. It emphasizes the immediate requirement for bridging wildlife and public health domains to address the threats posed by the antibiotic-resistant pathogens. [ABSTRACT FROM AUTHOR]

Enterotoxigenic Escherichia coli (ETEC) cause hundreds of millions of diarrheal illnesses annually ranging from mildly symptomatic cases to severe, life-threatening cholera-like diarrhea. Although ETEC are associated with long-term sequelae including malnutrition, the acute diarrheal illness is largely self-limited. Recent studies indicate that in addition to causing diarrhea, the ETEC heat-labile toxin (LT) modulates the expression of many genes in intestinal epithelia, including carcinoembryonic cell adhesion molecules (CEACAMs) which ETEC exploit as receptors, enabling toxin delivery. Here, however, we demonstrate that LT also enhances the expression of CEACAMs on extracellular vesicles (EV) shed by intestinal epithelia and that CEACAM-laden EV increase in abundance during human infections, mitigate pathogen-host interactions, scavenge free ETEC toxins, and accelerate ETEC clearance from the gastrointestinal tract. Collectively, these findings indicate that CEACAMs play a multifaceted role in ETEC pathogen-host interactions, transiently favoring the pathogen, but ultimately contributing to innate responses that extinguish these common infections. [ABSTRACT FROM AUTHOR]

The Shiga toxin producing Escherichia coli (STEC) are considered to be one of the most important groups of emerging public health pathogens with cattle being the main reservoir. The objective of this study was to isolate and characterise Escherichia coli Shiga toxins in dairy cattle farms. A total of 252 faeces samples were collected from healthy cows belonging to 37 farms. PCR screening of samples for the common sequences of stx1/stx2 genes and stx1 and stx2 genes resulted in a STEC faecal excretion prevalence of 59.5% at the farm level and 26.6% at the individual level. Among positive animals, 85.1% carried STEC with a single stx1 gene and 14.9% with the stx1 and stx2 genes. Immunomagnetic separation was performed on 40 PCR-positive samples (10/10 positive for the stx1 and stx2 genes and 30/57 positive only for stx1). Biochemical identification revealed the presence of 66 E. coli strains (27.5%). The search for virulence genes on these strains by PCR showed that only twenty-two (33.33%) were STEC. The presence of the stx1, stx2, ehx and eae genes was characterised in 30.3%, 4.54%, 13.63% and 1.51% of the strains, respectively, indicating that the virulotype with stx alone was dominant. Serological identification showed the absence of O157 sero-groups and the presence of O1(2), O2, O18(2), O128 sero-groups. The susceptibility testing of STEC showed 68.18% resistance to chloramphenicol, 63.64% to neomycin, 59.1% to ampicillin, 22.73% to trimethoprim + sulfamethoxazole and 9.1% to amoxicillin + clavulanic acid and nalidixic acid. Four strains showed multi-resistance. Bovine carriage of STEC constitutes a public health risk by contamination of milk and meat. To protect human health, it is necessary to limit the bovine STEC shedding. [ABSTRACT FROM AUTHOR]

In this study, two Mediterranean coastal lagoons (Lesina and Varano) of southern Italy, located in the north of the Apulia region, were investigated for the presence of Shiga toxin Escherichia coli (STEC) and potentially enteropathogenic Vibrio species in parallel with norovirus (NoV), hepatitis A virus (HAV), hepatitis E virus (HEV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This study aimed to evaluate the presence of potentially pathogenic bacteria and viruses in the water and sediments of these ecosystems. From March 2022 to February 2023, a total of 98 samples were collected: 49 water samples and 49 sediment samples. STEC strains were isolated in three samples (3.1%), of which one (2%) was water (stx1 and stx2 positive) and two (4.1%) were sediment (both stx2 positive) samples. Vibrio spp. were detected in twenty samples (20.4%), of which nine were water (18.4%) and eleven were sediment (22.4%) samples. The species detected included V. parahaemolyticus, V. cholerae, and V. vulnificus. NoV was detected in 25 (25.5%) samples, while none of the water or sediment samples were positive for HAV, HEV, and SARS-CoV-2. The results of this study provide an overview of the presence of potentially pathogenic microorganisms in areas influenced by anthropogenic pressure. Monitoring the circulation of these pathogens could be useful to evaluate the water flowing into the lagoons, in particular discharge waters (i.e., urban, agricultural, and livestock runoff), considering the presence of fish and shellfish farms in these sites. [ABSTRACT FROM AUTHOR]

Aim: Trafficking, membrane retention, and signal‐specific regulation of the Na+/H+ exchanger 3 (NHE3) are modulated by the Na+/H+ Exchanger Regulatory Factor (NHERF) family of PDZ‐adapter proteins. This study explored the assembly of NHE3 and NHERF2 with the cGMP‐dependent kinase II (cGKII) within detergent‐resistant membrane microdomains (DRMs, "lipid rafts") during in vivo guanylate cycle C receptor (Gucy2c) activation in murine small intestine. Methods: Small intestinal brush border membranes (siBBMs) were isolated from wild type, NHE3‐deficient, cGMP‐kinase II‐deficient, and NHERF2‐deficient mice, after oral application of the heat‐stable Escherichia coli toxin (STa) analog linaclotide. Lipid raft and non‐raft fractions were separated by Optiprep density gradient centrifugation of Triton X‐solubilized siBBMs. Confocal microscopy was performed to study NHE3 redistribution after linaclotide application in vivo. Results: In the WT siBBM, NHE3, NHERF2, and cGKII were strongly raft associated. The raft association of NHE3, but not of cGKII, was NHERF2 dependent. After linaclotide application to WT mice, lipid raft association of NHE3 decreased, that of cGKII increased, while that of NHERF2 did not change. NHE3 expression in the BBM shifted from a microvillar to a terminal web region. The linaclotide‐induced decrease in NHE3 raft association and in microvillar abundance was abolished in cGKII‐deficient mice, and strongly reduced in NHERF2‐deficient mice. Conclusion: NHE3, cGKII, and NHERF2 form a lipid raft‐associated signal complex in the siBBM, which mediates the inhibition of salt and water absorption by Gucy2c activation. NHERF2 enhances the raft association of NHE3, which is essential for its close interaction with the exclusively raft‐associated activated cGKII. [ABSTRACT FROM AUTHOR]

Plasmid-encoded toxin (Pet) is an autotransporter protein of the serine protease autotransporters of Enterobacteriaceae (SPATE) family, important in the pathogenicity of Escherichia coli. The pet gene was initially found in the enteroaggregative E. coli (EAEC) virulence plasmid, pAA2. Although this virulence factor was initially described in EAEC, an intestinal E. coli pathotype, pet may also be present in other pathotypes, including extraintestinal pathogenic strains (ExPEC). The complement system is an important defense mechanism of the immune system that can be activated by invading pathogens. Proteases produced by pathogenic bacteria, such as SPATEs, have proteolytic activity and can cleave components of the complement system, promoting bacterial resistance to human serum. Considering these factors, the proteolytic activity of Pet and its role in evading the complement system were investigated. Proteolytic assays were performed by incubating purified components of the complement system with Pet and Pet S260I (a catalytic site mutant) proteins. Pet, but not Pet S260I, could cleave C3, C5 and C9 components, and also inhibited the natural formation of C9 polymers. Furthermore, a dose-dependent inhibition of ZnCl2-induced C9 polymerization in vitro was observed. E. coli DH5a survived incubation with human serum pre-treated with Pet. Therefore, Pet can potentially interfere with the alternative and the terminal pathways of the complement system. In addition, by cleaving C9, Pet may inhibit membrane attack complex (MAC) formation on the bacterial outer membrane. Thus, our data are suggestive of a role of Pet in resistance of E. coli to human serum. [ABSTRACT FROM AUTHOR]

Even though differences in methodology (e.g., sample volume and detection method) have been shown to affect observed microbial water quality, multiple sampling and laboratory protocols continue to be used for water quality monitoring. Research is needed to determine how these differences impact the comparability of findings to generate best management practices and the ability to perform metaanalyses. This study addresses this knowledge gap by compiling and analyzing a data set representing 2,429,990 unique data points on at least one microbial water quality target (e.g., Salmonella presence and Escherichia coli concentration). Variance partitioning analysis was used to quantify the variance in likelihood of detecting each pathogenic target that was uniquely and jointly attributable to non-methodological versus methodological factors. The strength of the association between microbial water quality and select methodological and non-methodological factors was quant ified using conditional forest and regression analysis. Fecal indicator bacteria concentrations were more strongly associated with non-methodological factors than methodological factors based on conditional forest analysis. Variance partitioning analysis could not disentangle non-methodological and methodological signals for pathogenic Escherichia coli, Salmonella, and Listeria. This suggests our current perceptions of foodborne pathogen ecology in water systems are confounded by methodological difference s between studies. For example, 31% of total variance in likelihood of Salmonella detection was explained by methodological and/or non-methodological factors, 18% was jointly attributable to both methodological and non-methodological factors. Only 13% of total variance was uniquely attributable to non-methodological factors for Salmonella, highlighting the need for standardization of methods for microbiological water quality testing for comparison across studies. [ABSTRACT FROM AUTHOR]

Plasmid-encoded toxin (Pet) is an autotransporter protein of the serine protease autotransporters of Enterobacteriaceae (SPATE) family, important in the pathogenicity of Escherichia coli. The pet gene was initially found in the enteroaggregative E. coli (EAEC) virulence plasmid, pAA2. Although this virulence factor was initially described in EAEC, an intestinal E. coli pathotype, pet may also be present in other pathotypes, including extraintestinal pathogenic strains (ExPEC). The complement system is an important defense mechanism of the immune system that can be activated by invading pathogens. Proteases produced by pathogenic bacteria, such as SPATEs, have proteolytic activity and can cleave components of the complement system, promoting bacterial resistance to human serum. Considering these factors, the proteolytic activity of Pet and its role in evading the complement system were investigated. Proteolytic assays were performed by incubating purified components of the complement system with Pet and Pet S260I (a catalytic site mutant) proteins. Pet, but not Pet S260I, could cleave C3, C5 and C9 components, and also inhibited the natural formation of C9 polymers. Furthermore, a dose-dependent inhibition of ZnCl2-induced C9 polymerization in vitro was observed. E. coli DH5a survived incubation with human serum pre-treated with Pet. Therefore, Pet can potentially interfere with the alternative and the terminal pathways of the complement system. In addition, by cleaving C9, Pet may inhibit membrane attack complex (MAC) formation on the bacterial outer membrane. Thus, our data are suggestive of a role of Pet in resistance of E. coli to human serum. [ABSTRACT FROM AUTHOR]

The article explores the translation of toxin genes in bacteria's toxin-antitoxin systems (TA). These systems consist of a toxic protein and its antitoxin antidote. The study focuses on the timP-TimR T1TA system in Salmonella and reveals that a pseudoknot structure in the timP mRNA activates translation. The antitoxin TimR destabilizes the pseudoknot structure, inhibiting translation. The findings shed light on the role of RNA dynamics in gene expression and offer insights into alternative ways toxin mRNA can be activated. Understanding the control of toxin and antitoxin synthesis can provide valuable information about their physiological functions. [Extracted from the article]

Edema disease (ED) is a severe and lethal infectious ailment in swine, stemming from Shiga-toxin-producing Escherichia coli (STEC). An efficient, user-friendly, and safe vaccine against ED is urgently required to improve animal welfare and decrease antibiotic consumption. Recombinant attenuated Salmonella vaccines (RASV) administered orally induce both humoral and mucosal immune responses to the immunizing antigen. Their potential for inducing protective immunity against ED is significant through the delivery of STEC antigens. rSC0016 represents an enhanced recombinant attenuated vaccine vector designed for Salmonella enterica serotype Choleraesuis. It combines sopB mutations with a regulated delay system to strike a well-balanced equilibrium between host safety and immunogenicity. We generated recombinant vaccine strains, namely rSC0016 (pS-FedF) and rSC0016 (pS-rStx2eA), and assessed their safety and immunogenicity in vivo. The findings demonstrated that the mouse models immunized with rSC0016 (pS-FedF) and rSC0016 (pS-rStx2eA) generated substantial IgG antibody responses to FedF and rStx2eA, while also provoking robust mucosal and cellular immune responses against both FedF and rStx2eA. The protective impact of rSC0016 (pS-FedF) against Shiga-toxin-producing Escherichia coli surpassed that of rSC0016 (pS-rStx2eA), with percentages of 83.3%. These findings underscore that FedF has greater suitability for vaccine delivery via recombinant attenuated Salmonella vaccines (RASVs). Overall, this study provides a promising candidate vaccine for infection with STEC. [ABSTRACT FROM AUTHOR]

Many bacteria encode multiple toxin-antitoxin (TA) systems targeting separate, but closely related, cellular functions. The toxin of the Escherichia coli hipBA system, HipA, is a kinase that inhibits translation via phosphorylation of glutamyl-tRNA synthetase. Enteropathogenic E. coli O127:H6 encodes the hipBA-like, tripartite TA system; hipBST, in which the HipT toxin specifically targets the tryptophanyl-tRNA synthetase, TrpS. Notably, in the tripartite system, the function as antitoxin has been taken over by the third protein, HipS, but the molecular details of how activity of HipT is inhibited remain poorly understood. Here, we show that HipBST is structurally different from E. coli HipBA and that the unique HipS protein, which is homologous to the N-terminal subdomain of HipA, inhibits the kinase through insertion of a conserved Trp residue into the active site. We also show how auto-phosphorylation at two conserved sites in the kinase toxin serve different roles and affect the ability of HipS to neutralize HipT. Finally, solution structural studies show how phosphorylation affects overall TA complex flexibility. [ABSTRACT FROM AUTHOR]

Molecular cloning and controlled expression remain challenging when the target gene encodes a protein that is toxic to the host. We developed a set of multi-layer control systems to enable cloning of genes encoding proteins known to be highly toxic in Escherichia coli and other bacteria. The different multi-layer control systems combine a promoter–operator system on a transcriptional level with a riboswitch for translational control. Additionally, replicational control is ensured by using a strain that reduces the plasmid copy number. The use of weaker promoters (such as PBAD or PfdeA) in combination with the effective theophylline riboswitch is essential for cloning genes that encode notoriously toxic proteins that directly target translation and transcription. Controlled overexpression is possible, allowing the system to be used for evaluating in vivo effects of the toxin. Systems with a stronger promoter can be used for successful overexpression and purification of the desired protein but are limited to toxins that are more moderate and do not interfere with their own production. [ABSTRACT FROM AUTHOR]

Shiga toxin-producing strains represent pathogenic group that is of concern in food production. The present study evaluated forty-eight E. coli isolates (11 with intact stx gene, while remaining isolates presented only stx-fragments) for Shiga toxin production. The four most expressive stx-producers (O26, O103, O145, and O157) were selected to evaluate effects of pH (3.5, 4.5, and 7) and temperature (35, 40, and 50°C). After determining acid stress effects in media on Stx-induction, we mimicked "in natura" conditions using milk, apple, and orange juices. Only isolates that showed the presence of intact stx gene (11/48) produced Shiga toxin. In addition, acid pH had a role in down-regulating the production of Shiga toxin, in both lactic acid and juices. In contrast, non-lethal heating (40°C), when in neutral pH and milk was a favorable environment to induce Shiga toxin. Lastly, two isolates (O26 and O103) showed a higher capacity to produce Shiga toxin and were included in a genomic cluster with other E. coli involved in worldwide foodborne outbreaks. The induction of this toxin when subjected to 40°C may represent a potential risk to the consumer, since the pathogenic effect of oral ingestion of Shiga toxin has already been proved in an animal model. [ABSTRACT FROM AUTHOR]

This document is an editorial for a special issue of the journal Microorganisms on bacterial toxin-antitoxin systems (TAS). TAS are found in bacterial chromosomes and mobile genetic elements and play a key role in maintaining genetic materials. They are also involved in various physiological activities in bacteria, such as stress response, virulence, biofilm formation, antibiotic tolerance, and bacteriophage propagation. The special issue includes manuscripts on the identification and characterization of novel TAS, their biological functions, and their applications in biotechnology and medicine. The document highlights specific contributions, including the genetic and functional characterization of a TAS in the pathogenic bacterium Agrobacterium tumefaciens and the development of a novel method for gene deletion in bacteria using a toxin as a counter-selectable marker. The document also emphasizes the need for further research to understand the functions of uncharacterized TAS and orphan toxin or antitoxin genes. Overall, the document provides an overview of the importance and diversity of TAS in bacteria. [Extracted from the article]

Multidrug resistance (MDR) pathogens are usually associated with higher morbidity and mortality rates. Flavonoids are good candidates for the development of new potential antimicrobials. This research investigated whether luteolin 4′-neohesperidoside (L4N) has antibacterial and synergistic activities against four antibiotic-resistant pathogens: methicillin-resistant Staphylococcus aureus (MRSA), Klebsiella pneumoniae, fosA-positive shiga toxin producing the Escherichia coli serogroup O111 (STEC O111), and Bacillus cereus. In vitro antimicrobial susceptibility testing revealed highly potent anti-MRSA (MIC of 106.66 ± 6.95 µg/mL), anti-K. pneumoniae (MIC of 53.33 ± 8.47 µg/mL) and anti-STEC O111 (MIC of 26.66 ± 5.23 µg/mL) activities. Significant synergistic combination was clearly noted in the case of gentamycin (GEN) against Gram-negative bacteria. In the case of B. cereus, the combination of vancomycin (VAN) with L4N could efficiently inhibit bacterial growth, despite the pathogen being VAN-resistant (MIC of 213.33 ± 7.9 µg/mL). In vivo evaluation of L4N showed significant decreases in K. pneumoniae and STEC shedding and colonization. Treatment could significantly diminish the levels of pro-inflammatory markers, tumor necrosis factor-alpha (TNF-α), and immunoglobulin (IgM). Additionally, the renal and pulmonary lesions were remarkably enhanced, with a significant decrease in the bacterial loads in the tissues. Finally, this study presents L4N as a potent substitute for traditional antibiotics with anti-STEC O111 and anti-K. pneumoniae potential, a finding which is reported here for the first time. [ABSTRACT FROM AUTHOR]

Shiga toxin producing Escherichia coli (STEC) serotypes are recognized as potentially important food-borne pathogens for humans. Ingestion of E. coli contaminated food is largely known to originate from livestock. Cattle and sheep herds hold the majority of agricultural revenue in Türkiye but Anatolian water buffaloes have often been underestimated for foodborne pathogens. The aim of this study is to determine virulence genes harboring E. coli O157 and six major non-O157 STEC (O26, O45, O103, O111, O121, and O145) serotypes in feces of healthy Anatolian water buffaloes by using multiplex PCR (mPCR) method. Of the collected 458 fecal samples from healthy live animals, we have performed virulence and serotype targeting mPCR following direct DNA extraction from collected samples. Results indicate that there is 0.9% of O157 prevalence while six major non-O157 E. coli have not been identified. The characterization results of the virulence genes also showed that eae is most prevalent (5.7%) followed by ehxA (3.9%) and stx1 (3.1%). In this study, we have shown Anatolian buffaloes might have a relationship with other O-type E. coli strains. Non-O157 STECs, which are often disregarded in both animals and humans, should be investigated. As a consequence, gaining regional or national data collection will allow to implement better effective diagnosis and treatment options. [ABSTRACT FROM AUTHOR]

Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens implicated in diseases including hemorrhagic colitis and hemolytic uremic syndrome. In the last years, an increasing number of STEC infections associated with the consumption of raw milk cheeses have been reported, contributing to raise the public awareness. The aim of this study was to assess the occurrence of STEC biotypes in raw milk cheeses from Italian Alpine region and evaluate the influence of different scalding temperatures on their development. Six out 82 (7.3%) cheeses led to the isolation of E. coli biotypes with different virulence traits (stx1 , stx2 and eae genes). To evaluate the impact of scalding temperatures on STEC growth, three E. coli biotypes were spiked into milk (<50 CFU/mL) according to the FAO and WHO recommendation. An increase in STEC counts of 3 log occurred in uncooked and semi-cooked cheeses (scalding temperature of 38 and 45 °C) while in the cooked-cheeses (54 and 56 °C) E. coli content does not exceed the 2.5 log 10 CFU/g. These findings showed that raw milk cheese safety is strictly related to their production technology, highlighting the importance of the control measures at farm and dairy level to preserve the safety of these products. • Raw milk cheeses might merit concern regarding the risk of STEC infection. • 7.3% of raw milk cheeses from Alpine region of Italy were positive for STEC. • STEC isolates were able to growth in the early stages of cheese-making process. • Cheese-making process of uncooked and semi-cooked cheeses support the STEC growth. • Higher scalding temperatures (56 °C) hamper the STEC development. [ABSTRACT FROM AUTHOR]

We report fatal meningitis in 2 neonates in France caused by Shiga toxin 1-producing Escherichia coli. Virulence factors capsular K1 antigen and salmochelin were present in both strains, potentially representing a new hybrid pathotype. Clinicians should remain aware of emerging pathotypes and design therapeutic strategies for neonatal E. coli infections. [ABSTRACT FROM AUTHOR]

Background: Shiga toxin producing Escherichia coli O157:H7 (STEC) is considered the most prevalent food borne pathogen that has gained increasing attention worldwide in recent years. Methods: A cross-sectional study was carried out at Bedele Municipal abattoir on cattle that were reported healthy from detailed ante-mortem inspections and having various body conditions scores. A total of 516 samples were collected and examined after enriched in modified peptone water. Following an enrichment, the samples were plated onto MacConkey agar and then onto Eosin methylene blue agar. Finally after a few similar procedures, 14 E. coli O157:H7 (STEC) isolates were confirmed through latex agglutination test. The collected data were analyzed using SPSS version 20 statistical software. Results: This study finding revealed that the overall prevalence of E. coli O157:H7 out of 516 samples was found to be 2.7%. However, on sample type basis, the prevalence of E. coli O157:H7 from feacal samples, carcass swabs, butcher hand swabs and knife swabs were 4.7%, 3.3%, 1.1% and 1.1%, respectively. It was also found that that the prevalence of E. coli O157:H7 was significantly affected by age groups of slaughtered cattle (p< 0.05). Moreover, in vitro antimicrobial susceptibility test result on average showed that almost all of E. coli O157:H7 isolates were highly susceptible to kanamycin and no resistance was shown to ciprofloxacin and gentamicin. Finally, the conventional PCR detection of stx1, st2 and hylA genes revealed that only 21.4% and 14.3% were found to contain stx1 and hylA genes respectively. Conclusion: To wrap up, this study showed that Shiga toxin producing E. coli O157:H7 (STEC) isolates were found with almost low overall prevalence rate from all sample sources in this study site. Therefore, improving abattoir facilities and slaughter house workers' personal hygiene are recommended to curtail E. coli O157:H7 meat contamination in this abattoir. [ABSTRACT FROM AUTHOR]

Enterotoxigenic E. coli (ETEC) produce heat-labile (LT) and/or heat-stable (ST) enterotoxins, and commonly cause diarrhea in resource-poor regions. ETEC have been linked repeatedly to sequelae in children including enteropathy, malnutrition, and growth impairment. Although cellular actions of ETEC enterotoxins leading to diarrhea are well-established, their contributions to sequelae remain unclear. LT increases cellular cAMP to activate protein kinase A (PKA) that phosphorylates ion channels driving intestinal export of salt and water resulting in diarrhea. As PKA also modulates transcription of many genes, we interrogated transcriptional profiles of LT-treated intestinal epithelia. Here we show that LT significantly alters intestinal epithelial gene expression directing biogenesis of the brush border, the major site for nutrient absorption, suppresses transcription factors HNF4 and SMAD4 critical to enterocyte differentiation, and profoundly disrupts microvillus architecture and essential nutrient transport. In addition, ETEC-challenged neonatal mice exhibit substantial brush border derangement that is prevented by maternal vaccination with LT. Finally, mice repeatedly challenged with toxigenic ETEC exhibit impaired growth recapitulating the multiplicative impact of recurring ETEC infections in children. These findings highlight impacts of ETEC enterotoxins beyond acute diarrheal illness and may inform approaches to prevent major sequelae of these common infections including malnutrition that impact millions of children. Enterotoxigenic Escherichia coli infections have been linked to non-diarrheal sequelae however, the reasons for this are unclear. Here, the authors present an additional role of heat-labile toxin in disrupting the structure and function of intestinal epithelial cells. [ABSTRACT FROM AUTHOR]

ppGpp is an intracellular sensor that, in response to different types of stress, coordinates the rearrangement of the gene expression pattern of bacteria to promote adaptation and survival to new environmental conditions. First described to modulate metabolic adaptive responses, ppGpp modulates the expression of genes belonging to very diverse functional categories. In Escherichia coli, ppGpp regulates the expression of cellular factors that are important during urinary tract infections. Here, we characterize the role of this alarmone in the regulation of the hlyCABDII operon of the UPEC isolate J96, encoding the toxin α-hemolysin that induces cytotoxicity during infection of bladder epithelial cells. ppGpp is required for the expression of the α-hemolysin encoded in hlyCABDII by stimulating its transcriptional expression. Prototrophy suppressor mutations in a ppGpp-deficient strain restore the α-hemolysin expression from this operon to wild-type levels, confirming the requirement of ppGpp for its expression. ppGpp stimulates hlyCABDII expression independently of RpoS, RfaH, Zur, and H-NS. The expression of hlyCABDII is promoted at 37 °C and at low osmolarity. ppGpp is required for the thermoregulation but not for the osmoregulation of the hlyCABDII operon. Studies in both commensal and UPEC isolates demonstrate that no UPEC specific factor is strictly required for the ppGpp-mediated regulation described. Our data further support the role of ppGpp participating in the coordinated regulation of the expression of bacterial factors required during infection. [ABSTRACT FROM AUTHOR]

Persistence is the non-heritable state of dormancy of a small subset of cells that allows these cells to survive a wide range of stresses. The persister cells wake in a heterogeneous manner based on the number of active ribosomes; this was first shown with single cells by converting the whole population into persister cells and labelling ribosomes with the green fluorescent protein (Kim et al., 2018b). Other mechanisms for persister cell formation and resuscitation have been suggested, primarily as a result of not studying the correct cell population, since many groups fail to wait for persistence to be obtained and instead, study the far more populous dying-cell population (Song & Wood, 2021a). Persister cells form primarily as a result of stress, but unlike the bulk of the population of cells that actively respond via a stress response (e.g. via sigma factor RpoS), a small population of cells (<1%) enter a dormant state. [Extracted from the article]

Free-living amoebae (FLA) are widely distributed protozoa in nature, known to cause severe eye infections and central nervous system disorders. There is growing attention to the potential role that these protozoa could act as reservoirs of pathogenic bacteria and, consequently, to the possibility that, the persistence and spread of the latter may be facilitated, by exploiting internalization into amoebae. Shiga toxin-producing strains of Escherichia coli (STEC) are zoonotic agents capable of causing serious diseases, such as hemorrhagic colitis (HC) and hemolytic uremic syndrome (HUS). Cattle represent the main natural reservoir of STEC, which are frequently found also in other domestic and wild ruminants, often without causing any evident symptoms of disease. The aspects related to the ecology of STEC strains in animal reservoirs and the environment are poorly known, including the persistence of these microorganisms within niches unfavorable to survival, such as soils or waters. In this study we investigated the interaction between STEC strains of serotype O157: H7 with different virulence gene profiles, and a genus of a wild free-living amoeba, Acanthamoeba sp. Our results confirm the ability of STEC strains to survive up to 20 days within a wild Acanthamoeba sp., in a quiescent state persisting in a non-cultivable form, until they reactivate following some stimulus of an unknown nature. Furthermore, our findings show that during their internalization, the E. coli O157 kept the set of the main virulence genes intact, preserving their pathogenetic potential. These observations suggest that the internalization in free-living amoebae may represent a means for STEC to resist in environments with non-permissive growth conditions. Moreover, by staying within the protozoa, STEC could escape their detection in the vehicles of infections and resist to the treatments used for the disinfection of the livestock environment. [ABSTRACT FROM AUTHOR]

Shiga toxin-producing Escherichia coli (STEC) strains are considered the most common food-borne zoonotic pathogen and are highly pathogenic to humans in low infectious doses, causing food-borne diseases through consumption of contaminated water or food. Resistance against antibiotics by STEC is a big concern nowadays. Two hundred farm water samples (Cattle-40, Buffaloes-40, Sheep-30, Goat-20, Pig-20 and poultry-50) were collected aseptically from different livestock farms in and around Proddatur, Andhra Pradesh, India and Processed for E. coli isolation, identification with culture method and molecular characterization by PCR. E. coli was characterized as STEC with two genes i.e. stx1 and stx2. All the STEC isolates were subjected to an antibiotic sensitivity test by disc diffusion method against ten antibiotics. Results showed that out of 200 farm water samples, 196 were positive for E. coli with an overall prevalence of 98% (196/200) and 62.2% (122/196) for STEC by PCR. Antimicrobial susceptibility test by disc diffusion method against ten antibiotics revealed the higher resistance to Cepahlothin (100%) followed by Tetracyclin (98.4%), Ampicillin (96.7%), Streptomycin (95%), Sulphonamides (91.8%), Trimethoprim (84.4%), Kanamycin (34.4%), Chloromphenicol (17.2%), Colistin (9.0%) and least resistance to Gentamycin (4.9%). [ABSTRACT FROM AUTHOR]

Raw milk is a nutrient-rich food that is considered a high-quality nutritional medium for many microorganism, including Escherichia coli. The aim of the present work was the diagnosis, by molecular methods, of Shiga toxins produced by E. coli strains isolated from cow milk samples collected from different farms in Al-Mahmmodia, Al yoosifya, Al lattiffya, Al howasha, and Arab Jboor in the government of Baghdad during the summer season. Milk samples were incubated in media for bacterial isolation. Isolates were identified using Gram staining and biochemical tests. Out of 50 samples, 15 (30%) showed the presence of E. coli. To confirm the identity of the isolates, their 16S rRNA genes were amplified using specific primers. The results showed that all isolates were E. coli. Shiga toxin-producing E. coli (STE C) were detected among the samples. The prevalence of stx1 genes was higher than that of stx2 among them. No STE Cs were found among six of the sample isolates, and none of these isolates was positive for stx1 and stx2. SDS-PAGE was used to determine the molecular weight of the toxin, and four selected E. coli bacteria producing Shiga-like toxins showed a clear band of approximately 70 kDa. [ABSTRACT FROM AUTHOR]

Background Giant cell myocarditis (GCM) is a rare but well-known cause of fulminant myocarditis. Despite optimal medical therapy, many patients progress to orthotopic heart transplant (OHT). We present a case of recurrent GCM following OHT, including complex considerations in patient management and infectious sequelae. Case summary A 33-year-old previously healthy male presented with 2 months of worsening shortness of breath. Transthoracic echocardiogram (TTE) demonstrated a left ventricular ejection fraction of 30–35%. After ruling out an ischaemic aetiology, he was discharged on guideline-directed medical therapy and later presented with productive cough, worsening dyspnoea on exertion, and diarrhoea. He was found to have elevated troponins and N-terminal pro-brain natriuretic peptide, lactic acidosis, progression of severe bi-ventricular dysfunction on TTE and right heart catheterization, and low cardiac index (1.0 L/min/m2) requiring inotropes. He then required left ventricular assist device as a bridge to OHT. Pathology of the apical core diagnosed GCM as the cause of his fulminant heart failure. He eventually underwent heart transplantation, which was complicated by recurrent GCM. Treatment required intensification of his immunosuppressive regimen, which led to multiple infectious sequelae including norovirus, Shiga-like toxin producing Escherichia coli , and disseminated nocardia of the lung and brain. As of the most recent follow-up, the patient is currently clinically stable. Discussion Although recurrent GCM after OHT has been reported in the literature, the prognosis is not well understood and there are no clear guidelines regarding management. This case summarizes clinical considerations, treatment strategies, and adverse effects of recurrent GCM treatment. [ABSTRACT FROM AUTHOR]

Influenza virus infects the host and transmits through the respiratory tract (i.e., the mouth and nose); therefore, the development of intranasal influenza vaccines that mimic the natural infection, coupled with an efficient mucosal adjuvant, is an attractive alternative to current parenteral vaccines. However, with the withdrawal of cholera toxin and Escherichia coli heat-labile endotoxin from clinical use due to side effects, there are no approved adjuvants for intranasal vaccines. Therefore, safe and effective mucosal adjuvants are urgently needed. Previously, we reported that one derivative of α-Galactosylceramide (α-GalCer), 7DW8-5, could enhance the protective efficacy of split influenza vaccine by injection administration. However, the mucosal adjuvanticity of 7DW8-5 is still unclear. In this study, we found that 7DW8-5 promotes the production of secret IgA antibodies and IgG antibodies and enhances the protective efficacy of the split influenza vaccine by intranasal administration. Furthermore, co-administration of 7DW8-5 with the split influenza vaccine significantly reduces the virus shedding in the upper and lower respiratory tract after lethal challenge. Our results demonstrate that 7DW8-5 is a novel mucosal adjuvant for the split influenza vaccine. [ABSTRACT FROM AUTHOR]

Tuberculosis (TB) is one of the top 10 leading causes of death worldwide. The recombinant BCG strain expressing the genetically detoxified A subunit of the thermolabile toxin from Escherichia coli (LTAK63) adjuvant (rBCG-LTAK63) has previously been shown to confer superior protection and immunogenicity compared to BCG in a murine TB infection model. To further investigate the immunological mechanisms induced by rBCG-LTAK63, we evaluated the immune responses induced by rBCG-LTAK63, BCG, and Mycobacterium tuberculosis (Mtb) H37Rv strains in experimental infections of primary human M1 and M2 macrophages at the transcriptomic and cytokine secretion levels. The rBCG-LTAK63-infected M1 macrophages more profoundly upregulated interferon-inducible genes such as IFIT3, OAS3, and antimicrobial gene CXCL9 compared to BCG, and induced higher levels of inflammatory cytokines such as IL-12(p70), TNF-β, and IL-15. The rBCG-LTAK63-infected M2 macrophages more extensively upregulated transcripts of inflammation-related genes, TAP1, GBP1, SLAMF7, TNIP1, and IL6, and induced higher levels of cytokines related to inflammation and tissue repair, MCP-3 and EGF, as compared to BCG. Thus, our data revealed an important signature of immune responses induced in human macrophages by rBCG-LTAK63 associated with increased inflammation, activation, and tissue repair, which may be correlated with a protective immune response against TB. [ABSTRACT FROM AUTHOR]

The heat-labile enterotoxins of Escherichia coli and cholera toxin of Vibrio cholerae are related in structure and function. Each of these oligomeric toxins is comprised of one A polypeptide and five B polypeptides. The B-subunits bind to gangliosides, which are followed by uptake into the intoxicated cell and activation of the host's adenylate cyclase by the A-subunits. There are two antigenically distinct groups of these toxins. Group I includes cholera toxin and type I heat-labile enterotoxin of E. coli ; group II contains the type II heat-labile enterotoxins of E. coli. Three variants of type II toxins, designated LT-IIa, LT-IIb and LT-IIc have been described. Earlier studies revealed the crystalline structure of LT-IIb. Herein the carbohydrate binding specificity of LT-IIc B-subunits was investigated by glycosphingolipid binding studies on thin-layer chromatograms and in microtiter wells. Binding studies using a large variety of glycosphingolipids showed that LT-IIc binds with high affinity to gangliosides with a terminal Neu5Acα3Gal or Neu5Gcα3Gal, e.g. the gangliosides GM3, GD1a and Neu5Acα3-/Neu5Gcα3--neolactotetraosylceramide and Neu5Acα3-/Neu5Gcα3-neolactohexaosylceramide. The crystal structure of LT-IIc B-subunits alone and with bound LSTd/sialyl-lacto- N -neotetraose d pentasaccharide uncovered the molecular basis of the ganglioside recognition. These studies revealed common and unique functional structures of the type II family of heat-labile enterotoxins. [ABSTRACT FROM AUTHOR]

The objective of the experiment was to evaluate a commercial Saccharomyces cerevisiae fermentation product (SCFP; NaturSafe, Dimond V, Cedar Rapids, IA) on performance, plasma metabolites, rumen parameters, and fecal pathogen shedding in feedlot steers. Crossbred steers (n = 61; 271 ± 48.1 kg) were sourced from 3 different research units at Oklahoma State University and assigned to 1 of 3 experimental treatments in a randomized complete block design. Steers were group housed in pens equipped with an automated individual feed intake system. Treatments consisted of a negative control (CON), CON with tylosin phosphate (TYL; 9.9 mg ⋅ kg-1 of diet DM), or CON with SCFP (1.63 g ⋅ kg-1 of diet DM). Body weight and feed efficiency did not differ among treatments (P ≥ 0.12). However, steers fed SCFP tended to have greater ADG from d 35 to 56 (P = 0.10). In addition, steers fed SCFP exhibited increased DMI from d 56 to 112 (P = 0.03), a tendency for increased DMI from d 112 to final (P = 0.07), and increased DMI overall (P = 0.05). There was no treatment x time interaction for pH, pH change, or lactate concentrations in rumen fluid (P ≥ 0.11), but all were impacted by time (P < 0.01). A treatment x time interaction (P < 0.01) was detected for fecal pH and fecal pH change, with the pH being greater in TYL fed steers on d 56 and in CON fed steers on the final day. Steers that consumed CON had a reduced fecal pH (P < 0.01) and steers consuming SCFP tended to have a reduced fecal pH (P = 0.08) from d 35 to 56 compared with TYL fed steers, while both CON and SCFP fed steers experienced an increase in fecal pH from d 56 to 112 (P < 0.01). A time effect was detected (P ≤ 0.04) for plasma glucose, lactate, urea nitrogen, and non-esterified fatty acid concentrations among treatment groups. A treatment x time interaction was observed (P < 0.02) for stx2 genes of Shiga toxin producing Escherichia coli (STEC) in individual fecal samples where SCFP and TYL fed steers tended to exhibit an increased concentration compared with CON fed steers on d 14, while on d 190 CON and SCFP fed steers exhibited an increased concentration compared with TYL fed steers. Overall, STEC, E. coli O157:H7, and Salmonella concentrations were all impacted by time (P < 0.001). This experiment suggests that SCFP could be utilized as a potential alternative to TYL in feedlot cattle without adversely affecting cattle performance. However, SCFP did not impact pathogen shedding. In addition, CON fed steers performed similarly to both SCFP and TYL fed steers. [ABSTRACT FROM AUTHOR]

Purpose: In the present study, whole diphtheria toxin (dt) and fragment B (dtb) genes from Corynebacterium diphtheriae Park William were cloned into Escherichia coli, the purified expressed proteins were evaluated for ultimately using as a candidate vaccine. Materials and Methods: The dt and dtb genes were isolated from bacterial strain ATCC (American Type Culture Collection) no. 13812. Plasmid pET29a+ was extracted by DNA-spin TM plasmid purification kit where genes were inserted using BamHI and HindIII-HF. Cloned pET29a+dt and pET29a+dtb plasmids were transformed into E. coli BL21(DE3)PlysS as expression host. The identity of the sequences was validated by blasting the sequence (BLASTn) against all the reported nucleotide sequences in the NCBI (National Center for Biotechnology Information) GenBank. Production of proteins in high yield by different types and parameters of fermentation to determine optimal conditions. Lastly, the purified concentrated rdtx and rdtb were injected to BALB/c mice and antibody titers were detected. Results: The genetic transformation of E. coli DH5α and E. coli BL21 with the pET-29a(+) carrying the dt and dtb genes was confirmed by colony polymerase chain reaction assay and were positive to grow on Luria-Bertani/kanamycin medium. The open reading frame of dt and dtb sequences consisted of 1,600 bp and 1,000 bp, were found to be 100% identical to dt and dtb sequence of C. diphtheriae (accession number KX702999.1 and KX702993.1) respectively. The optimal condition for high cell density is fed-batch fermentation production to express the rdtx and rdtb at 280 and 240 Lf/mL, dissolved oxygen was about 24% and 22% and the dry cell weight of bacteria was 2.41 g/L and 2.18 g/L, respectively. Conclusion: This study concluded with success in preparing genetically modified two strains for the production of a diphtheria vaccine, and to reach ideal production conditions to achieve the highest productivity. [ABSTRACT FROM AUTHOR]

Background: Previous reports have differed as to whether nitric oxide inhibits or stimulates the SOS response, a bacterial stress response that is often triggered by DNA damage. The SOS response is an important regulator of production of Shiga toxins (Stx) in Shiga-toxigenic E. coli (STEC). In addition, the SOS response is accompanied by hypermutation, which can lead to de novo emergence of antibiotic resistance. We studied these effects in vitro as well as in vivo. Results: Nitric oxide donors inhibited induction of the SOS response by classical inducers such as mitomycin C, ciprofloxacin, and zidovudine, as measured by assays for E. coli RecA. Nitric oxide donors also inhibited Stx toxin protein production as well as stx2 RNA in vitro and in vivo. In vivo experiments were performed with ligated ileal segments in the rabbit using a 20 h infection. The NO donor S-nitroso-acetylpenicillamine (SNAP) reduced hypermutation in vitro and in vivo , as measured by emergence of rifampin resistance. SNAP blocked the ability of the RecA protein to bind to single-stranded DNA in an electrophoretic mobility shift assay (EMSA) in vitro , an early event in the SOS response. The inhibitory effects of SNAP were additive with those of zinc acetate. Conclusions: Nitric oxide donors blocked the initiation step of the SOS response. Downstream effects of this blockade included inhibition of Stx production and of hypermutation. Infection of rabbit loops with STEC resulted in a downregulation, rather than stimulation, of nitric oxide host defenses at 20 h of infection. [ABSTRACT FROM AUTHOR]

The emergence of clinically significant antimicrobial resistance (AMR) in bacteria is frequently attributed to the use of antimicrobials in humans and livestock and is often found concurrently with human and animal pathogens. However, the incidence and natural drivers of antimicrobial resistance and pathogenic virulence in the environment, including waterways and ground water, are poorly understood. Freshwater monitoring for microbial pollution relies on culturing bacterial species indicative of faecal pollution, but detection of genes linked to antimicrobial resistance and/or those linked to virulence is a potentially superior alternative. We collected water and sediment samples in the autumn and spring from three rivers in Canterbury, New Zealand; sites were above and below reaches draining intensive dairy farming. Samples were tested for loci associated with the AMR-related group 1 CTX-M enzyme production (blaCTX-M) and Shiga toxin producing Escherichia coli (STEC). The blaCTX-M locus was only detected during spring and was more prevalent downstream of intensive dairy farms. Loci associated with STEC were detected in both the autumn and spring, again predominantly downstream of intensive dairying. This cross-sectional study suggests that targeted testing of environmental DNA is a useful tool for monitoring waterways. Further studies are now needed to extend our observations across seasons and to examine the relationship between the presence of these genetic elements and the incidence of disease in humans. [ABSTRACT FROM AUTHOR]

Inhomogeneous structure occurring in biological membranes being rich in glycosphingolipids (GSL) has been proposed as an important phenomenon involved in the cellular endocytosis process. However, little is known about the correlation between the formation of microdomains and the GSL-dependent biogenesis for tubular endocytic pits occurred on the surface of the cellular membrane. In the present work, the interaction between the bacterial Shiga toxin from Escherichia coli (STxB) and its cellular receptor GSL globotriaosylceramide (Gb3) were studied using small unilamellar vesicle (SUV). The model membrane invagination induced by STxB was determined by the contrast variation small-angle neutron scattering (SANS) and the synchrotron radiation facility based small-angle X-ray scattering (SR-SAXS). The results revealed that Gb3 molecules provided the binding sites for STxB, inducing increased membrane fluctuation. The formation of protein–lipid complex (STxB-Gb3) apparently induced the thinning of model membrane with the thickness decreased from 3.10 nm to 2.50 nm. It is the first time to successfully characterize the mesoscopic change on membrane thickness upon GSL-dependent endocytic process using a small-angle scattering technique. Overall, this paper provided a practical method to quantify the inhomogeneous biological membrane structures, which is important to understand the cellular endocytosis process. [ABSTRACT FROM AUTHOR]

Despite the prevalence of co-infections and the association of over 50 viral and 46 bacterial pathogens with pig diseases, little is known about their simultaneous occurrence, particularly in commercial pig farming environments where health programs are in place. To address this knowledge gap, this study aimed to evaluate the pathogen threshold of respiratory and enteric pathogens in pig herds using the Pork MultiPath™ (PMP1 and PMP2, respiratory and enteric respectively) technology, which detects multiple pathogens simultaneously in a single reaction with high sensitivity and specificity. In this study the most prevalent respiratory pathogens, Mycoplasma hyrohinis , Pasteurella multocida , and Haemophilus parasuis detected by PMP1 were effectively controlled during the nursery stage through strategic treatment with tiamulin. Even though the major respiratory incidences were reduced, the recorded coughing and sneezing rates were associated with the levels of H. parasuis and M. hyrohinis, which were set at 1356 and 1275 copies/reaction, respectively. In addition, one of the identified co-infection patterns indicated a strong relationship between the occurrence of H. parasuis and M. hyorhinis at the sample and pen levels, highlighting the high likelihood of detecting these two pathogens together. Testing with enteric panel PMP2 revealed that the most frequently detected virulence factors during the early nursery stage were Escherichia coli genes for toxins - ST1, ST2, and fimbriae – F4 and F18. Moreover, a co-infection with Rotavirus B and C was often observed during the nursery stage, and a strong positive correlation between these two markers has been identified. Additionally, the levels of several markers, namely E. coli F4, F5, F18, LT, ST1, and ST2, have been associated with a higher likelihood of sickness in pig populations. In addition, the onset of Brachyspira pilosicoli during the nursery and grower stages was found to be associated with an increased risk of diarrhoea, with a set threshold at around 500 copies/reaction. Although simultaneous detection of multiple pathogens is not yet widely used in the pig industry, it offers a significant advantage in capturing the diversity and interactions of co-infections. Testing pooled samples with Pork MultiPath™ is cost-effective and practical to regularly monitor the health status of pig populations. [ABSTRACT FROM AUTHOR]

The Hok/Gef family consists of structurally similar, single-span membrane peptides that all contain a positively charged N-terminal domain, an α-helix and a periplasmic C-terminal domain. Hok/Gef peptides have previously been described to play distinct physiological roles. Indeed, while HokB has been implicated in bacterial persistence, other members of the Hok/Gef family are known to induce cell lysis. However, the generalizability of previously published studies is problematic, as they have all used different expression systems. Therefore, we conducted a systematic study of the nine Hok/Gef peptides of Escherichia coli. We observed rapid cell death following expression of hokA , hokC , hokD , hokE , pndA1 , hok or srnB , while expression of hokB or pndA2 does not result in cell lysis. A remarkable feature of Hok/Gef peptides is the presence of conserved periplasmic tyrosine and/or cysteine residues. For the HokB peptide, one of these residues has previously been implicated in intermolecular dimerization, which is essential for HokB to exert its role in persistence. To assess the role of the periplasmic cysteine and tyrosine residues in other Hok/Gef peptides and to decipher whether these residues determine peptide toxicity, an array of substitution mutants were constructed. We found that these residues are important activators of toxicity for Hok, HokA and HokE peptides. Despite the loss of the cell killing phenotype in HokS31_Y48, HokAS29_S46 and HokES29_Y46, these peptides do not exert a persister phenotype. More research is needed to fully comprehend why HokB is the sole peptide of the Hok/Gef family that mediates persistence. [ABSTRACT FROM AUTHOR]

Ta nia A. T. Gomes 1 *, Ulrich Dobrindt 2, Mauricio J. Farfan 3,4 and Roxane M. F. Piazza 5 1 Departamento de Microbiologia, Imunologia e Parasitologia, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, Brazil, 2 Institute of Hygiene, University of Muenster, Muenster, Germany, 3 Laboratorio Cli 'nico, Hospital Dr. Luis Calvo Mackenna, Santiago, Chile, 4 Departamento de Pediatri 'ayCirugi 'a Infantil, Hospital Dr. Luis Calvo Mackenna, Facultad de Medicina, Universidad de Chile, Santiago, Chile, 5 Laborato 'riodeBacteriologia,InstitutoButantan,SãoPaulo,Brazil Keywords: Escherichia coli, virulence factors, genome, antibiotic resistance, evolution Editorial on the Research Topic Interaction of Pathogenic Escherichia coli With the Host: Pathogenomics, Virulence and Antibiotic Resistance Escherichia coli live as commensals in the intestines of humans and warm-blooded animals (Leimbach et al., 2013). HYBRID- AND HETERO-PATHOGENIC E. COLI STRAINS It has been reported that certain pathogenic E. coli strains combine different pathotypes' main virulence traits, encompassing potentially more virulent hybrid strains (Dobrindt et al., 2003; Bielaszewska et al., 2007; Khan et al., 2018). Valiatti et al. characterized geno- and phenotypically uropathogenic E. coli isolate 252 (UPEC 252) and demonstrated that this strain represents an atypical enteropathogenic E. coli (EPEC) strain. Although most E. coli strains rarely produce disease in healthy individuals, pathogenic strains can cause a wide range of diseases in the gastrointestinal tract or extraintestinal sites in healthy and immunocompromised individuals (Kaper et al., 2004). [Extracted from the article]

Shiga toxin (Stx) is the main virulence factor of Shiga toxin-producing Escherichia coli (STEC), and ruminants are the main reservoir of STEC. This study assessed the abundance and expression of Stx genes and the expression of host immune genes, aiming to determine factors affecting these measures and potential gene markers to differentiate Stx gene expression in the recto-anal junction of feedlot beef cattle. Rectal tissue and content samples were collected from 143 feedlot steers of three breeds (Angus, Charolais, and Kinsella Composite) over 2 consecutive years 2014 (n=71) and 2015 (n=72). The abundance and expression of stx1 and stx2 were quantified using qPCR and reverse-transcription-qPCR (RT-qPCR), respectively. Four immune genes (MS4A1, CCL21, CD19 , and LTB), previously reported to be down-regulated in super-shedder cattle (i.e., > 104 CFU g-1) were selected, and their expression was evaluated using RT-qPCR. The stx1 gene abundance was only detected in tissue samples collected in year 2 and did not differ among breeds. The stx2 gene was detected in STEC from all samples collected in both years and did not vary among breeds. The abundance of stx1 and stx2 differed (P < 0.001) in content samples collected across breeds (stx1 :AN>CH>KC, stx2 : AN=CH>KC) in year 1, but not in year 2. Expression of stx2 was detected in 13 RAJ tissue samples (2014: n=6, 2015: n=7), while expression of stx1 was not detected. Correlation analysis showed that the expression of stx2 was negatively correlated with the expression of MS4A1 (R=-0.56, P=0.05) and positively correlated with the expression of LTB (R=0.60, P=0.05). The random forest model and Boruta method revealed that expression of selected immune genes could be predictive indicators of stx2 expression with prediction accuracy of MS4A1 > LTB > CCL21 > CD19. Our results indicate that the abundance of Stx could be affected by cattle breed and sampling year, suggesting that host genetics and environment may influence STEC colonization of the recto-anal junction of feedlot cattle. Additionally, the identified relationship between expressions of host immune genes and stx2 suggests that the host animal may regulate stx2 expression in colonizing STEC through immune functions. [ABSTRACT FROM AUTHOR]

Non-O157 STEC are increasingly linked to foodborne infections, yet little is known about the diversity and molecular epidemiology across locations. Herein, we used whole genome sequencing to examine genetic variation in 894 isolates collected from Michigan patients between 2001 and 2018. In all, 67 serotypes representing 69 multilocus sequence types were identified. Serotype diversity increased from an average of four (2001–2006) to 17 (2008–2018) serotypes per year. The top six serogroups reported nationally caused > 60% of infections in 16 of the 18 years; serogroups O111 and O45 were associated with hospitalization as were age ≥ 65 years, diarrhea with blood and female sex. Phylogenetic analyses of seven multilocus sequence typing (MLST) loci identified three clades as well as evidence of parallel evolution and recombination. Most (95.5%) isolates belonged to one clade, which could be further differentiated into seven subclades comprising isolates with varying virulence gene profiles and serotypes. No association was observed between specific clades and the epidemiological data, suggesting that serogroup- and serotype-specific associations are more important predictors of disease outcomes than lineages defined by MLST. Molecular epidemiological studies of non-O157 STEC are important to enhance understanding of circulating strain distributions and traits, genetic variation, and factors that may impact disease risk and severity. [ABSTRACT FROM AUTHOR]

Shiga-toxin-producing Escherichia coli (STEC) represents a significant cause of foodborne disease. In the last years, an increasing number of STEC infections associated with the consumption of raw and pasteurized milk cheese have been reported, contributing to raise the public awareness. The aim of this study is to evaluate the main genomic features of STEC strains isolated from a semi-hard raw milk cheese, focusing on their pathogenic potential. The analysis of 75 cheese samples collected during the period between April 2019 and January 2020 led to the isolation of seven strains from four stx -positive enrichment. The genome investigation evidenced the persistence of two serotypes, O174:H2 and O116:H48. All strains carried at least one stx gene and were negative for eae gene. The virulence gene pattern was homogeneous among the serogroup/ST and included adherence factors (lpfA , iha , ompT , papC , saa , sab , hra , and hes), enterohemolysin (ehxA), serum resistance (iss , tra), cytotoxin-encoding genes like epeA and espP , and the Locus of Adhesion and Autoaggregation Pathogenicity Islands (LAA PAIs) typically found in Locus of Enterocyte Effacement (LEE)-negative STEC. Genome plasticity indicators, namely, prophagic sequences carrying stx genes and plasmid replicons, were detected, leading to the possibility to share virulence determinants with other strains. Overall, our work adds new knowledge on STEC monitoring in raw milk dairy products, underlining the fundamental role of whole genome sequencing (WGS) for typing these unknown isolates. Since, up to now, some details about STEC pathogenesis mechanism is lacking, the continuous monitoring in order to protect human health and increase knowledge about STEC genetic features becomes essential. [ABSTRACT FROM AUTHOR]