Characterization of diarrheagenic Escherichia coli from different cattle production systems in Brazil.

Diarrheagenic E. coli (DEC) can cause severe diarrhea and is a public health concern worldwide. Cattle are an important reservoir for this group of pathogens, and once introduced into the abattoir environment, these microorganisms can contaminate consumer products. This study aimed to characterize the distribution of DEC [Shiga toxin-producing E. coli (STEC), enteroinvasive E. coli (EIEC), enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), and enteroaggregative E. coli (EAEC)] from extensive and intensive cattle production systems in Brazil. Samples (n = 919) were collected from animal feces (n = 200), carcasses (n = 600), meat cuts (n = 90), employee feces (n = 9), and slaughterhouse water (n = 20). Virulence genes were detected by PCR in 10% of animal samples (94/919), with STEC (n = 81) as the higher prevalence, followed by EIEC (n = 8), and lastly EPEC (n = 5). Animals raised in an extensive system had a higher prevalence of STEC (average 48%, sd = 2.04) when compared to animals raised in an intensive system (23%, sd = 1.95) (Chi-square test, P < 0.001). From these animals, most STEC isolates only harbored stx2 (58%), and 7% were STEC LEE-positive isolates that were further identified as O157:H7. This study provides further evidence that cattle are potential sources of DEC, especially STEC, and that potentially pathogenic E. coli isolates are widely distributed in feces and carcasses during the slaughter process.

Determinação do potencial do polissacarídeo O157 como antígeno vacinal contra a Escherichia coli produtora de toxina Shiga (STEC)

Diarrheal diseases persist as a global concern due to their significant impact on health. Diarrhea, responsible for the death of 370,000 children worldwide in 2019, ranks as the second leading cause of mortality in children under five. Diverse factors, such as infections by parasitic, viral, or bacterial pathogens, trigger episodes of diarrhea, with Escherichia coli standing out, associated with hemolytic-uremic syndrome (HUS). The E. coli O157:H7 serotype, known for its Shiga toxin production, is linked to gastrointestinal infections and HUS, particularly in children. Treatment primarily involves supportive measures, as antibiotic exposure can exacerbate symptoms, increasing the risk of HUS and other complications. Due to this serotype's association with foodborne outbreaks, strict surveillance in food production, sanitation practices, and processes is essential to prevent infections caused by these pathogens. This study evaluates the potential of O157 polysaccharide as a vaccine candidate against Shiga toxin-producing E. coli (STEC). Results indicate that antibodies generated against O157 polysaccharide could recognize E. coli O157:H7 even in the presence of a capsule. They also assisted the complement system in lysing these bacteria and inhibited their adherence to human epithelial cells. In summary, the findings suggest that O157 polysaccharide could be considered a promising antigen in vaccine formulations against Shiga toxin-producing E. coli.

As doenças diarreicas persistem como uma preocupação global devido ao seu impacto significativo na saúde. A diarreia, responsável pela morte de 370 mil crianças em 2019 no mundo, é a segunda principal causa de óbito em crianças menores de cinco anos. Fatores diversos, como infecções por patógenos parasitários, virais ou bacterianos, desencadeiam episódios de diarreia, destacando-se Escherichia coli, associada à síndrome hemolítico-urêmica (SHU). O sorotipo E. coli O157:H7, reconhecido por sua produção de toxinas Shiga, está associado a infecções gastrointestinais e SHU, especialmente em crianças. O tratamento envolve principalmente medidas de suporte, pois a exposição a antibióticos pode intensificar os sintomas, aumentando o risco de SHU e outras complicações. Devido à associação deste sorotipo com surtos alimentares, é essencial uma vigilância rigorosa na produção de alimentos, práticas sanitárias e processos que resultem na prevenção de infeções causadas por esses patógenos. Esse estudo, portanto, avalia o potencial do polissacarídeo O157 como agente vacinal contra E. coli produtora de toxina Shiga (STEC). Os resultados mostraram que anticorpos gerados contra o polissacarídeo O157 foram capazes de reconhecer E. coli O157:H7 mesmo na presença de cápsula. Eles também auxiliaram o sistema complemento na lise dessas bactérias bem como inibiram a adesão das mesmas a células epiteliais humanas. Em suma, os resultados sugerem que o polissacarídeo O157 pode ser considerado um bom candidato a antígeno em formulações vacinais contra E. coli produtoras de toxina Shiga.

Molecular and Genetic Characterization of Colicinogenic Escherichia coli Strains Active against Shiga Toxin-Producing Escherichia coli O157:H7.

The objective of this work was to molecularly and genotypically characterize and test the inhibitory activity of six colicinogenic Escherichia coli strains (ColEc) and their partially purified colicins against STEC O157:H7 isolated from clinical human cases. Inhibition tests demonstrated the activity of these strains and their colicins against STEC O157:H7. By PCR it was possible to detect colicins Ia, E7, and B and microcins M, H47, C7, and J25. By genome sequencing of two selected ColEc strains, it was possible to identify additional colicins such as E1 and Ib. No genes coding for stx1 and stx2 were detected after analyzing the genome sequence. The inhibitory activity of ColEc against STEC O157:H7 used as an indicator showed that colicins are potent growth inhibitors of E. coli O157:H7, being a potential alternative to reduce the presence of pathogens of public health relevance.

The Cantaloupe Farm Environment Has a Diverse Genetic Pool of Antibiotic-Resistance and Virulence Genes.

Cantaloupes contaminated with pathogens have led to many high-profile outbreaks and illnesses. Since bacterial virulence genes (VGs) can act in tandem with antibiotic-resistance and mobile genetic elements, there is a need to evaluate these gene reservoirs in fresh produce, such as cantaloupes. The goal of this study was to assess the distribution of antibiotic-resistance, virulence, and mobile genetic elements genes (MGEGs) in cantaloupe farm environments. A total of 200 samples from cantaloupe melons (n = 99), farm workers' hands (n = 66), and production water (n = 35) were collected in México. Each sample was assayed for the presence of 14 antibiotic-resistance genes, 15 VGs, and 5 MGEGs by polymerase chain reaction. Our results indicated that tetracycline (tetA and tetB) (18% of cantaloupe, 45% of hand samples) and sulfonamide (sul1) (30% of cantaloupe, 71% of hand samples) resistance genes were frequently detected. The colistin resistance gene (mcr1) was detected in 10% of cantaloupe and 23% of farm workers' hands. Among VGs, Salmonella genes invA and spiA were the most abundant. There was a significantly higher likelihood of detecting antibiotic-resistance, virulence, and MGEGs on hands compared with water samples. These results demonstrate a diverse pool of antibiotic-resistance and VGs in cantaloupe production.

Antimicrobial and Antibiofilm Activities of Essential Oils against Escherichia coli O157:H7 and Methicillin-Resistant Staphylococcus aureus (MRSA).

The emergence of multidrug resistant microorganisms represents a global challenge due to the lack of new effective antimicrobial agents. In this sense, essential oils (EOs) are an alternative to be considered because of their anti-inflammatory, antiviral, antibacterial, and antibiofilm biological activities. Therefore, multiple efforts have been made to consider the potential use of EOs in the treatment of infections which are caused by resistant microorganisms. In this study, 15 EOs of both Colombian and introduced aromatic plants were evaluated against pathogenic strains of E. coli O157:H7 and methicillin resistant Staphylococcus aureus (MRSA) in planktonic and sessile states in order to identify relevant and promising alternatives for the treatment of microbial infections. Forty different compounds were identified in the 15 EO with nine of them constituted mainly by oxygenated monoterpenes (OM). EOs from Lippia origanoides, chemotypes thymol, and carvacrol, displayed the highest antibacterial activity against E. coli O157:H7 (MIC50 = 0.9 and 0.3 mg/mL, respectively) and MRSA (MIC50 = 1.2 and 0.6 mg/mL, respectively). These compounds from EOs had also the highest antibiofilm activity (inhibition percentage > 70.3%). Using scanning electron microscopy (SEM), changes in the size and morphology of both bacteria were observed when they were exposed to sub-inhibitory concentrations of L. origanoides EO carvacrol chemotype. EOs from L. origanoides, thymol, and carvacrol chemotypes represented a viable alternative for the treatment of microbial infections; however, the Selectivity Index (SI ≤ 3) indicated that it was necessary to study alternatives to reduce its in vitro cytotoxicity.

Potent and Specific Antibacterial Activity against Escherichia coli O157:H7 and Methicillin Resistant Staphylococcus aureus (MRSA) of G17 and G19 Peptides Encapsulated into Poly-Lactic-Co-Glycolic Acid (PLGA) Nanoparticles.

Antimicrobial peptides constitute an excellent alternative against conventional antibiotics because of their potent antimicrobial spectrum, unspecific action mechanism and low capacity to produce antibiotic resistance. However, a potential use of these biological molecules as therapeutic agents is threatened by their low stability and susceptibility to proteases. In order to overcome these limitations, encapsulation in biocompatible polymers as poly-lactic-glycolic-acid (PLGA) is a promising alternative for increasing their stability and bioavailability. In this work, the effect of new synthetic antimicrobial peptides GIBIM-P5S9K (G17) and GAM019 (G19) encapsulated on PLGA and acting against methicillin resistant Staphylococus aureus (MRSA) and Escherichia coli O157:H7 was studied. PLGA encapsulation allowed us to load around 7 µg AMPs/mg PLGA with an efficiency of 90.5%, capsule sizes around 290 nm and positive charges. Encapsulation improved antimicrobial activity, decreasing MIC50 from 1.5 to 0.2 (G17NP) and 0.7 (G19NP) µM against MRSA, and from 12.5 to 3.13 µM for E. coli O157:H7. Peptide loaded nanoparticles could be a bacteriostatic drug with potential application to treat these bacterial E. coli O157:H7 and MRSA infections, with a slow and gradual release.

The use of High-Pressure Processing (HPP) to improve the safety and quality of raw coconut (Cocos nucifera L) water.

This research investigated the use of high-pressure processing (HPP) for inactivating vegetative pathogens and spoilage microbiota in fresh unfiltered coconut water (Cocos nucifera L) from nuts obtained from Florida and frozen CW from Brazil with pH >5.0 and storage at 4 °C. Additionally, CW was evaluated to determine if it supported the growth and toxin production of Clostridium botulinum with or without the use of HPP when stored at refrigeration temperatures. Samples of fresh unfiltered CW were inoculated to 5.5 to 6.5 logs/mL with multiple strain cocktails of E. coli O157:H7, Salmonella spp. and Listeria monocytogenes and HPP at 593 MPa for 3 min at 4 °C. HPP and inoculated non-HPP controls were stored at 4 °C for 54 and 75 days for Florida CW and Brazil CW, respectively. Results of analyses showed HPP samples with <1 CFU/mL and no detection (negative/25 mL) with enrichment procedures for the 3 inoculated pathogens for all analyses. The non-HPP control samples did not show growth of the pathogens but a gradual decrease in levels to ca. 3-Logs/mL by day 54 in the fresh Florida CW and similarly in frozen Brazil CW by Day 75. Microbial spoilage of uninoculated samples was evaluated for normal spoilage microbiota through 120 days storage at 4 °C. Microbial counts remained at ca. 2-logs with no detectable signs of spoilage for HPP samples through 120 d. The non-HPP control samples spoiled within 2 weeks of storage at 4 °C with gas production, cloudiness, and off-odors. To evaluate if CW supports the growth and toxin production of C. botulinum, samples of unfiltered and filtered (0.2 µm) CW were inoculated with either proteolytic or non-proteolytic C. botulinum spores at 2 log CFU/mL that were processed at 593 MPa for 3 min and stored at 4 °C and 10 °C for 45 days. Inoculated positive and non-inoculated negative controls were prepared and stored as the HPP treated and non-HPP samples. No growth of C. botulinum or toxin production was detected in either the unfiltered or filtered CW regardless if products were HPP treated or not. All inoculated samples with C. botulinum spores were enriched at Day-45 in PYGS media to determine the viability of the inoculated spores at the end of shelf-life and screened for C. botulinum toxins. In all samples, C. botulinum toxin Types A, B and E were detected indicating spores were viable throughout the storage. Type F toxin was not detected possibly due to inherent conditions in the samples that may affected toxin screening.

First report of Escherichia coli O157:H7 in ready-to-eat sushi.

AIMS: The aim of this study was to evaluate the microbiological quality of commercially prepared ready-to-eat (RTE) sushi by enumerating aerobic mesophilic bacteria (AMB) and thermotolerant coliforms (TC) and detecting Escherichia coli and Salmonella ssp. An isolate was identified as E. coli O157:H7 which was evaluated for its virulence and antimicrobial resistance profiling as well as its ability to form biofilms on stainless steel. METHODS AND RESULTS: There were four sampling events in seven establishments, totalling 28 pools of sushi samples. Mean AMB counts ranged between 5·2 and 7·7 log CFU per gram. The enumeration of TC varied between 2·1 and 2·7 log MPN per gram. Salmonella ssp. were not detected, and one sample was positive for E. coli and was identified as E. coli O157:H7. To the best of our knowledge, this is the first report of E. coli O157:H7 in sushi samples in the world literature. This isolate presented virulence genes stx1, stx2, eae and hlyA. It was also susceptible to 14 antimicrobials tested and had the ability to form biofilms on stainless steel. CONCLUSIONS: There is a need to improve the good hygiene practices adopted in establishments selling sushi in the city of Pelotas, Brazil. In addition, the isolated E. coli O157:H7 carries a range of important virulence genes being a potential risk to consumer health, as sushi is a RTE food. This isolate also presents biofilm formation ability, therefore, may trigger a constant source of contamination in the production line of this food. SIGNIFICANCE AND IMPACT OF THE STUDY: The increase in the consumption of sushi worldwide attracts attention regarding the microbiological point of view, since it is a ready-to-eat food. To our knowledge, this was the first time that E. coli O157:H7 was identified in sushi samples.

Survival of Lactobacillus acidophilus LA-5 and Escherichia coli O157:H7 in Minas Frescal cheese made with oregano and rosemary essential oils.

The effects of the incorporation of the essential oils from Origanum vulgare L. (OVEO; 0.07 µL/g) and Rosmarinus officinalis L. (ROEO; 2.65 µL/g) in combination in Minas Frescal cheese on the counts of the probiotic Lactobacillus acidophilus LA-5 and Escherichia coli O157:H7 were evaluated during refrigerated storage (7 ±â€¯0.5 °C). The terpenes of OVEO and ROEO, survival of the probiotic strain during in vitro digestion, as well as the physicochemical and sensory aspects were also monitored in Minas Frescal cheese. All terpenes decreased in cheese when the storage time increased. The incorporation of OVEO and ROEO delayed the increase in L. acidophilus LA-5 counts in cheese, but did not affect its ability to survive in cheese under simulated gastrointestinal conditions. The decreases in counts of E. coli O157:H7 observed in the first 15 days of refrigerated storage were strongly correlated (r ≥ 0.82) with the terpenes detected in cheese. Scores attributed for aroma, flavor, overall impression and purchase intention of cheese with OVEO and ROEO increased with the increase of the storage time. The incorporation of OVEO and ROEO in combination could be a strategy to control E. coli O157:H7 in probiotic Minas cheese during storage; however, the amounts of these substances should be cautiously selected considering possible negative sensory impacts in this product.

Evaluación de Lactobacillus plantarum microencapsulado y su viabilidad bajo condiciones gastrointestinales simuladas e inhibición frente a Escherichia coli O157:H7 / Evaluation of microencapsulated Lactobacillus plantarum and its viability under simulated gastrointestinal conditions and inhibition against Escherichia coli O157: H7

RESUMEN Se evaluó la viabilidad de Lactobacillus plantarum microencapsulado, su efecto de inhibición sobre Escherichia coli O157:H7 y su crecimiento en condiciones gastrointestinales simuladas. Se utilizaron L. plantarum ATCC 8014® y E. coli ATCC 43888®. Se realizaron pruebas de inhibición de L. plantarum sobre E. coli y test de susceptibilidad a CTX (30 µg), P (10 IU), GN (10 µg), DCX (1 µg), CIP (5 µg) y KF (30 µg) para ambas cepas. En la bacteria láctica se determinó la cinética de fermentación y la presencia de péptidos y aminoácidos por HPLC en ambas cepas. Se evaluó el crecimiento a 37 y 45°C de L. plantarum y se valoró la viabilidad de su microencapsulación mediante condiciones gastrointestinales (bilis, sales biliares y pH ácido), al igual que la supervivencia y estabilidad de preparado y sus características físicas y morfológicas. Los resultados indicaron que L. plantarum inhibió a E. coli y el microencapsulado, resultados positivos con una viabilidad del 83,3%; eficiencia de 88,4%; humedad de 7,79%; actividad de agua 0,4; humectabilidad de 1 min, 56 s; solubilidad del 96%; morfología esférica y tamaño entre 15,18 a 35,68 pm. Finalmente, se observó un alto potencial de L. plantarum como agente inhibidor para E. coli O157:H7.

ABSTRACT The viability of microencapsulated Lactobacillus plantarum, its inhibition effect on Escherichia coli O157: H7 and growth in simulated gastrointestinal conditions was evaluated. L. plantarum ATCC 8014® and E. coli ATCC 43888® were used. Inhibition tests of L. plantarum on E. coli and susceptibility test to CTX (30 µg), P (10 IU), GN (10 µg) DCX (1 µg), CIP (5 µg) and KF (30) were performed µg) for both strains. In the lactic bacterium the fermentation kinetics were determined, and the presence of peptides and amino acids by HPLC in both strains. The growth at 37 ° C and 45 ° C of L. plantarum was evaluated and the viability of its microencapsulation was assessed by gastrointestinal conditions (Bile, Bile salts and acidic pH), as well as the survival and stability of the preparation and its physical characteristics and morphological. The results indicated that L. plantarum inhibited E. coli and the microencapsulated positive results with a viability of 83.3%, efficiency 88.4%, Humidity 7.79%, water activity 0.4, wettability of 1 min, 56 s, 96% solubility, spherical morphology and size between 15.18 to 35.68 pm. Finally, a high potential of L. plantarum was observed as an inhibitory agent for E. coli O157: H7.

New Synthetic Peptides Conjugated to Gold Nanoclusters: Antibiotic Activity Against Escherichia coli O157:H7 and Methicillin-Resistant Staphylococcus aureus (MRSA).

Gold nanoclusters protected with bovine serum albumin (AuNC) can be used in multiple biomedical applications through functionalization with two new and bioactive peptides. Both cationic peptides sequences of 17 amino acids in length and the cysteine residue at its C-terminus were designed and synthesized. Peptides were obtained by solid phase synthesis using the Fmoc strategy. Peptides may be coupled via disulfide bonds to AuNC with hydrodynamic size ~ 2 nm ± 0.3 determined by dynamic light scattering and it had a zeta potential value equal to - 42 mV. Peptides named NBC2253 and NBC2254 were attached to the AuNC using N-succinimidyl-3-(2-pyridyl-dithiol) propionate as crosslinker agent. AuNC@NBC2253 was more active against methicillin-resistant Staphylococcus aureus (MIC50 6.5 µM) and AuNC@NBC2254 exhibited higher antimicrobial activity than the free peptides on Escherichia coli O157:H7 (MIC50 3.5 µM). No hemolysis was detected for any of the peptides. It is evidenced that these antimicrobial peptides conjugated to AuNC serve as promising agents to combat some multi-resistant bacterial strains and that the specific binding of these antimicrobial peptides to gold nanoclusters improves the interaction of these nanostructured systems with the biological target.

Síndrome hemolítico urémico asociado a Shigatoxina: ¿Cómo prevenirlo? / Shiga-toxin associated hemolytic uremic syndrome: how to prevent it?

Resumen: El síndrome hemolítico urémico (SHU) asociado a infección intestinal por bacterias productoras de Shigatoxina, que afecta principalmente a población infantil, puede causar morbilidad aguda grave, secuelas crónicas en varios órganos, y la muerte prematura en algunos de ellos. Dado su carácter zoonótico, adecuadas medidas de manejo agropecuario y correcta higiene de lo que consumimos es indispensable a la hora de prevenir la infección. Actualmente, una vez gatillado el SHU el manejo es médico y, principalmente, de soporte. En los últimos años diversas estrategias terapéuticas se han ido desarrollando para evitar que esta enfermedad ocurra, o, al menos, que pueda ser atenuada en sus consecuencias de morbi-mortalidad. El presente artículo describe acciones específicas a diferentes niveles de prevención de esta patología.

Abstract Hemolytic uremic syndrome (HUS) associated with intestinal infection by Shiga toxin-producing bacteria, which mainly affects children, can cause severe acute morbidity, chronic sequelae in seve ral organs, and premature death in some of them. Given its zoonotic nature, adequate measures of agricultural management and proper hygiene of what we consume are essential to prevent infection. Once the HUS is triggered, medical management is currently mainly supportive. In recent years, va rious therapeutic strategies have been developed to prevent this disease from occurring or, at least, to mitigate its morbidity and mortality consequences. This article describes specific actions at different levels of prevention of this pathology.

Foods introduced into Brazil through the border with Argentina and Uruguay: Pathogen detection and evaluation of hygienic-sanitary quality.

This study aimed to evaluate the presence of pathogens in, and the hygienic-sanitary quality of, commercialized foods of animal origin at the international border region of Rio Grande do Sul, Brazil. In total, 270 samples of raw and processed foods of animal origin were collected in Paso de los Libres, Argentina (n = 65 raw meat, n = 47 dairy products, n = 28 processed meat) and Rivera, Uruguay (n = 60 raw meat, n = 31 dairy products, n = 29 processed meat), or were seized by the Brazilian International Agricultural Surveillance System (Brazil-Argentina border) (n = 9 raw meat, n = 1 bush meat). The samples were subjected to the enumeration of aerobic mesophilic bacteria, enterobacteria, and coagulase-positive staphylococci, and were tested for Salmonella spp., Listeria monocytogenes, and Escherichia coli O157:H7. The virulence genes for Salmonella spp. (hilA, invA, spvC, pefA, and sefA), L. monocytogenes (prs, inlA, inlC, and inlJ) and E. coli O157:H7 (uspA, eae, rfbO157, fliCH7, stx1, stx2, and hlyA) were investigated using PCR assays. Raw products showed higher counts of aerobic mesophiles and enterobacteria compared to processed products (P < 0.05). There were no significant differences in aerobic mesophile or in enterobacterial counts between identical products according to origin (Argentina vs. Uruguay, P > 0.05). Escherichia coli O157:H7 was not detected in any of the samples tested. Salmonella spp. was detected in six (8%) raw products from Argentina. Listeria monocytogenes was isolated from five (6.66%) raw products originating in Argentina and 20 (16.66%) raw products from Uruguay. All 52 E. coli isolates carried the uspA gene, but only one carried the eae gene. The rfbO157, fliCH7, stx1, stx2, and hlyA genes were not detected. All Salmonella spp. isolates carried hilA and invA genes, but spvC, pefA, and sefA were not found. All L. monocytogenes isolates carried the prs gene; however, inlA, inlC, and inlJ genes were found in 20% of the isolates from Argentina and 95% of those from Uruguay. To our knowledge, this is the first microbiological study into the hygienic-sanitary quality of animal products in Brazil's land border region. Salmonella spp. and L. monocytogenes were detected in products of animal origin, constituting a public health concern and emphasizing the need for an active surveillance system to reduce the risk of foodborne pathogen introduction into Brazil.

Growth kinetics of Escherichia coli O157: H7 on the epicarp of fresh vegetables and fruits

Despite the increasing reports on the incidence of fresh vegetables and fruits as a possible vehicle for human pathogens, there is currently limited knowledge on the growth potential of Escherichia coli O157:H7 on different plant substrates. This study analyzed the selective adhesion and growth of E. coli O157:H7 on chili habanero (Capsicum chinense L.), cucumber (Cucumis sativus), radish (Raphanus sativus), tomato (Lycopersicon esculentum), beet (Beta vulgaris subsp. vulgaris), and onion (Allium cepa L.) under laboratory conditions. The Gompertz parameters were used to determine the growth kinetics. Scanning electron microscopy was used to visualize the adhesion of E. coli O157:H7 on the epicarp of the samples. Predictive models were constructed to compare the growth of E. coli O157:H7 on the samples with different intrinsic factors and to demonstrate the low selectivity of the pathogen. No significant difference was observed in the lag-phase duration (LPD), generation time (GT), and exponential growth rate (EGR) of the pathogen adhered to the samples. The interaction between the microorganism and the substrate was less supportive to the growth of E. coli O157:H7 for onion, whereas for tomato and cucumber, the time for the microorganism to attain the maximum growth rate (M) was significantly longer than that recorded for other samples.(AU)

Growth kinetics of Escherichia coli O157: H7 on the epicarp of fresh vegetables and fruits

ABSTRACT Despite the increasing reports on the incidence of fresh vegetables and fruits as a possible vehicle for human pathogens, there is currently limited knowledge on the growth potential of Escherichia coli O157:H7 on different plant substrates. This study analyzed the selective adhesion and growth of E. coli O157:H7 on chili habanero (Capsicum chinense L.), cucumber (Cucumis sativus), radish (Raphanus sativus), tomato (Lycopersicon esculentum), beet (Beta vulgaris subsp. vulgaris), and onion (Allium cepa L.) under laboratory conditions. The Gompertz parameters were used to determine the growth kinetics. Scanning electron microscopy was used to visualize the adhesion of E. coli O157:H7 on the epicarp of the samples. Predictive models were constructed to compare the growth of E. coli O157:H7 on the samples with different intrinsic factors and to demonstrate the low selectivity of the pathogen. No significant difference was observed in the lag-phase duration (LPD), generation time (GT), and exponential growth rate (EGR) of the pathogen adhered to the samples. The interaction between the microorganism and the substrate was less supportive to the growth of E. coli O157:H7 for onion, whereas for tomato and cucumber, the time for the microorganism to attain the maximum growth rate (M) was significantly longer than that recorded for other samples.

Immunization of pregnant cows with Shiga toxin-2 induces high levels of specific colostral antibodies and lactoferrin able to neutralize E. coli O157:H7 pathogenicity.

E. coli O157:H7 is a foodborne pathogen responsible for bloody diarrhea, hemorrhagic colitis and hemolytic uremic syndrome (HUS). The objective of the present work was to evaluate the ability of colostral IgG obtained from Stx2-immunized cows to prevent against E. coli O157:H7 infection and Stx2 cytotoxicity. Hyperimmune colostrum (HC) was obtained from cows intramuscularly immunized with inactivated Stx2 or vehicle for controls. Colostral IgG was purified by affinity chromatography. Specific IgG antibodies against Stx2 and bovine lactoferrin (bLF) levels in HC and the corresponding IgG (HC-IgG/bLF) were determined by ELISA. The protective effects of HC-IgG/bLF against Stx2 cytotoxicity and adhesion of E. coli O157:H7 and its Stx2-negative mutant were analyzed in HCT-8 cells. HC-IgG/bLF prevention against E. coli O157:H7 was studied in human colon and rat colon loops. Protection against a lethal dose of E. coli O157:H7 was evaluated in a weaned mice model. HC-IgG/bLF showed high anti-Stx2 titers and high bLF levels that were able to neutralize the cytotoxic effects of Stx2 in vitro and in vivo. Furthermore, HC-IgG/bLF avoided the inhibition of water absorption induced by E. coli O157:H7 in human colon and also the pathogenicity of E. coli O157:H7 and E. coli O157:H7Δstx2 in rat colon loops. Finally, HC-IgG/bLF prevented in a 100% the lethality caused by E. coli O157:H7 in a weaned mice model. Our study suggests that HC-IgG/bLF have protective effects against E. coli O157:H7 infection. These beneficial effects may be due to specific anti-Stx2 neutralizing antibodies in combination with high bLF levels. These results allow us to consider HC-IgG/bLF as a nutraceutical tool which could be used in combination with balanced supportive diets to prevent HUS. However further studies are required before recommendations can be made for therapeutic and clinical applications.

Growth kinetics of Escherichia coli O157:H7 on the epicarp of fresh vegetables and fruits.

Despite the increasing reports on the incidence of fresh vegetables and fruits as a possible vehicle for human pathogens, there is currently limited knowledge on the growth potential of Escherichia coli O157:H7 on different plant substrates. This study analyzed the selective adhesion and growth of E. coli O157:H7 on chili habanero (Capsicum chinense L.), cucumber (Cucumis sativus), radish (Raphanus sativus), tomato (Lycopersicon esculentum), beet (Beta vulgaris subsp. vulgaris), and onion (Allium cepa L.) under laboratory conditions. The Gompertz parameters were used to determine the growth kinetics. Scanning electron microscopy was used to visualize the adhesion of E. coli O157:H7 on the epicarp of the samples. Predictive models were constructed to compare the growth of E. coli O157:H7 on the samples with different intrinsic factors and to demonstrate the low selectivity of the pathogen. No significant difference was observed in the lag-phase duration (LPD), generation time (GT), and exponential growth rate (EGR) of the pathogen adhered to the samples. The interaction between the microorganism and the substrate was less supportive to the growth of E. coli O157:H7 for onion, whereas for tomato and cucumber, the time for the microorganism to attain the maximum growth rate (M) was significantly longer than that recorded for other samples.

Immunogenic peptide mimotopes from an epitope of Escherichia coli O157 LPS.

Escherichia coli O157:H7 is a subtype of Shiga toxin-producing E. coli that is associated with haemorrhagic colitis and haemolytic uremic syndrome (HUS). Studies of populations in endemic areas have reported that the presence of specific antibodies against the O157 lipopolysaccharide (LPS) is associated with a lower incidence of diarrhoea and HUS. Phage display and IgG anti-O157 LPS antibodies were used in the present study to select peptide mimotopes of O157 LPS expressed in protein III of the M13 phage. Synthetic peptides (SP) were designed using the derived amino acid sequences obtained from DNA nucleotides of 63 selected phagotopes. The LxP/YP/SxL motif was identified in five of the phagotope amino acid sequences. Antibody responses against the phagotopes and their corresponding SPs were evaluated. SP12, one of the designed SP, induced the production of antibodies against the homologous peptide (1:800) and O157 LPS (1:200). The specificity of anti-SP12 antiserum was confirmed by analyzing its response to SP3, an SP with a different amino acid sequence than that of SP12, as well as against an E. coli LPS different from O157. Competitive studies with SP12 and O157 LPS showed a significant decrease in anti-SP12 and anti-LPS O157 antiserum responses against SP12 and O157 LPS, respectively. Eighteen (82%) of the 22 human serum samples with positive reactivity against E coli O157 LPS reacted with SP12 SP (cut-off >0.4). These results support the idea that SP12 is an immunogenic mimotope of O157 LPS.

A New Synthetic Peptide with In vitro Antibacterial Potential Against Escherichia coli O157:H7 and Methicillin-Resistant Staphylococcus aureus (MRSA).

In this work, we performed the rational design of a cationic antimicrobial peptide, GIBIMPY4, using the software DEPRAMPs developed at the GIBIM research group. GIBIMPY4 has a length of 17 amino acids, it is amphipathic, its structure is α-helix and it has a net charge of (+5). Solid-phase peptide synthesis was performed using the Fmoc strategy in acid medium. The primary structure was confirmed by MALDI-TOF mass spectrometry. The antimicrobial activity of the peptide was evaluated by broth microdilution method by measuring optical density in 96-well microplates. The minimal inhibitory concentration of GIBIMPY4 to kill 50 % of the bacterial cells (MIC50) was 6.20 ± 0.02 µM for MRSA and 4.55 ± 0.02 µM for E. coli O157:H7, while also reporting a bacteriostatic effect for the later. GIBIMPY4 activity was sensitive to salt concentration in E. coli but insignificant effect in its activity against MRSA. The peptide seems to be a broad-spectrum antimicrobial agent based on the results against Gram-positive and Gram-negative bacteria and was specific for bacterial cells E. coli O157:H7 with index of specificity equal to 9.01 in vitro assays.