Lactic acid bacteria isolated from women' breast milk and infants' faeces have appreciable immunogenic and probiotic potentials against diarrheagenic E. coli strains.

Diarrheal diseases remain the leading cause of high mortality among the infants, particularly in the developing countries; Probiotic intervention for diarrhea has been an ongoing novel approach to diarrheal prevention and treatment. This study aims to characterize immunogenic and probiotic properties of lactic acid bacteria (LAB) isolated from human breast milk and neonates' faeces. The LAB isolates from 16 mothers' breast milk and 13 infants' faeces were screened and identified by 16 S rRNA gene partial sequencing. Their antimicrobial activities against 5 strains of diarrheagenic Escherichia coli were tested. Organic acids production was quantified by HPLC, and antibiotic resistance pattern were determined by VITEK®. Autoaggregation, co-aggregation and hydrophobicity properties were assessed by UV spectrophotometry and immunomodulatory effect was determined in mouse model. Ninety-three LAB of five genera were identified. The most abundant species was Lactiplantibacillus plantarum with inhibition zones ranged from 8.0 to 25.0 ± 1 mm. Lacticaseibacillus rhamnosus A012 had 76.8 mg/mL lactic acid, (the highest concentration), was susceptible to all antibiotics tested. L. plantarum A011 and L. rhamnosus A012 were highly resistance to gastrointestinal conditions. L. rhamnosus A012 produced hydrophobicity of 25.01% (n-hexadecane), 15.4% (xylene) and its autoaggregation was 32.52%. L. rhamnosus A012 and L. plantarum A011 exert immunomodulatory effects on the cyclophosphamide-treated mice by upregulating anti-inflammatory cytokine and downregulating proinflammatory cytokines. Lactobacillus sp. demonstrated good probiotic and immunomodulatory properties. Further works are ongoing on the practical use of the strains.

The genetic resistance of sows to Escherichia coli F4 adhesion reduces their response to a vaccine containing F4 fimbriae but does not affect the preweaning performance of their susceptible piglets.

INTRODUCTION: Pigs without intestinal receptors for F4 fimbriae are congenitally resistant to F4 fimbriae-bearing enterotoxigenic Escherichia coli (ETEC F4). In general, 50 % and 100 % of piglets born to resistant (RR) sows crossed with hetero- or homozygous susceptible (SR, SS) boars, respectively, are susceptible but do not receive colostral antibodies against F4 fimbriae unless the sows have been vaccinated. The question arises as to whether resistant sows produce protective amounts of F4 antifimbrial antibodies after vaccination. The serum and colostrum antibody titres of 12 resistant and 12 susceptible vaccinated gilts were compared. The effect of the receptor status of the dam and sire on the preweaning performance of 5027 piglets was evaluated using Agroscope's recordings. The sows of the experimental herd, where ETEC F4 was circulating, were vaccinated against ETEC twice during the first pregnancy and once during each following pregnancy. The log2 transformed F4 antibody titres in the serum obtained after the second vaccine injection as well as in the colostrum of the 12 resistant animals were lower than the titres of the susceptible animals (serum: F4ab 11,19 ± 1,44 vs. 12,18 ± 1,33, P = 0,096; F4ac 10,03 ± 1,58 vs. 11,59 ± 1,43, P = 0,019; colostrum: F4ab 12,20 ± 2,41 vs. 14,02 ± 1,31, P = 0,033; F4ac 10,93 ± 2,46 vs. 13,03 ± 5,21, P = 0,006). The heat labile enterotoxin (LT) antibody titres after vaccination did not differ between susceptible and resistant animals (p > 0,10). Preweaning mortality in the offspring of RR sows × SS boars was slightly lower than in the offspring of SS sows × RR boars (P = 0,04), suggesting that the disease risk of susceptible piglets born to vaccinated resistant sows was not increased, even though they received colostrum with a slightly reduced content of antibody against F4 fimbriae.

INTRODUCTION: Les porcs dépourvus de récepteurs intestinaux pour les fimbriae F4 sont congénitalement résistants aux Escherichia coli entérotoxinogènes porteurs de fimbriae F4 (ETEC F4). En général, 50 % et 100 % des porcelets nés de truies résistantes (RR) croisées avec des verrats hétéro- ou homozygotes sensibles (SR, SS), respectivement, sont sensibles mais ne reçoivent pas d'anticorps colostraux contre les fimbriae F4, à moins que les truies n'aient été vaccinées. La question se pose de savoir si les truies résistantes produisent des quantités protectrices d'anticorps antifimbriae F4 après la vaccination. Les titres d'anticorps dans le sérum et le colostrum de 12 truies reproductrices vaccinées résistantes et de 12 truies reproductrices vaccinées sensibles ont été comparés et l'effet du statut récepteur de la mère et du père sur les performances avant sevrage de 5027 porcelets a été évalué. Les truies du troupeau expérimental, où circulait ETEC F4, ont été vaccinées deux fois au cours de la première gestation et une fois au cours de chaque gestation suivante contre ETEC. Les titres d'anticorps F4 transformés en log2 dans le sérum obtenu après la deuxième injection de vaccin ainsi que dans le colostrum des 12 animaux résistants étaient inférieurs aux titres des animaux sensibles (sérum : F4ab 11,19 ± 1,44 vs. 12,18 ± 1,33, P = 0,096 ; F4ac 10,03 ± 1,58 vs. 11,59 ± 1,43, P = 0,019 ; colostrum : F4ab 12,20 ± 2,41 vs. 14,02 ± 1,31, P = 0,033 ; F4ac 10,93 ± 2,46 vs. 13,03 ± 5,21, P = 0,006). Les titres d'anticorps contre l'entérotoxine thermolabile (LT) après la vaccination ne différaient pas entre les animaux sensibles et résistants (p > 0,10). La mortalité avant sevrage dans la progéniture des truies RR × verrats SS était légèrement inférieure à celle de la progéniture des truies SS × verrats RR (P = 0,04), ce qui suggère que le risque de maladie des porcelets sensibles nés de truies résistantes vaccinées n'a pas été augmenté, même s'ils ont reçu du colostrum avec une teneur légèrement réduite en anticorps contre les fimbriae F4.

Exploiting membrane vesicles derived from avian pathogenic Escherichia coli as a cross-protective subunit vaccine candidate against avian colibacillosis.

Avian pathogenic Escherichia coli (APEC) is a notable pathogen that frequently leads to avian colibacillosis, posing a substantial risk to both the poultry industry and public health. The commercial vaccines against avian colibacillosis are primarily inactivated vaccines, but their effectiveness is limited to specific serotypes. Recent advances have highlighted bacterial membrane vesicles (MV) as a promising candidate in vaccine research. How to produce bacterial MVs vaccines on a large scale is a significant challenge for the industrialization of MVs. The msbB gene encodes an acyltransferase and has been implicated in altering the acylation pattern of lipid A, leading to a decrease in lipid A content in lipopolysaccharides (LPS). Here, we evaluated the immunoprotective efficacy of MVs derived from the LPS low-expressed APEC strain FY26ΔmsbB, which was an APEC mutant strain with a deletion of the msbB gene. The nitrogen cavitation technique was employed to extract APEC MVs, with results indicating a significant increase in MVs yield compared to that obtained under natural culture. The immunization effectiveness was assessed, revealing that FY26ΔmsbB MVs elicited an antibody response of laying hens and facilitated bacterial clearance. Protective efficacy studies demonstrated that immunization with FY26ΔmsbB MVs conferred the immune protection in chickens challenged with the wild-type APEC strain FY26. Notably, LPS low-carried MVs recovered from the mutant FY26ΔmsbB also displayed cross-protective capabilities, and effectively safeguarding against infections caused by O1, O7, O45, O78, and O101 serotypes virulent APEC strains. These findings suggest that MVs generated from the LPS low-expressed APEC strain FY26ΔmsbB represent a novel and empirically validated subunit vaccine for the prevention and control of infections by various APEC serotypes.

Probiotic Limosilactobacillus reuteri KUB-AC5 decreases urothelial cell invasion and enhances macrophage killing of uropathogenic Escherichia coli in vitro study.

Introduction: Bacterial urinary tract infections (UTI) are among the most common infectious diseases worldwide. The rise of multidrug-resistant (MDR) uropathogenic Escherichia coli (UPEC) UTI cases is a significant threat to healthcare systems. Several probiotic bacteria have been proposed as an alternative to combat MDR UTI. Lactic acid bacteria in the genus Limosilactobacillus are some of the most studied and used probiotics. However, strain-specific effects play a critical role in probiotic properties. L. reuteri KUB-AC5 (AC5), isolated from the chicken gut, confers antimicrobial and immunobiotic effects against some human pathogens. However, the antibacterial and immune modulatory effects of AC5 on UPEC have never been explored. Methods: Here, we investigated both the direct and indirect effects of AC5 against UPEC isolates (UTI89, CFT073, and clinical MDR UPEC AT31) in vitro. Using a spot-on lawn, agar-well diffusion, and competitive growth assays, we found that viable AC5 cells and cell-free components of this probiotic significantly reduced the UPEC growth of all strains tested. The human bladder epithelial cell line UM-UC-3 was used to assess the adhesion and pathogen-attachment inhibition properties of AC5 on UPEC. Results and discussion: Our data showed that AC5 can attach to UM-UC-3 and decrease UPEC attachment in a dose-dependent manner. Pretreatment of UPEC-infected murine macrophage RAW264.7 cells with viable AC5 (multiplicity of infection, MOI = 1) for 24 hours enhanced macrophage-killing activity and increased proinflammatory (Nos2, Il6, and Tnfa) and anti-inflammatory (Il10) gene expression. These findings indicate the gut-derived AC5 probiotic could be a potential urogenital probiotic against MDR UTI.

Induction of trained immunity in broiler chickens following delivery of oligodeoxynucleotide containing CpG motifs to protect against Escherichia coli septicemia.

Oligodeoxynucleotides containing CpG motifs (CpG-ODN) can promote antimicrobial immunity in chickens by enriching immune compartments and activating immune cells. Innate memory, or trained immunity, has been demonstrated in humans and mice, featuring the absence of specificity to the initial stimulus and subsequently cross-protection against pathogens. We hypothesize that CpG-ODN can induce trained immunity in chickens. We delivered single or multiple administrations of CpG-ODN to birds and mitochondrial oxidative phosphorylation (OXPHOS) and glycolysis of peripheral blood mononuclear cells were quantified using Seahorse XFp. Next, chickens were administered with CpG-ODN twice at 1 and 4 day of age and challenged with Escherichia coli at 27 days of age. The CpG-ODN administered groups had significantly higher mitochondrial OXPHOS until 21 days of age while cellular glycolysis gradually declined by 14 days of age. The group administered with CpG-ODN twice at 1 and 4 days of age had significantly higher survival, lower clinical score and bacterial load following challenge with E. coli at 27 d of age. This study demonstrated the induction of trained immunity in broiler chickens following administration of CpG-ODN twice during the first 4 days of age to protect birds against E. coli septicemia at 27 days of age.

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

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

Dietary supplementation with a silicate clay mineral (palygorskite) alleviates inflammatory responses and intestinal barrier damage in broiler chickens challenged with Escherichia coli.

This experiment aimed to explore the protective effects of dietary palygorskite (Pal) supplementation on inflammatory responses and intestinal barrier function of broiler chickens challenged with Escherichia coli (E. coli). A 2 × 2 factorial arrangement was designed to assess the effects of Pal administration (0 or 5 g/kg of feed) and E. coli challenge (E. coli or bacterial culture medium) on broilers in a 21-d feeding trial. Birds were randomly assigned into one of the 4 groups, and each group had 8 replicates with ten birds each. The challenged chickens were orally gavaged with E. coli suspended in Luria-Bertani broth on 14 d of age, while unchallenged birds were administrated with an equivalent amount of culture medium. The sampling was performed at 21 d of age. Compared with the normal birds, an oral E. coli challenge reduced final body weight, and decreased feed intake, weight gain, and feed efficiency during the challenge period (P < 0.05). E. coli challenge promoted colonization of E. coli in cecal content and their translocation to internal organs (heart, liver, and spleen) (P < 0.05). E. coli infection also increased levels of pro-inflammatory cytokines in jejunum and ileum possibly through activating the toll-like receptor-4-mediated signaling pathway (P < 0.05). Moreover, E. coli administration increased intestinal mucosal permeability (higher serum D-lactate level and diamine oxidase activity, and lower intestinal mucosal disaccharidase activities), altered intestinal morphology, and downregulated the gene expression of intestinal tight junction proteins (P < 0.05). In contrast, Pal supplementation enhanced growth performance, inhibited colonization of E. coli, reduced intestinal inflammation, decreased intestinal permeability, restored intestinal morphology, and normalized the expression of genes responsible for inflammatory processes and maintenance of intestinal mucosal barrier (P < 0.05), and most of these beneficial effects resulting from Pal administration were independent of bacterial challenge. The results indicated dietary Pal incorporation was effective in improving growth performance and alleviating inflammation and intestinal mucosal barrier damage in broilers challenged with E. coli.

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

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

Biogenic amine tryptamine in human vaginal probiotic isolates mediates matrix inhibition and thwarts uropathogenic E. coli biofilm.

Probiotics offer a promising prophylactic approach against various pathogens and represent an alternative strategy to combat biofilm-related infections. In this study, we isolated vaginal commensal microbiota from 54 healthy Indian women to investigate their probiotic traits. We primarily explored the ability of cell-free supernatant (CFS) from Lactobacilli to prevent Uropathogenic Escherichia coli (UPEC) colonization and biofilm formation. Our findings revealed that CFS effectively reduced UPEC's swimming and swarming motility, decreased cell surface hydrophobicity, and hindered matrix production by downregulating specific genes (fimA, fimH, papG, and csgA). Subsequent GC-MS analysis identified Tryptamine, a monoamine compound, as the potent bioactive substance from Lactobacilli CFS, inhibiting UPEC biofilms with an MBIC of 4 µg/ml and an MBEC of 8 µg/ml. Tryptamine induced significant changes in E. coli colony biofilm morphology, transitioning from the Red, Dry, and Rough (RDAR) to the Smooth and White phenotype, indicating reduced extracellular matrix production. Biofilm time-kill assays demonstrated a four-log reduction in UPEC viability when treated with Tryptamine, highlighting its potent antibacterial properties, comparable to CFS treatment. Biofilm ROS assays indicated a significant elevation in ROS generation within UPEC biofilms, suggesting a potential antibacterial mechanism. Gene expression studies with Tryptamine-treated samples showed a reduction in expression of curli gene (csgA), consistent with CFS treatment. This study underscores the potential of Tryptamine from probiotic Lactobacilli CFS as a promising antibiofilm agent against UPEC biofilms.

Exposure behaviour to Escherichia coli among households in Imvepi refugee settlement, Terego district Uganda.

INTRODUCTION: Exposure to Escherichia coli (E. coli) is a risk factor for diarrhoeal diseases, which pose a significant problem in refugee settlements. Refugee populations are exposed to faecal microorganisms through multiple pathways including sub-optimal sanitary facilities, contaminated drinking water, produce and food, flood water, bathing water, and soil among others. While these pathways are well-documented, specific exposure behaviours remain underexplored. We assessed exposure behaviour to E. coli among households in Imvepi refugee settlement, Uganda, and provided evidence-based recommendations for the design of interventions to reduce excreta-related disease in refugee settlements. METHODS: Guided by the Sanitation Safety Planning approach, we surveyed 426 households in Imvepi refugee settlement, Uganda, using a digitized questionnaire and an observation checklist. We collected data on the background characteristics and exposure behaviour of women and emancipated girls (minors living on their own, having borne a child, married, or pregnant). The outcome variable, E. coli exposure behaviour, was measured using a five-point Likert scale, assessing behaviours that increase the risk of exposure. Data were cleaned in Microsoft Excel and analyzed in Stata version 17. Descriptive statistics were performed to summarize the data. We used modified Poisson regression to determine the factors associated with the outcome. RESULTS: Over 59.4% (253) exhibited high-risk exposure behaviour. Residing in compound homes (Adjusted Prevalence Ratio (APR) = 0.72, 95% Confidence interval (CI): 0.58-0.90), being aged 35-49 years (APR = 0.76, 95% CI: 0.60-0.97), having household heads with post-primary education (APR = 0.54, 95% CI: 0.38-0.77), high knowledge (APR = 0.69, 95% CI: 0.59-0.80), and high-risk perceptions regarding exposure to E. coli (APR = 0.75, 95% CI: 0.64-0.88) were associated with a lower prevalence of high-risk E. coli exposure behaviours. Conversely, having sanitary facilities with excreta overflowing from the squat hole (APR = 1.26, 95% CI: 1.08-1.48) was associated with a higher prevalence of high-risk exposure behaviours. CONCLUSION: The study indicates a substantial prevalence of high-risk E. coli exposure behaviours in the refugee settlement.. There's a need to implement behaviour change interventions targeted at preventing or minimizing exposure, especially among households whose heads have low education attainment, those with young caretakers and those with limited knowledge and low-risk perceptions regarding exposure to E. coli.

Vaccinating parent flocks against colibacillosis reduces broiler mortality - A retrospective observational study from 2016 to 2019 in Finland.

Colibacillosis is one of the most important infectious diseases in modern poultry production. The complex nature of colibacillosis has made it challenging to produce an effective vaccine. As a control measure for colibacillosis outbreak in Finland, a vaccination program with a commercial colibacillosis vaccine and later also an autogenous vaccine was started for parent flocks in 2017. In this retrospective observational study from years 2016-2019, we evaluated first week and total mortality of broiler flocks (n= 6969) originating from parents with different colibacillosis vaccination status. Broiler flocks were divided into three groups according to vaccination status of their parent flocks. First group were flocks from parents with no colibacillosis vaccines; second group was flocks from parents vaccinated with commercial vaccine only; and third group was flocks from parents with both commercial and autogenous vaccine. Bayesian modelling was used to predict posterior distributions of first week mortality and total mortality of the broiler flocks. Results of the modelling revealed that broiler flocks from unvaccinated parents had the highest mortality rates (mean first week mortality 1.40 % and mean total mortality 4.33 %, respectively) whereas flocks from parents with a combination of commercial and autogenous vaccinations had the lowest mortality rates (mean first week mortality 0,91 % and mean total mortality 3,14 %). The mortalities from broilers flocks from parents with only commercial vaccine fell in between these groups. Also, standard deviations of mortality rates were lower in broilers from parents with commercial or both vaccines. This demonstrates that in addition to lowering the mean mortality rates, the vaccinations made high mortality broiler flocks less common. Best performance was obtained when autogenous vaccine was combined to the commercial vaccine. The autogenous vaccine consists of the same type of Escherichia coli strain that was causing most colibacillosis cases during the study period in Finland. This study adds to the evidence of benefits of colibacillosis vaccines during outbreaks. It also demonstrates the importance of the knowledge of the types of APEC strains causing outbreaks to produce effective autogenous vaccines.

Putative new combination vaccine candidates identified by reverse vaccinology and genomic approaches to control enteric pathogens.

OBJECTIVES: The pathogenic microorganisms that cause intestinal diseases can significantly jeopardize people's health. Currently, there are no authorized treatments or vaccinations available to combat the germs responsible for intestinal disease. METHODS: Using immunoinformatics, we developed a potent multi-epitope Combination (combo) vaccine versus Salmonella and enterohemorrhagic E. coli. The B and T cell epitopes were identified by performing a conservancy assessment, population coverage analysis, physicochemical attributes assessment, and secondary and tertiary structure assessment of the chosen antigenic polypeptide. The selection process for vaccine development included using several bioinformatics tools and approaches to finally choose two linear B-cell epitopes, five CTL epitopes, and two HTL epitopes. RESULTS: The vaccine had strong immunogenicity, cytokine production, immunological properties, non-toxicity, non-allergenicity, stability, and potential efficacy against infections. Disulfide bonding, codon modification, and computational cloning were also used to enhance the stability and efficacy of expression in the host E. coli. The vaccine's structure has a strong affinity for the TLR4 ligand and is very durable, as shown by molecular docking and molecular modeling. The results of the immunological simulation demonstrated that both B and T cells had a heightened response to the vaccination component. CONCLUSIONS: The comprehensive in silico analysis reveals that the proposed vaccine will likely elicit a robust immune response against pathogenic bacteria that cause intestinal diseases. Therefore, it is a promising option for further experimental testing.

Antimicrobial lipids loaded on lectin display reduced MIC, curtail pathogenesis and protect zebrafish from reinfection by immunomodulation.

Antibiotic resistance and re-emergence of highly resistant pathogens is a grave concern everywhere and this has consequences for all kinds of human activities. Herein, we showed that N-palmitoylethanolamine-derived cationic lipid (cN16E) had a lower minimum inhibitory concentration (MIC) against both Gram-positive and Gram-negative bacteria when it was loaded with Butea monosperma seed lectin (BMSL). The analysis using lectin-FITC conjugate labelling indicated that the improved antibacterial activity of BMSL conjugation was due to bacterial cell surface glycan recognition. Live and dead staining experiments revealed that the BMSL-cN16E conjugate (BcN16E) exerts antibacterial activity by damaging the bacterial membrane. BcN16E antimicrobial activity was demonstrated using an infected zebrafish animal model because humans have 70 % genetic similarity to zebrafish. BcN16E therapeutic potential was established successfully by rescuing fish infected with uropathogenic Escherichia coli (UPEC). Remarkably, the rescued infected fish treated with BcN16E prevented reinfection without further therapy, indicating BcN16E immunomodulatory potential. Thus, the study examined the expression of immune-related genes, including tnfα, ifnγ, il-1ß, il-4, il-10, tlr-2, etc. There was a significant elevation in the expression of all these genes compared to control and fish treated with BMSL or cN16E alone. Interestingly, when the rescued zebrafish were reinfected with the same pathogen, the levels of expression of these genes were many folds higher than seen earlier. Radial immune diffusion analyses (RIA) using zebrafish serum revealed antibody production during the initial infection and treatment. Interestingly, reinfected fish had significant immunoprecipitation in RIA, a feature absent in the groups treated with cN16E, BMSL, and control. These results clearly show that the BcN16E complex not only rescued infected zebrafish but also conferred long-lasting protection in terms of immunomodulation that protects against multiple reinfections. The findings support that BcN16E has immense potential as a novel immunostimulant for various biomedical applications.

A systematic review on the role of biosecurity to prevent or control colibacillosis in broiler production.

This systematic review aimed at investigating the role that biosecurity can have in preventing or controlling colibacillosis in broiler production. Primary studies with natural or experimental exposure to avian pathogenic Escherichia coli, evaluating any biosecurity measure to prevent or control colibacillosis in broiler chickens with at least one of the following outcomes: feed conversion ratio (FCR), condemnations at slaughter, and mortality due to colibacillosis, were included. A systematic search was carried out in 4 databases according to the Cochrane handbook and reported following the PRISMA 2020 directions. Studies (n = 3,886) were screened in a 2-phase process and data matching the inclusion criteria were extracted. Risk of bias assessment was performed. Four studies reporting biosecurity measures to prevent or control colibacillosis in broiler production were included. In all studies, only disinfection during either the pre-hatching period (n = 3) or the post-hatching period (n = 1) was evaluated as biosecurity measure in broiler production, as well as its effect on FCR (n = 2) and mortality (n = 4) due to colibacillosis. No studies with effects on condemnations at slaughter were found. Due to the heterogeneity of studies in regard to interventions and outcomes, meta-analysis was not carried out. The limited findings of this systematic review do not provide a comprehensive evidence to statistically evaluate the efficacy of biosecurity to prevent or control colibacillosis in broiler production. The scarcity of evidence found suggests that further and deeper investigations on the topic are needed, considering the variety of interventions related to biosecurity.

Vaccines against extraintestinal pathogenic Escherichia coli (ExPEC): progress and challenges.

The emergence of antimicrobial resistance (AMR) is a principal global health crisis projected to cause 10 million deaths annually worldwide by 2050. While the Gram-negative bacteria Escherichia coli is commonly found as a commensal microbe in the human gut, some strains are dangerously pathogenic, contributing to the highest AMR-associated mortality. Strains of E. coli that can translocate from the gastrointestinal tract to distal sites, called extraintestinal E. coli (ExPEC), are particularly problematic and predominantly afflict women, the elderly, and immunocompromised populations. Despite nearly 40 years of clinical trials, there is still no vaccine against ExPEC. One reason for this is the remarkable diversity in the ExPEC pangenome across pathotypes, clades, and strains, with hundreds of genes associated with pathogenesis including toxins, adhesins, and nutrient acquisition systems. Further, ExPEC is intimately associated with human mucosal surfaces and has evolved creative strategies to avoid the immune system. This review summarizes previous and ongoing preclinical and clinical ExPEC vaccine research efforts to help identify key gaps in knowledge and remaining challenges.

Preclinical validation of an Escherichia coli O-antigen glycoconjugate for the prevention of serotype O1 invasive disease.

A US collection of invasive Escherichia coli serotype O1 bloodstream infection (BSI) isolates were assessed for genotypic and phenotypic diversity as the basis for designing a broadly protective O-antigen vaccine. Eighty percent of the BSI isolate serotype O1 strains were genotypically ST95 O1:K1:H7. The carbohydrate repeat unit structure of the O1a subtype was conserved in the three strains tested representing core genome multi-locus sequence types (MLST) sequence types ST95, ST38, and ST59. A long-chain O1a CRM197 lattice glycoconjugate antigen was generated using oxidized polysaccharide and reductive amination chemistry. Two ST95 strains were investigated for use in opsonophagocytic assays (OPA) with immune sera from vaccinated animals and in murine lethal challenge models. Both strains were susceptible to OPA killing with O1a glycoconjugate post-immune sera. One of these, a neonatal sepsis strain, was found to be highly lethal in the murine challenge model for which virulence was shown to be dependent on the presence of the K1 capsule. Mice immunized with the O1a glycoconjugate were protected from challenges with this strain or a second, genotypically related, and similarly virulent neonatal isolate. This long-chain O1a CRM197 lattice glycoconjugate shows promise as a component of a multi-valent vaccine to prevent invasive E. coli infections. IMPORTANCE: The Escherichia coli serotype O1 O-antigen serogroup is a common cause of invasive bloodstream infections (BSI) in populations at risk such as newborns and the elderly. Sequencing of US BSI isolates and structural analysis of O polysaccharide antigens purified from strains that are representative of genotypic sub-groups confirmed the relevance of the O1a subtype as a vaccine antigen. O polysaccharide was purified from a strain engineered to produce long-chain O1a O-antigen and was chemically conjugated to CRM197 carrier protein. The resulting glycoconjugate elicited functional antibodies and was protective in mice against lethal challenges with virulent K1-encapsulated O1a isolates.

Differences in P-Type and Type 1 Uropathogenic Escherichia coli Urinary Anti-Adhesion Activity of Cranberry Fruit Juice Dry Extract Product and D-Mannose Dietary Supplement.

BACKGROUND: Urinary tract infection (UTI) prevention benefits of cranberry intake are clinically validated, especially for women and children. To ensure the benefits of cranberry dietary supplement products, the anti-adhesion activity (AAA) against uropathogenic bacteria is routinely used in in vitro bioassays to determine the activity in whole product formulations, isolated compounds, and ex vivo bioassays to assess urinary activity following intake. D-mannose is another dietary supplement taken for UTI prevention, based on the anti-adhesion mechanism. OBJECTIVE: Compare the relative AAA of cranberry and D-mannose dietary supplements against the most important bacterial types contributing to the pathogenesis of UTI, and consider how certain components potentially induce in vivo activity. METHODS: The current study used a crossover design to determine ex vivo AAA against both P- and Type 1-fimbriated uropathogenic Escherichia coli of either D-mannose or a cranberry fruit juice dry extract product containing 36 mg of soluble proanthocyanidins (PACs), using bioassays that measure urinary activity following consumption. AAA of extracted cranberry compound fractions and D-mannose were compared in vitro and potential induction mechanisms of urinary AAA explored. RESULTS: The cranberry dietary supplement exhibited both P-type and Type 1 in vitro and ex vivo AAA, while D-mannose only prevented Type 1 adhesion. Cranberry also demonstrated more robust and consistent ex vivo urinary AAA than D-mannose over each 1-week study period at different urine collection time points. The means by which the compounds with in vitro activity in each supplement product could potentially induce the AAA in urines was discussed relative to the data. CONCLUSIONS: Results of the current study provide consumers and healthcare professionals with additional details on the compounds and mechanisms involved in the positive, broad-spectrum AAA of cranberry against both E. coli bacterial types most important in UTIs and uncovers limitations on AAA and effectiveness of D-mannose compared to cranberry.

Protecting the piglet gut microbiota against ETEC-mediated post-weaning diarrhoea using specific binding proteins.

Post-weaning diarrhoea (PWD) in piglets presents a widespread problem in industrial pig production and is often caused by enterotoxigenic E. coli (ETEC) strains. Current solutions, such as antibiotics and medicinal zinc oxide, are unsustainable and are increasingly being prohibited, resulting in a dire need for novel solutions. Thus, in this study, we propose and evaluate a protein-based feed additive, comprising two bivalent heavy chain variable domain (VHH) constructs (VHH-(GGGGS)3-VHH, BL1.2 and BL2.2) as an alternative solution to manage PWD. We demonstrate in vitro that these constructs bind to ETEC toxins and fimbriae, whilst they do no affect bacterial growth rate. Furthermore, in a pig study, we show that oral administration of these constructs after ETEC challenge reduced ETEC proliferation when compared to challenged control piglets (1-2 log10 units difference in gene copies and bacterial count/g faeces across day 2-7) and resulted in week 1 enrichment of three bacterial families (Prevotellaceae (estimate: 1.12 ± 0.25, q = 0.0054), Lactobacillaceae (estimate: 2.86 ± 0.52, q = 0.0012), and Ruminococcaceae (estimate: 0.66 ± 0.18, q = 0.049)) within the gut microbiota that appeared later in challenged control piglets, thus pointing to an earlier transition towards a more mature gut microbiota. These data suggest that such VHH constructs may find utility in industrial pig production as a feed additive for tackling ETEC and reducing the risk of PWD in piglet populations.

Redox balance and immunity of piglets pre- and post-E. coli challenge after treatment with hemp or fish oil, and vitamin E.

This study investigated the influence of polyunsaturated fatty acid composition and vitamin E supplementation on oxidative status and immune responses in weanling piglets pre- and post-E. coli challenge. Suckling piglets (n = 24) were randomly selected from two litters for an oral supplementation (1 mL/day) with fish oil or hemp oil and vitamin E supplementation (60 mg natural vitamin E/mL oil) from day 10 to 28 of age. At day 29 and 30 of age, each piglet was orally inoculated with 6.7 × 108 and 3.96 × 108 CFU of F4 and F18 E. coli, respectively. Blood was sampled from all piglets on day 28 before E. coli challenge and on day 35 of age to investigate immunological and oxidative stress markers in plasma. One week after weaning and exposure to E. coli, a general reduction in the α-tocopherol concentration and activity of GPX1 was obtained. Vitamin E supplementation lowered the extent of lipid peroxidation and improved the antioxidative status and immune responses after E. coli challenge. Hemp oil had the greatest effect on antioxidant enzyme activity. Provision of hemp oil and vitamin E to suckling piglets may reduce the incidence of post-weaning diarrhea.