Avian Colibacillosis Research Articles

The response regulator OmpR contributes to the pathogenicity of avian pathogenic Escherichia coli

Avian colibacillosis is a serious systemic infectious disease in poultry and caused by avian pathogenic Escherichia coli (APEC). Previous studies have shown that 2-component systems (TCSs) are involved in the pathogenicity of APEC. OmpR, a response regulator of OmpR/EnvZ TCS, plays an important role in E. coli K-12. However, whether OmpR correlates with APEC pathogenesis has not been established. In this study, we constructed an ompR gene mutant and complement strains by using the CRISPR-Cas9 system and found that the inactivation of the ompR gene attenuated bacterial motility, biofilm formation, and the production of curli. The resistance to environmental stress, serum sensitivity, adhesion, and invasion of DF-1 cells, and pathogenicity in chicks were all significantly reduced in the mutant strain AE17ΔompR. These phenotypes were restored in the complement strain AE17C-ompR. The qRT-PCR results showed that OmpR influences the expression of genes associated with the flagellum, biofilm formation, and virulence. These findings indicate that the regulator OmpR contributes to APEC pathogenicity by affecting the expression and function of virulence factors.

Impact of commercial and autogenous Escherichia coli vaccine combination on broiler breeder stock performance, gross pathology, and diversity of Escherichia coli isolates

Avian colibacillosis is one of the main causes of economic losses in the poultry industry worldwide. Vaccination could help to prevent infection during the laying period on broiler breeder farms. Effective vaccination against avian pathogenic Escherichia coli (APEC) may be an essential step for protection of poultry flocks depending on the region where they are raised. The aim of this study was to investigate the additive protective effect of an autogenous E. coli vaccine in broiler breeders pre-vaccinated with a licensed E. coli vaccine (Poulvac®). Our field study was partially blinded and parallel group designed. Group 1 included 24 000 laying hens vaccinated by Poulvac®. Group 2 comprised 12 000 laying hens vaccinated by Poulvac® and additionally, by an autogenous E. coli vaccine via intramuscular application before transfer. The effectiveness of vaccination in both groups was evaluated according to the results of gross pathology, bacteriology (isolation and characterization of E. coli) and utility indicators. Based on the pathology, the occurrence of E. coli polyserositis syndrome (EPS), salpingoperitonitis syndrom (SPS), and haemorrhagic septicaemia was decreased in Group 2 compared to Group 1. The difference in the occurrence of EPS (P < 0.001) and SPS (P = 0.0342) was significant. The proportion of serotype O78 among E. coli isolated from Group 1 and Group 2 was also significant (P = 0.0178). The effective and multi-serotype vaccination program in order to expand heterologous protection of laying hens and combination of commercial and autogenous vaccines seems to be a promising preventive management tool.

Study on antibiotic resistance and phylogenetic comparison of avian-pathogenic Escherichia coli (APEC) and uropathogenic Escherichia coli (UPEC) isolates.

Avian pathogenic Escherichia coli (APEC) and uropathogenic E. coli (UPEC) can cause vast infections in humans and poultry. The present study was conducted to compare the isolates of the APEC and UPEC pathotypes on the basis phenotypic and genotypic features of antibiotic resistance and phylogenetic differences. Total number of 70 identified E. coli strains, including 35 APEC and 35 UPEC isolates, were isolated from avian colibacillosis and human urinary tract infection (UTI), and were subjected to the antimicrobial susceptibility testing, polymerase chain reaction (PCR) detection of the resistance genes, phylogenetic grouping and DNA fingerprinting with enterobacterial repetitive intergenic consensus PCR (ERIC - PCR) to survey the variability of the isolates. The most resistance rates among all E. coli isolates were, respectively, obtained for Ampicillin (84.20%) and sulfamethoxazole-trimethoprim (65.70%). The APEC and UPEC isolates showed the most susceptibility to imipenem and gentamycin,respectively. Among 70 APEC and UPEC isolates 34.20%, 32.80%, 20.00%, and 12.80% belonged to the A, B2, D, and B1 phylogenetic groups, respectively. Analysis of the DNA fingerprinting phylogenetic tree showed 10 specific clusters of APEC and UPEC isolates. According to the results, the most effective antibiotics and the phenotypic and genotypic predominant resistance patterns of the APEC and UPEC isolates were different. Moreover, APECs and UPECs showed various dominant phylogenetic groups. With all descriptions, the APEC isolates still are potential candidates for carrying important resistance genes and can be one of the possible strains related to human infections.

AVIAN ESCHERICHIA COLI INFECTION IN MEAT-TYPE CHICKEN: PREVALENCE AND PATHOLOGY

Avian colibacillosis is considered the major bacterial disease in the poultry industry worldwide. This study was conducted to determine prevalence and pathology by avian pathogenic Escherichia coli (APEC) in broiler chicken. Various organs of affected broilers were checked for the E. coli infection. One hundred and forty commercial poultry farms were divided into seven age groups (A, B, C, D, E, F, and G). Group A (1st week), B (2nd week), C (3rd week), D (4th week), E (5th week), F (6th week), G (7th to 9th week). Investigations were conducted from day 1 until the marketing of birds based on clinical findings. A total of 2491 (23.71% of sick birds) dead birds were observed positive for E. coli infection. Three forms of infection were observed i.e. omphalitis, colisepticaemia, and colibacillosis at the rate of 1.55, 17 and 56.65%, respectively. Omphalitis was confirmed by microbial culture and revealed in only groups A and B with 76.24 and 23.76 respectively. While, Colisepticaemia was observed at the rate of 3.11, 6.74, 20.18, 29.63, 19.16, and 22.18 in groups B, C, D, E, F, and G respectively. Whereas Colibacillosis was noticed as 0.075, 3.35, 5.99, 19.20, 26.70, 16.11 and 28.55% in groups A, B, C, D, E, F, and G respectively. Petechiation of heart, kidney, and gizzard besides hemorrhagic enteritis, hemorrhagic tracheitis, and necrotic foci on the liver and caseous exudates in air sacs were the prime gross lesions noticed in colibacillosis. Microscopically, no lesion was observed in serosa, muscularis externa, and submucosa. However, loss of epithelial tissue and breaches in the mucosal layer of the small intestine were observed. Furthermore, characteristic pink and grayish colonies were observed in E. coli cultures on McConkey’s and Blood agar respectively.

An Integrated Perspective on Virulence-Associated Genes (VAGs), Antimicrobial Resistance (AMR), and Phylogenetic Clusters of Pathogenic and Non-pathogenic Avian Escherichia coli.

Avian pathogenic Escherichia coli (APEC) is an important bacterial pathogen that causes avian colibacillosis and leads to huge economic losses in the poultry industry. Different virulence traits contribute to pathogenesis of APEC infections, and antimicrobial resistance (AMR) has also been an overwhelming issue in poultry worldwide. In the present study, we aimed to investigate and compare the presence of virulence-associated genes (VAGs), AMR, and phylogenetic group's distribution among APEC and avian fecal E. coli (AFEC) strains. E. coli from birds with colisepticemia and yolk sac infection (YSI) (APEC) plus E. coli strains from the feces of healthy birds (AFEC) were compared by the aforementioned traits. In addition, the clonal relatedness was compared using Enterobacterial repetitive intergenic consensus PCR (ERIC-PCR). Although all strains were susceptible to fosfomycin, ceftriaxone, and cefixime, almost all strains (98%) were multi-drug resistant (MDR). All strains (except two) harbored at least three or more VAGs, and the virulence scores tended to be higher in pathogenic strains especially in the colisepticemic group. All phylogenetic groups were found in isolates from YSI, colisepticemia, and the feces of healthy birds; however, the frequency of phylogroups varied according to the source of the isolate. B1 and C phylogroups were statistically more likely to be found among APEC from YSI and colisepticemic E. coli groups, respectively, while phylogroup A was the most frequently occurring phylogroup among AFEC strains. Our findings also revealed that AMR and VAGs are not essentially co-evolved traits as in some instances AMR strains were more prevalent among AFEC. This reflects the divergent evolutionary pathways of resistance acquisition in pathogenic or non-pathogenic avian E. coli strains. Importantly, strains related to phylogenetic group C showed higher virulence score and AMR that requires further attention. To some extent, ERIC-PCR was able to group strains by isolation source, phylogroup, or virulence genes. Further integrated studies along with assessment of more detailed genotypic and phenotypic features could potentially lead to better understanding of virulence, resistance, and evolution of ExPEC.

Combining pangenome analysis to identify potential cross-protective antigens against avian pathogenic Escherichia coli

ABSTRACT Avian pathogenic Escherichia coli (APEC) is the bacterial pathogen of poultry colibacillosis, which causes significant economic losses to the poultry industry. The lack of an effective vaccine against multiple serotypes and the emergence of multi-resistant isolates have made the control of avian colibacillosis troublesome. To identify conserved potential vaccine candidates, 58 genomes of APEC were obtained (54 sequenced by our laboratory and four downloaded from NCBI). A reverse vaccinology (RV) method based on the pangenome – called Pan-RV analysis – was performed in APEC-protective protein mining for the first time. Finally, four proteins were selected, and their immunoreactivity with anti-O1, O2, and O78 serum was verified by western blotting. Our in silico method of analysis will pave the way for rapid screening of vaccine candidates and will lay the foundation for the development of a highly effective subunit vaccine controlling APEC infection. RESEARCH HIGHLIGHTS Pan-RV analysis was used for the first time in the discovery of APEC-protective proteins. A total of 53 potential protective proteins were screened out. Four proteins were verified as potential vaccine candidates using western blotting.

Prevalence and Genetic Relationship of Predominant Escherichia coli Serotypes Isolated from Poultry, Wild Animals, and Environment in the Mekong Delta, Vietnam.

Avian pathogenic Escherichia coli (APEC) is the main causative agent of avian colibacillosis, which is an important systemic disease of profound economic and clinical consequences for the poultry industry worldwide. In this study, 975 E. coli strains were isolated from 2,169 samples collected from cloacal swabs of chickens, in-farm wild animals (ants, geckos, flies, and rats), and environment. The highest proportion of E. coli isolation was obtained from chicken cloacal swabs with 71.05% (95% confidence interval (CI) 66.69–75.05%) followed by the proportions of 38.15% (95% CI 35.41–40.97%) and 38.11% (95% CI 34.15–42.24%) from wild animals or environment, respectively. Distribution of O-antigen serotypes of the E. coli isolates, including O1, O2, O18, and O78, was determined by PCR. The most predominant serotype was O18 (10.56%) followed by O2 (9.44%), O1 (7.79%), and O78 (6.56%). Of note, serotype O18 was more likely distributed in the examined wild animals, especially in geckos. Polymorphic DNA fingerprints, generated by ERIC-PCR, of representative E. coli strains of each serotype revealed genetic heterogeneity of the examined E. coli, and O18 was more divergent with 63 clusters formed from 66 isolates. Furthermore, several E. coli strains from different sample sources shared high DNA fingerprint relatedness, suggesting that there exists complex transmission of E. coli from chickens to wild animals and environment and vice versa in poultry husbandry settings. Although pathotypes of the examined E. coli were not determined in this study, our results provided important findings of epidemiological and genetic characteristics of E. coli in the Mekong Delta and highlighted the prerequisite of stricter biocontainment to reduce the prevalence and consequences of APEC in poultry production.

Baicalin ameliorates APEC-induced intestinal injury in chicks by inhibiting the PI3K/AKT-mediated NF-κB signaling pathway

Avian pathogenic Escherichia coli (APEC) is the causative agent of avian colibacillosis. Baicalin (BA) possesses multiple pharmacological effects, but the mechanism underlying its activity in APEC-induced intestinal injury remains unknown. This study aims to investigate the protective effects and possible mechanism of BA against APEC-induced intestinal injury. Sixty 1-day-old chicks were randomly divided into 4 groups: the control group (basal diet), E. coli group (basal diet), BAI10 group (10 mg/kg BA), and BAI20 group (20 mg/kg BA). After pretreatment with BA for 15 d and subsequent induction of APEC infection by pectoralis injection, the ileum was collected and analyzed. The results showed that BA-pretreatment demonstrated an alleviation of chicks in diarrhea rate, mortality, and histopathological changes in intestinal tissues after APEC infection. Additionally, following APEC infection, BA improved the intestinal barrier by elevating zona occludens (ZO)s (ZO-1, 2, 3), Claudins (Claudin1, 2, 3), Occludin, avian β-defensin (AvBD)s (AvBD1, 2, 4), lysozyme (Lyz) mRNA levels and ZO-1, Claudin1, and Occludin protein levels. Besides, the activities of total superoxide dismutase (T-SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) and the SOD-1 and CAT mRNA levels and SOD-1 protein level were elevated by BA pretreatment. BA pretreatment also decreased the malondialdehyde (MDA) content, heme oxygenase-1 (HO-1) and NADH quinone oxidoreductase 1 (NQO1) mRNA levels, and HO-1 protein level after APEC infection. BA alleviated the APEC-induced inflammatory response, including downregulating the mRNA levels of proinflammatory cytokines (tumor necrosis factor-α (TNF-α), interleukin [IL]-1β, IL-6, IL-8) and upregulating the mRNA levels of anti-inflammatory cytokines (IL-4, IL-10, IL-13, transforming growth factor-β [TGF-β]). Furthermore, BA decreased the mRNA and protein levels of phosphatidylinositol 3 kinase (PI3K), protein kinase B (AKT), and nuclear factor kappa-B (NF-κB) as well as the expression of the phosphorylated forms of these proteins after APEC infection. Collectively, our findings indicate that BA exerts a protective effect against APEC-induced intestinal injury in chicks by inhibiting the PI3K/AKT-mediated NF-κB pathway, suggesting that BA may be a potential therapeutic approach for avian colibacillosis.

Interleukin gene expression in broiler chickens infected by different Escherichia coli serotypes.

Background and Aim:Escherichia coli is the cause of avian colibacillosis, a significant threat to the poultry industry and public health. Thus, this study investigated the prevalence of E. coli in diseased chicken broilers, pathological effects of these bacteria, and interleukin (IL) gene expression of different serotypes of E. coli (O78, O26, O44, and O55) on experimentally infected chickens.Materials and Methods:A total of 295 organ samples (liver, lungs, heart, and spleen) from 59 diseased broiler chickens were used for conventional identification of E. coli. Chickens were orally infected with one of the following E. coli serotypes (O78, O26, O44, or O55) and examined for clinical signs, mortality, macroscopic and microscopic lesions, and IL gene expression using real-time quantitative polymerase chain reaction.Results:E. coli was isolated from 53.2% of broiler chicken organs with a high prevalence in lungs (26.1%). The most prevalent serotypes were O78, O26, O44, O55, O157, and O127 prevalence of 27.8, 22.2, 16.7, 16.7, 5.6, and 5.6%, respectively. In the experimental design, five groups (G1-G5) of birds were established. G1 served as the negative control group, while G2-G5 were challenged orally with E. coli O78, O26, O55, or O44, respectively. Chickens infected with E. coli O78 or O26 showed significant clinical signs in comparison to the other infected birds. Mortality (13.3%) was only observed in birds infected with E. coli O78. Necropsy of dead birds after E. coli O78 infection showed pericarditis, enteritis, airsacculitis, and liver and lung congestion. More severe histopathological changes were observed in intestines, spleen, liver, and lung from chickens infected with either E. coli O78 or O26 than for birds infected with other serotypes. On the 2nd day post-infection, E. coli challenge, particularly with E. coli O78, displayed significantly upregulated levels of ileal IL-6 and IL-8, but ileal IL-10 level tended to be downregulated in comparison to the control group.Conclusion:This study assessed the application of cytokines as therapeutic agents against infectious diseases, particularly colibacillosis.

Quorum sensing-1 signaling of N-hexanoyl-L-homoserine lactone contributes to virulence in avian pathogenic Escherichia coli.

Avian pathogenic E. coli (APEC) caused avian colibacillosis is mostly common in poultry industry worldwide. APEC virulence factors lead to pathogenesis and the quorum sensing (QS) system is actively involved in the regulation of these virulence factors. Signaling molecules in QS are known as autoinducers (AIs). In QS-1, E. coli encodes a single LuxR homolog, i.e., SdiA, but does not express the LuxI homolog, an acyl-homoserine lactone (AHL) synthase of producing AI-1. Avian pathogenic E. coli (APEC) regulates its virulence genes expression in response to exogenous AHLs, but regulatory mechanisms of AHL and QS-1 are still unknown. This study targeted the APEC CE129 isolate as the reference strain, and the Yersinia enterocolitica yenI gene was expressed into APEC CE129. CE129/pyenI was conferred the ability to produce AHL signal. The CE129 SdiA mutant strain with an in-frame sdiA (AHL receptor) gene deletion was constructed by a λRed recombination system, which lost the ability to sense AHL. The goal of this study was to explore the function of QS-1 upon virulence and elucidate the regulatory effect of QS-1/AHL signals in the APEC strain. Adherence and invasion assays revealed that QS-1 affected APEC adherence and survival ability. APEC biofilm formation was also suppressed under C6HSL. Interestingly, APEC exhibited different phenotypes of acid tolerance and flagella expression when compared to enterotoxigenic E. coli or enterohemorrhagic E. coli (ETEC and EHEC, respectively). These findings enhance our understanding of the QS mechanism.

Multidrug resistant E. coli recovered from household reared female budgerigar pet bird (Melopsittacus undulatus) in Ibadan, Oyo State Nigeria: a case report

BackgroundPet birds are housed and reared exclusively for ornamental use. These include psittaciformes (parrots, parakeets, budgerigars, love birds) and passeriformes (e.g. canaries, finches, sparrows, also called songbirds). E. coli is a Gram negative bacterium. In birds, it is called Avian Pathogenic E. coli and is a causative agent of avian colibacillosis. Antimicrobial resistance is the process through which bacteria evade the activity of antibiotics. According to WHO (World Health Organization), antibiotic resistance is the result of indiscriminate use of these drugs which are used both in Veterinary and human medicine.Case presentationA one and half year old yellow female budgerigar pet bird (Melopsittacus undulatus) with patches of black and white on wings and back, kept as companion (one out of four) was presented dead at the avian clinic section of the Veterinary Teaching Hospital, University of Ibadan, Nigeria. It was reported to have presented signs of anorexia and diarrhea for two days before death. Provisional diagnosis was colibacillosis. Intestinal samples were sent for microbial culture and sensitivity. The samples yielded growth of E. coli. Antimicrobial susceptibility revealed that the organism showed resistance to all tested antibiotics.ConclusionsMultidrug-resistant Escherichia coli constitutes a major public health concern. The constant interaction between household companion birds and humans gives room for disease transmission. Wild birds kept as pets or companions harbor pathogenic and zoonotic pathogens, hence a threat to public health.

Flagellar rotor protein FliG is involved in the virulence of avian pathogenic Escherichia coli

Avian pathogenic Escherichia coli (APEC), a type of extraintestinal pathogenic E. coli, causes avian colibacillosis, a disease of significant economic importance to poultry producers worldwide, which is characterized by systemic infection. However, the pathogenesis of avian pathogenic E. coli strains is not well defined. Here, the role of a flagellar rotor protein encoded by the fliG gene of avian pathogenic E. coli strain AE17 was investigated. To study the role of FliG in the pathogenicity of APEC, fliG mutant and complemented strains were constructed and characterized. The inactivation of fliG had no effect on APEC growth, but significantly reduced bacterial motility. Compared with the wild type, the fliG mutant was highly attenuated in a chick infection model and showed severe defects in its adherence to and invasion of chicken embryo fibroblast DF-1 cells. The fliG mutant also showed reduced resistance to serum in chicks. The expression of the inflammatory cytokines interleukin 1β (IL1β), IL6, and IL8 was reduced in HD-11 macrophages infected with the fliG mutant strain compared with their expression in the wild-type strain. These results demonstrate that the FliG contributes to the virulence of APEC.

Avian pathogenic Escherichia coli (APEC) and uropathogenic Escherichia coli (UPEC): characterization and comparison

Avian pathogenic E. coli (APEC) and uropathogenic E. coli (UPEC) are responsible for avian colibacillosis and human urinary tract infections, respectively. There are genetic similarities between the APEC and UPEC pathotypes, suggesting the APEC strains could be a potential reservoir of virulence and antimicrobial-resistance genes for the UPEC strains. This study aimed to characterize and compare APEC and UPEC strains regarding the phylogroup classification, pathogenicity and antimicrobial susceptibility. A total of 238 APEC and 184 UPEC strains were selected and characterized. The strains were assayed for antimicrobial susceptibility and classified into phylogenetic groups using a multiplex-PCR protocol. In addition, the APEC strains had previously been classified according to their in vivo pathogenicity. The results showed that both pathotypes had variation in their susceptibility to most of the antimicrobial agents evaluated, with few strains classified as multidrug resistant. The highest resistance rate for both pathotypes was to amoxicillin. Classifying the APEC and UPEC strains into phylogenetic groups determined that the most frequently frequencies were for groups D and B2, respectively. These results reflect the pathogenic potential of these strains, as all the UPEC strains were isolated from unhealthy patients, and most of the APEC strains were previously classified as pathogenic. The results indicate that distribution into phylogenetic groups provided, in part, similar classification to those of in vivo pathogenicity index, as it was possible to adequately differentiate most of the pathogenic and commensal or low-pathogenicity bacteria. However, no relationship could be found between the specific antimicrobial agents and pathogenicity or phylogenetic group for either pathotype.

A strategy for mitigating avian colibacillosis disease using plant growth promoting rhizobacteria and green synthesized zinc oxide nanoparticles.

Avian colibacillosis caused by the zoonotic pathogen Escherichia coli is a common bacterial infectionthat causes major losses in the poultry sector. Extracts of different medicinal plants and antibiotics have been used against poultry bacterial pathogens. However, overuse of antibiotics and extracts against pathogenic strains leads to the proliferation of multi-drug resistant bacteria. Due to their environmentally friendly nature, nanotechnology and beneficial bacterial strains can be used as effective strategies against poultry infections. Green synthesis of zinc oxide nanoparticles (ZnO-NPs) from Eucalyptus globulus leaves was carried out in this study. Their characterization was done by UV-vis spectroscopy, X-ray diffraction (XRD), and Fourier transmission infrared spectroscopy (FT-IR) which confirmed their synthesis, structure, and size. In vitro, antimicrobial activities of plant leaf extract, ZnO-NPs, and plant growth-promoting rhizobacteria (PGPR) were checked against E. coli using well diffusion as well as disc diffusion method. Results proved that the antimicrobialactivity of ZnO-NPs and PGPR strains was more enhancedwhen compared to eucalyptus leaf extract at 36 h. The maximum relative inhibition shown by ZnO-NPs, PGPR strains and eucalyptus leaf extracts was 88%, 67% and 58%, respectively. The effectiveness of ZnO-NPs was also increased with an increase in particle dose and treatment time. The 90 mg/ml of ZnO-NPs was more effective. PGPR strains from all over the tested strains, Pseudomonas sp. (HY8N) exhibited a strong antagonism against the E. coli strain as compared to other PGPR strains used in this study. However, combined application of PGPR (Pseudomonas sp. (HY8N)) and ZnO-NPs augment antagonistic effects and showed maximum 69% antagonism. The study intends to investigate the binding affinity of ZnO-NPs with the suitable receptor of the bacterial pathogen by in silico methods. The binding site conformations showed that the ligand ZnO binds with conserved binding site of penicillin-binding protein 6 (PBP 6) receptor. According to the interactions, ZnO-NPs form the same interaction pattern with respect to other reported ligands, hence it can play a significant role in the inhibition of PBP 6. This research also found that combining ZnO-NPs with Pseudomonas sp. (HY8N) was a novel and effective technique for treating pathogenic bacteria, including multidrug-resistant bacteria.

Therapeutic effect of Schisandrin A on avian colibacillosis through gut-liver axis

This study evaluated the therapeutic efficacy of Schisandrin A on systemic colibacillosis of chickens. One hundred and eighty, 1-day-old Hailan Brown chickens were divided into 6 groups of 30 chickens each and assigned to the following treatments: 1) uninfected/untreated control; 2) infected Escherichia coli; 3) infected-plus low dose of Schisandrin A therapy (50 mg/kg); 4) infected-plus medium dose of Schisandrin A therapy (100 mg/kg); 5) infected-plus high dose of Schisandrin A therapy (200 mg/kg) and 6) infected-plus antimicrobial therapy (florfenicol). Each group of chickens was placed in cages with a photoperiod of 12 h of light and 12 h of dark. Feed and water for all groups were provided ad libitum for the duration of the study. On d 14, all the chickens except the uninfected control group were intraperitoneally inoculated with a fresh culture of E. coli containing 1 × 108 CFU/mL. The parameters measured included: average daily weight gain (ADG), percent survivability, liver index, serum activity of enzymes (ALT and AST), hepatic and intestinal concentrations of TNF-α, IL-1β, IL-6, IL-8, and LPS, expression of tight junction proteins (occludin, ZO-1, and claudin-1), relative abundance of bacterial species and histopathological changes in hepatic and intestinal tissue. The results showed that the medium and high doses of Schisandrin A ameliorated the detrimental effects of colibacillosis on weight gain. Regarding organ indexes, E. coli infection induced a significant increase in liver index, all the doses of Schisandrin A produced a significant reduction of liver index in comparison to the E. coli infected control. Serum activity of ALT and AST enzymes significantly increased due to E. coli infection, with the exception of the low dose of Schisandrin A for AST enzyme activity, all the Schisandrin A treatments significantly lowered enzyme activity in comparison to the E. coli infected control. Regarding concentrations of inflammatory markers in hepatic and intestinal, E. coli infection caused a significant increase in TNF-α, IL-1β, IL-6, and IL-8, except the lowest dose of Schisandrin A for IL-1β, the rest of the doses tested were able to significantly reduced the concentrations of inflammatory markers. Concentrations of LPS in hepatic and intestinal tissues were significantly increased by E. coli infection, all doses of Schisandrin A significantly reduced the concentration of LPS in hepatic and intestinal tissue. E. coli infection significantly reduced the expression of 2 tight junction proteins (ZO-1 and Claudin-1), the higher doses of Schisandrin A were effective in significantly increasing the expression of these tight junction proteins when compared with the E. coli infected control. Taken together, these results show that Schisandrin A has potential as an alternative therapy for the treatment of colibacillosis in chickens.

Zoonotic Potential of Escherichia Coli in Poultry Intestinal Contents in Ismailia City, Egypt with Special Reference to Shiga Toxin-Producing (STEC) Strains

Avian colibacillosis can affect birds of all ages. It is now one of the major causes of economic losses in the poultry industry, as well as a public health risk all over the world.This study aimed to determine the occurrence of E. coli in market-age poultry and the risk of its zoonotic infections in Ismailia city, Egypt. A total of 350 samples were collected from the intestinal contents of the slaughtered poultry (100 farm chickens, 100 backyard chickens, 50 ducks, 50 geese, and 50 turkeys) in Ismailia City. On the other hand, a total of 200 samples were collected from humans at outpatient clinics (100 stool samples and 100 urine samples) from the same investigated area. Samples were examined using bacteriological and molecular examination for genus-specific and virulence gene detection. Results revealed that the isolation rates of E. coli were 70% in farm chickens and turkeys, 88% in backyard chickens, 86% in domestic ducks, 72% in geese. Of avian E. coli isolates, the rates of Shiga toxin-producing E. coli (STEC) strains were 91.4%, 96.6%, 90.9%, 41.7%, and 51.4%% in farm chickens, backyard chickens, ducks, geese, and turkey respectively. In humans, the isolation rate of E. coli was 42% in urine samples and 66% in stool samples. The human STEC isolates were higher in stool samples (26%) than in urine samples (6%). The isolation rates of E. coli were significantly higher in persons who were in contact with poultry than in persons who were not in contact with poultry (p ≤ 0.01). In conclusion, the high isolation rates of STEC and detection with similarities of some E. coli virulence genes (It, St, eaeA, Stx1, and Stx2 genes) from poultry intestinal contents and human samples indicated a significant risk of zoonotic transmission of E. coli via food chain in the investigated area.

Detection of virulence genes of APEC (avian pathogenic Escherichia coli) isolated from poultry in Noakhali, Bangladesh

Avian colibacillosis, caused by avian pathogenic Escherichia coli (APEC), is one of the major infectious diseases of poultry that bring about great economic loss for the Bangladesh poultry industry. The present study aimed to determine the virulence genes of avian pathogenic Escherichia coli (APEC) from cases of colibacillosis in poultry at the Noakhali district of Bangladesh. Currently, virulence-associated gene profiles of APEC isolates were investigated by polymerase chain reaction (PCR). A total of 24 (twenty-four) Escherichia coli isolates were collected and presumptively identified from 8 (eight) colibacillosis cases from 4 commercial broiler poultry farms (2 broilers per farm) in Noakhali, Bangladesh. The pathogenesis of Escherichia coli involves a wide range of different virulence genes. At this point, four virulence genes, iutA, hlyF, iroN, and iss were detected by PCR analysis. It has been observed that iutA, iss, hlyF, and iroN genes were found in 7(29.16%), 20(83.33%), 22(91.66%), and 24(100%) APEC isolates respectively. Furthermore, out of the twenty-four APEC isolates, six (25%) isolates had four virulence genes, fourteen (58.33%) isolates carried at least three virulence genes, three (12.5%) isolates carried two genes and one (4.16%) isolates had one virulence gene. Most importantly. six types of virulence gene profiles existed within the APEC isolates from which profile number 3 (hlyF, iroN, iss) having 13 (54.16%) isolates were predominant. The occurrence of APEC isolates of this region which is responsible for avian colibacillosis cases can be a matter of concern from the public health point of view. Future investigations will be able to utilize these virulence genes to identify APEC in Bangladesh helping in the diagnosis and prevention of colibacillosis in poultry.
 Bioresearch Commu. 7(1): 967-972, 2021 (January)

Development of multiplex PCR for rapid and simultaneous detection of E. coli (APEC), Salmonella, Mycoplasma gallisepticum and Mycoplasma Synoviae

The Escherichia coli (E. coli), Salmonella, Mycoplasma synoviae (MS) and Mycoplasma gallisepticum (MG) are the major avian bacterial pathogens closely related to avian colibacillosis, salmonellosis, and mycoplasmosis respectively. In present study a single-tube multiplex PCR (m-PCR) assay was developed to allow the rapid, specific and simultaneous detection of these pathogenic species of major avian bacterial diseases. For this purpose, species specific genes of the relevant pathogens were first identified and utilized to obtain similar thermal profile conditions in singleplex PCR against the species-specific target genes “vat” of E. coli, “FimH” of Salmonella, “DNA polymerase III PolC” of MS and “mgc2” gene of MG. The primer pairs amplified specifically DNA fragment of the target genes in singleplex PCR, of 200 bp (vat), 300 bp (fimH), 380 bp (DNA polymerase III PolC) and 450 bp (mgc2). The validated species-specific primers from the similar thermal profile of singleplex PCR were then simultaneously subjected to optimize multiplex PCR condition at Tm 63 °C and amplified the same size products on agarose gel. The m-PCR was able to detect and differentiate simultaneously the aforementioned four major pathogenic species. The assay was corroborated for validation against field trial at the laboratory. The developed multiplex PCR for simultaneous identification of the aforementioned avian bacterial pathogens will help to enhance the diagnostic procedures of different poultry laboratories for field samples.

Antibiotic Resistance and Virulence Gene Patterns Associated with Avian Pathogenic Escherichia coli (APEC) from Broiler Chickens in Qatar.

Avian Pathogenic Escherichia coli (APEC) is the contributing agent behind the avian infectious disease colibacillosis, which causes substantial fatalities in poultry industries that has a significant impact on the economy and food safety. Several virulence genes have been shown to be concomitant with the extraintestinal survival of APEC. This study investigates the antibiotic resistance patterns and APEC-associated virulence genes content in Escherichia coli isolated from non-healthy and healthy broiler chickens from a commercial poultry farm in Qatar. A total of 158 E. coli strains were isolated from 47 chickens from five different organs (air sac, cloacal, kidney, liver, and trachea). Based on genetic criteria, 65% were APEC strains containing five or more virulence genes, and 34% were non-pathogenic E. coli (NPEC) strains. The genes ompT, hlyF, iroN, tsh, vat, iss, cvi/cva, and iucD were significantly prevalent in all APEC strains. E. coli isolates showed 96% resistance to at least one of the 18 antibiotics, with high resistance to ampicillin, cephalothin, ciprofloxacin, tetracycline, and fosfomycin. Our findings indicate high antibiotic resistance prevalence in non-healthy and healthy chicken carcasses. Such resistant E. coli can spread to humans. Hence, special programs are required to monitor the use of antibiotics in chicken production in Qatar.

Studies of Virulence Factors of Avian Pathogenic Escherichia coli in Avian Colibacillosis by In vitro Method and PCR

Background: Avian colibacillosis is considered as major cause of morbidity and mortality in poultry. It is a common bacterial disease of poultry and many virulence factors of E. coli are associated with the disease. The current study was aimed to investigate the presence of some virulence factors of E. coli isolated from the cases of colibacillosis.Methods: In present study, total 150 samples (liver, heart, lungs, air sacs and feaces) of chicken exhibiting pathological conditions of colibacillosis were collected from various poultry farms (organized and backyard) situated in and around Mhow and Indore cities. E.coli was isolated and identified from the samples on the basis of cultural characteristics and biochemical test. All E. coli isolates were further subjected to evaluate the presence of virulence factors such as biofilm production, haemolysis, invasiveness and molecular detection of fimH and stx1 gene.Result: Out of these 51.33% of incidence of E. coli was recorded. E. coli O84 and O149 serotypes were found most prevalent. Out of 77 isolates, 46 (59.7%) and 45 (58.4%) were positive for biofilm formation by tube method and modified CRA method, respectively. All E. coli isolates were showing invasiveness in congo red binding assay while none of the isolates was found haemolytic. Molecular detection revealed the presence of fimH (508bp) gene in 33.3% of tested samples while stx1 gene could not be detected in any isolates.