The Campylobacter jejuni Response Regulator and Cyclic-Di-GMP Binding CbrR Is a Novel Regulator of Flagellar Motility

A leading cause of bacterial gastroenteritis, Campylobacter jejuni is also associated with broad sequelae, including extragastrointestinal conditions such as reactive arthritis and Guillain-Barré Syndrome (GBS). CbrR is a C. jejuni response regulator that is annotated as a diguanylate cyclase (DGC), an enzyme that catalyzes the synthesis of c-di-GMP, a universal bacterial second messenger, from GTP. In C. jejuni DRH212, we constructed an unmarked deletion mutant, cbrR−, and complemented mutant, cbrR+. Motility assays indicated a hyper-motile phenotype associated with cbrR−, whereas motility was deficient in cbrR+. The overexpression of CbrR in cbrR+ was accompanied by a reduction in expression of FlaA, the major flagellin. Biofilm assays and scanning electron microscopy demonstrated similarities between DRH212 and cbrR−; however, cbrR+ was unable to form significant biofilms. Transmission electron microscopy showed similar cell morphology between the three strains; however, cbrR+ cells lacked flagella. Differential radial capillary action of ligand assays (DRaCALA) showed that CbrR binds GTP and c-di-GMP. Liquid chromatography tandem mass spectrometry detected low levels of c-di-GMP in C. jejuni and in E. coli expressing CbrR. CbrR is therefore a negative regulator of FlaA expression and motility, a critical virulence factor in C. jejuni pathogenesis.

Evaluation of Three Isolation Experiments for Campylobacter Bacteriophages from Chicken Skin: A Comparative Study

Background: Campylobacter strains are of the leading pathogens causing bacterial gastroenteritis, whose infections are generally considered to be one of the most common foodborne illnesses of animal origin. The etiology of this infection often goes back to eating contaminated raw meat or infected poultry. The bacteria are present in abundance in chicken skin. The use of appropriate bacteriophages is one of the most effective experiments in eliminating Campylobacter strains. Phage therapy refers to the use of bacteriophages to treat bacterial infections. Aim: Accordingly, the present study aimed to compare three experiments of bacteriophage isolation in chicken skin. Experiments: Thus, 15 samples of chicken skin were collected from five different fresh chicken suppliers in Ghaemshahr, Iran. The samples were transported to the laboratory aseptically in the vicinity of ice, and then cultured in blood agar medium, and the isolates were identified by various tests including gram staining, catalase and oxidase tests. Results: The results were compared before and after three bacteriophage isolation experiments. Out of 15 chicken skin samples tested in all three experiments, 6 (40%) strains were identified in the first experiment, 8 (53.4%) strains in the second experiment and 12 (20%) strains in the third experiment after bacteriophage therapy. Conclusion: The bacteriophage isolation experiments alone or in combination with other intervention strategies are recommended as promising tools for greater food safety. These experiments can be useful to increase food safety and reduce the risk of infection in humans through the consumption of potentially infected edible parts of chicken. According to the results of this study, among the three proposed experiments, the experiment of chicken skin enrichment in Bolton selective media containing target isolates was the most efficient approach, which showed a high limit of detection at low concentrations and the highest rate of phage recovery. This can be a more reliable way to isolate the Campylobacter bacteriophages and eliminate the Campylobacter strains.

Low Frequency of Adenovirus, Rotavirus, and Norovirus in Pediatric Diarrheal Samples from Central Iran

Background: Acute viral gastroenteritis is a disorder that affects children globally but mostly in developing countries. Adenoviruses, rotaviruses, and noroviruses are the leading viral causes of childhood gastroenteritis. Objectives: This study is the first to investigate the frequency of these viruses in diarrheal samples from pediatric patients living in central Iran. Methods: A total of 173 samples of pediatric diarrhea, from May 2015 to May 2016, were included in this descriptive cross-sectional study. The samples were analyzed using in-house developed PCR and reverse transcription (RT)-PCR methods to investigate the frequency of adenoviruses, rotaviruses, and noroviruses. Results: Out of 173 samples of pediatric diarrhea, eight were shown to contain enteric viruses (4.6%): (1) four with adenoviruses (2.3%); (2) three with rotaviruses (1.7%); and (3) one with a genogroup II norovirus (0.6%). Most of the positive samples were obtained from children under the age of seven. The most common additional clinical symptoms in pediatric patients with viral agents were fever, vomiting, and abdominal pain. Conclusions: In central Iran, adenoviruses and rotaviruses were rarely found as agents responsible for gastroenteritis. Although viral gastroenteritis in this area had less frequency than bacterial gastroenteritis, we need to monitor all enteropathogenic agents for longer periods to understand better real endemicity and the possibility of unexpected viral enteritis outbreaks.

Multi-virulence and phenotypic spread of Campylobacter jejuni carried by chicken meat in Brazil

Campylobacter jejuni is the most incriminated pathogen in bacterial gastroenteritis, and therefore, characteristics of its epidemiology must be continuously investigated to support possible mitigating measures. This is particularly important when evaluating representative strains of the world's leading chicken meat exporter, Brazil. We evaluated a panel of 14 virulence genes in 359 strains of C. jejuni isolated from chilled broiler carcasses of Brazil. The genes were classified into five virulence categories (B: biofilm/motility; SS: secretion/cytotoxicity system; CI: invasion/colonization; GB: Guillain-Barré and AE: adaptation to stress). The percentage of strains with stress adaptation genes (86.07%) indicates the potential to adapt to unfavorable environmental conditions and hcp gene in 97.77%, indicates the ability to cause serious infections in humans. Genes related to GBS in 77.44% of strains are an additional concern, which must be monitored. The gene panel showed the presence of 124 virulence profiles. Individual analyzes by carcass, slaughter establishment, and municipalities where they were located showed high I.Var., of 0.82, 0.87 and 0.78, respectively. Georeferencing indicated state A as a hotspot for virulent strains. Higher levels of isolation and multi-virulence were identified in the summer, which in Brazil is hot and humid. Proteomics was able to discriminate the strains, but due to the high heterogeneity between them, it did not allow to explain their dissemination. Together, our results showed that the studied strains are a potential danger to public health and that there is an urgent need for their surveillance and the adoption of control measures, especially in state A.

Campylobacter Biofilms: Potential of Natural Compounds to Disrupt Campylobacter jejuni Transmission

Microbial biofilms occur naturally in many environmental niches and can be a significant reservoir of infectious microbes in zoonotically transmitted diseases such as that caused by Campylobacter jejuni, the leading cause of acute human bacterial gastroenteritis world-wide. The greatest challenge in reducing the disease caused by this organism is reducing transmission of C. jejuni to humans from poultry via the food chain. Biofilms enhance the stress tolerance and antimicrobial resistance of the microorganisms they harbor and are considered to play a crucial role for Campylobacter spp. survival and transmission to humans. Unconventional approaches to control biofilms and to improve the efficacy of currently used antibiotics are urgently needed. This review summarizes the use plant- and microorganism-derived antimicrobial and antibiofilm compounds such as essential oils, antimicrobial peptides (AMPs), polyphenolic extracts, algae extracts, probiotic-derived factors, d-amino acids (DAs) and glycolipid biosurfactants with potential to control biofilms formed by Campylobacter, and the suggested mechanisms of their action. Further investigation and use of such natural compounds could improve preventative and remedial strategies aimed to limit the transmission of campylobacters and other human pathogens via the food chain.

Immunization with Leishmania tarentolae-derived norovirus virus-like particles elicits high humoral response and stimulates the production of neutralizing antibodies

Background Noroviruses are a major cause of epidemic and sporadic acute non-bacterial gastroenteritis worldwide. Unfortunately, the development of an effective norovirus vaccine has proven difficult and no prophylactic vaccine is currently available. Further research on norovirus vaccine development should be considered an absolute priority and novel vaccine candidates are needed. One of the recent approaches in safe vaccine development is the use of virus-like particles (VLPs). VLP-based vaccines show great immunogenic potential as they mimic the morphology and structure of viral particles without the presence of the virus genome. Results This study is the first report showing successful production of norovirus VLPs in the protozoan Leishmania tarentolae (L. tarentolae) expression system. Protozoan derived vaccine candidate is highly immunogenic and able to not only induce a strong immune response (antibody titer reached 104) but also stimulate the production of neutralizing antibodies confirmed by receptor blocking assay. Antibody titers able to reduce VLP binding to the receptor by > 50% (BT50) were observed for 1:5–1:320 serum dilutions. Conclusions Norovirus VLPs produced in L. tarentolae could be relevant for the development of the norovirus vaccine.

Aetiology and risk factors of bacterial gastroenteritis among febrile outpatients at the Dschang District Hospital, West Region of Cameroon: a cross-sectional study

ObjectivesTo investigate the bacterial aetiologies and associated risk factors of gastroenteritis among typhoid suspected cases.DesignCross-sectional study.SettingThis study was conducted at Dschang District Hospital of the Menoua Division, West Region of Cameroon, between April–November 2019 and June 2020.ParticipantsParticipants aged ≥2 years (mean 34±18.77 years) and of both sex suspected of having typhoid fever were included, while non-suspected typhoid cases were excluded. Self-reported sociodemographic and health information at recruitment was obtained from 556 participants.MethodsCollected stool samples were examined macroscopically and microscopically and subjected to culture. After culture, Gram staining was performed, followed by biochemical testing and characterisation using the Analytical Profile Index (API-20E) test kit.Interventions’No intervention was done during the period of study.Outcome measuresWe identified bacterial causing gastroenteritis, and associated risk factors calculated using binary regression, adjusting for sociodemographic and health variables.ResultsOf 556 patients, 74.28% tested positive for gastroenteritis. Among pathogens responsible for gastroenteritis, Escherichia coli was found to be the main cause (21.1%), followed by Salmonella typhi (10.4%), Citrobacter diversus (8.2%), and Proteus mirabilis (8.2%), Proteus vulgaris (7.3%), whereas Citrobacter spp and Yersinia enterocolitica were less represented among pathogens causing the disease among patients. A significant difference (p=0.002) was observed between abdominal pain and all the micro-organisms isolated from the patients. Patients having primary level of education were significantly associated (p=0.017; 3.163 (95% CI 1.228 to 8.147)) with the prevalence of gastroenteritis. Consumption of beverages (Wald statistic: 4.823; OR: 2.471; 95% CI (1.102 to 5.539); p=0.028), use of modern toilet (Wald statistic: 4.471; OR: 1.723; 95% CI (1.041 to 2.852); p=0.034) were strongly associated with gastroenteritis and rearing of bird (Wald statistic: 4.880; OR: 0.560; 95% CI (0.335 to 0.937); p=0.027), was found to be protective.ConclusionAcute bacterial gastroenteritis is a significant cause of morbidity in Dschang, with the prevalence of 74.28%. Many pathogens accounted for gastroenteritis, and E. coli (21.1%) could be a major cause, followed by S. typhi (10.4%), C. diversus (8.2%), P. mirabilis (8.2%), P. vulgaris (7.3%), whereas Citrobacter spp and Y. enterocolitica were less represented. Gastroenteritis was highly associated with primary level of education, consumption of beverages, use of modern toilet while rearing of birds was unexpectedly found to be protective against Gastroenteritis. Further characterisation is planned.

Vitamin D Reverses Disruption of Gut Epithelial Barrier Function Caused by Campylobacter jejuni

Infections by the zoonotic foodborne bacterium Campylobacter jejuni (C. jejuni) are among the most frequent causes of bacterial gastroenteritis worldwide. The aim was to evaluate the relationship between epithelial barrier disruption, mucosal immune activation, and vitamin D (VD) treatment during C. jejuni infection, using intestinal epithelial cells and mouse models focused on the interaction of C. jejuni with the VD signaling pathway and VD treatment to improve C. jejuni-induced barrier dysfunction. Our RNA-Seq data from campylobacteriosis patients demonstrate inhibition of VD receptor (VDR) downstream targets, consistent with suppression of immune function. Barrier-preserving effects of VD addition were identified in C. jejuni-infected epithelial cells and IL-10−/− mice. Furthermore, interference of C. jejuni with the VDR pathway was shown via VDR/retinoid X receptor (RXR) interaction. Paracellular leakiness of infected epithelia correlated with tight junction (TJ) protein redistribution off the TJ domain and apoptosis induction. Supplementation with VD reversed barrier impairment and prevented inhibition of the VDR pathway, as shown by restoration of transepithelial electrical resistance and fluorescein (332 Da) permeability. We conclude that VD treatment restores gut epithelial barrier functionality and decreases bacterial transmigration and might, therefore, be a promising compound for C. jejuni treatment in humans and animals.