Award details

Functional characterisation of a Campylobacter flagellin glycosylation island important in avian adaptation

Reference	BB/D521849/1
Principal Investigator / Supervisor	Professor Brendan Wren
Co-Investigators / Co-Supervisors
Institution	London Sch of Hygiene and Trop Medicine
Department	Infectious and Tropical Diseases
Funding type	Research
Value (£)	187,158
Status	Completed
Type	Research Grant
Start date	01/01/2006
End date	31/12/2008
Duration	36 months

Abstract

Campylobacter jejuni is the principal bacterial cause of human gastroenteritis worldwide. The main source of transmission is the consumption and handling of contaminated poultry, but the underlying reasons why most chickens are particularly susceptible to colonisation by C. jejuni are unknown. To investigate genetic determinants in C. jejuni that are involved in this process, we compared the genomes of 91 C. jejuni strains of diverse origin. The resulting phylogenetic tree was made up of two distinct clades - one comprising the chicken-associated strains, and one comprising the strains that were not chicken-associated. We found that the most prominent genetic marker shared by the chicken-associated strains was a cluster of six genes within the O-linked flagellum glycosylation locus (coding sequences cj1321 to cj1326). It is known that flagellin glycan structures, which are exposed on the cell surface, are important in determining the immune response of the host - they have been found to be immunodominant in C. jejuni and to be important in host specificity in other bacterial pathogens. We hypothesise that the cj1321-26 island enables C. jejuni strains that possess it to persist in poultry, by encoding a specific glycoform of the flagellin that plays an integral role in the bacterium's interaction with its avian host. The presence of variant flagellin glycoforms encoded by this island may confer a combination of altered properties relating to colonisation, survival and immune response. To investigate this hypothesis, we propose to thoroughly characterise the genetics, structure, function and immunology of the O-linked flagellin modification system in C. jejuni 11168H, including the newly identified cj1321-26 island. First, we will mutate and complement each of the six genes in the cj1321-26 glycosylation island, as well as generating defined and spontaneous deletion mutants of the island. We will then carry out studies to test the ability of the different strains tocolonise and survive in chickens, as well as histological and avian immune response studies. To test whether selective pressure is required to maintain the cj1321-26 glycosylation island in the avian host, we will compare the rate of spontaneous loss of the island in vitro and in vivo. To assess how the cj1321-26 genes affect cell surface properties and may be relevant in human disease, we will test the mutants in a range of in vitro assays including hydrophobicity, autoagglutination, serum resistance, motility, biofilm formation and bile salt resistance. We will also test the parent and mutant strains ability to adhere to and invade CaCo2 human tissue culture cells, as well as their ability to trigger human innate immune and cytokine responses. In addition, we will use NMR and mass spectroscopy to determine the structure of the flagellin glycan in the parent 11168H strain and in each of the mutants. This research programme will improve our understanding of the interactions between Campylobacter and chickens, and may make it possible to select chickens that are less susceptible to colonisation by this bacterium. In the longer term, this approach will help inform strategies to reduce C. jejuni from the food chain. Joint with BB/D521965/1.

Summary

Campylobacter jejuni is a type of bacterium that causes severe food poisoning. It is the most common cause of bacterial food poisoning in the world and it is usually caught from handling or eating contaminated poultry, especially chickens. Large numbers of C. jejuni bacteria can live inside chickens without causing any symptoms, but we don¿t understand how they survive in chickens, and why they cause disease in people but not in birds. The overall aim of this research is to find ways to reduce the number of C. jejuni bacteria in the food chain, especially in chickens, to reduce the number of cases of food poisoning. Antibiotic resistance is a growing problem, so we need to find clever alternatives to antibiotics by understanding exactly how the bacteria work. We hope that if we can understand how C. jejuni survives in chickens, we can work out how to block the process, perhaps by feeding the birds harmless bacteria that stop the disease-causing ones from surviving, or by selecting chickens that the bacteria aren¿t able to live in. Before writing this grant proposal, we compared the genomes of 91 different strains of C. jejuni. These strains had been isolated from all over the place: chickens, cows, sheep and humans, as well as from the general environment. We found a set of six genes that were only present in the strains that came from chickens, and we think that these genes may be key to explaining how the bacteria are able to survive inside birds. This is exciting, because it is the first time that anyone has found a gene or set of genes that is specific to strains of C. jejuni from one particular source. The genes we found are within a region of the genome that is involved in modifying a particular protein, called flagellin, which appears on the surface of the bacteria. The genes modify the flagellin protein by adding sugar structures to it. We know that in order disease-causing bacteria, this kind of sugar modification is important because it affects they way the bacteria bind to cells in the animals they infect, and it affects the kind of immune response they trigger in the animals. In the proposed programme of research, we went to investigate these six genes (we¿ve called them cj1321-26) in detail, to find out exactly what they do, and just how they help the bacteria to survive in the chicken. Joint with BB/D521965/1.

Committee	Closed Committee - Agri-food (AF)
Research Topics	Animal Health, Immunology, Microbial Food Safety, Microbiology, Structural Biology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

Associated awards:

BB/D521965/1 Functional characterisation of a Campylobacter flagellin glycosylation island important in avian adaptation

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