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Mechanisms behind the host-specificity of bacteria, investigated by use of host-specific salmonella serotypes

Contract nr: FAIR-CT96-1743
Project nr: 1743
Project type: SC
Starting date: 01/02/1997
Duration: 48 months
Total cost: 1,415,000 EUR
EC Contribution: 1,056,000 EUR
Scientific Officer: Isabel MINGUEZ-TUDELA
Research topic: Animal health
Acronym: HOST-SPECIFIC SALMONELLAS

Background:
Host-specific bacteria cause significant economic loss due to disease in all production animals, a reality that is to the detriment of production economy and to the welfare of animals. The mechanisms responsible for this host-specificity are only poorly understood, and due to the lack of knowledge, it has never been possibly to evaluate whether host-specificity factors can be utilised for specific prevention and control of diseases. Salmonellosis is currently the most important zoonosis within the EU. Host-speciflc Salmonella serotypes cause economically significant diseases in all common production animals, as well as life threatening, systemic human salmonellosis.

Objectives:
In the present project, a systematic investigation will be conducted into the molecular mechanisms behind the host-specificity of host-specific Salmonella serotypes, i.e. S. Dublin, S. Gallinarum, S.abortusovis, and S. Choleraesuis. The project has six participants, two of which have one sub-contractor each. The overall objective is to establish a basis for utilising host-range determining factors of bacteria as a means for controlling disease at the pre-harvest level, and to investigate if host-specific Salmonella are good candidates as vectors in live-vaccine constructs.

Description:
In sub-project 1(four tasks with eight sub-tasks), the specific objectives are to describe the biology of the host-specificity, and to investigate if the host-immune response towards a hostspecific serotype is different in the host to which it is adapted, compared with other animals. Challenge experiments in cattle, pigs, sheep and poultry, and assays with cell and tissue cultures from the same animals, are employed.

Specific tasks aim to identify the stages during infection where the host-specific phenotypes are manifested, to investigate if the serotype-specific virulence plasmids are significant for host-specificity, and to investigate if challenged animals are protected against challenge with the other host-specific serotypes. In sub-project 2 (six tasks with two sub-tasks), molecular methods are used to identify and characterise the factor(s) responsible for the host-specificity of the host-specific Salmonella serotypes. The factors can be traits of the bacteria as well as of the host, and are probably different for different serotypes. Factors are identified from studies of mutant and recombinant strains that show an altered host restriction phenotype.

Characterisation of the factors include identification of the relevant DNA fragments, mapping of the fragments, genetic analysis of the nature of mutations, identification of the relevant genes, sequence and expression analysis, and analysis of the ability of the DNA to change the host-specificity of other host-specific Salmonella serotypes. In sub-project 3 (two tasks), the ability of S. Abortusovis to act as a vector for heterologaus antigen presentation in a live vaccine is investigated. The gene for E. Coli maltose binding protein (male) is cloned into the attenuated strain Rv6 and the immune response towards this protein is measured in mice and sheep. Next, small fragments of CAEV (Caprine Arthritis Encephalitis Virus) genome are cloned into the male and the protective ability of these antigens is measured by challenging with the virus.

The project is carried out over a four year period. Major deliverables will be in the form of publication in international journals from all sub-projects, sets of well-characterised strains with different genetic manipulations (which will be in the public domain), a quantitative assay for measurement of cytokine profiles, fragments of DNA encoding, one or more factors that are responsible for host-specificity, an evaluation report on the possible use of host-restriction factors in control of diseases, and an evaluation report on the possible use of host-specific serotypes as carriers of heterologous antigens. Commercialisation will be considered before publication of results.

Current situation/results:
After 36 months of studies, the biology of Salmonella host-specificity and host-adaptation has been investigated by challenge experiments, studies of invasion in intestinal loop assays - some of which have been developed for the project - and by in vitro methods. It has been firmly established that there is no correlation between the ability to invade in the intestine and the host adapted/host-specific phenotypes seen with these bacteria.

The host-specific and host-adapted serotypes are characterised by the ability to reach high counts at systemic sites following challenge. They differ in their ability to invade and survive in cell cultures and primary cultures of phagocytic cells, but not in a host-specific manner. Nor do they seem to induce a host-specific oxidative or nitric oxide response from phagocytic cells. The role of the virulence plasmid is being investigated.

The host response towards the host-specific infection has been investigated in mice and sheep, in terms of antibody response and cytokine profiles. For this purpose, a number of cytokine-RT-PCR methods have been developed. The possibility that immunity towards one serotype may protect against infection caused by other host-specific serotypes has also been considered. Mutants of the four serotypes S. abortusovis, S. choleraesuis, S. dublin and S. gallinarum have been constructed by signature-tagged transposon mutagenesis, and libraries of strains - where the chromosome of other serotypes have been cloned-in - are under construction. The study of the host-specificity and host-adaptation of mutant strains is ongoing, while recombinant strains will be tested when produced. Putative host-specificity genes have already been obtained.

With a view to vaccine development, it has been shown that the host-specific serotype S. abortusovis can be used to prime the immune system of sheep with foreign antigen. Antibodies were detected against MalE from E. coli and the Gag-protein from CAEV virus, when genes encoding parts of these proteins were cloned and expressed in S. abortusovis. However, no protection against CAEV-virus was obtained.

Website: http://www.vetmi.kvl.dk/fair3-ct96.html


Coordinator
John Elmerdahl OLSEN
Royal Veterinaryand Agricultural University
Bülowsvej 13
DK-1870 Frederiksberg C
Tel.: +45 35 28 27 84
Fax: +45 35 28 27 57
E-mail: John.E.Olsen@vetmi.kvl.dk


Partners

  • Guido LEORI
    Istituto Zooprofilattico Sperimentale della Sardegna g. Pegreffi
    Via Duca Degli Abruzzi 8
    I-07100 Sassari
    Tel.: +39 079 28 92 31
    Fax: +39 079 27 50 40
    E-mail: Guido-Leori@sardegna.izs.it

  • Frederic LANTIER
    I.N.R.A. - Centre de Recherche de Tours-Nouzilly
    F-37380 Nouzilly
    Tel.: +33 2 47 42 78 68
    Fax: +33 2 47 42 77 79
    E-mail: frederic.lantier@tours.inra.fr

  • Salvatore RUBINO
    Università degli Studi di Sassari
    Viale San Pietro 43b
    I-07100 Sassari
    Tel.: +39 079 22 83 02
    Fax: +39 079 21 23 45
    E-mail: microb@ssmain.uniss.it

  • Timothy Stephen WALLIS
    Institute for Animal Health
    Compton Laboratory
    UK-RG20 7NN Compton - NR Newbury
    Tel.: +44 1635 57 84 11
    Fax: +44 1635 57 72 37
    E-mail: wallis@bbsrc.ac.uk

  • Josep CASADESUS
    Universidad de Sevilla
    Apartado 1095
    E-41080 Sevilla
    Tel.: +34 95 455 71 05
    Fax: +34 95 455 71 04
    E-mail: genbac@cica.es
 
 
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