Analyse der antigenen Proteine von Mycoplasma mycoides subsp. mycoides SC

• Forschungsstelle: BLV
• Projektnummer: 1.01.13
• Projekttitel Englisch: Analysis of antigenic proteins of Mycoplasma mycoides subsp. mycoides SC

Schlüsselwörter (Englisch)

Contagious bovine pleuropneumonia, Mycoplasma, animal health, diagnostics, gene expression, site directed mutagenesis

Kurzbeschreibung (Englisch)

Mycoplasma mycoides subsp. mycoides small colony type (SC) is the etiological agent of contagious bovine pleuropneumonia, a severe respiratory disease of cattle, which represents a major constraint in cattle raising and livestock production in Africa and which threatens also the European continent, as proved the reemerging outbreaks in the last 15 years in several countries. The early diagnosis of the disease, which is essential to prevent outbreaks and spread of CBPP, requests more specific and more sensitive serological assays. The current project aims at a genetic, biochemical and antigenic analysis of major membrane proteins of M. mycoides subsp. mycoides SC, which play a central role in antigenicity. Genes encoding membrane proteins will be cloned and mutated using site specific mutagenesis in order to allow their expression in non-pathogenic microorganisms used as standard gene expression systems. The antigens will subsequently be purified and analyzed biochemically and immunologically for their suitability in developments of serological diagnostic methods.

Projektziele (Englisch)

The objective of the current research on the analysis of antigenic proteins of Mycoplasma mycoides subsp. mycoides SC is to detect and characterise those antigens, particularly surface antigens which are stable and elicit a specific immune response during natural infection of cattle with M. mycoides subsp. mycoides SC. Such antigens will be used for further developments of serological methods for the early detection of CBPP. Particular attention will be given to those antigens which are present in both, the European cluster and the African/Australian cluster strains. Preliminary results obtained from field sera of cattle which were naturally infected with M. mycoides subsp. mycoides SC revealed a large number of seroreactive proteins. The nature of only a few of these proteins is meanwhile identified, but their function in pathogenicity is yet unclear. Certain of these factors are expected to be ideal antigens as specific markers for M. mycoides subsp. mycoides SC infections, while others are expected to give cross reactions. It will reveal which of the antigens of M. mycoides subsp. mycoides SC can be considered as components for developments of sero-diagnostic and sero-epidemiological methods to detect CBPP and which of them cross react with related Mycoplasma species. Furthermore the current study aims in concentrating on those specific antigens for M. mycoides subsp. mycoides SC which would be potentially useful to serologically distinguish animals which were vaccinated with modern vaccines from those which were infected with M. mycoides subsp. mycoides SC. A tight collaboration with the 5th EU Frameworkprogramme INCO (Protective immunity against M. mycoides subsp. mycoides SC infections) is planned.

Abstract (Deutsch)

Das in diesem Projekt entdeckte und näher charakterisierte Antigen LppC ist ein immuno-dominantes Oberflächenprotein von M. mycoides subsp. mycoides SC. Das Gen lppC, welches für dieses Lipoprotein kodiert, ist in allen geprüften Stämmen von M. mycoides subsp. mycoides SC vorhanden und scheint auch in verwandten Mycoplasma Spezies vorzukommen. Das lppC Gen wird aber in diesen Spezies nicht exprimiert. LppC dürfte sich deshalb zusammen mit dem Lipoprotein LppQ für serodiagnostische Zwecke eignen. Die genetischen Kenntnisse der Lipoprotein-Gene und anderer genetischen Elemente von M. mycoides subsp. mycoides SC dienten zur Entwicklung molekularer Methoden für die Detektion und Identifikation von Mykosplasmen die bei Ruminanten Infektionen erzeugen.

Publikationen / Ergebnisse (Deutsch)

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Publikationen / Ergebnisse (Englisch)

Bruderer, U., Regalla, J., El-Mostafa, A., Huebschle, O. J.B., Frey, J. (2002): Serodiagnosis and monitoring of contagious bovine pleuropneumonia (CBPP) with an indirect ELISA baded on the specific lipoprotein LppQ of Mycoplasma mycoides subsp. mycoides SC. Veterinary Microbiology 84, 195-205.

Vilei, E.M., Abdo, E-M, Nicolet, J., Botelho, A., Gonçalves, R., Frey, J. (2000): Genomic and antigenic differences between the European and African/Australien clusters of Mycoplasma mycoides subs. mycoides SC. Microbiology 146, 477-486.

Monnerat, M., Thiaucourt, F., Nicolet, J., Frey, J. (1999): Comparative analysis of the lppA locus in Mycoplasma capricolum subsp. capricolum and Mycoplasma capricolum subsp. capripneumoniae. Veterinary Microbiology 69, 157-172.

Monnerat, M., Thiaucourt, Poveda, J.B., Nicolet, J., Frey, J. (1999): Genetic and Serological Analysis of Lipoprotein LppA in Mycoplasma mycoides subsp. mycoides LC and Mycoplasma mycoides subsp. capri. Clinical and Diagnostic Laboratory Immunology Vol. 6, No. 2, 224-230.

Vilei, E.M., Nicolet, J., Frey, J. (1999): IS1634, a Novel Insertion Element Creatin Long, Variable-Length Direct Repeats Which Is Specific for Mycoplasma mycoides subsp. mycoides Small-Colony Type. Journal of Bacteriology, Vol. 181, No. 4, 1319-1323.

Bruderer, U., Regalla, J., El-Mostafa, A., Huebschle, O. J.B., Frey, J. (2002): Serodiagnosis and monitoring of contagious bovine pleuropneumonia (CBPP) with an indirect ELISA baded on the specific lipoprotein LppQ of Mycoplasma mycoides subsp. mycoides SC. Veterinary Microbiology 84, 195-205.

Vilei, E.M., Abdo, E-M, Nicolet, J., Botelho, A., Gonçalves, R., Frey, J. (2000): Genomic and antigenic differences between the European and African/Australien clusters of Mycoplasma mycoides subs. mycoides SC. Microbiology 146, 477-486.

Monnerat, M., Thiaucourt, F., Nicolet, J., Frey, J. (1999): Comparative analysis of the lppA locus in Mycoplasma capricolum subsp. capricolum and Mycoplasma capricolum subsp. capripneumoniae. Veterinary Microbiology 69, 157-172.

Monnerat, M., Thiaucourt, Poveda, J.B., Nicolet, J., Frey, J. (1999): Genetic and Serological Analysis of Lipoprotein LppA in Mycoplasma mycoides subsp. mycoides LC and Mycoplasma mycoides subsp. capri. Clinical and Diagnostic Laboratory Immunology Vol. 6, No. 2, 224-230.

Vilei, E.M., Nicolet, J., Frey, J. (1999): IS1634, a Novel Insertion Element Creatin Long, Variable-Length Direct Repeats Which Is Specific for Mycoplasma mycoides subsp. mycoides Small-Colony Type. Journal of Bacteriology, Vol. 181, No. 4, 1319-1323.

Djordjevic, S.P., Vilei, E.M., Frey, J. (2003): Characterization of a chromosomal region of Mycoplasma sp. bovine group 7 strain PG50 encoding a glycerol transport locus (gtsABC). Microbiology 149, 195-204

Pilo, P., Martig, S., Frey, J., and Vilei, E.M. (2003) Antigenic and genetic characterisation of lipoprotein LppC from Mycoplasma mycoides subsp. mycoides SC. Vet. Res. 34: 761-775.

Mbulu, R.-S., Tjipura-Zaire, G., Lelli, R. Frey, J. Pilo, P., Vilei, E.M., Mettler, F., Nicholas R.A.J., and Huebschle, O.J.B. (2004) Contagious bovine pleuropneumonia (CBPP) caused by vaccine strain T1/44 of Mycoplasma mycoides subsp. mycoides SC. Vet. Microbiol. 98: 229-234.

Vilei, E.M. and Frey, J. (2004) Differential clustering of Mycoplasma mycoides subsp. mycoides SC strains by PCR-REA of the bgl locus. Vet. Microbiol. 100: 283-288.

Frey, J. (2002) Mycoplasmas of Animals. In: Molecular Biology and Pathogenicity of Mycoplasmas. S. Razin and R. Herrmann (Eds). Chapter Kluwer Academic/Plenum Publishers, New York, Boston, Dordrecht, London, Moscow pp 73-90.

Hübschle, O., Lelli, R., Frey, J. and Nicholas, R. (2002) Contagious bovine pleuropneumonia and vaccine strain T1/44. Letters, Vet. Rec. 150 (19): 615.

Hotzel, H., Frey, J., Bashiruddin, J. and Sachse, K. (2002) Detection and Differentiation of Ruminant Mycoplasmas. In: Methods in Molecular Biology Volume 216: PCR Detection of Microbial Pathogens. K. Sachse &
J. Frey (Eds). Humana Press Totowa, New Jersey. pp 231-245.

Pilo, P., (2004) Characterization of antigenic surface proteins of Mycoplasma mycoides subsp. mycoides Small Colony type. PhD Thesis, University of Bern.