Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 1/21/2003
Publication Date: 1/21/2003
Citation: BEARSON, S.M., COLLIER, S.D., BRANTON, S.L., BEARSON, B.L., MONTGOMERY, R.D. INDUCTION OF THE MYCOPLASMA GALLISEPTICUM PMGA1.2 GENE IN THE CHICKEN TRACHEAL RING ORGAN CULTURE MODEL. INTERNATIONAL POULTRY SCIENTIFIC FORUM. 2003. ABSTRACT P. 52.
Technical Abstract: Mycoplasma gallisepticum (MG), the causative agent of chronic respiratory disease in poultry, must adhere to tracheal epithelial cells to establish infection. To identify MG genes involved in respiratory tract colonization, an in vitro model system was developed utilizing chicken Tracheal Ring Organ Cultures (TROC) and the lacZ gene of Escherichia coli to serve as a reporter for mycoplasma gene expression. The chromosome of the MG S6 strain was randomly mutagenized following transformation with the plasmid pBSC5 containing the promoterless lacZ gene in the Tn4001 transposon. The MG mutants were independently screened for expression of the lacZ gene in the absence and presence of chicken TROC using the fluorescent substrate, 5-acetylaminofluorescein di-beta-D-galactopyranoside. Preliminary data from this screen revealed a mutant of MG that exhibited a 2-fold increase in beta-galactosidase production over a 22-h period in the presence of the TROC. To identify the mycoplasma gene promoter driving the increased expression of the lacZ gene in the presence of the TROC, the chromosome of the MG p12G6 mutant was digested, cloned and transformed into E. coli with selection for kanamycin resistance. DNA sequence analysis revealed that the Tn4001 transposon inserted into the MG gene pMGA 1.2, a member of a large gene family of cell surface adhesins that experience frequent phase variation and are associated with hemagglutination. Further analysis of the MG p12G6 mutant revealed a decrease in the ability to hemagglutinate chicken red blood cells compared to the wild-type strain.