Analysis of in vivo expressed genes in Mannheimia haemolytica A1

Abstract

The expression of Mannheimia haemolytica A1 genes during in vivo growth was examined by reverse transcriptase-polymerase chain reaction (RT-PCR) using total RNA extracted directly from M. haemolytica A1 recovered from pneumonic lungs of cattle. Primers specific for three groups of genes were used. Group 1 includes virulence-related genes: lktC, tbpB, ahs, nmaA, gs60 and gcp. Group 2 includes genes that code for putative two-component regulatory systems: narP, narQ, ttrR, ttrS, phoB and phoR. Group 3 includes genes involved in regular cellular functions such as plp4, thiL and rrf. The RT-PCR data were examined in conjunction with the percent pneumonic lesion in each lung scored during necropsy. The analysis showed that lungs with a higher percent pneumonic score exhibit expression of more M. haemolytica A1 genes. For group 1 genes, lktC was expressed in the majority of samples, whereas the other genes were only expressed in some samples. This was not unexpected as the leukotoxin is a major virulence factor of the bacterium. The genes encoding the response regulators for the putative two-component regulatory systems were found to be expressed in more samples than the genes encoding the sensor proteins. The regulator proteins may be required in higher levels to regulate expression of target genes.