The response of Mannheimia haemolytica to iron limitation: implications for the acquisition of iron in the bovine lung.

Mannheimia haemolytica is the major causative agent of shipping fever, a severe pneumonia in cattle causing high morbidity and mortality. A prerequisite of successful lung colonization by M. haemolytica is the necessity to adapt to the paucity of iron. The lack of genome information has precluded an assessment of the genetic repertoire available to M. haemolytica to adapt to low iron environments. To close this knowledge-gap, we have determined 90% of a virulent M. haemolytica serotype A1 genome sequence and produced a microarray in order to study gene expression under iron-limiting growth for 15, 30 and 60 min. M. haemolytica responded to iron limitation by the up-regulation of transcripts coding for receptors and ABC-type transporters of transferrin, haemoglobin, haem and siderophores. Real time PCR analysis of lung tissue from Mannheimia-infected calves demonstrated the in vivo transcription of two potential haemoglobin receptors, hmbR1 and hmbR2. The relative hmbR1 and hmbR2 transcript levels in the infected lung tissue were comparable to the induced levels observed under iron-limiting growth, demonstrating in vivo induction of receptor transcription in the context of an infection. When the iron response of M. haemolytica was compared to the iron response of Pasteurella multocida, another pathogen colonizing the bovine lung, only few homologous genes were induced in both organisms. These included the haemoglobin receptor hmbR2 and the periplasmic transport systems yfeABCD and fbpABC. The comparative analysis suggests that the two pathogens use different strategies to adapt to the iron-limiting environment in the bovine host.

Microarray analysis of Haemophilus parasuis gene expression under in vitro growth conditions mimicking the in vivo environment.

Haemophilus parasuis is the causative agent of polyserositis in pigs, a mostly fatal disease on the rise especially in early-weaned pigs and in pig herds with a high-health status. The mechanisms by which H. parasuis propagates through the body and colonizes the serous membranes are unknown. We have used an H. parasuis microarray to identify virulence genes involved in host adaptation. H. parasuis gene expression was analysed under in vitro growth conditions mimicking the environmental conditions encountered during an infection. These included iron-limitation, acidic and temperature stress and growth under microaerobic conditions. A kinetic impression of the gene regulation was obtained by analysing the transcription 10, 30 and 60 min after induction of the altered growth conditions. A total of 75 regulated H. parasuis genes were identified, most of which coded for transporters of iron and sugar metabolites, metabolic enzymes, DNA metabolism and hypothetical proteins with unknown functions. Furthermore, H. parasuis genes were identified that have homology to known virulence factors in other pathogenic bacteria. Homologues of some of the identified H. parasuis genes are known to be expressed during natural and experimental infections in pathogens of the Pasteurellaceae family.