Global gene expression profiling of Ehrlichia ruminantium virulent and attenuated strains

The obligate intracellular bacterium, Ehrlichia ruminantium (ER) is the causal agent of Heartwater, a fatal disease in ruminants. It is transmitted by ticks of the genus Amblyomma. Here, we report the genomic comparative and the global transcriptional profile of 4 strains of ER, Gardel and Senegal, two distant virulent strains with their corresponding attenuated strains. Our results showed a higher metabolic activity in attenuated strains compared to virulent strains, suggesting a better adaptation in vitro of attenuated strains to the host cells. There was a strong modification of membrane protein encoding genes expression for the 4 strains. A major over-expression of map1-related genes was observed for virulent strains, whereas attenuated strains over-expressed genes encoding for hypothetical membrane proteins. This result suggests that in vivo, MAP-1 related proteins could induce non-protective immune responses for virulent strains. For the attenuated strains, the lack of expression of map1-related genes and over-expression of other membrane proteins encoding genes could be important in induction of efficient immune responses.The diminution of expression of many genes in attenuated Senegal was caused by severe mutation. One of them, the gene recO is involved in DNA repair and its mutation could explain the higher proportion of mutated genes in attenuated Senegal, inducing the faster attenuation of Senegal compared to Gardel. Three biological replicates for each strain were used for the microarrays experiments and correspond to samples Gp38, Gp43, Gp44 (virulent gardel), Gp238, Gp243, Gp251 (attenuated Gardel), Sp7b, Sp8a, Sp8b (virulent Senegal) and Sp70, Sp75, Sp77 (attenuated Senegal). When 80% cell lysis was observed, samples containing supernatant and cellular debris were harvested and then ultra-centrifuged at 20 000 × g for 15 min at 4°C to collect elementary bodies (EB). Total RNA was extracted from the pellets using TRIzol reagent (Invitrogen). One color experiment with 4 experimental conditions (n=3) : virulent Gardel, attenuated Gardel, virulent Senegal, attenuated Senegal for a total of 12 samples.