Analysis of "Anaplasma marginale" strains grown "in vitro"

<i>Anaplasma marginale</i> is a tick-borne pathogen that affects ruminants worldwide, causing a disease called anaplasmosis. The disease is endemic in tropical and subtropical regions of the New World, Europe, Africa, Asia and Australia where it causes large economic losses in the cattle industry. <br> <i>A. marginale</i> is an obligatory intracellular bacterium that multiplies only within tick cells or ruminants' erythrocytes. Many differences among <i>A. marginale</i> strains have emerged, which were probably driven by continuous exposure to different host immune systems during the transition of bacteria between ticks and vertebrates. The vast majority of studies aiming at elucidating differences between strains were conducted on the genomic level, and little is known about protein expression. Thus, this thesis investigates differences in protein regulation among <i>A. marginale</i> strains. <br> <i>A. marginale</i> cultivated in vitro are in general an excellent source of organisms for experimentation. Furthermore, culture-derived organisms offer an alternative to the use of experimental animals. <br> Many studies require intracellular organisms free from host cell debris. Therefore the use of Percoll gradients for the separation of <i>A. marginale</i> was evaluated. Bacteria isolated in this way contained only minimal amounts of IDE8 cell stroma but most importantly they retained their viability. <i>A. marginale</i> purified this way can be used directly for proteomic studies or for vaccination trials. <br> In this thesis three geographical <i>A. marginale</i> strains grown <i>in vitro</i> have been partially characterized by gene and serological analyses. The differences on the proteomic level have been assessed by the 2D-DIGE technique, indicating that many antigenic membrane proteins are differentially regulated among the strains examined. Some of these proteins are also known to be virulence-associated. <br> Increasing the number of strains in continuous <i>in vitro</i> cultivation, and improving purification methods for rickettsia, allow researchers to investigate differences in protein expression between <i>A. marginale</i> strains, and therefore identify proteins which could be incorporated into an improved vaccine against anaplasmosis.

Thèse de doctorat : Université de Neuchâtel, 2015