Identification of IgG2-specific antigens in Mexican Anaplasma marginale strains.

To identify novel antigens with immunoglobulin G2 (IgG2) specificity and immunostimulant properties for bovine Th1 cells, humoral and cellular responses were studied in cattle inoculated with initial bodies from a Mexican isolate of Anaplasma marginale and challenged with a heterologous strain. Analysis of post-immunization sera by ELISA and assaying of in vitro cellular responses in peripheral blood mononuclear cells (PBMCs) cultured in the presence of protein extracts from three Anaplasma marginale strains showed positive values of optical density ELISA readings and stimulation indices in the immunized but not control cattle. Post-immunization and post-challenge sera recognized in Western blots several proteins with molecular weights ranging from 15 to 209 kDa, twelve of which were recognized by IgG2 in the three Anaplasma marginale strains. Seven of these are novel and have not been previously reported for their IgG2 specificity; three are confirmed to be major surface proteins (MSP-1a, MSP-2, and MSP-5); and the others correspond to other well-studied MSPs but were not confirmed. Partially purified fractions of protein extracts of the Mex-17 strain were tested against PBMCs cultured in vitro. One out of the seven novel proteins induced detectable lymphoproliferation (LP) of PBMCs, and interferon-gamma was detected in supernatants of PBMC cultured in the presence of two protein fractions, including the one that caused LP. It is concluded that novel antigens, particularly the 28-kDa protein, played an additional role in the protection of immunized cattle and should be considered vaccine candidates after in vivo immunization experiments are concluded.

Anaplasma marginale inactivated vaccine: dose titration against a homologous challenge.

The present study was performed to dose-titrate an Anaplasma marginale experimental immunogen derived from partially purified initial bodies from three geographically different Mexican strains. Three five-bovine groups were inoculated twice on days zero and 21 with A. marginale initial bodies equivalent to 1.5 x 10(10) (group I), 3 x 10(10) (group II) or 6 x 10(10) (group III) infected erythrocytes mixed with STDCM adjuvant. A similar group served as non-vaccinated controls. All four groups were challenged with 1 x 10(8) infected erythrocytes from a donor cow with an increasing rickettsemia of strain MEX-15 on day 87 post-vaccination. The prepatent period was very similar for all four groups. All five non-vaccinated controls presented typical acute anaplasmosis syndrome reaching a mean of 30.9% rickettsemia and a loss of 73.4% in the packed cell volume (PCV). Two of five controls died of acute anaplasmosis. Within the vaccinated groups only one animal (group II) suffered acute disease and died. Although all the other vaccinated animals were free of clinical signs, they developed very low rickettsemias (3.2, 3.8 and 4.3%) and PCV losses of 49.9, 47.8, and 49.3% for groups I, II and III. The starting mean weight was very similar for all four groups. All animals lost weight following challenge but losses for groups I and II were lower and significantly different from group IV losses (P < or = 0.1). Although there were no significant differences among vaccinated groups, group III was more severely affected. Taken altogether, these results show a 93.3% protection against both illness and death for all groups; and 100% protection for groups I and III, and 80% for group II.

Bovine anaplasmosis prevalence in northern Veracruz state, Mexico.

In order to learn more about the presence of bovine anaplasmosis in northern Veracruz state, México, paired blood and serum samples from 368 cattle were subjected to polymerase chain reaction (PCR) and complement-fixation test (CFt). The overall prevalence of Anaplasma marginale by PCR was 69.2% and seroprevalence by CFt 54.6%. Age-specific prevalence was calculated for each test. Sixty-eight percent of animals from 0 to 3 months of age already were infected (PCR-positive), compared to only 42.4% positive by serology. CFt results suggested that presence of antibody increases with age up to 18 to 36 months, decreasing thereafter. Presence of the rickettsia seems to follow the same early pattern but with a new increase in animals 36 months or older. Serology results provided a biased picture of the true prevalence of anaplasmosis. Calculated specificity and sensitivity (63.5% and 68.2%) for CFt using PCR values as true values, appear very low and unreliable. The data generated by DNA-based surveys seem more appropriate to help design and implement control or eradication programs for bovine anaplasmosis.