Submicroscopic levels of Anaplasma marginale rickettsemia in persistently infected cattle were determined by using nucleic acid hybridization. Within individuals, the rickettsemia levels steadily increased from less than 104 infected erythrocytes per ml to a peak of more than 106 infected erythrocytes per ml and then rapidly declined. This logarithmic variation parallels the variation of the rickettsemia level seen in acute infection and suggests that cyclic emergence of antigenic variants is a mechanism of rickettsial persistence.

Cattle which have recovered from acute infection with Anaplasma marginale, a rickettsial hemoparasite of cattle, frequently remain persistently infected with a low-level parasitemia and serve as reservoirs for disease transmission. To fully understand the role of these carriers in disease prevalence and transmission, it is essential that low levels of parasitemia can be accurately detected and quantitated. We have developed a nucleic acid probe, derived from a portion of a gene encoding a 105,000-molecular-weight surface protein, that can detect A. marginale-infected erythrocytes. The probe is specific for A. marginale and can detect 0.01 ng of genomic DNA and 500 to 1,000 infected erythrocytes in 0.5 ml of blood, which is equivalent to a parasitemia of 0.000025%. This makes the probe at least 4,000 times more sensitive than light microscopy. Hybridization of the probe with treated blood from animals proven to be carriers of anaplasmosis showed that parasitemia levels were highly variable among carriers, ranging from >0.0025 to <0.000025%. Parasitemia levels of individual animals on different dates were also variable. These results imply that, at any given time, individuals within a group of cattle may differ significantly in their abilities to transmit disease.

Anaplasma mapignak, an intraerythrocytic rickettsia of cattle, is transmitted biologically by ticks. Because ofthe brevity of acute A. marginake infection, transmission may rely on the tick's ability to acquire the organism from persistently infected cattle with low rickettsemia levels. By using a nucleic acid probe to quantitate low-level infection, we found that rickettsemia levels in persistently infected cattle fluctuated at approximately 5-week intervals during a 24-week period, from <104 infected erythrocytes per ml of blood to high levels of approximately 107 infected erythrocytes per ml of blood. Cattle maintained very low rickettsemia levels (<10-3 infected erythrocytes per ml of blood) for approximately 4 to 8 days of every 5-week cycle. The effect of fluctuations in rickettsemia in persistently infected cattle on acquisition by Dermacentor andersoni nymphal and adult male ticks was examined. A positive correlation was observed between rickettsemia levels in cattle and the resulting infection rates of ticks. At high rickettsemia levels, up to 80%o of ticks acquired infection, but even at extremely low rickettsemia levels, 27% of adult male ticks became infected. Moreover, once ticks acquired infection, biological replication of the organism within the ticks appeared to make up for initial differences in the infecting dose. The high infection rates in adult males, combined with their intermittent feeding behavior and the observation that only a few infected ticks were required for transmission to a susceptible host, suggest that adult male D. andersoni ticks are epidemiologically important in A. mai nale transmission. Because cattle with all levels of rickettsemia were capable of efficient transmission to ticks, population control efforts must include decreasing transmission from persistently infected individuals.

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Abstract. Anaplasma marginale, a rickettsial hemoparasite of cattle and other ruminants, results in significant economic losses worldwide. Distinct strains of A. marginale have been identified based on differences in tick transmissibility, molecular size of surface proteins and DNA restriction fragments, and reactivity to a panel of monoclonal antibodies. These different strains vary considerably in their virulence, antigenic composition, and ability to protect against heterologous challenge. In this paper, we report on the molecular characterization of a newly isolated strain of A. marginale, designated St. Maries, recovered from an acutely infected cow in northern Idaho. Dermacentor andersoni ticks taken from the infected animal were tested for infection by RNA probe analysis. The infection rate of male ticks (as determined by midgut infection) was 100%, and the infection rate of female ticks was 83%. Infected male ticks were able to transmit the St. Maries strain to a susceptible calf. The high infection rate in male ticks may be particularly relevant, given that male ticks are believed to be epidemiologically important in transmission of A. marginale because of their intermittent feeding behavior, which promotes interhost transfer. The newly isolated strain differs from other US strains, including strains previously isolated in Idaho and Washington, based on reactivity to a panel of monoclonal antibodies and restriction fragment length polymorphisms. These results imply that antigenically distinct strains of A. marginale may arise within the same region. However, because epitopes previously demonstrated to be conserved in other A. marginale strains were also present in the St. Maries strain, it is possible to develop diagnostic tests against conserved epitopes that could be used for early identification of infected cows or ticks.