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PubMed1985

High-yield preparation of purified Anaplasma marginale from infected bovine red blood cells.

Purification of Anaplasma marginale from infected bovine RBC was achieved through enzyme treatment and density-gradient centrifugation. A relative yield of 41.6% was obtained by dividing the number of organisms in the final purified preparation by the number of A marginale-infected RBC. Purified parasites were verified as A marginale by light microscopy, electron microscopy, and immunologic tests. The purified parasites reacted positively with calf and rabbit anti-A marginale sera in interfacial and slide agglutination tests. Anti-bovine RBC serum did not agglutinate purified A marginale, indicating absence of any contaminating RBC stroma. Anaplasma marginale was antigenic, but did not cause infection when the preparation was inoculated into a susceptible calf. The density of A marginale was determined to be 1.19 g/ml and cell diameters ranged from 0.25 to 0.63 micron. This method provided procedures for obtaining A marginale free of bovine RBC antigens for accurate biochemical assays and vaccine production.

Agglutination Tests↗

PubMed1985

Modified indirect fluorescent antibody test for the serodiagnosis of Anaplasma marginale infections in cattle.

A modified indirect fluorescence antibody technique was used for the serodiagnosis of Anaplasma marginale infections in cattle. Nonspecific antibodies adherent to infected erythrocytes were removed, using acidic glycine buffer. Evans blue was used as a counterstain.

Anaplasmosis↗

PubMed1986

Isolate of Anaplasma marginale not transmitted by ticks.

The tick-borne transmissibility of 2 isolates of Anaplasma marginale was compared. Dermacentor variabilis were exposed to A marginale as nymphs by feeding on 1 of 4 splenectomized calves during periods of ascending parasitemia (maximum 49% to 81% parasitized erythrocytes) induced by injection of a stabilate. Tick-borne transmission was attempted, using 26 to 224 adult ticks within 30 to 220 days after molting. Adult D variabilis did not transmit an Illinois isolate of A marginale in 7 tick-borne transmission experiments (P = 0.0047), including 2 experiments in which calves were inoculated IV with homogenates of adult ticks. In contrast, a Virginia isolate of A marginale was readily transmitted by the same tick colony. Thus, previously reported morphologic and immunologic differences among A marginale isolates may extend to tick-borne transmissibility. The Virginia and Illinois A marginale isolates had an inclusion appendage that was not a marker for tick transmissibility.

Anaplasmosis↗

PubMed1986

Identification of antigens of two isolates of Anaplasma marginale, using a western blot technique.

Antigens of the Illinois (IAM) and Florida (FAM) isolates of Anaplasma marginale were analyzed, using the western blot technique and antiserum from A marginale-infected calves. Crude antigens were prepared from the parasitemic blood of each. Antiserum was collected after the primary and recrudescent parasitemias. Antigens were separated, using sodium dodecyl sulfate polyacrylamide gel electrophoresis. Antigens were then transferred onto nitrocellulose membranes and exposed to test sera. Antibodies attached to the membrane-bound antigens were detected, using an avidin/biotin peroxidase assay and biotinylated rabbit anti-goat immunoglobulin G. Antigens detected were of a high molecular weight group (108 to 91 kilodaltons [kd]) or of a low molecular weight group (47 to 27 kd). The IAM antigens were 100 kd, 96 kd, 47 kd, 38 to 43 kd, and 27 kd; these antigens were detected, using anti-IAM and anti-FAM antibodies, but the anti-FAM antibodies had a strong reaction to only the 100-kd and 38- to 43-kd antigens of IAM. The FAM antigens were 108 kd, 91 kd, 47 kd, 38 to 43 kd, and 27 kd; these antigens were detected, using anti-FAM antibodies and, except the 91 kd antigen, anti-IAM antibodies. Because the 91-kd antigen was detected only in the FAM antigen and detected only by sera from FAM-infected calves, this isolate-specific antigen may be associated with the ability of FAM to induce disease in an IAM-immune animal. Sheep anti-A ovis antibodies reacted only to the 38- to 43-kd antigens of each isolate, indicating that these antigens may be genus-specific.(ABSTRACT TRUNCATED AT 250 WORDS)

Anaplasma↗

PubMed1970