Diagnosis and treatment of A. marginale infection
The blood samples were found positive for
Anaplasma spp. infection by using microscopic examination of blood smears (Fig 1A). It was again confirmed by PCR, as evident by the presence of expected 576 bp product in agarose gel under UV transilluminator (Fig 1B). Partial gene sequencing of
rpoB gene revealed that the samples are identical (99.2-100%) to other sequences of
A. marginale available at GenBank (GenBank accession number: MT755603 and MT755604). The blood samples were found negative for commonly occurring haemoprotozoan diseases such as babesiosis, theileriosis and trypanosomosis. All the animals were treated with intravenous oxytetracycline at the dose rate of 10 mg/kg, daily for 3 days along with supportive therapy comprising anti-pyretic, multivitamins and haematinics. Intravenous oxytetracycline can be used as an effective drug against bovine anaplasmosis
(Radostits et al., 2007). Unfortunately, two animals did not respond to treatment and succumbed to infection after two days of initiation of therapy. The primary effect of
A. marginale infection is the induction of extravascular hemolytic anaemia and the resultant anaemic anoxia is presumed to be the cause of death
(Abba et al., 2016). The remaining four animals showed uneventful recovery.
Gross lesions
A varying degree of changes were noticed in different organs of the body. Gross pathological observation revealed emaciated carcass with yellowish discoloration of serosal surfaces of the abdominal organs (Fig 2A). Icteric condition observed in the present study could be due to excessive destruction of red blood cells and similar findings were also reported by previous reports
(Jaswal et al., 2015; Abba et al., 2016). Paintbrush haemorrhages and severely congested blood vessels on epicardial surface (Fig 2B) and presence of endocardial haemorrhages (Fig 2C) were the major gross findings noticed in heart. These findings are in consistent with the reports published earlier
(Coetzee et al., 2005). The post mortem examination also revealed presence of severely congested lungs (Fig 2D), hepatomegaly with blunt edges and necrosis (Fig 2E), splenomegaly with discoloration (Fig 2F) and distended gall bladder with dark colored bile (Fig 2G). Pulmonary congestion, distension of gallbladder, enlargement of liver and spleen has been documented in both natural and experimental bovine anaplasmosis
(Jaswal et al., 2015; Abba et al., 2016; Lima et al., 2019). Splenomegaly with congestion and discoloration reported in the present study may be due to erythrophagocytosis that results in excessive destruction of defective erythrocytes by macrophages within the spleen
(Devos et al., 2006). No gross lesions were observed in kidney (Fig 2H). A dead foetus was found in the uterus of one animal (Fig 2I), whereas the other animal had a recent history of abortion. Deprivation of oxygen due to excess haemolysis may cause foetal death and abortion in pregnant cows
(Whittier et al., 2009).
Histopathological observations
Histological examination of heart revealed presence of haemorrhage in the myocardium along with degeneration and necrosis (Fig 3A). These findings may be secondary to severe anaemia induced by
A. marginale infection and similar findings were also documented in previous reports
(Jaswal et al., 2015). Examination of lung tissues showed interstitial pneumonic and oedematous changes, thickening of interalveolar septum and infiltration of mononuclear cells (Fig 3B). Degeneration and necrosis of hepatocytes, congestion and haemorrhage, mild infiltration with mononuclear cells and enlargement of sinusoids were the major findings noticed in liver tissues (Fig 3C). Hepatic necrosis was more prominent around the central vein. Hepatic necrosis is also an important feature of bovine anaplasmosis and is generally more severe than in babesiosis
(Devos et al., 2006). Histological examination of spleen revealed enlargement of red pulp area, histiocytic proliferation with hemosiderosis, serofibrinous exudation in the red pulp area and lymphoid proliferation in white pulp area (Fig 4A). Accumulation of hemosiderin pigment in the cells of reticuloendothelial system has already been reported in bovine anaplsmosis
(DeVos et al., 2006). Centrilobular hypoxic hepatic necrosis and splenic hemosiderosis may be due to enhanced destruction of red blood cells (
Anderson and Hurtado, 1989). Splenic changes are similar to those occurring in babesiosis except that there is generally more extensive accumulation of plasma cells in the red pulp in case of anaplasmosis
(DeVos et al., 2006). Tissue section of kidneys obtained from
A. marginale infected cattle showed degeneration and necrosis of tubular epithelial cells and infiltration of mononuclear cells in the interstitial space of renal parenchyma (Fig 4B). Congestion and haemorrhages were also observed. In overall, the histopathological changes observed in the present study are in agreement with previous reports
(Jaswal et al., 2015; Abba et al., 2016; Lima et al., 2019). It is well known that anoxia causes degenerative changes and necrosis in parenchymatous organs. During
A. marginale infection, the oxygen content of the blood is lowered due to less haemoglobin content which could lead to degenerative changes in liver, spleen and kidney.
