The protozoan parasite Theileria annulata bestowing the tick-borne disease bovine tropical theileriosis (BTT), intricate enormous economic losses about US$ 384.3 million annually in Indian livestock sector (Minjauw and McLeod, 2003). It is transmitted through Hyalomma anatolicum ticks which survive better and multiply faster in hot and humid climatic conditions like Odisha.

Oxidative stress is a very good indicator of health status in animals. There is excessive production of reactive oxygen species (ROS) when the body undergoes certain state of disease condition which is countered by natural antioxidant mechanism exist in the body (Samal et al., 2017). This includes antioxidant enzymes like SOD, Catalase, GSH-PX and antioxidant vitamins like vitamin-A, E and C. (Sugino, 2006). Lipid peroxidation is a general mechanism whereby ROS induce tissue damage and are implicated under several diverse pathological conditions (Samal et al., 2016; Halliwela and Gutteridge, 1999). The increased level of oxidative stress and lipid peroxidation in erythrocytes of cattle infected with T. Annulata causes increased erythrocyte fragility and membrane lysis which leads to severe anaemia and death (Grewal et al., 2005).

Keeping in view the above facts, the present study was undertaken to study the changes in erythrocytic oxidative stress indices and haemato-biochemical parameters in the BTT affected cattle with respect to healthy cattle.

Materials and Methods

Present study was carried out in the cattle affected with BTT of Bhubaneswar, Khurda, Puri and the presented cases of Teaching Veterinary Clinical Complex, College of Veterinary Science & Animal Husbandry, OUAT. Clinico-pathological investigation for detection of the blood protozoa Theileria annulata was carried out for the affected animals showing typical clinical symptoms. 10ml blood was collected from jugular vein with EDTA from 35 suspected cows and blood smear examination was carried out by standard staining method (Kelly’s method, 1979) to differentiate the positive cases. The study was carried out by taking 10 number of positive animals into Group II and 10 apparently healthy animal showing blood smear negative for protozoa in Group I. Different investigations like detection of blood protozoa, estimation oxidative stress (MDA, SOD, Catalase), haematological parameters (Hb, PCV, Neutrophil, Lymphocyte, TLC, TEC) and biochemical parameters (AST, ALT, Total protein, A-G ratio) were carried out from collected serum samples. Estimation of oxidative stress parameters MDA, SOD, Catalase was carried out by manual method using double beam UV-VIS spectrophotometer (Samal et al., 2016). The haematobiochemical parameters were estimated by fully automatic analyzer using standard kit method. Data were analyzed by one-way analysis of variance (ANOVA) with post-hoc analysis by Duncan’s multiple comparison tests using SPSS 16 software. Results were expressed as mean ± SE with p<0.05 considered statistically significant.

Results

The haemato-biochemical parameters like Hb, PCV, TEC, TLC, neutrophil, total protein and albumin- globulin ratio (Table 1) are found to be significantly (P≤0.05) lower in Group II animals than Group I animals, whereas lymphocyte count and AST values were significantly higher (P≤0.05) in Group II than Group I. no significant difference was found in the value of ALT in Group I and Group II.

Table 1: Haematobiochemical changes in Group II with respect to Group I

Group I: Healthy control group; Group II: Cattle affected with bovine tropical theileriosis. Values are expressed as Mean ± S.E. values. Mean values with different superscripts (^{a, b, c}) differs in a column between the groups are significant (P≤0.05)

The values of SOD and catalase were found to be significantly lower (P≤0.05) in Group II than Group I whereas the value of MDA was significantly higher (P≤0.05) in Group II than Group I (Table 2).

Table 2: Oxidative stress parameters of animals in different groups

Group I: Healthy control group; Group II: Cattle affected with bovine tropical theileriosis. Values are expressed as Mean ± S.E. values. Mean values with different superscripts (^{a, b, c}) differs in a column between the groups are significant (P≤0.05)

Discussion

Theileriosis is one of the most important blood protozoan disease in cattle as there is every chance of death along with reduced quantity and quality of milk yield for a prolonged period, reduced reproductive performances leading to heavy economic losses to dairy farmers, immuno-suppression leading to high susceptibility to various diseases (Velusamy et al., 2014). In this study, screening by blood smear examination revealed all forms of Theileria annulata piroplasms (cocci, rod, comma, signet-ring, and pear-shaped) with abnormalities in erythrocyte structure which was similar to the findings of Al-Emarah et al. (2012). The haematobiochemical and oxidative stress parameters of all animals in Group I and Group II are thoroughly investigated.

The haemoglobin (Hb) concentration in theileriosis affected group was lower (p<0.05) than the healthy control group (Ganguly et al., 2015; Al-Emarah et al., 2012). The mean packed cell volume and total erythrocyte count in Group II were at a significant (p<0.05) lower level than Group I which is in agreement with Kumar et al. (2009). In contrast to present study, Vahora et al. (2012) recorded normal values of PCV in along with decrease in RBC count and Hb levels. The reduced PCV and TEC as reported in present study may be due to the extensive hemorrhages, abdominal ulcers and persistence of parasitic stages in erythrocytes leading to lower values of PCV, TEC and Hb (Madder et al., 2001) or due to the removal of infected erythrocytes by spleen and liver and not due to the destruction of erythrocytes by the parasite (Modi et al., 2015) or may be due to production of pro-inflammatory cytokines, especially tumor necrosis factor-α (TNF- α), which is implicated in the pathogenesis of anemia in bovine theileriosis by suppressing haemopiotic progenitors (Boulter and Hall, 2000; Kumar et al., 2009) stated lysis of RBC may be due to the lipid peroxidation of membrane leading to low TEC count. Lower level of TLC was found in Group II, which was in accordance with the findings of Hasanpour et al. (2013) and Hussein et al. (2007) and in contrast Modi et al. (2015) and Ugalmugle et al. (2010) found significant (P<0.01) increase in the White blood cell (WBC) counts in infected crossbred cattle as compared to healthy control group whereas Tehrani et al. (2013) demonstrated that the leucocyte count increased immediately following theileria infection and then significantly decreased within several days. Such changes in leucogram might be attributed to persistent harmful effects of toxic metabolites of Theileria annulata on the haemopiotic organs especially on bone marrow and their interference with the process of leucogenesis. Neutrophils level was also lowered (P<0.05) in group-II and group-I which is supported by the findings of Al-Emarah et al. (2012) and Ananda et al. (2009) and contrasted by Ugalmugle et al. (2010) as they found neutrophilia with mild left shift in the theileriosis affected cattle. The mean lymphocyte count in Group II was at a significantly higher level (p<0.05) than healthy controll group, which is in agreement with the findings of Ganguly et al. (2015) and Muraleedharan et al. (2005) whereas several other researchers (Osman and Al-Gaabary 2007, El-Deeb and Younis 2009) have reported a pronounced lymphocytopenia in theileriosis affected cattle. Increase in the number of lymphocytes reflects compensatory mechanism as target cells in response to their invasion with Theileria protozoan. According to Yamaguchi et al. (2010), the lymphocytosis may be due to intra-lymphocytic theilerial parasites transforming the host cells, leading to clonal growth of lymphocytes.

The mean AST values in theileriosis affected cattle had significantly (p<0.05) higher level which is in agreement with the findings of Hasanpour et al. (2013) and Saber et al. (2008) and Ganguly et al. (2015). Increase in serum AST level in theileriosis affected animals may indicate the hepatic tissue damage that included coagulation necrosis, distortion of hepatic cords with heavy infiltration of lymphocytes in the periportal areas indicating severe damage to hepatobiliary system, (Modi et al., 2015). Lower level of serum total protein and albumin level is in agreement with Ganguly et al. (2015), Saber et al. (2008), Hussein et al., 2004, Omer et al., 2003 and Singh et al., 2001. Decreased albumin-globulin ratio are in accordance to Hussein et al. (2007) and Singh et al. (2001). The hypoproteinaemia and hypoalboumineamia is possibly due to the harmful effect of toxic metabolites of Theileria.

In this present study, the oxidative stress was measured as lipid peroxidase (LPO) in form of malondialdehyde (MDA) n mol/mg of Hb. It was revealed that the cattle suffering from BTT in Group II were having significantly higher level (p<0.05) of mean MDA value as compared to healthy control Group I. This proved that there was oxidative stress at a significant higher level in tropical theileriosis affected cattle. This finding is in agreement with Kumar et al. (2009), Radwan et al. (2013) and El-deeb et al. (2009). Theileriosis affected cattle undergoes oxidative stress and there is reduction in antioxidant enzymes due to their neutralization by the excessive production of free radicals which was revealed lower level of mean erythrocytic catalase values in Group II with respect to healthy cattle in Group I. There was significant (p<0.05) decrease in mean erythrocytic superoxide dismutase concentration in Group II indicating another clue on reduction in antioxidant enzyme concentration, which reflects that theileriosis causes severe oxidative stress, which is in agreement with Rezaei and Dalir-Naghadeh (2006) and Razavi et al. (2011).

Conclusion

Based on the above finding, it was concluded that Bovine Tropical Theileriosis (BTT) produces significant oxidative stress along with amemia and alterations in other haematobiochemical parameters in affected cattle as compared to healthy cattle.

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