Comparative analysis of hematological changes in bovine species infected with Haemoprotozoans in Bikaner region of Rajasthan, India

Introduction

Haemoprotozoan diseases, the majority of which are transmitted by tick vectors, represent a major challenge in the dairy sector. Among these bovine babesiosis, tropical theileriosis, and bovine anaplasmosis are of particular importance in India. Overall the clinicopathology associated with these infections lead not only to high mortality rates but also affect physiological functions, feed consumption and productive and reproductive performance. Genetically improved cattle breeds are particularly vulnerable to these infections and often have lower survival rates (Radostitis et al., 1994) [26] , resulting in increased costs associated with control measures (Makala et al., 2003) [16] . Although advanced diagnostic techniques are now available, conventional diagnostic methods still primarily rely on the microscopic identification of infective parasite stages in blood or tissue fluids (Terkawi et al., 2011) [31] . The widespread epidemiology of these diseases is of particular importance in Indian subcontinent mainly due to high prevalence of vector population owning to subtropical climatic conditions. (Vahora et al., 2012; Velusamy et al., 2014; Bhatnagar et al., 2015; Maharana et al., 2016) [32,33,5,15] . In their mammalian hosts all the above mentioned haemoparasites have tropism towards Red blood cells so they proliferate inside them and due to their repeated invasion cause their phagocytosis or extravascular lysis by immune system of host that lead to haemolytic anaemia and decrease in blood indices, further complex pathogenic mechanisms also involve various erythropoietic and lymphoid organs. These parasites often induce clinical signs such as pyrexia, lymphadenopathy, nasal and ocular discharges, anorexia, weakness, haemoglobinuria and severity of disease vary with host susceptibility and stage of infection (El-Deeb and Younis, 2009) [9] . With the exception of trypanosomiasis, which typically causes leukopenia, most haemoprotozoan infections are characterized by decreased hemoglobin (Hb), packed cell volume (PCV), total erythrocyte count (TEC), and elevated total leukocyte count (TLC) as a consequence of the aforementioned pathological processes (Maxie et al., 1976; Aziz et al., 2020) [17,3] . In past studies low haemoglobin, packed cell volume has been reported from naturally infected bovines (Nazifi et al., 2012) [20] . The magnitude of haematological changes vary with species of parasite, its tissue tropism and host factors such as systemic immune response, nutritional status, age, breed. Also concurrent infection with multiple parasites can further complicate disease pathology and clinical signs. Haematogram analysis of both infected and non infected animals help in comparative analysis of physiological impacts of haemoparasites in host's body. Hematological and coagulation parameters serve as valuable tools for diagnosis, prognosis, and therapeutic monitoring, providing critical information regarding the severity and progression of infections. Consequently, the present study focused on evaluating changes in these parameters to better understand the pathogenesis of haemoprotozoan infections in dairy animals. Earlier studies conducted on bovine haemoprotozaons in arid region of Rajasthan have reported the prevalence rate, various epidemiological factor associated with it were mainly focused on a particular haemoprotozoan species. However the limited data is available on the comparative analysis of haemtogram and leukogram of infected and non infected animals. We have documented data on prevalence of individual protozoans provided information about the prevalence rate of different haemoprotozoan in Bovines however haematological changes associated with the infections were not discussed in detail (Bharti et al., 2022) [4] . Thus present study was undertaken to evaluate the erythrocytic and leucocytic profiles of naturally infected animals ( Kachhawa et al., 2016; Goyal et al., 2018; Bharti et al., 2022) [14,11,4] .

Materials and Methods Study Area and Sample Collection

The study was conducted in Bikaner, located in arid western plain zone of Rajasthan. A total of 200 samples were collected from bovines (cattle and buffaloes) at selected Gaushalas, polyclinics, and the TVCC of the College of Veterinary and Animal Science (CVAS), Rajasthan University of Veterinary and Animal Sciences (RAJUVAS), Bikaner. Both infected as well as apparently healthy animals were selected based on their availability during study period from field and as well as clinically presented animals. Aseptical blood collection was done using disposable syringes and immediately transferred into EDTA-containing vials and gently mixed to prevent clot formation. The samples were properly labelled and transported to the laboratory for further examination. If required, fresh blood smears were prepared from peripheral circulation for microscopic examination.

Haematological Analysis

Haemoglobin (Hb) was estimated by Sahli's acid haematin method (Cohen and Smith, 1919) [7] and expressed as g/dL.

Packed cell volume (PCV) was determined by Wintrobe's method (Wintrobe, 1929) [34] and expressed as percentage. Total leucocyte count (TLC) was estimated using a haemocytometer as described by Coles (1974) and expressed as cells/mm³. Differential leucocyte count (DLC) was determined by counting 100 leucocytes in stained blood smears following Parida et al. (2012) [21] and expressed as percentage.

Statistical Analysis

Data were classified according to month, area, age, sex, and breed. Statistical analysis was performed using IBM SPSS Statistics version 20.0 following Snedecor and Cochran (1994) [30] .

Results

Alterations in Haemoglobin (Hb)

In animals infected with haemoprotozoa, lower average haemoglobin values were observed, with the lowest (7 g/dl) recorded in Babesia spp. infection. Healthy animals had an average Hb of 10.42 g/dl, while infected animals had 8.39 g/dl, although the difference was statistically non-significant (p > 0.05).

Alterations in Packed Cell Volume (PCV)

PCV values were lower in infected animals (26.6%) compared to healthy animals (31.2%), with the lowest PCV (23.6%) recorded in babesiosis cases. The difference was statistically non-significant (p > 0.05).

Alterations in Total Leucocyte Count (TLC):

A highly significant increase (p<0.01) in TLC was observed in infected animals compared to healthy ones. Although values remained within the normal range, the highest leucocytosis was found in animals infected with Anaplasma spp. (Table 1, Figure 1).

Alterations in Differential Leucocyte Count (DLC):

In infected animals, a highly significant decrease (p<0.01) in neutrophil count was observed. A significant increase in lymphocyte count and a significant decrease in monocyte count (p<0.05) were recorded, while eosinophil count showed a non-significant (p > 0.05) increase. Notably, neutrophil and monocyte counts were higher in co-infected animals, whereas the highest lymphocyte and eosinophil counts were observed in Theileria spp. and Anaplasma spp. infections, respectively (Table 2, Figure 2).

Discussion

Haemtological analysis revealed variation in blood parameters of infected animals as compared to healthy. The value of haemoglobin and packed cell volume were declined in parasitic animals as compared to non parasitimic animals, though the observation was non significant. This could be due to multiplication of blood protozoans in red blood cells which were phagocytised by reticuloendothelial cells and cause haemolytic anaemia (Jayalakshmi et al., 2019) [13] . A highly significant increase was observed in total leucocyte count of infected animals as compared to healthy ones. In diferential leucocyte count a significant decrease was seen in values of Neutophill (p ≤ 0.01) and monocytes (p ≤ 0.05) while the values of lymphocytes (p ≤ 0.05) were increased. These results may be due to immune response of body of animlas to ongoing parasitic infection. Across the world

Conclusion

The present study highlighted the impact of haemoprotozoan infection on the physiology of animals.

The parameters included for the study provided the valuable information on how the presence of infection can cause changes in animal body and how they can act as supportive tools for assessing health status of animals under field conditions. This regular screening and early detection can help timely management and reduction of adverse impact on bovine health and productivity.

References

1. from Karnataka, Aulakh and Singla (2006) [2] from Punjab, Ananda et al. (2009) [1] from Karanataka, Shahnawaz et al. (2011) [27] from Pakistan, Chaudhri et al. (2013) [6] from Eastern Haryana, Sitotaw et al. (2014) [29] from Central Eithopia, Paul et al. (2016) [25] from Nigeria, Ganguly et al. (2017) [10] from Haryana, Patel et al. (2017) [23] from Gujarat, Jaiswal et al. (2019) [12] from Rajasthan, Jayalakshmi et al. (2019) [13] from Tamil Nadu, Parmar and Chandra (2019) [22] from Uttar Pradesh, Patil and Satbige (2019) [24] from Karnatka, Aziz et al. (2020) [3] from
2. Rajasthan, Sharma et al. (2020) [28] from Rajasthan and Muniraja et al. (2021) [18] from Andhra Pradesh. References
3. Ananda KJ, D'Souza PE, Puttalakshmamma GC. Prevalence of haemoprotozoan diseases in crossbred cattle in Bangalore north. Vet World. 2009;2(1):15.
4. Aulakh GS, Singla LD. Clinico-haematobiochemical observations on bovines naturally infected with Theileria annulata. J Vet Parasitol. 2006;20(1):49-52.
5. Aziz PR, Marodia S, Ganesan PI, Sharma CS. A clinical study on hemato-biochemical changes in cows affected with babesiosis. Pharma Innov J. 2020;9(2):242-245.
6. Bharti V, Pilania PK, Choudhary P, Joshi SP. Prevalence rate of haemoprotozoan infection and assessment of associated risk factors in dairy animals from Bikaner region of Rajasthan, India. J Anim Res. 2022;12(1):69-74.
7. Bhatnagar CS, Bhardwaj B, Sharma DK, Meena SK. Incidence of haemoprotozoan diseases in cattle in southern Rajasthan, India. Int J Curr Microbiol Appl Sci. 2015;4(3):509-514.
8. Chaudhri SS, Bisla RS, Bhanot V, Singh H. Prevalence of haemoprotozoan infections in pyretic dairy animals of eastern Haryana. Indian J Anim Res. 2013;47(4):344-347.
9. Cohen B, Smith AH. The colorimetric determination of hemoglobin: A practical procedure. J Biol Chem. 1919;39(3):489-496.
10. Coles EH. Veterinary clinical pathology. 4th ed. Philadelphia: W.B. Saunders; 1974.
11. El-Deeb WM, Younis EE. Clinical and biochemical studies on Theileria annulata in Egyptian buffaloes (Bubalus bubalis) with particular emphasis on oxidative stress and ketosis relationship. Cercet Agric Moldova. 2009;42(3):63-73.
12. Ganguly A, Bisla RS, Singh H, Bhanot V, Kumar A, Kumari S, et al. Prevalence and haematobiochemical changes of tick-borne haemoparasitic diseases in crossbred cattle of Haryana, India. Indian J Anim Sci. 2017;87(5):552-557.
13. Goyal P, Chahar A, Tanwar RK, Fakhruddin, Shringi BN. Hospital-prevalence of Theileria annulata infection in cattle-calves determined by blood smear and lymph node aspirate smear examination in Bikaner, Rajasthan. Int J Curr Microbiol Appl Sci. 2018;7(6):877-884. doi:10.20546/ijcmas.2018.706.103.
14. Jaiswal M, Singh VK, Saini K. Concurrent infection of theileriosis and anaplasmosis in crossbred cattle. Pharma Innov J. 2019;8(6):624-626.
15. Jayalakshmi K, Sasikala M, Veeraselvam M, Venkatesan M, Yogeshpriya S, Ramkumar PK, et al. Prevalence of haemoprotozoan diseases in cattle of Cauvery delta region of Tamil Nadu. J Parasit Dis. 2019;43(2):308-312.
16. Kachhawa JP, Kumar S, Sharma A, Singh AP, Ahuja A. Studies on alterations of clinical and hematobiochemical parameters before and after treatment in calves naturally infected with theileriosis. Vet World. 2016;9(12):1381-1385. doi:10.14202/vetworld.2016.1381-1385.
17. Maharana BR, Kumar B, Prasad A, Patbandha TK, Sudhakar NR, Joseph JP, et al. Prevalence and assessment of risk factors for haemoprotozoan infections in cattle and buffaloes of south-west Gujarat, India. Indian J Anim Res. 2016;50(5):733-739.
18. Makala L, Mangani P, Fujisaki K, Nagasawa H. The current status of major tick-borne diseases in Zambia. Vet Res. 2003;34(1):27-45.
19. Maxie MG, Losos GJ, Tabel H. A comparative study of the hematological aspects of the diseases caused by Trypanosoma vivax and Trypanosoma congolense in cattle. In: Pathophysiology of parasitic infection. New York: Academic Press; 1976. p. 183-198.
20. Muniraja K, Subapriya S, Sangaran A, Vairamuthu S. Incidence of blood parasites in bovine in Chittoor District of Andhra Pradesh. Int J Sci Res Rev. 2021;10(4):18-23.
21. Muraleedharan K, Ziauddin KS, Hussain PM, Puttabyattappa R, Seshadri SJ. Haematological International Journal of Advanced Biochemistry Research https://www.biochemjournal.com observations on Theileria annulata infection in cattle and buffaloes. J Vet Parasitol. 2005;19(1):71-72.
22. Nazifi S, Razavi SM, Kaviani F, Rakhshandehroo E. Acute phase response in cattle infected with Anaplasma marginale. Vet Microbiol. 2012 Mar 23;155(2-4):267271. doi:10.1016/j.vetmic.2011.08.024.
23. Parida SP, Dutta SK, Pal A. Hematological and plasma biochemistry in Psammophilus blanfordanus (Sauria: Agamidae). Comp Clin Pathol. 2012;21(6):1387-1394.
24. Parmar D, Chandra D. Studies on comparative haematological parameters in Theileria annulata infected cattle. J Entomol Zool Stud. 2019;7(3):167170.
25. Patel MD, Kumar N, Rathod PH, Tyagi KK. Incidence and haematological changes in haemoprotozoan infections in bovines of south Gujarat. Indian J Vet Sci Biotechnol. 2017;13(2):62-66.
26. Patil NA, Satbige AS. Molecular detection, haematological and therapeutic studies on theileriosis in buffaloes. Buffalo Bull. 2019;38(1):141-146.
27. Paul BT, Bello AM, Ngari O, Mana HP, Gadzama MA, Abba A, et al. Risk factors of haemoparasites and some haematological parameters of slaughtered trade cattle in Maiduguri, Nigeria. J Vet Med Anim Health. 2016;8(8):83-88.
28. Radostitis MO, Gay CC, Hinchcliff KW, Constable PD. Veterinary medicine: a textbook of the diseases of cattle, horses, sheep, pigs and goats. 10th ed. London: Bailliere Tindall; 2007.
29. Shahnawaz S, Ali M, Aslam MA, Fatima R, Chaudhry ZI, Hassan MU, et al. A study on the prevalence of a tick-transmitted pathogen, Theileria annulata, and hematological profile of cattle from southern Punjab (Pakistan). Parasitol Res. 2011;109(4):1155-1160.
30. Sharma DK, Singathia R, Bhatnagar CS, Manne V, Meena S. Acute anaplasmosis in a Holstein-Friesian crossbreed cattle: a case report. Int J Chem Stud. 2020;8(1):1154-1156.
31. Sitotaw T, Regassa F, Zeru F, Kahsay AG. Epidemiological significance of major hemoparasites of ruminants in and around Debre-Zeit, Central Ethiopia. J Parasitol Vector Biol. 2014;6(2):16-22.
32. Snedecor GW, Cochran WG. Statistical methods. 6th ed. Iowa: Iowa State University Press; 1994.
33. Terkawi MA, Thekisoe OMM, Katsande C, Latif AA, Mans BJ, Matthee O, et al. Serological survey of Babesia bovis and Babesia bigemina in cattle in South Africa. Vet Parasitol. 2011;182(2-4):337-342.
34. Vahora SP, Patel JV, Patel BB, Patel SB, Umale RH. Seasonal incidence of haemoprotozoal diseases in crossbred cattle and buffalo in Kaira and Anand districts of Gujarat, India. Vet World. 2012;5(4):223225.
35. Velusamy R, Rani N, Ponnudurai G, Harikrishnan TJ, Anna T, Arunachalam K, et al. Influence of season, age and breed on prevalence of haemoprotozoan diseases in cattle of Tamil Nadu, India. Vet World. 2014;7(8):574578.
36. Wintrobe MM. A simple and accurate hematocrit. J Lab Clin Med. 1929;15:287-289