Flow cytometry based profiling of leukocytes: A new method for diagnosis of tropical theileriosis in crossbred cattle

Abstract

Aim:In India, dairy industries are important for the livelihood of small scale farmers and dairy owners. Tropical theileriosis, mostly affecting dairy cattle and buffaloes is a major threat to dairy and related industries. Tropical theileriosis is caused by Theileria annulata, a hemoprotozoan parasite transmitted by Ixodid ticks of Hyalomma spp. In the present study, we examined the clinical signs, hematological parameters and flow cytometric profile of whole blood in 30 theileriosis affected crossbred cattle. The aim of our study is to analyze, in comparison with clinical and hematological diagnosis, whether flow cytometry based profiling of leukocytes could be used as better, quick and alternative method for diagnosis and screening of bovine tropical theileriosis in crossbred cattle.Materials and Methods:In this study, we screened parasites in 30 peripheral blood samples from clinical cases of theileriosis by Giemsa’s staining technique in crossbred cattle. Hematological analysis was done to estimate hemoglobin (Hb) content, total red blood cell (RBC) count, total leukocyte count and differential leukocyte count. Further, flow cytometric analysis of whole blood was carried out to study leukocytes profile in affected cattle.Results:Microscopic examination of stained blood films revealed the presence of piroplasms in erythrocytes and schizonts in lymphocytes. Hematological examination revealed significant (p<0.05) decrease of Hb percent (Hb %), reduced total RBC and total leukocytes, lymphocytosis, eosinopenia, and neutropenia compared to that of apparently healthy cattle. Flow cytometric profiling of leukocytes revealed the severe effect on shape, size, and granularity of leukocytes, marked decrease in granulocytes and 3-5 fold increase in lymphocytes count compared to clinically healthy cattle. Thus, in both methods, namely conventional and flow cytometric analysis, marked lymphocytosis and decrease in other blood cell counts were observed compared to clinically healthy cattle.Conclusions:From results, it can be concluded that though conventional staining techniques and hematology are efficient in diagnosis of theileriosis, leukocytes profiling based on flow cytometry combined with clinical examination could be a quick, novel and alternative method for diagnosis and screening of clinical tropical theileriosis in crossbred cattle. Thus, there is potential to offer a flow cytometry based diagnostic service for tropical theileriosis in crossbred cattle.

Development and validation of a flow cytometry method to examine circulating leukocyte subpopulations in barramundi (Lates calcarifer)

“…Certain cell-based dyes have been shown to stain leukocytes and RBCs as distinct populations in flow cytometry analysis [13] . Cytometric analysis can also be used to elucidate the structural and functional properties namely, percentage of cells, their size and granularity, and their various expressed proteins [14] . Here we report a new stain-based technique which enables differentiating leukocytes from whole blood with subsequent differentiation of the major leukocyte populations—granulocytes, monocytes, and lymphocytes—using flow cytometry.…”

Application of Multiparametric Flow Cytometry Panels to Study Lymphocyte Subpopulations in Tuberculin-Positive Cattle

“…This condition results in a poor characterization of the different cell populations in the sample and a longer processing time, since more samples per individual need to be handled to identify the cell markers of interest [ 8 ]. In cattle, FC was used for counting total leukocytes in a blood sample [ 9 ], in the study of neutrophil adhesion receptors (CD11/CD18) [ 10 , 11 ], in the analysis of the immune response to M. bovis [ 12 , 13 , 14 , 15 ] and other pathogens [ 16 , 17 ], in the evaluation of vaccine effectiveness [ 18 , 19 , 20 ], and in the diagnosis of theileriosis [ 21 ]. However, all these panels used a limited number of fluorochromes.…”

Evaluation of Therapeutic Regimens for Theileriosis-affected Buffaloes in the Malwa Region of Madhya Pradesh, India

“…Theileriosis in cattle, particularly in crossbred cattle, has been documented by numerous researchers from around the world (Preston et al, 1992;Omer et al, 2002;Col and Uslu, 2006;Kohli et al, 2014;Jagtap et al, 2015;Devadevi et al, 2018). A few reports of theileriosis in buffaloes, however, have been documented (Singh et al, 2012;Vahora et al, 2012;Chaudhari et al, 2013).…”