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Bovine anaplasmosis is endemic in Pakistan where it reduces livestock productivity and leads to high mortality, especially in young animals. This study was aimed to identify the potential risk factors responsible for the occurrence and spread of anaplasmosis in cattle and buffaloes for the first time in Pakistan. A total of 900 (cattle = 479, buffalo = 421) blood samples were collected irrespective of age and sex from three distinct zones of Khyber Pakhtunkhhwa (KP) province of Pakistan. Polymerase chain reaction (PCR) technique was used for the molecular detection of anaplasmosis. Data collected on a piloted questionnaire including 11 predicting variables which were analyzed using R-statistical software, and association between the dependent and independent variables was assessed using univariable analysis. Automated and manual approaches were exercised, producing comparable models. Key risk factors identified in all the approaches included species of the animal, breed of animal, sex of animal, tick infestation status, previous tick history, tick control status, and acaricides used (odds ratio > 1). The 611 bp DNA fragment specific for 16S rRNA gene of Anaplasma spp. was produced from 165 samples. The samples were confirmed for anaplasmosis through sequencing and BLAST queries. The findings of the current study conclude that by enhancing the protective measures to control the identified risk factors can reduce the spread of anaplasmosis in Pakistan.

The dairy industry in Pakistan is booming, and investors are anxious to fund dairy farms that are using high-milk-producing (exotic) cattle breeds such as Holstein Friesians that are not native to the country. Unfortunately, the benefits of increased milk production do not provide resistance to pathogens present in regions where the exotic breeds are introduced. Therefore, the current study was conducted to evaluate the economic impact of Theileria annulata on a commercial Holstein Friesian dairy farm in the District of Ranjanpur, in the Province of Punjab, Pakistan. The economic impact of T. annulata infection was calculated for cattle with subclinical and clinical theileriosis. Losses were estimated based on milk production, morbidity, mortality, and tick control costs (organophosphate sprays). Animals were classified into groups after screening for mastitis, teat abnormality, abnormal parturition, intestinal parasites, and hemoparasites ( T. annulata, Babesia spp., and Anaplasma spp.). Microscopy was done for hemoparasites and intestinal parasites. PCR was used to confirm microscopic identification of T. annulata. Animals were classified into 3 groups: group A (normal), group B (subclinical theileriosis), and group C (acute theileriosis). Hemoparasites were observed microscopically in 28.7% of cows. Theileria annulata was found in 8%, and the herd incidence (new cases) of T. annulata was 2.8%. Milk production, animal rectal temperature, and body condition scores between group A and groups B and C were significantly different ( P < 0.05). But the enlargement of sub-scapular lymph node and interval of body condition score of the 3 groups were not significant ( P > 0.05). The total expenditure incurred due to theileriosis was US $74.98 per animal and 13.83% of total farm costs. Hence theileriosis caused significant economic loss of US $18,743.76 (0.02 million) on this Holstein Friesian dairy.

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Authors' Contribution SHF, MI, MIR conceived and designed the projecr and conducted research work. MHS, AA, AIA arranged and analyzed the data statistically. SHF, AIA wrote and reviewed the manuscript. SHF, SM arranged and performed GIS Mapping.

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