Abstract Coxiellosis, also known as Q fever, is a zoonotic disease caused by Coxiella burnetii , a gram-negative bacterium that exerts a significant deleterious impact on the productive and reproductive capabilities of livestock, severely effecting the economics of this sector. In this study, 448 sera samples from cattle (n = 224) and buffalo (n = 224) were collected from 112 farms in Pakistan and examined for antibodies against C. burnetii using an indirect ELISA. Ticks were also collected from these animals. Serological analysis revealed a 23.66% and 27.23% seroprevalence of Q fever in cattle and buffalo, respectively. Odds ratio (OR) analysis of the factors associated with C. burnetii seropositivity was performed, and a multivariable logistic model identified five main variables associated with the seropositivity for coxiellosis. These were: (i) the absence of acaricide use (OR 5.61; 95% CI 2.97–10.94); (ii) the presence of ticks (OR 3.23; 95% CI 1.87–5.69); (iii) the abortion history during the preceding year on the farm (OR 14.96; 95% CI 8.09–29.34); (iv) the presence of sheep and goats (OR 2.47; 95% CI 1.20–5.35); and (v) the absence of a separate parturition area (OR 3.17; 95% CI 1.76–5.86). This study provides new insights into the seroprevalence of Q fever in large ruminants across seven studied districts of Punjab, Pakistan, also providing baseline data to inform improved herd management and on-farm practices for the prevention and control of Q fever in large ruminants in the region. Results of this work suggest that further molecular investigation of coxiellosis is warranted to provide a more thorough evaluation of C. burnetii epidemiology in Pakistan.