Serological and Molecular Investigation of Coxiella burnetii Infection and Associated Risk Factors in Selected Sites of Ethiopian Livestock

Abstract

Coxiella burnetii (C. burnetii) infection is a significant global public health, veterinary, and economic concern. Despite the global public health and veterinary importance, information regarding its distribution, prevalence and risk factors is scarce in Ethiopia. Hence, a comprehensive sero-epidemiological and molecular investigation was conducted to characterize C. burnetii infection, the causative agent of Q fever, and its associated risk factors in livestock from Ethiopia. We carried out this study to bridge some of the knowledge and information gaps by focusing on the geographical distribution of serological evidence of infection in livestock slaughtered at major abattoirs in Addis Ababa, Adama, and Modjo, as well as pastoral areas of Oromia, Ethiopia. A total of 4,140 whole blood samples were collected from livestock species at three abattoirs (Addis Ababa, Adama, and Modjo) and three pastoral districts (Mega, Dubuluk, and Yabello) from January 2021 to May 2022. In addition, 264 biological samples consisting of milk, urine, vaginal swabs, aborted fetal tissues and whole blood were collected from livestock species in three pastoral districts (Moyale, Boku Luboma and Elweye) where there is evidence of reproductive problems. Moreover, 292 ticks were collected from infested livestock during the biological sample collection. During sample collection geographical data was collected using GPS for mapping the origin of livestock species sampled. Georeferencing was performed to accurately delineate the sample locations. The sera samples were analyzed for the presence of anti-C. burnetii antibodies using an indirect Enzyme Linked Immunosorbent Assay (iELISA) kit. Additionally, buffy coat from iELISA serum positive livestock, milk, urine, vaginal swabs, aborted fetal tissues, and ticks were analyzed by real time quantitative polymerase chain reaction (RT-qPCR) targeting cytochrome c oxidase subunit 1 (COX1) gene. Logistic regression analysis was used to quantify the association between seroprevalence of C. burnetii infection and various potential risk factors. Out of the 4,140 serum samples tested, 777 (18.77%; 95% CI: 17.59, 19.99) were found positive for C. burnetii antibody. The sero-prevalence estimate was 27.17% at Addis Ababa abattoir, 19.41% at Adama abattoir, 19.13% at Modjo abattoir and 12.1% in livestock tested from Mega, Dubuluk, Yabello pastoral areas. Sera analysis at the animal species level showed that cattle exhibited the lowest sero-prevalence estimate (11.83%; 95% CI, 10.27 13.53%), while the highest was observed in camels (28.39%; 95% CI, 25.16–31.80%). The sero-prevalence estimate was 21.34% (95% CI, 18.86–23.99%) in goats and 20.17% (95% CI, 17.49–23.07%) in sheep. Female livestock were observed to have a higher seroprevalence (26.65%; 95% CI, 23.37 30.13%) than male livestock (17.21%; 95% CI, 15.97–18.51%). The prevalence was also higher in adult livestock (25.02%; 95% CI, 23.26–26.85%) than their younger counterparts (11.03%; 95% CI, 9.64–12.54%). Higher seroprevalence of C. burnetii infection was also observed in livestock infested with ticks (carry ticks on their body parts) (80.99%; 95% CI, 73.55–87.08%) than those livestock which were not infested by ticks at all (7.05%; 95% CI, 6.52–7.59%). The results of multivariable logistic regression analysis showed that species, age, and tick infestation were important risk factors for C. xv burnetii infection. The odds of infection were 3.22 times higher in camels and almost twice as high in goats and sheep compared to cattle. Adult livestock were infected more likely (OR = 3.23) than young ones. Interestingly, a significant difference was observed in the sero prevalence of infection between livestock that were infested with ticks (OR = 16.32) and those which were tick-free. Furthermore, out of 384 buffy coat samples randomly selected from the iELISA sera positive livestock and tested by real-time quantitative PCR (RT-qPCR), none of them showed positive result. Similarly, none of the 264 biological samples (aborted fetal tissues, urine, milk, buffy coat, and vaginal swabs) tested by RT-qPCR showed positive result. The prevalence of tick infestation was highest in camels, with 143 ticks (49.0% of the total collection) while cattle had the lowest prevalence of infestation (23 ticks, 7.9%). The tick fauna was dominated by Rhipicephalus pulchellus (30.5%) and Amblyomma variegatum (15.4%), but Rhipicephalus pravus (1.0%) and Hyalomma truncatum (2.7%) being the least frequently observed tick species. Similar to the biological samples collected from livestock, all ticks (292) analyzed using RT-qPCR showed negative results. This suggests a potential low-level infection, intermittent pathogen shedding, or challenges with detection using the chosen gene target. In conclusion, this study provides the first comprehensive sero-epidemiological data for C. burnetii infection in a multi-species livestock population in Ethiopia, while also highlighting the challenge of molecular detection. The serological study revealed the widespread exposure of livestock to the C. burnetii (seropositive in the absence of vaccination), identified important risk factors, and warrants the need for enhanced surveillance and the implementation of biosecurity measures to safeguard both animal and public health.