Brucellosis represents one of the world's most widespread zoonotic diseases, causing significant human illness and substantial economic losses in livestock production (1, 2). The World Health Organization (WHO) has recognized brucellosis as a major neglected zoonotic disease that continues to pose significant public health challenges in endemic areas due to its impact on human health and livestock productivity, as well as difficulties in effective diagnosis, surveillance, and control measures (3).

The complexity of brucellosis is reflected in its diverse causative agents, with 12 recognized Brucella species classified according to host preference, pathogenicity, and phenotypic characteristics (4) Among these, six “core” species are particularly significant: B. abortus primarily affecting cattle, B. melitensis in goats and sheep, B. suis in pigs, B. ovis in sheep, B. canis in dogs, and B. neotomae in desert woodrats (5, 6). While these species show host preferences, cross-species transmission occurs frequently, with B. melitensis notably demonstrating broad host adaptability (7, 8).

Human infection primarily stems from three Brucella species: B. abortus, B. melitensis, and B. suis (7). Transmission to humans typically occurs through multiple pathways, including direct contact with infected animals, consumption of unpasteurized dairy products, and occupational exposure among veterinarians, livestock handlers, and abattoir workers. Airborne transmission also poses a significant risk (4). The World Health Organization estimates 500,000 new human cases annually (3) though this figure likely underestimates the true disease burden, particularly in malaria-endemic regions where symptoms may be misattributed (9).

Recent decades have seen successful control of B. abortus in many developed nations. However, the increasing prevalence of B. melitensis and B. suis in livestock has expanded both the geographical range and overall prevalence of human brucellosis (2, 10, 11). In developing countries like Rwanda, limited and inconsistent epidemiological data on Brucella species hampers effective disease control efforts. Comprehensive studies are essential to generate reliable incidence estimates, assess impact, understand transmission dynamics, identify reservoirs, and develop targeted control strategies (12).

To address these knowledge gaps, we conducted a cross-sectional study in Nyagatare District, a strategically important region bordering Tanzania and Uganda that contains 30% of Rwanda's cattle and goat populations (13). This district was specifically selected due to its high livestock density, pastoralist communities with frequent human-animal contact, and its transboundary location which increases risk of disease transmission across national borders. Our study had three primary objectives: 1) to estimate brucellosis seroprevalence in both human and livestock populations, 2) to identify risk factors associated with Brucella seropositivity, and 3) to assess awareness and practices related to brucellosis among high-risk occupational groups.

This research aims to provide comprehensive epidemiological data to inform evidence-based strategies for brucellosis prevention and control in Rwanda and similar settings, contributing to the broader goal of reducing the global burden of this zoonotic disease. By focusing on the human-animal interface in a high-risk region, our findings offer valuable insights for implementing effective One Health approaches to disease control in endemic areas.

Brucellosis remains an endemic disease affecting both livestock and humans in developing countries, posing significant public health and economic burdens (22, 23). Despite its classification as a neglected zoonotic disease without pandemic potential (24), brucellosis can lead to severe complications in humans, including encephalitis, orchitis, arthritis, and myelitis (25). This study provides comprehensive insights into the seroprevalence and risk factors associated with brucellosis in both human and livestock populations in Nyagatare District, Rwanda, using a One Health approach that integrates human, animal, and environmental factors. The findings underscore the endemic nature of brucellosis in this region and highlight critical areas for intervention.

The human seroprevalence observed in this study, using i-ELISA (19.9%, 95% CI: 17.3–22.6), represents the highest ELISA-based prevalence reported to date in Rwanda. This finding is particularly concerning as it suggests a substantial underdiagnosis of brucellosis in human populations in this region, potentially leading to mismanagement of febrile illnesses. This prevalence is comparable to the prevalence previously reported in humans in Uganda (21.2%) (26), likely reflecting the heightened exposure risk among participants, most of whom are part of high-risk populations with regular livestock contact. A recent study conducted in the Kagera region of Tanzania reported a much higher seroprevalence of 41.1% among individuals in a multi-herd ranch system, with the elevated risk attributed to close contact with animals during parturition and handling aborted fetuses (27). This comparison highlights the critical role of occupational exposure and livestock handling practices as significant contributors to human brucellosis risk in endemic regions. Consistently, occupation was a significant risk factor associated with brucellosis seropositivity in our study. Particularly, animal health care practitioners exhibited higher odds of brucellosis infection (OR = 2.90, 95% CI: 1.38–6.06), compared to other occupational groups. Notably, the prevalence of brucellosis observed among veterinary professionals in this study (35.2%) surpassed that reported in India (7.04%) and Turkey (11.8%) in animal health care practitioners (28, 29). This occupation-associated prevalence observed in the current study could be attributed to the heightened risk of exposure inherent to close and regular contact with infected animals. Furthermore, this exposure risk may be exacerbated by a lack of awareness regarding the zoonotic nature of brucellosis among veterinarians and para-veterinarians in Rwanda.

Indeed, although no significant exposure risk factors were identified for veterinarians and butchers, the high seroprevalence observed in these groups suggests inadequate biosafety precautions like slaughtering with bare hands and the occurrence of accidental self-injections during blood sampling and vaccinations against brucellosis (Supplementary Tables S4A, S4B). Similar findings have been reported in studies from Tanzania and Uganda (24–26), emphasizing the need for occupation-specific preventive measures. Thus, it is crucial to continually educate veterinarians, butchers, and their supervisors on biosafety and biosecurity practices when sampling, vaccinating, handling, and treating livestock. The absence of statistically significant risk factors in our study may be attributed to the limited sample size for these two groups (veterinarians and butchers). Therefore, future large-scale investigations focused on these occupational groups are necessary to better understand the specific predictors of brucellosis risk in Rwanda. These findings highlight the urgent need for targeted interventions in high-risk occupational groups and emphasize the importance of biosecurity practices.

In the current study, age and gender were demographic potential risk factors associated with brucellosis in humans. This agreed with findings reported in Uganda by (26). Both studies indicated that males and older individuals demonstrate a stronger association with higher odds of brucellosis seropositivity compared to females and young individuals. In Rwanda, livestock farming remains an important economic activity, with a significant proportion of farms management activities and services being handled by men, compared to women who are predominantly involved in small-scale and/or zero-grazing farming operations (19, 30, 31). This gendered pattern in livestock management and activity distribution could contribute to the observed gender disparity in brucellosis seropositivity, where males seem to be associated with higher odds of infection. These observations demonstrate the importance of considering gender dynamics in understanding and addressing the risk factors associated with brucellosis transmission in both human and livestock populations. The observed AUC values for both the human model (0.76) and livestock model (0.78) further demonstrate good discriminatory performance, supporting the reliability of the identified risk factors. In addition to demographic and occupational factors, questionnaire findings provided important contextual insights. Despite the lack of statistical significance in our dataset, the high prevalence of unpasteurized milk consumption remains a major epidemiological concern, as raw milk is a well-documented transmission route in East Africa. The absence of an observed association here may reflect sampling bias or underreporting, but warrants attention for public health education. Similarly, awareness gaps about brucellosis transmission underscore the need for targeted outreach interventions to improve knowledge and prevention practices in pastoral communities.

This study showed that the Brucella seroprevalence increases with age, possibly due to cumulative exposure to risk environments over time. Indeed, older individuals, particularly those working in high-risk occupations such as livestock farming or veterinary services, are more likely to have repeated contact with infected animals or contaminated materials, increasing their odds of seropositivity. The strong age-related gradient observed (OR = 7.39, 95% CI: 3.82–14.8 for those >55 years) highlights the importance of targeting prevention strategies toward long-term livestock workers who may have developed habitual unsafe practices over decades of animal contact. This has been confirmed in studies where middle-aged adults had significantly higher odds of being seropositive compared to younger individuals (32, 33). Similarly, as seen in this and other studies (33, 34) older livestock tend to have a higher prevalence of brucellosis, suggesting that both humans and animals face increased risk due to prolonged exposure.

In livestock, seroprevalence rates of 11.9% (95% CI: 9.4–14.5) in cattle and 11.3% (95% CI: 7.1–15.4) in goats were found, aligning closely with recent reports of 8.2% in cattle (19) and 10.7% in goats (30) within the same district. The similarity in prevalence between cattle and goats is particularly noteworthy as it suggests potential cross-species transmission or common environmental sources of infection, which has important implications for control strategies. However, other studies have reported higher brucellosis seroprevalence rate, such as 22% in cattle and 51.6% in bulk milk from both open grazing and zero-grazing systems (35). These findings collectively suggest that brucellosis is endemic in this transboundary region. The variations in reported prevalence rates across studies may be attributed to differences in sample sizes and other potential factors, such as herd composition and sampling period (23). Additionally, factors such as differences in management practices, geographical variability, and livestock-wildlife interactions could also contribute to these disparities, warranting further investigation into the regional epidemiology of brucellosis.

Our study also provides, the first report of brucellosis seroprevalence in sheep in Rwanda, with a prevalence of 1.4% (95% CI: 0.0–4.2). This finding is comparable to the 1.23% reported in Uganda in 2022 (36). Although the prevalence is low, this finding is significant as sheep may serve as potential reservoirs for Brucella infection within mixed herds, especially given that small ruminants are not vaccinated against brucellosis in Rwanda. The significantly lower odds of seropositivity in sheep compared to cattle (OR = 0.12, 95% CI: 0.01–0.60, p = 0.042) suggests species-specific differences in susceptibility or exposure that could inform targeted control strategies. However, this prevalence is lower than the average rate of 13.8% reported across the entire East African Community (EAC) (37) and the 12.8% prevalence reported in sheep in South Africa (38). The relatively lower prevalence observed in Rwanda could be attributed to country's small sheep population, reported to be 331,748 (39), as well as the limited sample size in our study. These findings highlight the critical need for enhanced surveillance and control strategies for brucellosis in small ruminants, particularly in Rwanda where these species are not vaccinated. While the lower seroprevalence in sheep (1.4%) compared to goats (11.3%) suggests species-specific transmission dynamics, both species can serve as maintenance hosts in mixed herding systems. The combination of lack of vaccination programs in small ruminants, and the practice of mixed livestock farming in Nyagatare District, create conditions that could facilitate inter-species transmission. Additionally, the identification of shared risk factors such as communal grazing and water points highlights the need for integrated control approaches that consider the interconnected nature of small ruminant management practices in this region.

In the current study, we observed that improper disposal of aborted materials and keeping aborting animals within a herd led to significant increased odds of brucellosis seropositivity (OR = 3.15, 95% CI: 1.18–7.99 and OR = 10.0, 95% CI: 2.27–49.2, respectively) consistent with previous reports (40, 41). Indeed, these inadequate biosecurity practices are well-documented practices that significantly increase the risk of brucellosis transmission (40, 41). This is because Brucella bacteria are highly concentrated in aborted tissues, placental fluids, and vaginal discharges of infected animals, making these materials potent sources of infection (40). In our study, we observed that these poor biosecurity practices led to significantly increased odds of brucellosis seropositivity. Similar findings have been reported in other studies, where improper management of aborted materials and the presence of aborting animals in herds were strongly associated with increased seroprevalence of brucellosis. These results highlight the importance of implementing proper disposal methods, such as burying or incinerating aborted materials, and promptly isolating aborting animals to prevent further transmission within herds and to human handlers.

Lastly, there were geographical significant difference in Brucella seropositivity in the current study with Rwimiyaga sector exhibiting the highest prevalence (OR = 3.06, 95% CI: 1.37–7.45, p = 0.009) compared to the Karangazi sector. Interestingly, Brucella melitensis was isolated from a goat flock presenting a storm abortion in this sector in 2018 (42). Thus, this difference may be attributed to the previous exposure and persistence of brucellosis infection in animals of Rwimiyaga sector. The identification of this spatial cluster of brucellosis is a critical finding as it suggests potential environmental or management factors specific to this area that facilitate transmission. This indicates that brucellosis control strategies including vaccination, surveillance and education campaign should prioritize this sector to effectively mitigate brucellosis. Geographic targeting of interventions based on spatial risk assessment can optimize resource utilization in control programs, particularly in resource-limited settings.

This study demonstrates high seroprevalence of brucellosis among humans (19.9%) and livestock (10.9%) in Nyagatare District. Key risk factors included male gender, older age, and animal health occupations in humans, and poor abortion management practices in livestock. The Rwimiyaga sector emerged as a hotspot of transmission.

Based on these findings, we recommend three specific interventions:

These evidence-based recommendations require a coordinated One Health approach involving veterinary, medical, and public health authorities to effectively reduce the burden of brucellosis in this region.

Our findings provide evidence to guide Rwanda's national zoonotic disease control program and emphasize the importance of One Health approaches for brucellosis control in East Africa. Integrating these interventions into Rwanda's National Zoonotic Disease Control Program would strengthen surveillance, prevention, and cross-sectoral coordination to reduce the burden of brucellosis.

Our study has several limitations that should be considered when interpreting the results. First, the small sample sizes of high-risk occupational groups (veterinarians and butchers) limited our ability to detect occupation-specific risk factors. While this sampling distribution reflects the actual occupational demographics in the district, it reduced statistical power for between-groups comparisons. Although we found elevated seroprevalence among veterinarians (35.2%; Supplementary Table S4), these results should be interpreted conservatively given the wide confidence intervals stemming from the limited sample size.

Second, our human sampling strategy focusing on febrile patients and high-risk occupational groups introduces potential selection bias, and the reported 19.9% seroprevalence likely overestimates the true prevalence in the general population. Third, the cross-sectional design prevents us from establishing temporal relationships between risk factors and infection, and potential seasonal variations in brucellosis transmission could not be assessed. Fourth, while i-ELISA provides high sensitivity and specificity for detecting anti-Brucella antibodies, it cannot distinguish between active and past infections, potentially overestimating current disease burden. Finally, we hope that future studies using molecular techniques and longitudinal designs, focusing on veterinary professionals across multiple districts in Rwanda, will better elucidate the epidemiology of brucellosis and help inform targeted interventions.