We detected the presence of Brucella DNA in pig sera that also screened positive using the RBT. According to our knowledge, this is the first report of molecular detection of Brucella spp. in pig serum in East Africa. Brucellosis is a widespread zoonotic disease and serological evidence of pig brucellosis has previously been described in Africa [15, 18,19,20,21,22]. However, information on B. spp. circulating in the pig population remains limited. South America and Southeast Asia are considered to have a higher prevalence of porcine brucellosis, as compared to the other regions, including Africa [1]. The sero-positivity of 0.57% detected in this study is consistent with findings from different serological studies conducted in other African countries such as Nigeria [19, 20, 22], Zambia [22] and Uganda [20], which reported prevalence of between 0 and 0.6%. However, our results differ with one study that reported a high sero-prevalnce of 30.6% in Benue State of Nigeria [21]. The high prevalence reported in Benue State, however, could be due to the clustering of positive animals in the targeted region, as other studies in Nigeria also reported lower prevalence rates [18, 23]. Despite the low prevalence of 0.57% detected from the pig population in this study, the risk of transmission in the pig population may grow with the rapidly growing pig production in Kenya, especially if control interventions are not put into place.
In this study, we found that four out of the six positive samples detected were of B. abortus, as they amplified with B. abortus primers [14]. The other two samples that did not amplify with the species identification primers could belong to a different species, given that the assay is designed to distinguish between B. abortus and B. melitensis [14]. The detection of zoonotic Brucella spp. in pig sera, and more so B. abortus, is of importance given that pigs are traditionally associated with B. suis, while cattle are considered to be the preferred host for B. abortus [1]. This finding therefore highlights the possibility for novel transmission dynamics within pigs and the need for further investigations to inform appropriate control strategies for brucellosis in Kenya and similar settings. The number of studies that have used serum samples for extraction of genomic DNA, and subsequent detection of Brucella DNA, has increased in the recent past [13, 15, 20]. This could be due to the fact that serum samples can be used for routine testing of brucellosis in both animals and humans since, unlike other samples such as abortion materials, swabs, hygromas and milk, they are not affected by the age, sex or physiological stage (such as pregnancy). Even though Brucella spp. are known to have host preference, cross-infection has previously been reported to be common in areas where mixed husbandry systems are practised [11, 24, 25].
Similarly, the presence of B. melitensis in pigs’ serum was recently reported in Egypt [15]. In that study, the prevalence was comparable to a previous report from Latin America where B. melitensis was detected in pigs [26]. In the current study, the positive pigs were sourced from peri-urban and slum areas, where small-scale farmers keep mixed herds that facilitates close contact between the different animal species. Free-range systems practised in the informal settlements, and feeding of pigs on waste from the market [27] could also contribute to the cross-transmission of B. abortus from cattle to pigs. The presence of B. abortus in pigs may not only present a zoonotic risk to non-suspecting farmers, slaughterhouse workers and pork consumers but also raises the need for further investigation on the epidemiology and pathogenicity of B. abortus in pigs, as well as their contribution to human infection.
The Rose Bengal Test (RBT), Buffered Plate Agglutination Test (BPAT) and ELISA are recommended serological tests for screening brucellosis in pigs [1]. However, the results from these tests should be confirmed by reference serology tests or confirmatory bacteriology and molecular techniques [10, 11, 28, 29]. Variability between the performance of different serological tests, or in the sensitivities and specificities for the same tests using pig serum in different studies, have been recorded [28, 30, 31]. This study also observed a poor agreement between the RBT and cELISA, which is similar to the findings in other studies [30, 31]. Further investigation should be carried out to evaluate the performance of these two assays on pig samples. There was a much better agreement between the conventional and qPCR. All four positive samples detected by conventional PCR (B4/B5 primers) were further confirmed by a multi-level genus- and species-specific RT-PCR. Brucella abortus was identified in four of the six positive samples. The agreement between molecular and serological tests raises an important consideration for these tests to be used for routine testing (surveillance) of brucellosis in livestock and wildlife.
This study had several limitations; first, not all 700 sera screened by the RBT and cELISA were tested by PCR. This could downplay the positivity proportion detected in the molecular assays used. Secondly, the time lapse between the serology (September 2018) and molecular testing (November 2019) implies that study personnel were not entirely blinded to the results of the initial screening when conducting PCR. Finally, the limited scope of the species identification technique used (B. abortus and B. melitensis) could imply that other Brucella spp., including a possibly novel species in this pig population, could have been missed in this study. Future studies should consider the suitability of different assays for the detection of pig brucellosis since it is an emerging area of research. All the assays used in this study are not explicitly designed for pig host testing, rather the assays are generally applied to the detection of Brucella antibodies and DNA. This could affect the sensitivity or specificity of detecting brucellosis in pigs, especially when a single test is used. Therefore, future studies should evaluate a range of tests for utility in testing for brucellosis within this crucial population.