Comparing the two groups of people, one living near an ecosystem of bat in habited caves and forest reserves in western Uganda and another in savanna rangeland in central Uganda, we see that the risk of being filovirus positive is higher in the former group than the later by at least four times. Geographical differences that favour a filovirus reservoir in different countries in Africa could explain the variation in seroprevalences. We cannot clearly explain why Marburg virus seroprevalence is lower than that of Ebola, but this is consistent with other studies where the two pathogens have been tested. One of the explanations could be that the antibodies for Marburg virus are not as long-lasting compared to those of Ebola virus, but this needs to be explored in further studies.
We also see a higher filovirus seroprevalence in our high-risk exposure group in Ibanda and Kamwenge district in Western Uganda at 3.7% (16/433) compared to low-risk exposure group in Central Uganda at 1.1% (3/291). Artisanal gold miners who enter bat-inhabited caves had an even higher prevalence of 5.6% (9/161) compared to non-miners in central Uganda at 1% (3/291) As has been reported before, the Kitaka mines where the exposed population is centred is inhabited by bats of species Rousettus aegyptiacus that are the known reservoirs for Marburg virus (7–10). We expected a higher seroprevalence against Marburg virus than Ebola virus, but the opposite was observed with Sudan virus seroprevalence being higher than Marburg virus. Whereas it has been confirmed during previous investigations that bats occupying the mines are actively infected with Marburg virus and had been associated with two MVD outbreaks (6, 7), no outbreak of EVD has been reported in this region. It was therefore surprising to find higher seroprevalence to Sudan ebolavirus instead of the expected Marburg virus. Another possibility for this finding is there could be a reservoir for Sudan ebolavirus or another closely related filovirus in Kitaka mines and/or inhabiting the area around the Kasyoho-Kitomi reserve ecosystems to which these individuals were exposed, especially the gold miners. This area is near Queen Elizabeth National Park, and so there is a possibility of having an unknown reservoir of Ebola virus in the game reserve that has not been previously identified. This contrasts with our low-risk exposure group from Central Uganda, Luweero district where only three people were identified as being seropositive for Sudan ebolavirus, and none was positive for any other species of filoviruses. The only positive MVD case was in a miner from the high-risk exposure region, and he did not show seroprevalence for any other filovirus species.
Our findings are also consistent with another filovirus serological study by Nkoghe et al.(2011) in rural Cameroon and Gabonese populations where the prevalence of Ebola virus was higher in populations near forests (21, 24, 34). Although no other risk factors were identified in Gabonese study, we find in our study that being a miner is highly associated with being seropositive for Sudan ebolavirus. It is important to note the miners in Kitaka cave live in forested ecosystem near a national park that is comparable to the central African forest.
Another study in the Gabon found that pygmies, who are forest dwellers, had a higher percentage of ebolavirus seroprevalence than other populations at 7.02% compared to non-pygmies (4.2%) (45). This further indicates that communities that live in the forested areas, like the ones we studied in western Uganda are at higher risk of infection with filoviruses compared to those living in more developed or non-forested areas. Forested areas tend to have a greater abundance of fruiting trees that provide food to the fruit bats, the hypothesized reservoirs of Ebola virus. However, in this study, going into the forest was not shown to be a risk factor for individuals being seropositive for filoviruses.
Gold mining has been previously described as a risk factor for Marburg virus infection in a study in DRC (46) with OR = 13.9, 95% CI;3.1–62.1 but not for Ebola virus. We report artisanal mining and going inside the mines as risk factors for being seropositive for filovirus in Uganda (AOR = 3.4, 1.3–8.5). In fact, the very first cases of Ebola virus were reported in mining communities in DRC in 1976. Other factors that we identified as risk factors were being a family member of a miner. Since filoviruses are spread by contact when miners fall sick, they are primarily taken care of by family members, and hence are likely to be at greater risk of acquiring the infection. The four seropositive individuals we found in the Luweero district group could be due to travel and migration from high-risk areas but may also be due to the movement of reservoirs such as bats that are known to travel long distances hence spreading the infection. Ebola virus seropositivity has before been reported in a grassland savanna-like ecosystem in Nigeria similar to the grassland savannah ecosystem of Luweero where the four EVD (4) seropositive cases were identified (36). However, frequent travels outside high-risk areas were protective (0.3; 0.1–0.7). This may be because those who frequently travel away from risk areas are less likely to be exposed to the putative reservoir. Being male was associated with a high risk of being seropositive (3.1;1.01–9.5) compared to being female, likely partly due to men being more likely to be miners and go inside the mines and the forests for manual work and become exposed and hence acquire infection. Cleaning a dead body was significantly associated with being seropositive for a filovirus. This has been widely reported in outbreaks of filoviruses as burials and funeral rites amplify these outbreaks. Unlike the study by Nkonghe et al (2011) in Gabon, receiving injections was not a risk factor in this study simply because of a possible higher level of infection control practiced in hospitals in Uganda. Contact with EVD/MVD suspect was a predictor of seropositivity with a filovirus and has been reported in a partial meta-analysis done on the risk of Ebola transmissions (47).
Looking at the overall seroprevalence reported in this study, the findings suggest that there may be filovirus infections that occur in Sub- Saharan African countries and go undetected by the health care systems. Also, our findings suggest that people who are involved in artisanal gold mining and live close to caves are at higher risk of infection with filoviruses, likely because of their increased risk of exposure to bats that inhabit those mines and caves. This could possibly lead to large epidemics as was seen in West Africa (16). Additionally, our findings do not rule out the possibility that there could be cross-reactivity for filoviruses in our diagnostic assays caused by either another filovirus infection or a non-filovirus infection.
Here, we report 19 individuals that were seropositive for IgG antibodies against filoviruses representing 2.6% of the people tested. Out of these 19, 18 were seropositive for Sudan ebolavirus, and one was seropositive for Marburg virus. Of the 18 seropositive for Sudan ebolavirus, one was also seropositive for Bundibugyo ebolavirus, likely a cross-reactivity rather than representing previous exposure to both Bundibugyo virus species, as has been described previously. However, a similar unpublished study was carried out by CDC and the Ministry of Health following 2007 MVD outbreak in Kamwenge and Ibanda district in the same area. In that study, they found seroprevalence of Marburg virus at 1.2% (7/564) and Sudan virus at 1.2% (7/564). The seroprevalence of Marburg virus was slightly lower in our study whereas that of ebolaviruses was higher. We do not have a clear explanation for these differences in seropositivity between the two studies conducted in the same area eight years apart. Also, the seroprevalence in our study is lower than 3% and 8% for pooled seroprevalences reported in meta-analyses of seroprevalence of ebolaviruses performed in other parts of the world (1, 17). Following the West Africa EVD outbreak, reports of asymptomatic infection in West African populations has been suggested in populations who had contact with EVD patients at 12%, and at 2.6% in non-contacts (18, 19).
Our study also reported a lower seroprevalence of Ebolaviruses (2.5%) than pooled seroprevalences reported in other studies in neighbouring Democratic Republic of Congo (DRC) at 10% (20–25), Central African Republic and Gabon at 11% (26–30), Sudan at 22% (31), Madagascar at 4% (32), Liberia at 13% (33) and Cameroon at 7% (34, 35), However, our study showed higher seroprevalence than that reported in Nigeria at 2% (36), Germany at 1% (37) and Kenya at 1% (38). Only one Marburg virus seropositive person was confirmed in our study and this was much lower than has been reported in other studies (32, 37–42).
We also report for the first time seroprevalence of Bundibugyo ebolavirus in one individual, while no individuals showed seroprevalence for Ebola Zaire virus. These variations in seroprevalence could be due to differences in filovirus ELISA testing protocols and potential cross-reactivity caused by a non-filoviral infection. The test, developed by CDC that was used in this study has been shown to be more specific than other filovirus serological tests used in previous filovirus seroprevalence studies (15). This serological test was developed and validated by US Centres for Disease Control and Prevention (CDC) on known positive and negative human samples with a sensitivity > 90% and specificity of > 90%. However, we still see serological cross-reactivity within filovirus species even with this test. In this study, for example, one Sudan ebolavirus IgG-positive blood sample was also positive for Bundibugyo ebolavirus IgG antibodies. This cross-reactivity has been reported in several other studies (43, 44). These findings should not be over interpreted as the study could be biased towards high-risk groups. In addition, testing for filoviruses using serological tests can potentially overestimate the true level of seropositivity and therefore overestimate risk and exposure due to varying cross reactivity between differing and unvalidated serological assays used in previous studies. We are continuing to classify these serological results with validation assays including viral neutralization in order to confirm if our findings represent true undetected filovirus infections in these communities, or through cross-reaction with other viral infections, or variability in serologic assays performed.