Risk factors of African swine fever virus in suspected infected pigs in smallholder farming systems in South-Kivu province, Democratic Republic of Congo

Pig farming has become very popular across Africa continent as a means of increasing food supply, income, and employment. The economic potentials of this farming sector are related to some of its characteristics that are distinct from farming of other domestic animals, including prolificacy (up to 20 piglets per litter), relatively short generation time, and high feed to flesh conversion rate, resulting in high production turnover for sale and consumption [1]. However, this sector faces several constraints, with the associated disease burden being a major problem [2]. One of the most common swine diseases is African swine fever (ASF), a complex, contagious, lethal, and economically important viral disease of domestic pigs that causes hemorrhagic fever and leads to considerable economic loss due to morbidity and death of animals, and for which no effective vaccine is available [3, 4]. In addition, the disease threatens food supply security by limiting the availability of animal protein, particularly in regions where alternative livestock production is a challenge.

ASF is endemic in Sub-Saharan Africa and has been reported in more than 26 countries [5]. Outbreaks of ASF have serious social and economic implications, both in areas where it is endemic and where it has been introduced, due to its high mortality rate, which can approach 100 percent. In several endemic African countries, ASF affects smallholder pig producers primarily due to a lack of biosecurity infrastructure and a lack of effective prevention and biosecurity measures [6].

The clinical presentation of ASF in domestic pigs depends mainly on the viral isolate's virulence, the pig breed, the infective dose, and the route of infection [7, 8]. Typical signs in infected animals include severe depression, high fever (40–42°C), cyanotic skin on ears, bleeding from bodily orifices, breathing difficulty, nasal discharge, abortion in pregnant sows, and coma and death [9].

Susceptible animals are infected through either the sylvatic or domestic cycles or between interaction between the sylvatic cycle and domestic pigs [10]. In the sylvatic cycle, the ASF virus circulates between wild pigs, the natural reservoir, and soft ticks of the Ornithodoros spp. [11]. Ticks transmit the virus to common warthogs (Phacochoerus spp.) while the ticks feed on the wild pig's blood. The domestic cycle occurs when the virus is transmitted from one infected domestic pig to another, either by direct contact or through ingestion of an infected pork product, and without the involvement of sylvatic cycle hosts or arthropod vectors [11, 12]. Transmission between the sylvatic cycle and domestic pigs occurs when domestic pigs are infected through tick bites, which occurs in those areas of Africa where warthogs are present. Furthermore, an infection can occur when domestic pigs are fed ASF virus (ASFV)-contaminated warthog carcasses or come in contact with warthog feces [13]. Prevention and control methods rely mainly on early diagnosis, restriction of the movement of pigs and pig products, strict application of biosecurity measures, and culling of infected swine herds [14]. Some of the mitigative measures against ASF spread in domestic pig farms are the adoption of biosecurity practices, proper disposal of carcasses, and efficient reporting of suspected outbreaks.

The Democratic Republic of Congo (DRC) has the second-largest pig population in central Africa, after Cameroon, with a standing pig population of 1 million [15]. These are distributed unequally among the different provinces of the country. Although pig farming is an important livestock activity among farmers in several rural areas of South Kivu province, this region is one of the provinces with a small pig population. The relatively low population is because of the frequent occurrence of confirmed and suspected ASF outbreaks and the resulting high mortality. To date, most of the ASF outbreak reports have been based on clinical diagnosis without the use of any confirmatory testing; moreover, there is a reluctance among farmers to report suspected outbreaks [15].

The South Kivu provinces of the DRC have a pig population of approximately 66,636 [15]. Smallholder pig farmers have faced a serious challenge due to the regular occurrence of hemorrhagic fever outbreaks, leading to high domestic pig mortality and a notable negative effect on productivity. Pigs are raised mostly by smallholders in various farming environments ranging from municipal to rural areas [16], and such farmers can easily sell their pigs to cover school fees, medical bills, and other family requirements [17]. In this region, 3 pig-farm management systems, intensive, tethering, and extensive, are used [16, 17]. Intensive pig farming involves housing pigs in pens to restrict their movement and provide them with feed and water. In the tethering system, pigs are tied by a rope to a post to restrict their movement within a specified radius. In the extensive system, pigs are left to scavenge in the open to obtain food and water, which places them at a very high risk of infections.

A previous study conducted in South Kivu province focused mainly on genetic characterization of isolated ASFV strains, based on sequence analysis of the major capsid protein (p72) [18]. That study reported the circulation of ASFV genotype IX, which was identical to the isolates responsible for outbreaks in other DRC provinces with ASF outbreaks [19]. The same genotype has been reported in Tanzania, which borders the South Kivu province of DRC. This indicates that ASFV can plausibly move between DRC provinces, and movements between Tanzania and South Kivu might have occurred. Despite the devastating effects of swine diseases, the lack of routine diagnostic and the poor husbandry practices observed in smallholder pig farms, limited studies have been conducted in South Kivu to determine the infection rate and prevalence of ASF in suspected clinically infected pig farms by using a sensitive diagnostic method (such as real-time polymerase chain reaction [RT-PCR]). Such information is important when differentiating ASFV infection from other pig diseases with similar clinical manifestations and when investigating epidemiology in South Kivu. Therefore, this study aimed to identify the risk factors associated with the occurrence of ASF and its spread among smallholder pig farmers. The results will contribute to the elucidation of ASF transmission, which is necessary for the implementation of proper control measures.

This study was conducted from November 2018 to January 2019, a period in which there was no official report of an ASF outbreak in the sampled districts. The purpose of the study was to determine the proportion of ASFV-positive animals among the symptomatic domestic pigs sampled and determine the associated potential risk factors for ASFV spread among domestic pigs.

The limited access to formal education for the majority of farmers interviewed during this investigation and the lack of good husbandry practices might negatively affect pig-farm management and may favor disease occurrence. This finding agrees with previous studies conducted in Uganda [2, 14], which showed that insufficient knowledge of husbandry practices and pig management were major constraints on pig farmers.

An average ASFV PCR-positive rate of 18.4% was detected in this study, close to that in a study conducted in Kenya, where 28% were positive [24] but much lower than the 42.8% reported in another study in symptomatic pigs in Cameroon [25]. Identification of symptomatic pigs that were ASFV PCR-positive in South Kivu indicates current or previous infections in the province, suggesting that outbreaks might be occurring in the region; however, a proper reporting system is lacking, so outbreaks may not be officially recorded. Similar results were obtained in previous studies conducted in the same region and in some other parts of the DRC where ASFV viral DNA was detected in domestic pigs with ASF-specific clinical signs [17, 26].

The highest level of PCR-positive pigs was observed in the Uvira district, followed by that in the Walungu district. This may be related to the use of a free-range system, which is practiced in Uvira and some parts of Walungu and allows pigs to move around the farm area. Pigs in a free-range system are far more likely to have more frequent contact with infected animals or materials that may facilitate infection. Logistic regression analysis revealed that the feeding of pigs with swill in pen, the presence of pigs of mixed ages in the same pen, and the introduction of a new animal to a farm were the most prominent risk factors associated with ASFV positivity in South Kivu province. The swill feeding practice, common in most pig-keeping households, increases the risk of consuming ASFV-contaminated products. This linkage has been suggested in previous studies conducted in Uganda and Kenya [14], which showed that swill feeding is a potential risk factor for ASF transmission, particularly where many farms in peri-urban areas practice the same system, probably because of their proximity to many restaurants and hotels. However, a study conducted by Allaway et al. [27] found that feeding kitchen leftovers was not a potential risk factor in the spread of ASF in Malawi. Regardless, treating swill before feeding can be an efficient way of preventing the introduction of ASFV in pig farms.

Mixing of pigs of different ages in the same pen as a factor influencing ASFV transmission is likely due to direct pig-to-pig contact or spreading through fomites, especially in confined farms with poor or no biosecurity measures. Similar results were reported in Sardinia and Madagascar [28, 29]. The introduction of new pigs from another farm to an existing herd (without a quarantine area) is an obvious risk factor, as ASFV can be introduced through such a route. The surveys conducted in this study revealed that pig keepers usually buy pigs from sellers or neighboring farms without knowing the animals' health status, and such transactions are followed by the introduction of the newly purchased pig into their herd. This practice is also conducted for breeding purposes. The farmer may borrow a boar, or a sow in heat may be sent to a boar for mating, leading to the spread of ASF. This approach was also reported in studies in Uganda [30] and Tanzania [31]. Logistic regression analysis revealed that the presence of male pigs and the used of an in-pen breeding system are less likely to be infected by ASF; thus, these were considered protective factors. These findings support those in a previous study in Uganda where farm perimeter fencing was one of the protective factors against ASF infection [32].

However, the epidemiology and risk factor information from this study are different from findings in a similar study carried out in Uganda [32], which revealed that prompt disposal of dead pigs on farms, wild animals present in the village, and farmers obtaining drugs from stockists to be the major risk factors associated with ASF outbreaks in Uganda. These differences could be attributed to diversity among the 6 geographical districts included in the present study compared to the previous study in Uganda, in which samples were from only 3 districts. Furthermore, this study was conducted over a shorter period (3 months) than that of the study in Uganda (5 months).

Our findings are also different from those reported in Nigeria [33], where the sharing of tools was negatively associated with the spread of infection, whereas it was positively associated with ASF infection in the current study. This could be due to differences in the implementation of biosecurity measures between the 2 countries. For example, it is possible that tools shared between pig farms in Nigeria are regularly washed and disinfected; hence, the observation of a negative association (ASF protection).

Despite the informal approach to most livestock farming, particularly for small-scale pig farming in rural areas, the security issues in several villages in the study region, the lack of transport to reach some areas due to road damage, and the lack of up to date animal registration lists, the data for the 6 districts included in this study may not be representative of all pig farms in the study region. Therefore, the results obtained may not be conclusive. Further study covering all of the districts of South Kivu is recommended.

In conclusion, most ASF studies carried out in South Kivu province (DRC) have focused on the clinical diagnosis and molecular characterization of ASFV. This study, however, highlights molecular-based ASFV diagnosis and describes important potential risk factors for the spread of ASF. Such factors may contribute to the regular occurrence of ASF outbreaks and most importantly may result in maintenance of an ASFV population in South Kivu. The observed ASFV infectious rate in clinically infected pigs is evidence of the active circulation of ASFV in a domestic pig population. Improved biosecurity, including fencing, strict control of feed sources, foot baths, and remote quarantine areas for new pigs (with foot baths), are important recommended practices for the control of ASFV transmission in South Kivu province, DRC.

Supplementary Table 1

Demographic characteristics during the field survey and the number of farms and animals in each district

Supplementary Table 2

Characteristics of pigs screened according to sex, age, breed, breeding system and geographical region

Supplementary Fig. 1

PCR amplification of the B646L gene encoding the p72 protein. A 1.5% agarose gel showing the bands obtained using diagnostic primers targeting the central portion of the p72 gene of ASFV. Lane M is 1 Kb plus (Thermo Fisher Scientific) molecular weight DNA marker; Lanes 1, 2, 3, 4, 5, 6, 7, 8, and 9 are selected positive samples showing an approximately 257 bp band size, Lane NC is the negative control; and lane PC is a positive control.