African swine fever virus (ASFV) was first reported in eastern Europe/Eurasia in 2007. Continued spread of ASFV has placed central European countries at risk, and in 2014, ASFV was detected in Lithuania and Poland. Sequencing showed the isolates are identical to a 2013 ASFV from Belarus but differ from ASFV isolated in Georgia in 2007.

A highly sensitive and specific real-time PCR method was developed for the reliable and rapid detection of African swine fever virus (ASFV). The method uses a commercial Universal Probe Library (UPL) probe combined with a specifically designed primer set to amplify an ASFV DNA fragment within the VP72 coding genome region. The detection range of the optimized UPL PCR technique was confirmed by analysis of a large panel (n = 46) of ASFV isolates, belonging to 19 of the 22 viral p72 genotypes described. No amplification signal was observed when closely clinically related viruses, such as classical swine fever, or other porcine pathogens were tested by this assay. The detection limit of the UPL PCR method was established below 18 DNA copies. Validation experiments using an extensive collection of field porcine and tick samples (n = 260), coming from Eastern and Western African regions affected by ASF, demonstrated that the UPL PCR technique was able to detect over 10% more positive samples than the real-time TaqMan PCR test recommended in the OIE manual, confirming its superior diagnostic sensitivity. Clinical material collected during experimental infections with different ASFV p72 genotypes was useful for assuring both the capacity of the UPL PCR for an early viral DNA detection and the competence of the technique to be applied in any ASF diagnostic target sample. The reliability and robustness of the UPL PCR was finally verified with a panel of ASFV-infected clinical samples which was repeatedly tested at different times. Additionally, an internal control PCR assay was also developed and standardized using UPL probes within the endogenous β-actin gene. Finally, the complete study offers a new validated real-time PCR technique, by means of a standardized commercial probe, providing a simple, rapid and affordable test, which is ready for application in the routine diagnosis of ASF.

A frican swine fever (ASF) is a complex and lethal viral disease affecting swine and has a significant socioeconomic impact on both the developed and developing world. It has a major negative effect on national, regional, and international trade and constrains pig production in affected areas. The devastating acute form of the disease is characterized, among other features, by functional and congestive-hemorrhagic disorders of the digestive and respiratory systems and causes around 100% mortality in infected pigs (1). Both European wild boars (Sus scrofa) and feral pigs are susceptible and exhibit clinical signs and mortality rates similar to those of domestic pigs. In contrast, African wild pigs (Phacochoerus and Potamochoerus spp.) are resistant to the disease (2-10).The causative agent of the disease, the ASF virus (ASFV), is a large double-stranded DNA virus and the only member of the Asfarviridae family, genus Asfivirus (11, 12). The virus genome is 170 to 192 kb long (13-17). ASF is endemic in sub-Saharan Africa, where it was first described in 1921 (18). Several outbreaks have occurred since then in Europe and South and Central America. In most non-African countries, the disease has been successfully eradicated, the only exception being Sardinia (Italy), where the disease is still endemic (19,20). In April 2007, the disease spread from East Africa to the Republic of Georgia (21), and outbreaks occurred in Armenia, Azerbaijan, and the Russian Federation (22). The ongoing spread of ASFV into adjacent eastern European countries, such as Ukraine (23,24) and Belarus (25), and the situation in Russia affecting both wild boars and domestic pigs placed neighboring areas in the European Union (EU) at risk for the spread of ASFV. The first cases of ASF in wild boars in Lithuania and Poland were reported in early 2014 in areas bordering . According to the World Organisation for Animal Health (OIE), during 2014, nearly 260 ASF cases or outbreaks in wild boars and domestic pigs were detected in EU countries (Latvia, Lithuania, Estonia, and Poland). This situation, combined with the uncertainty present in Belarus, has created a permanent risk of reintroducing ASF into the EU via wild boars or the illegal trade of contaminated pork products and waste (31).No vaccine is available to prevent ASF infection. The control and eradication measures applicable are based on classical disease control methods, including surveillance, epidemiological investigation, tracing of pigs, and stamping out in infected holdings. Citation Gallardo C, Nieto R, Soler A, Pelayo V, Fernández-Pinero J, MarkowskaDaniel I, Pridotkas G, Nurmoja I, Granta R, Simón A, Pérez C, Martín E, Fernández-Pacheco P, Arias M. 2015. Assessment of African swine fever diagnostic techniques as a response to the epidemic outbreaks in eastern European Union countries: how to improve surveillance and control programs.

An experimental infection was conducted to evaluate horizontal transmission, clinical, virological and humoral response induced in domestic pigs infected with African swine fever (ASF) genotype II virus circulating in 2014 into the European Union (EU). Ten naive pigs were placed in contact with eight pigs experimentally inoculated with the Lithuanian LT14/1490 ASF virus (ASFV) responsible for the first ASF case detected in wild boar in Lithuania in January 2014. Clinical examination and rectal temperature were recorded each day. Blood sampling from every animal was carried out twice weekly. Blood samples were examined for presence of ASF virus-specific antibodies and for determining the ASFV viral load. From the obtained results, it was concluded that the Lithuanian ASFV induced an acute disease which resulted in 94, 5% mortality. The disease was easily detected by real-time PCR prior to the onset of clinical signs and 33% of the animals seroconverted. All findings were in accordance with observations previously made in domestic pigs and wild boar when infected with ASF genotype II viruses characterized by a high virulence. One in-contact pig remained asymptomatic and survived the infection. The role of such animals in virus transmission would need further investigation.

The role of the ancestral sylvatic cycle of the African swine fever virus (ASFV) is not well understood in the endemic areas of eastern Africa. We therefore analysed the ASF infection status on samples collected from 51 free-ranging warthogs (Phacocherus africanus) and 1576 Ornithodorus porcinus ticks from 26 independent warthog burrows at a single ranch in Kenya. Abattoir samples from 83 domestic pigs without clinical symptoms, originating from specific locations with no recent reported ASF outbreaks were included in this study. All samples were derived from areas of central Kenya, where ASF outbreaks have been reported in the past. Infection with ASFV was confirmed in 22 % of O. porcinus pools, 3.22 % of adult warthog serum samples and 49 % of domestic pig serum samples by using p72-based PCR. All of the warthog sera were positive for anti-ASFV antibodies, investigated by using ELISA, but none of the domestic pig sera were positive. Twenty O. porcinus-, 12 domestic pig-and three warthogderived viruses were genotyped at four polymorphic loci. The ASFV isolates from ticks and domestic pigs clustered within p72 genotype X. By contrast, ASF viruses genotyped directly from warthog sera, at same locality as the tick isolates, were within p72 genotype IX and genetically similar to viruses causing recent ASF outbreaks in Kenya and Uganda. This represents the first report of the co-existence of different ASFV genotypes in warthog burrow-associated ticks and adult wild warthogs. The data from this and earlier studies suggest transfer of viruses of at least two different p72 genotypes, from wild to domestic pigs in East Africa.

Summary African swine fever (ASF) has persisted in Eastern Europe since 2007, and two endemic zones have been identified in the central and southern parts of the Russian Federation. Moderate‐ to low‐virulent ASF virus isolates are known to circulate in endemic ASF‐affected regions. To improve our knowledge of virus transmission in animals recovered from ASF virus infection, an experimental in vivo study was carried out. Four domestic pigs were inoculated with the NH/P68 ASF virus, previously characterized to develop a chronic form of ASF. Two additional in‐contact pigs were introduced at 72 days post‐inoculation (dpi) in the same box for virus exposure. The inoculated pigs developed a mild form of the disease, and the virus was isolated from tissues in the inoculated pigs up to 99 dpi (pigs were euthanized at 36, 65, 99 and 134 dpi). In‐contact pigs showed mild or no clinical signs, but did become seropositive, and a transient viraemia was detected at 28 days post‐exposure (dpe), thereby confirming late virus transmission from the inoculated pigs. Virus transmission to in‐contact pigs occurred at four weeks post‐exposure, over three months after the primary infection. These results highlight the potential role of survivor pigs in disease maintenance and dissemination in areas where moderate‐ to low‐virulent viruses may be circulating undetected. This study will help design better and more effective control programmes to fight against this disease.

African swine fever (ASF) has had significant economic and social impact in Nigeria since 1997. However, there has been no effective national response to bring it under control. In this report, we confirm that ASF is still prevalent and widespread in Nigeria. Results from both serosurveillance and virological analyses indicated that ASF is present in most of the agro-ecological zones of the country. Nine per cent (9%) of serum samples and 48% of tissue samples were positive for ASF virus antibody and genome, respectively. Areas with high pig-related activities (marketing, consumption and farming) have higher prevalences compared with areas with less pig activities. Farm-gate buyers, marketing systems and transport of untested pigs within the country assist with the circulation of the virus. Only by putting in place a comprehensive routine surveillance and testing system, reorganizing the market and transportation systems for pigs, implementing on-farm bio-security protocols and considering the option of compensation will it be possible to achieve a significant reduction in ASF prevalence in Nigeria.

African swine fever virus p72 genotype IX, associated with outbreaks in eastern Africa, is cocirculating in the Republic of the Congo with West African genotype I. Data suggest that viruses from eastern Africa are moving into western Africa, increasing the threat of outbreaks caused by novel viruses in this region.