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In the case of African swine fever (ASF) outbreaks in pig farms, EU legislation requires a thorough epidemiological investigation to determine, among other tasks, the extent of infection in the affected farm. The main aim of this study was to implement a reliable sampling strategy to quickly obtain an overview of the extent of ASF virus spread in an affected pig farm. We developed and tested a three-step approach: (i) identification of sub-units within the affected farm, (ii) categorization of sub-units, and (iii) targeted selection of animals for testing. We used commercially available lateral flow devices (LFDs) to detect ASF antigen and antibodies under field conditions and compared them with routinely performed laboratory tests (qPCR, ELISA, IPT). The study was conducted in three commercial farms in Latvia that were affected by ASF in July 2020. One of the affected farms was relatively small with only 31 pigs, whereas the other two were large with 1800 and 9800 animals, respectively. The approach proved to be helpful and practical for efficient and reliably assess the ASF situation on the farm and to identify sub-units within a farm where infected animals are present and sub-units which might (still) be free of infection. This important epidemiological information helps to better estimate the high-risk period and to track the potential spread of infection outside the farm. It allows also to prioritize culling and, if appropriate, to pursue a partial culling strategy taking into account the absence of clinical signs, implemented biosecurity measures, quarantine and negative test results, among others. This might be of interest for large commercial farms where the infection was identified very early and has not yet spread widely. Due to its limited sensitivity, the antigen LFD test is useful for testing animals showing signs of disease., (© 2022 The Authors. Transboundary and Emerging Diseases published by Wiley-VCH GmbH.)

Infection with African swine fever virus (ASFV) causes a highly lethal haemorrhagic disease in domestic and Eurasian wild pigs. Thus, it is a major threat to pig populations worldwide and a cause of substantial economic losses. Recently, less virulent ASFV strains emerged naturally, which showed higher experimental virulence in wild boar than in domestic pigs. The reason for this difference in disease progression and outcome is unclear but likely involves different immunological responses. Unfortunately, besides the importance of CD8α + lymphocytes, little is known about the immune responses against ASFV in suids. Against this background, we used a multicolour flow cytometry platform to investigate the T-cell responses in wild boar and domestic pigs after infection with the moderately virulent ASFV strain 'Estonia2014' in two independent trials. CD4 - /CD8α + and CD4 + /CD8α + αβ T-cell frequencies increased in both subspecies in various tissues, but CD8α + γδ T cells differentiated and responded in wild boar only. Proliferation in CD8α + T cells was found 10 days post infectionem only. Frequencies of T-bet + T cells increased in wild boar but not in domestic pigs. Of note, we found a considerable loss of perforin expression in cytotoxic T cells, 5 and 7 dpi. Both subspecies established a regulatory T-cell response 10 dpi. In domestic pigs, we show increasing levels of ICOS + and CD8α + invariant Natural Killer T cells. These disparities in T-cell responses might explain some of the differences in disease progression in wild boar and domestic pigs and should pave the way for future studies., (© 2021 The Authors. Transboundary and Emerging Diseases published by Wiley-VCH GmbH.)

African swine fever (ASF) is a highly pathogenic viral disease affecting all Suidae, with Ornithodoros moubata complex soft ticks acting as the biological arthropod vectors of the causative agent, African swine fever virus (ASFV). While ASFV is also transmissible via direct contact, pig products and fomites, other arthropods may be involved in virus transmission and persistence. Therefore, we checked various groups of blood-feeding arthropods collected during summer 2017 in wild boar habitats on the Estonian Island of Saaremaa for the presence of ASFV. Saaremaa had the highest ASF infection prevalences in Estonia in 2017, with an incidence of 9% among hunted wild boar. In addition to ASFV, we tested for other selected pathogens. In total, 784 ticks, 6,274 culicoid biting midges, 77 tabanids and 757 mosquitoes were tested as individuals or pools. No ASFV-DNA was found in any of them although about 20% of the tick samples tested positive for swine DNA. By contrast, tick-borne encephalitis virus-RNA was detected in one out of 37 tick pools (2.7%) and Borrelia burgdorferi s.l.-DNA in 20 individual ticks and 17 tick pools (25.2% of all samples). No Schmallenberg virus was detected in the Culicoides specimens. In conclusion, we found no evidence for Ixodes ricinus ticks, Culicoides punctatus and Obsoletus complex biting midges, Aedes spp., Anopheles spp. and Culiseta annulata mosquitoes, and Haematopota pluvialis tabanids playing a role in ASFV transmission in the wild boar population in Estonia., (© 2021 Wiley-VCH GmbH.)

Since African swine fever (ASF) first appeared in the Caucasus region in 2007, it has spread rapidly and is now present in numerous European and Asian countries. In Europe, mainly wild boar populations are affected and pose a risk for domestic pigs. In Asia, domestic pigs are almost exclusively affected. An effective and safe vaccine is not available, and correlates of protection are far from being understood. Therefore, research on immune responses, immune dysfunction and pathogenesis is mandatory. It is acknowledged that T cells play a pivotal role. Thus, we investigated T-cell responses of domestic pigs and wild boar upon infection with the highly virulent ASF virus (ASFV) strain 'Armenia08'. For this purpose, we used a flow cytometry-based multicolour analysis to identify T-cell subtypes (cytotoxic T cells, T-helper cells, γδ T cells) and their functional impairment in ASFV-infected pigs. Domestic pigs showed lymphopaenia, and neither in the blood nor in the lymphoid organs was a proliferation of CD8 + effector cells observed. Furthermore, a T-bet-dependent activation of the remaining CD8 T cells did not occur. In contrast, a T-cell response could be observed in wild boar at 5 days post-inoculation in the blood and in tendency also in some organs. However, this cytotoxic response was not beneficial as all wild boars showed a severe acute lethal disease and a higher proportion died spontaneously or was euthanized at the humane endpoint., (© 2020 The Authors. Transboundary and Emerging Diseases published by Blackwell Verlag GmbH.)

Since the first introduction of African swine fever (ASF) into the European wild boar population in 1957, the question of virus survival in carcasses of animals that succumbed to the disease has been discussed. The causative African swine fever virus (ASFV) is known to be very stable in the environment. Thus, carcasses of infected wild boar could play a major role as ASFV reservoir and thereby help to locally maintain and spread the disease in wild boar populations. To minimize this risk, removal of wild boar carcasses in ASF affected areas is regarded to be crucial for effective disease control. If removal is not feasible, carcasses are usually disposed by burial on the spot to avoid direct contact of wild boar to the infection source. In this study, carcasses of ASFV infected wild boar buried in Lithuania at different time points and locations have been excavated and retested for the presence of infectious ASFV by in vitro assays and for viral genome by qPCR. Soil samples potentially contaminated by body fluids have been additionally tested for viral genome. In seventeen out of twenty burial sites, samples of excavated carcasses were positive for ASFV genome. However, in none of the carcass samples ASFV could be isolated. On seven sites soil samples contained ASF viral DNA. These results unexpectedly negate the long-term persistence of infectious ASFV in wild boar carcasses independent from the burial time. In this context, sensitivity of ASFV isolation from carcass samples versus susceptibility of animals and doses needed for oral inoculation has to be further investigated. Furthermore, research is required to consider alternative ASF infection sources and drivers in the infection cycle among wild boar., (© 2020 The Authors. Transboundary and Emerging Diseases published by Blackwell Verlag GmbH.)

This study targeted the assessment of a potential African swine fever virus (ASFV) carrier state of 30 pigs in total which were allowed to recover from infection with ASFV "Netherlands'86" prior exposure to six healthy sentinel pigs for more than 2 months. Throughout the whole trial, blood and swab samples were subjected to routine virological and serological investigations. At the end of the trial, necropsy of all animals was performed and viral persistence and distribution were assessed. Upon infection, a wide range of clinical and pathomorphological signs were observed. After an initial acute phase in all experimentally inoculated pigs, 66.6% recovered completely and seroconverted. However, viral genome was detectable in blood samples for up to 91 days. Lethal outcomes were observed in 33.3% of the pigs with both acute and prolonged courses. No ASFV transmission occurred over the whole in-contact phase from survivors to sentinels. Similarly, infectious ASFV was not detected in any of the tissue samples from ASFV convalescent and in-contact pigs. These findings indicate that the suggested role of ASFV survivors is overestimated and has to be reconsidered thoroughly for future risk assessments., (© 2018 Blackwell Verlag GmbH.)

African swine fever (ASF) is a notifiable disease with serious socio-economic consequences that has been present in wild boar in the Baltic States and Poland since 2014. An introduction of ASF is usually accompanied by increased mortality, making fallen wild boar and hunted animals with signs of disease the main target for early warning and passive surveillance. It is difficult, however, to encourage hunters and foresters to report and take samples from these cases. A pragmatic and easy sampling approach with quick-drying swabs could facilitate this. In this study, we further evaluated the use of dry blood swabs for the detection of ASFV antibody and genome with samples from animal trials and diagnostic submissions (blood, bone and organs) from Estonia. Compared to serum samples, dried blood swabs yielded 93.1% (95% confidence interval: [83.3, 98.1]) sensitivity and 100% [95.9, 100.0] specificity in a commercial ASFV antibody ELISA. Similarly, the swabs gave a sensitivity of 98.9% [93.4, 100.0] and a specificity of 98.1% [90.1, 100.0] for genome detection by a standard ASFV p72 qPCR when compared to EDTA blood. The same swabs were tested in a VP72-antibody lateral flow device, with a sensitivity of 94.7% [85.4, 98.9] and specificity of 96.1% [89.0, 99.2] compared to the serum ELISA. When GenoTube samples tested in ELISA and LFD were compared, the sensitivity was 96.3% [87.3, 99.5] and the specificity was 93.8% [86.0, 97.9]. This study demonstrates reliable detection of ASFV antibody and genome from swabs. A field test of the swabs with decomposed wild boar carcasses in an endemic area in Estonia also gave promising results. Thus, this technique is a practical approach for surveillance of ASF in both free and endemic areas., (© 2017 Blackwell Verlag GmbH.)

Due to its impact on animal health and pig industry, African swine fever (ASF) is regarded as one of the most important viral diseases of pigs. Following the ongoing epidemic in the Transcaucasian countries and the Russian Federation, African swine fever virus was introduced into the Estonian wild boar population in 2014. Epidemiological investigations suggested two different introductions into the southern and the north-eastern part of Estonia. Interestingly, outbreak characteristics varied considerably between the affected regions. While high mortality and mainly virus-positive animals were observed in the southern region, mortality was low in the north-eastern area. In the latter, clinically healthy, antibody-positive animals were found in the hunting bag and detection of virus was rare. Two hypotheses could explain the different behaviour in the north-east: (i) the frequency of antibody detections combined with the low mortality is the tail of an older, so far undetected epidemic wave coming from the east, or (ii) the virus in this region is attenuated and leads to a less severe clinical outcome. To explore the possibility of virus attenuation, a re-isolated ASFV strain from the north-eastern Ida-Viru region was biologically characterized in European wild boar. Oronasal inoculation led to an acute and severe disease course in all animals with typical pathomorphological lesions. However, one animal recovered completely and was subsequently commingled with three sentinels of the same age class to assess disease transmission. By the end of the trial at 96 days post-initial inoculation, all animals were completely healthy and neither virus nor viral genomes were detected in the sentinels or the survivor. The survivor, however, showed high antibody levels. In conclusion, the ASFV strain from north-eastern Estonia was still highly virulent but nevertheless, one animal recovered completely. Under the experimental conditions, no transmission occurred from the survivor to susceptible sentinel pigs., (© 2017 Blackwell Verlag GmbH.)