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ABSTRACTAfrican swine fever virus (ASFV), a large and complex DNA-virus circulating between soft ticks and indigenous suids in sub-Saharan Africa, has made its way into swine populations from Europe to Asia. This virus, causing a severe haemorrhagic disease (African swine fever) with very high lethality rates in wild boar and domestic pigs, has demonstrated a remarkably high genetic stability for over 10 years. Consequently, analyses into virus evolution and molecular epidemiology often struggled to provide the genetic basis to trace outbreaks while few resources have been dedicated to genomic surveillance on whole-genome level. During its recent incursion into Germany in 2020, ASFV has unexpectedly diverged into five clearly distinguishable linages with at least ten different variants characterized by high-impact mutations never identified before. Noticeably, all new variants share a frameshift mutation in the 3’ end of the DNA polymerase PolX gene O174L, suggesting a causative role as possible mutator gene. Although epidemiological modelling supported the influence of increased mutation rates, it remains unknown how fast virus evolution might progress under these circumstances. Moreover, a tailored Sanger sequencing approach allowed us, for the first time, to trace variants with genomic epidemiology to regional clusters. In conclusion, our findings suggest that this new factor has the potential to dramatically influence the course of the ASFV pandemic with unknown outcome. Therefore, our work highlights the importance of genomic surveillance of ASFV on whole-genome level, the need for high-quality sequences and calls for a closer monitoring of future phenotypic changes of ASFV.

Since September 2020, Germany has experienced the first ever outbreak of African swine fever (ASF). The first known cases occurred exclusively in wild boar in forest areas in Brandenburg and Saxony; in July 2021, infected domestic pigs were also confirmed for the first time. As wild boar are considered the main reservoir for the virus in the European region, an effective interruption of this infection chain is essential. In particular, the removal and safe disposal of infected carcasses and the direct disinfection of contaminated, unpaved ground are priorities in this regard. For the disinfection, highly potent as well as environmentally compatible disinfectants must be used, which are neither influenced in their effectiveness by the soil condition nor by increased organic contamination. Thus, in this study, slaked lime, milk of lime and quicklime (1% to 10% solutions) were selected for efficacy testing against the test virus recommended by the German Veterinary Society (DVG), Modified Vaccinia Ankara virus (MVAV), and ASF virus (ASFV) in conjunction with six different forest soils from Saxony in two different soil layers (top soil and mineral soil) each. In summary, 10% of any tested lime type is able to inactivate both MVAV and ASFV under conditions of high organic load and independent of the water content of the soil. At least a 4 log reduction of the virus titer in all tested forest soil types and layers and by all applied lime types was observed. In conclusion, the high efficacy and suitability of all tested lime products against both viruses and in the presence of high organic load in forest soil can be confirmed and will help to control ASF spread.

African swine fever (ASF) has evolved from an exotic animal disease to a threat to global pig production. An important avenue for the wide-spread transmission of animal diseases is their dissemination through boar semen used for artificial insemination. In this context, we investigated the role of male reproductive organs in the transmission of ASF. Mature domestic boars and adolescent wild boars, inoculated with different ASF virus strains, were investigated by means of virological and pathological methods. Additionally, electron microscopy was employed to investigate in vitro inoculated sperm. The viral genome, antigens and the infectious virus could be found in all gonadal tissues and accessory sex glands. The viral antigen and viral mRNAs were mainly found in mononuclear cells of the respective tissues. However, some other cell types, including Leydig, endothelial and stromal cells, were also found positive. Using RNAScope, p72 mRNA could be found in scattered halo cells of the epididymal duct epithelium, which could point to the disruption of the barrier. No direct infection of spermatozoa was observed by immunohistochemistry, or electron microscopy. Taken together, our results strengthen the assumption that ASFV can be transmitted via boar semen. Future studies are needed to explore the excretion dynamics and transmission efficiency.