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The first report of African swine fever virus (ASFV) genotype II in Italy in 2022 marked the beginning of a significant invasion in at least eight Italian regions with different infection clusters. In this study, we used the multi-gene approach to investigate the epidemiological associations between ASFV strains causing cases and outbreaks in wild boar and pigs in Italy from January 2022 to the end of 2023. Our results confirm that all the tested ASFV-positive Italian samples belonged to genotype II and show high homology with genotype II ASFV sequences previously collected in Eurasian countries. Molecular characterization revealed the presence of four genetic groups in Italy. The majority of African swine fever (ASF) samples analyzed in the current study (72%) belonged to genetic group 3, which was the most representative in Europe. The results also provide evidence of the prevalence of genetic group 19 (15.9%). In addition, we identified new putative genetic groups, genetic group 25 (9.1%) and genetic group 26 (3.0%), which have never been described before. This is the first detailed report on the molecular characterization of more than 130 ASFV strains circulating in Italy.

African swine fever virus (ASFV) causes a devastating disease affecting domestic and wild pigs. ASF was first introduced in Sardinia in 1978 and until 2019 only genotype I isolates were identified. A remarkable genetic stability of Sardinian ASFV isolates was described, nevertheless in 2019 two wild boar isolates with a sustained genomic deletion (4342 base pairs) were identified (7303WB/19, 7212WB/19). In this study, we therefore performed in vitro experiments with monocyte-derived macrophages (moMФ) to unravel the phenotypic characteristics of these deleted viruses. Both 7303WB/19 and 7212WB/19 presented a lower growth kinetic in moMФ compared to virulent Sardinian 26544/OG10, using either a high (1) or a low (0.01) multiplicity of infection (MOI). In addition, flow cytometric analysis showed that both 7303WB/19 and 7212WB/19 presented lower intracellular levels of both early and late ASFV proteins. We subsequently investigated whether deleted virus variants were previously circulating in wild boars in Sardinia. In the four years preceding the last genotype I isolation (February 2015-January 2019), other eight wild boar isolates were collected, all belonging to p72 genotype I, B602L subgroup X, but none of them presented a sustained genomic deletion. Overall, we observed the deleted virus isolates in Sardinia only in 2019, at the end of a strong eradication campaign, and our data suggest that it might possess an attenuated phenotype in vivo. A better understanding of ASFV evolution in endemic territories might contribute to development of effective control measures against ASF., (© 2024. The Author(s).)

African swine fever (ASF) is a disease that is a growing threat to the global swine industry. Regulations and restrictions are placed on swine movement to limit the spread of the virus. However, these are costly and time-consuming. Therefore, this study aimed to determine if high-pressure processing (HPP) sanitization techniques would be effective against the ASF virus. Here, it was hypothesized that HPP could inactivate or reduce ASF virus infectivity in tissue homogenates. To test this hypothesis, 30 aliquots of each homogenate (spleen, kidney, loin) were challenge-infected with the Turin/83 strain of ASF, at a 10 7.20 median hemadsorption dose (HAD) 50 /mL. Subsequently, eight aliquots of each homogenate were treated with 600 millipascal (600 MPa) HPP for 3, 5, and 7 min. Six untreated aliquots were used as the controls. Virological results showed a reduction in the viral titer of more than 7-log. These results support the validity of the study hypothesis since HPP treatment was effective in inactivating ASFV in artificially prepared samples. Overall, this study suggests the need for further investigation of other ASFV-contaminated meat products., Competing Interests: The authors declare no conflicts of interest.

African Swine Fever (ASF), a hemorrhagic disease with a high mortality rate in suids, is transmitted via direct and indirect contact with infectious animals and contaminated fomites, respectively. ASF reached Europe in 2014, affecting 14 of the 27 EU countries including, recently, the Italian peninsula. The fast and unprecedented spread of ASF in the EU has highlighted gaps in knowledge regarding transmission mechanisms. Fomites, such as contaminated clothing and footwear, farming tools, equipment and vehicles have been widely reported in the spread of ASF. The absence of available vaccines renders biosecurity measures, cleaning and disinfection procedures an essential control tool, to a greater degree than the others, for the prevention of primary and secondary introductions of ASF in pig farms. In this review, available data on the virucidal activity of chemical compounds as disinfectants against the ASF virus (ASFV) are summarized together with laboratory methods adopted to assess the virucidal activity.

A number of animal diseases can be transmitted to pigs via pork meat and pork products imported from infected areas. Therefore, feeding of swill to pigs is regulated or prohibited in many swine-rearing countries. African swine fever is one of the major porcine diseases recognized as significant in this transmission pathway. Assessment of disease risks associated with pork products requires knowledge about the viral load in the original material and for how long infectious virus can be recovered from the resulting product. In this work, we present the effect of the dry curing process on the inactivation of African swine fever virus (ASFV) in three different Italian dry-cured meat products prepared from experimentally infected pigs slaughtered at the peak of viremia. The meat products were processed using commercial methods and industrial procedures currently being followed in Italy. Samples collected at predetermined intervals during processing were analyzed for virus survival by virus isolation and animal inoculation. ASFV was detected by in vivo experiments for up to 18, 60, and 83 days of curing in Italian salami, pork belly, and loin, respectively. These data provide valuable information for the pork processing industry when planning the export of these products., (Copyright © 2018 Elsevier B.V. All rights reserved.)

African swine fever (ASF) virus is a DNA virus responsible for a severe haemorrhagic fever in pigs, which (still in the absence of vaccination strategies) results in high mortality rates. Herein, we present a biosensor-based method for the detection of ASF viral DNA in the blood of pigs. The biosensor exploits a single-strand DNA probe with locked nucleic acid nucleotides (LNA) substitutions as the complementary recognition element for the conserved region of vp72 gene of ASF virus. The biosensor was calibrated using qPCR-quantified ASF viral DNA extracted from the blood of pigs experimentally infected with the virulent Italian isolate 49/08, genotype I. Globally, the proposed biosensor showed good sensitivity and specificity, with the limits of detection (LOD) and quantification (LOQ) being 178 and 245 copies/μL of genomic ASF viral DNA, respectively. The reversible nature of the interaction between the DNA/LNA probe and the target DNA sequence granted multiple rapid analyses, with up to 40 analyses per single surface possible, and a single test requiring approximately 5 min. When applied to non-amplified DNA extracts from the blood of field-infected pigs, the assay discriminated between ASFV-infected and ASFV non-infected animals, and allowed the rapid quantification of ASF viral DNA, with values falling in the range 373-1058 copies/μL of genomic ASFV DNA. In this range, excellent correlation was observed between the results of this biosensor and OIE-approved qPCR. This method represents a promising screening assay for preliminary ASF diagnosis, having the major advantages in the relative rapidity, ease-of-use, the reusability of the sensing surface, and low cost per single test., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Three discrete regions of the African swine fever virus (ASFV) were analysed in the genomes of a wide range of isolates collected from wild and domestic pigs in Sardinia, over a 31-year period (1978-2009). The analysis was conducted by genotyping based on sequence data from three single copy ASF genes. The E183L gene encoding the structural protein p54 and part of the gene encoding the p72 protein were used to delineate genotypes, before intra-genotypic resolution of viral relationships by analysis of tetramer amino acid repeats within the hypervariable central variable region (CVR) of the B602L gene. The data revealed that these isolates did not show significant variation in their p72 and p54 sequence when compared between different isolates showing a remarkable genetic stability of these genome regions. In particular, the phylogeny revealed that all the Sardinian isolates belong to the same largest and most homogeneous p72 genotype I together with viruses from Europe, South America, the Caribbean and West Africa, and p54 genotype Ia which comprises viruses from Europe and America. The analysis of B602L gene revealed a minor difference in the number of tetramer repeats, placing the Sardinian isolates into two clusters, accordingly to their temporal distribution, namely sub-group III and sub-group X, this latter showing a deletion of 12 tetramer repeats located in the centre of the array. The genetic variation of this fragment suggests that one sub-group could be derived from the other supporting the hypothesis of a single introduction of ASFV in Sardinia.

Simple Summary: The African Swine Fever Virus (ASFV) is a DNA virus belonging to the Asfivirus genus and Asfarviridae family. In recent years, ASF has grown to be a significant concern for the pig industry worldwide. The ever-growing handling of ASFV pathological material in laboratories, for either diagnostic or research activities, requires particular caution to avoid accidental virus release from laboratories with consequent detrimental economic and environmental effects. This paper aims to describe the Italian National Reference Laboratory (NRL) efforts to develop a biological containment tool for ASF laboratories and animal facilities. The easy-to-apply recommendations focusing on the risk assessment system and internal audits used for enhancing Italian biosafety and biosecurity measures are reported. The African Swine Fever Virus (ASFV) is a DNA virus of the Asfarviridae family, Asfivirus genus. It is responsible for massive losses in pig populations and drastic direct and indirect economic impacts. The ever-growing handling of ASFV pathological material in laboratories, necessary for either diagnostic or research activities, requires particular attention to avoid accidental virus release from laboratories and its detrimental economic and environmental effects. Recently, the Commission Delegated Regulation (EU) 2020/689 of 17 December 2019 repealed the Commission Decision of 26 May 2003 reporting an ASF diagnostic manual (2003/422/EC) with the minimum and supplementary requirements for ASF laboratories. This decision generated a regulatory gap that has not been addressed yet. This paper aims to describe the Italian National Reference Laboratory (NRL) efforts to develop an effective and reliable biological containment tool for ASF laboratories and animal facilities. The tool consists of comprehensive and harmonized structural and procedural requirements for ASF laboratories and animal facilities that have been developed based on both current and repealed legislation, further entailing a risk assessment and internal audit as indispensable tools to design, adjust, and improve biological containment measures. [ABSTRACT FROM AUTHOR]

African swine fever (ASF) is responsible for important socio-economic effects in the global pig industry, especially for countries with large-scale piggery sectors. In January 2022, the African swine fever virus (ASFV) genotype II was identified in a wild boar population in mainland Italy (Piedmont region). This study describes the molecular characterization, by Sanger and next-generation sequencing (NGS), of the first index case 632/AL/2022 and of another isolate (2802/AL/2022) reported in the same month, in close proximity to the first, following multiple ASF outbreaks. Phylogenetic analysis based on the B646L gene and NGS clustered the isolates 632/AL/2022 and 2802/AL/2022 within the wide and most homogeneous p72 genotype II that includes viruses from European and Asian countries. The consensus sequence obtained from the ASFV 2802/AL/2022 isolate was 190,598 nucleotides in length and had a mean GC content of 38.38%. At the whole-genome level, ASF isolate 2802/AL/2022 showed a close genetic correlation with the other representative ASFV genotype II strains isolated between April 2007 and January 2022 from wild and domestic pigs in Eastern/Central European (EU) and Asian countries. CVR subtyping clustered the two Italian ASFV strains within the major CVR variant circulating since the first virus introduction in Georgia in 2007. Intergenic region I73R-I329L subtyping placed the Italian ASFV isolates within the variant identical to the strains frequently identified among wild boars and domestic pigs. Presently, given the high sequence similarity, it is impossible to trace the precise geographic origin of the virus at a country level. Moreover, the full-length sequences available in the NCBI are not completely representative of all affected territories. [ABSTRACT FROM AUTHOR]

African swine fever (ASF) is a highly contagious and lethal viral disease of pigs and wild boars, which is enzootic in many African countries and on the Italian island of Sardinia, where it has been present since 1978. Previous genetic analyses of Sardinian ASF virus (ASFV) isolates have revealed that they all belong to p72 genotype I, with only minor sequence variations. However, these studies examined only a few selected genes. To distinguish between these closely related isolates and better investigate ASFV evolution in Sardinia, we sequenced the complete genomes of 12 Sardinian ASFV isolates collected between 1978 and 2012, and compared them with 47/Ss/2008 and 26544/OG10. Most of the observed changes occurred in a time‐dependent manner; however, their biological significance remains unclear. As a whole, our results demonstrate the remarkable genetic stability of these strains, supporting a single‐source introduction of the virus. [ABSTRACT FROM AUTHOR]

African swine fever virus (ASFV), the cause of a devastating disease affecting domestic and wild pigs, has been present in Sardinia since 1978. In the framework of the regional ASF eradication plan, 4484 illegal pigs were culled between December 2017 and February 2020. The highest disease prevalence was observed in the municipality with the highest free-ranging pig density, and culling actions drastically reduced ASFV circulation among these animals. ASFV-antibody were detected in 36.7% of tested animals, which were apparently healthy, thus, the circulation of low-virulence ASFV isolates was hypothesized. ASFV genome was detected in 53 out of 2726 tested animals, and virus isolation was achieved in two distinct culling actions. Two ASFV haemadsorbing strains were isolated from antibody-positive apparently healthy pigs: 55234/18 and 103917/18. Typing analysis revealed that both isolates belong to p72 genotype I, B602L subgroup X; phylogenetic analysis based on whole genome sequencing data showed that they were closely related to Sardinian ASFV strains collected since 2010, especially 22653/Ca/2014. Our data suggested the absence of immune-escaped ASFV variants circulating among free-ranging pigs, indicating that other elements contributed to virus circulation among these animals. Understanding factors behind disease persistence in endemic settings might contribute to developing effective countermeasures against this disease. [ABSTRACT FROM AUTHOR]