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SUMMARYA haemadsorption microtest for African swine fever (ASF) virus is described. This assay is as sensitive and its response is faster than the conventional assay which uses huffy coat cultures in Leighton tubes. The method can also process a larger number of samples by using smaller amounts of swine blood and laboratory space. A plaque assay for ASF virus adapted to grow in VERO cells gives a titre similar to that obtained using the haemadsorption microtest. In both the micromethod and the plaque assay infection may be produced by a single infective particle.

African swine fever virus (ASFV) encodes a novel DNA polymerase, constituted of only 174 amino acids, belonging to the polymerase (pol) X family of DNA polymerases. Biochemical analyses of the purified enzyme indicate that ASFV pol X is a monomeric DNA-directed DNA polymerase, highly distributive, lacking a proofreading 3-5-exonuclease, and with a poor discrimination against dideoxynucleotides. A multiple alignment of family X DNA polymerases, together with the extrapolation to the crystal structure of mammalian DNA polymerase ␤ (pol ␤), showed the conservation in ASFV pol X of the most critical residues involved in DNA binding, nucleotide binding, and catalysis of the polymerization reaction. Therefore, the 20-kDa ASFV pol X most likely represents the minimal functional version of an evolutionarily conserved pol ␤-type DNA polymerase core, constituted by only the "palm" and "thumb" subdomains. It is worth noting that such an "unfingered" DNA polymerase is able to handle templated DNA polymerization with a considerable high fidelity at the base discrimination level. Base excision repair is considered to be a cellular defense mechanism repairing modified bases in DNA. Interestingly, the fact that ASFV pol X is able to conduct filling of a single nucleotide gap points to a putative role in base excision repair during the ASFV life cycle.Despite the variety of existing DNA polymerases, there are a few basic principles that are common to all these enzymes, irrespective of their role either in DNA replication or in DNA repair. The basic chemistry of each individual reaction always involves a pair of divalent metal ions that are coordinated by carboxylate residues. Such a two-metal ion mechanism, originally proposed by Beese and Steitz (1) and probably extrapolative to all nucleotidyltransferases (2-4), appears to be either evolutionarily conserved or acquired by convergent evolution of nonhomologous proteins. In addition to the general deoxynucleotidyl transfer mechanism, it appears that some structural convergency could apply also for the interaction with DNA, a common substrate; an overall view of the crystal structures available for DNA-dependent polymerases always shows a hand-shaped structure, with "thumb," "palm," and "fingers" subdomains, defining at least one cleft for holding DNA (5-12). Moreover, both DNA replicases and DNA repair enzymes are often multienzymatic proteins, having built-in nucleolytic activities that exist as individual structural modules, separated from the polymerization domain. Thus, the paradigmatic Escherichia coli DNA polymerase I (pol I) 1 has a proofreading 3Ј-5Ј-exonuclease, and a 5Ј-3Ј-exonuclease to remove the RNA from the Okazaki fragments (13). Most DNA replicases are also endowed with a proofreading 3Ј-5Ј-exonuclease domain, having an evolutionarily conserved pol I-type active site (14). Reverse transcriptases have an RNase H activity, required for second strand DNA synthesis (reviewed in Telesnitsky and Goff (15)). Even pol ␤, the smallest of the known DNA polymerases (39 kDa)...