Duck hepatitis A virus (DHAV), an avian picornavirus, causes high-mortality acute disease in ducklings. Among the three serotypes, DHAV-1 is globally distributed, whereas DHAV-2 and DHAV-3 serotypes are chiefly restricted to Southeast Asia. In this study, we analyzed the genomic evolution of DHAV-1 strains using extant GenBank records and genomic sequences of 10 DHAV-1 strains originating from a large disease outbreak in 2004–2005, in Hungary. Recombination analysis revealed intragenotype recombination within DHAV-1 as well as intergenotype recombination events involving DHAV-1 and DHAV-3 strains. The intergenotype recombination occurred in the VP0 region. Diversifying selection seems to act at sites of certain genomic regions. Calculations estimated slightly lower rates of evolution of DHAV-1 (mean rates for individual protein coding regions, 5.6286 × 10⁻⁴ to 1.1147 × 10⁻³ substitutions per site per year) compared to other picornaviruses. The observed evolutionary mechanisms indicate that whole-genome-based analysis of DHAV strains is needed to better understand the emergence of novel strains and their geographical dispersal.

A novel gyrovirus was detected in an intestinal specimen of a common pheasant that died due to poult enteritis and mortality syndrome. The genome of the pheasant-associated gyrovirus (PAGyV) is 2353 nucleotides (nt) long and contains putative genes for the VP1, VP2, and VP3 proteins in an arrangement that is typical for gyroviruses. Gyrovirus-specific motifs were identified in both the coding region and the intergenic region of the PAGyV genome. The VP1 of PAGyV shares up to 67.6% pairwise nt sequence identity with reference sequences and forms a distinct branch in the phylogenetic tree. Thus, according to the recently described species demarcation criteria, PAGyV belongs to a novel species in the genus Gyrovirus, family Anelloviridae, for which we propose the name "Gyrovirus phaco 1".

Fowl adenovirus 1 (FAdV-1) is the main cause of gizzard erosion in chickens. Whole genome sequencing and sequence analyses of 32 FAdV-1 strains from a global collection provided evidence that multiple recombination events have occurred along the entire genome. In gene-wise phylogenies, only the adenoviral pol gene formed a tree topology that corresponded to whole genome-based phylogeny. Virus genetic features that were clearly connected to gizzard erosion were not identified in our analyses. However, some genome variants tended to be more frequently identified from birds with gizzard erosion and strains isolated from healthy birds or birds with non-specific pathologies tended to form common clusters in multiple gene phylogenies. Our data show that the genetic diversity is greater, and the evolutionary mechanisms are more complex within FAdV-1 than previously thought. The implications of these findings for viral pathogenesis and epidemiology await further investigation.

Porcine reproductive and respiratory syndrome virus (PRRSV) is a major concern worldwide. Control of PRRSV is a challenging task due to various factors, including the viral diversity and variability. In this study, we evaluated an amplicon library preparation protocol targeting the ORF7 region of both PRRSV species, Betaarterivirus suid 1 and Betaarterivirus suid 2. We designed tailed primers for a two-step PCR procedure that generates ORF7-specific amplicon libraries suitable for use on Illumina sequencers. We tested the method with serum samples containing common laboratory strains and with pooled serum samples (n = 15) collected from different pig farms during 2019–2021 in Hungary. Testing spiked serum samples showed that the newly designed method is highly sensitive and detects the viral RNA even at low copy numbers (corresponding to approx. Ct 35). The ORF7 sequences were easily assembled even from clinical samples. Two different sequence variants were identified in five samples, and the Porcilis MLV vaccine strain was identified as the minor variant in four samples. An in-depth analysis of the deep sequencing results revealed numerous polymorphic sites along the ORF7 gene in a total of eight samples, and some sites (positions 12, 165, 219, 225, 315, 345, and 351) were found to be common in several clinical specimens. We conclude that amplicon deep sequencing of a highly conserved region of the PRRSV genome could support both laboratory diagnosis and epidemiologic surveillance of the disease.

PRRS elimination strategies often rely on depopulation-repopulation. However, this approach is accompanied by a long-term loss of production. With adequate control measures, such as well-designed immunization programs and technological changes along with prevalence-based laboratory testing, the virus-free status of the most vulnerable age groups in swine herds can be achieved. The most common reason for acquiring PRRSV at large farrow-to-finish swine farm units is that the previously settled fattening pigs serve as a source of infection for the newly reared PRRS-free animals. Following such unwanted events, PRRSV may persist in an affected establishment for several years. In this observational study, we selected four farrow-to-finish type swine herds. We implemented different laboratory testing protocols to find the most optimal solution for a successful PRRS elimination program. To aid our objectives, we used a DIVA PCR technique. The PRRS DIVA PCR assay is a fast, reliable method to identify sows shedding farm-specific PRRSV strain(s). As a result of elimination efforts at the sentinel pig herds, we found that reliable detection of wild-type PRRSV shedding among sows requires sampling at least three weaned piglets per litter. The strict adherence to this sampling protocol, the systematic use of laboratory methods that quickly detect the presence of wild virulent virus in the herd during the rearing period and the culling of DIVA PCR positive litters and their sows decreased the presence of the resident virus markedly. These procedures at Hungarian farrow-to-finish type farms successfully inhibited the wild-type PRRSV infection of different age groups. The results of this study demonstrate that applying this methodology together with strict biosecurity measures enabled us to reach PRRS-vaccinated-free status in large, farrow-to-finish herds within two years.

Porcine reproductive and respiratory syndrome (PRRS) is the cause of the most severe economic losses in the pig industry worldwide. PRRSV is extremely diverse in Europe, which poses a significant challenge to disease control within a country or any region. With the combination of phylogenetic reconstruction and network analysis, we aimed to uncover the major routes of the dispersal of PRRSV clades within Hungary. In brief, by analyzing >2600 ORF5 sequences, we identified at least 12 clades (including 6 clades within lineage 1 and 3 clades within lineage 3) common in parts of Western Europe (including Denmark, Germany and the Netherlands) and identified 2 novel clades (designated X1 and X2). Of interest, some genetic clades unique to other central European countries, such as the Czech Republic and Poland, were not identified. The pattern of PRRSV clade distribution is consistent with the route of the pig trade among countries, showing that most of the identified clades were introduced from Western Europe when fatteners were transported to Hungary. As a result of rigorous implementation of the national eradication program, the swine population was declared officially free from PRRSV. This map of viral diversity and clade distribution will serve as valuable baseline information for the maintenance of PRRSV-free status in the post-eradication era.