African swine fever (ASF) remains one of the most devastating transboundary diseases affecting domestic pigs and wild boars worldwide, making molecular epidemiology essential for understanding viral transmission and supporting disease control. This study aimed to characterise African swine fever virus (ASFV) isolates circulating in Romania through comparative analysis of the B646L (p72) gene and the B602L central variable region (CVR). Fifty-four ASFV-positive samples collected from domestic pigs and wild boars were confirmed by real-time PCR and subjected to molecular analysis. Following Sanger sequencing and sequence quality assessment, 32 reliable consensus sequences were retained for B602L and 33 for p72. Sequencing of the p72 gene demonstrated a very high degree of nucleotide conservation, confirming that all successfully sequenced isolates belonged to genotype II. In contrast, the B602L region exhibited limited intra-genotypic variability characterised by scattered single nucleotide polymorphisms and localised insertion/deletion events, providing additional molecular resolution compared to p72, although insufficient for fine-scale epidemiological tracking. No genetic segregation according to host species was observed for either genomic region, and sequences obtained from domestic pigs and wild boar were interspersed in the phylogenetic analyses, demonstrating a high degree of genetic similarity at the investigated loci. These findings highlight the complementary value of p72 and B602L for ASFV molecular characterisation, while also indicating that these partial genomic markers alone cannot resolve transmission pathways or epidemiological links between domestic and wild host populations.

Avian influenza is a highly contagious viral disease affecting domestic and wild birds, caused by influenza A viruses of the family Orthomyxoviridae. Continuous genetic surveillance is essential for monitoring viral evolution and identifying molecular determinants associated with pathogenicity and host adaptation. In this study, the PB1 and PB2 polymerase genes of highly pathogenic avian influenza (HPAI) H5N1 viruses detected in Romania between 2017 and 2024 were genetically characterized. A total of 18 organ samples collected from domestic and wild birds in several Romanian counties were analyzed by real-time RT-PCR, followed by conventional RT-PCR, Sanger sequencing, mutational analysis, and phylogenetic reconstruction. High-quality sequence data were obtained for six PB1 and eight PB2 gene fragments. Sequence analysis revealed a high degree of conservation within the PB1 gene, particularly across regions involved in viral replication, indicating strong evolutionary constraints. In contrast, the PB2 gene exhibited several amino acid substitutions, including changes at positions 464 and 678 in selected isolates. Notably, all analyzed strains lacked the mammalian adaptation marker E627K. Phylogenetic analysis clustered the Romanian isolates within Eurasian H5N1 lineages and demonstrated close genetic relationships with contemporary strains circulating in Eastern Europe and Russia. These findings highlight the genetic stability of PB1, the ongoing diversification of PB2, and the importance of continuous molecular surveillance for the early detection of viral variants with potential implications for host adaptation and zoonotic risk.