The aim of this study was to estimate substitution rate and imprints of natural selection on parvovirus B19 genotype 1. Studied datasets included 137 near complete coding B19 genomes (positions 665 to 4851) for phylogenetic and substitution rate analysis and 146 and 214 partial genomes for selection analyses in open reading frames ORF1 and ORF2, respectively, collected 1973–2012 and including 9 newly sequenced isolates from Serbia. Phylogenetic clustering assigned majority of studied isolates to G1A. Nucleotide substitution rate for total coding DNA was 1.03 (0.6–1.27) x 10−4 substitutions/site/year, with higher values for analyzed genome partitions. In spite of the highest evolutionary rate, VP2 codons were found to be under purifying selection with rare episodic positive selection, whereas codons under diversifying selection were found in the unique part of VP1, known to contain B19 immune epitopes important in persistent infection. Analyses of overlapping gene regions identified nucleotide positions under opposite selective pressure in different ORFs, suggesting complex evolutionary mechanisms of nucleotide changes in B19 viral genomes.

zevi c A. Comparison of HPV detection rate in formalin-fixed paraffin-embedded tissues of head and neck carcinoma using two DNA extraction kits and three amplification methods.

The presence of pseudorabies virus (PrV), porcine parvovirus (PPV) and porcine circovirus 2 (PCV2) was examined in sixty samples (spleen and lymph nodes) and thirty samples of sacral ganglia collected from non-vaccinated swine by virus isolation and polymerase chain reaction (PCR). Using PCR method PrV was detected in three samples, PPV in seven samples and six samples were found positive for PCV2. The phylogenetic analysis of the nucleotide sequences of three PrV isolates identifi ed in this study showed high similarity and signifi cant clustering within the PrV genotype I strains such as Kaplan and Bartha isolated from pigs in Hungary, strain Becker isolated in USA and strain Kolchis isolated in Greece. The nucleotide sequences of two PPV isolates showed high level of similarity with the strain Challenge isolated from pigs in UK, strain Kresse isolated in USA and strains 77 and LZ isolated in China. The phylogenetic analysis of the nucleotide sequences of two PCV2 isolates showed high level of similarity and signifi cant clustering within genotype PCV2b strains such as NIVS-3, NIVS-5 and NIVS-6 isolated in Serbia, strain 3959 isolated in Austria, strain PM165 isolated from pigs in Brasil, and strain XT2008 isolated in China. The results of our study present the molecular characterization of PrV, PPV and PCV2 identifi ed in swine in Republic of Montenegro. Besides that, these results confi rmed that PCR is a very useful method for rapid detection of these viruses in subclinically infected swine.

The presence of bovine parainfl uenza virus type 3 (BPIV3) was examined in 119 nasal swabs collected from cattle with severe respiratory infection. All samples were conducted for virus isolation on the MDBK cell line. The cytopathic effect was observed after 48h to 72h in cells inoculated with eight samples (8/119; 6.7%). The confi rmation of isolated strains of BPIV3 was done by the virus-neutralization test. In addition, all samples of bovine nasal swabs were also examined for the presence of BPIV3 virus using RT-PCR with primers specifi c for the part of HN gene. The presence of BPIV3 was detected in eight samples (8/119; 6.7%) that were also positive upon virus isolation. The molecular characterization based on nucleotide sequencing of the part of the HN gene showed that all BPIV3 isolates belonged to genotype C of BPIV3. They branched in one distinct cluster with three different branches, but these branches were very similar to each other (98.1% to 99.8%). Serbian BPIV3c isolates were most similar to the Chinese BPIV3c isolates SD0805, SD0809 and SD0835 (from 97.92% to 99.7%), and to South Korean (12Q061), Japanese (HS9) and American (TVMDL16 and TVMDL20) BPIV3c strains (from 97.1% to 98.8%), and distinct from American (TVMDL15and TVMDL17) and Australian (Q5592) BPI3V genotype B strains (only 79.9% to 82.3% similarity), as well as from the genotype A BPIV3 strains from different countries published in GenBank.

This study was endeavoured to contribute in furthering our understanding of the molecular epidemiology of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by sequencing and analysing the first full-length genome sequences obtained from 48 coronavirus disease-2019 (COVID-19) patients in five districts in Western Serbia in the period April 2020–July 2020. SARS-CoV-2 sequences in Western Serbia distinguished from the Wuhan sequence in 128 SNPs in total. The phylogenetic structure of local SARS-CoV-2 isolates suggested the existence of at least four distinct groups of SARS-CoV-2 strains in Western Serbia. The first group is the most similar to the strain from Italy. These isolates included two 20A sequences and 15−30 20B sequences that displayed a newly occurring set of four conjoined mutations. The second group is the most similar to the strain from France, carrying two mutations and belonged to 20A clade. The third group is the most similar to the strain from Switzerland carrying four co-occurring mutations and belonging to 20B clade. The fourth group is the most similar to another strain from France, displaying one mutation that gave rise to a single local isolate that belonged to 20A clade.