Acute respiratory infection (ARI) is the most common infection in children under 5 years of age and it is frequently caused by two pneumoviruses, human respiratory syncytial virus (HRSV) and human ...metapneumovirus (HMPV). Epidemic seasons of these viruses overlap and disease manifestations are highly similar, including severe lower ARI such as bronchiolitis or pneumonia. Reinfections with pneumoviruses are frequent and limited prevention treatment is available. Genetic diversity of HRSV and HMPV strains circulating in Croatia was monitored during four consecutive years (2014–2017). Co-circulation of multiple lineages was observed for both viruses. Within HRSV group A, ON1 strains gained strong predominance during the 4-year period, while previously dominant genotype NA1 was detected only sporadically. Similarly, newly occurring HMPV genotype A2c gained predominance over genotype A2b during this period, resulting in all infection in 2017 being caused by A2c. Along with phylogenetic analysis based on the commonly used fragments for detection and genotyping of these viruses, full length G and SH genes were also analysed. Evolutionary dynamics showed that inferred substitution rates of HRSV and HMPV are between 2.51 × 10−3 and 3.61 × 10−3 substitutions/site/year. This study established presence of recently described HMPV strains containing large duplications in the G gene in Croatia. Viruses with either of the two duplications belong to a subcluster A2c, which has completely replaced all other group A subclusters in 2017.
•Diversity of HRSV and HMPV in Croatia, 2014–2017, is described.•Co-circulation of multiple lineages was observed for both viruses.•Recently described HMPV strains containing large duplications in the G gene were discovered in Croatia.
A recent resurgence of mumps in doubly vaccinated cohorts has been observed, identifying genotype G as the current predominant genotype. In this study, the neutralization efficacy of guinea pig sera ...immunized with three vaccine viruses: L-Zagreb, Urabe AM9 and JL5, was tested against seven mumps viruses: three vaccine strains and four wild-type strains (two of genotype G, one of genotype C, one of genotype D) isolated during 1998–2011. All sera neutralized all viruses although at different levels. The neutralization efficiency of sera decreases several fold by temporal order of virus isolation. Therefore, we concluded that gradual evolution of mumps viruses, rather than belonging to a certain genotype, results in an antigenic divergence from the vaccine strains that decrease the neutralization capacity of vaccine-induced antibodies. Moreover, the amino-acid sequence alignment revealed three new potentially relevant regions for escape from neutralization, i.e. 113–130, 375–403 and 440–443.
Virus mumpsa uzročnik je zaušnjaka, bolesti koja se može prevenirati cijepljenjem. Mumps je RNK virus koji se u kliničkim uzorcima i u supernatantima inficiranih staničnih kultura nakon izolacije RNK ...može detektirati RT-PCR testom. Prednosti RT-PCR testa su brzina, specifičnost i mogućnost detekcije malog broja kopija virusa. Genskom karakterizacijom i genotipizacijom virusa omogućeno je epidemiološko praćenje distribucije i cirkuliranja divljih tipova virusa mumpsa koji uzrokuju epidemije i u procijepljenim populacijama. Molekularna detekcija i genska karakterizacija divljih i cjepnih virusa mumpsa provodi se u Republici Hrvatskoj od kraja 90-tih godina.
Replicative double-stranded RNA (dsRNA) is useful in preliminary identification of
Cucumber mosaic virus and its satellite RNA (satRNA). This plant pathogen complex yields sufficient quantity of the ...replicative RNA form that can be isolated by chromatography on chemically unmodified graded cellulose powder (CF-11). In this work, much faster and more efficient procedure using DEAE monoliths was developed in which dsRNA was separated from other species in total nucleic acids extract originating from the infected plant tissue. The developed chromatographic method revealed the pathogens’ presence in only 15
min, avoiding nucleic acid precipitation and electrophoretic analysis.
Restriction enzymes have proven to be among the most valuable tools in molecular biology. In this work, we demonstrate that the cleavage of fluorescently labeled, PCR-amplified DNA can be used as a ...simple and highly sensitive technique for detection of sequences present in a percentage as low as 0.6% in a DNA pool. Due to the fact that fluorescent labeling of DNA fragments enables such sensitive detection and quantification of restriction enzyme cleavage, the method was further exploited in monitoring of the enzymatic digestion completeness and in determination of factors that influence restriction enzyme effectiveness. We analyzed the activity of six restriction endonucleases; the percentage of uncleaved DNA fragments predominantly ranged between 2.0 and 2.5 and the highest value was 8.00%. We conclude that, since the enzymatic digestion completeness may not always be assured, each assay based on restriction enzyme cleavage that is intended to be used in investigations of heterogeneity in a DNA pool should be constructed so that the presence of cleaved sequences is the indication of pool nonuniformity. When the presence of uncleaved sequences indicates pool heterogeneity, the results could be misleading due to possible incompleteness of enzymatic cleavage.
