In sheep polymorphisms of the prion gene (PRNP) at the codons 136, 154 and 171 strongly influence the susceptibility to scrapie and bovine spongiform encephalopathy (BSE) infections. In goats a number of other gene polymorphisms were found which are suspected to trigger similar effects. However, no strong correlation between polymorphisms and TSE susceptibility in goats has yet been obtained from epidemiological studies and only a low number of experimental challenge data are available at present. We have therefore studied the potential impact of these polymorphisms in vitro by cell-free conversion assays using mouse scrapie strain Me7. Mouse scrapie brain derived PrPSc served as seeds and eleven recombinant single mutation variants of sheep and goat PrPC as conversion targets. With this approach it was possible to assign reduced conversion efficiencies to specific polymorphisms, which are associated to low frequency in scrapie-affected goats or found only in healthy animals. Moreover, we could demonstrate a dominant-negative inhibition of prion polymorphisms associated with high susceptibility by alleles linked to low susceptibility in vitro.
Dengue virus (DENV 1-4) represents the major emerging arthropod-borne viral infection in the world. Currently, there is neither
an available vaccine nor a specific treatment. Hence, there is a need of antiviral drugs for these viral infections; we describe the
prediction of short interfering RNA (siRNA) as potential therapeutic agents against the four DENV serotypes. Our strategy was to
carry out a series of multiple alignments using ClustalX program to find conserved sequences among the four DENV serotype
genomes to obtain a consensus sequence for siRNAs design. A highly conserved sequence among the four DENV serotypes,
located in the encoding sequence for NS4B and NS5 proteins was found. A total of 2,893 complete DENV genomes were
downloaded from the NCBI, and after a depuration procedure to identify identical sequences, 220 complete DENV genomes were
left. They were edited to select the NS4B and NS5 sequences, which were aligned to obtain a consensus sequence. Three different
servers were used for siRNA design, and the resulting siRNAs were aligned to identify the most prevalent sequences. Three
siRNAs were chosen, one targeted the genome region that codifies for NS4B protein and the other two; the region for NS5 protein.
Predicted secondary structure for DENV genomes was used to demonstrate that the siRNAs were able to target the viral genome
forming double stranded structures, necessary to activate the RNA silencing machinery.
Dengue fever is one of the most common viral infections in the world. Although a vaccine against dengue virus (DENV) has been approved in several countries, this disease is still considered a public health priority worldwide. The ability of three small interfering RNAs (FG-siRNAs) targeting conserved sequences in the NS4B and NS5 regions of the DENV genome to inhibit DENV replication was tested in vitro in both Vero and C6/36 cells. The FG-siRNAs were effective against DENV-1, -3, and -4, but not DENV-2. A fourth siRNA specifically targeting the NS5 region of the DENV-2 genome (SG-siRNA) was designed and tested against two different DENV-2 strains, showing high levels of inhibition in both mammalian and insect cells.
Emerging viruses have been constant since life arises on earth. These viruses could infect humans and produce pandemic such as the caused by SARS-COV-2. The emergence of viruses is inherent to nature and cannot been avoided; however, as it spread mainly by human activities, it is possible to develop strategies to detect and control outbreaks of emerging diseases and prevent new pandemics.
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