The neighborhood of the Spike gene is a hotspot for modular intertypic homologous and non-homologous recombination in Coronavirus genomes.

Coronaviruses (CoVs) have very large RNA viral genomes with a distinct genomic architecture of core and accessory open reading frames (ORFs). It is of utmost importance to understand their patterns and limits of homologous and non-homologous recombination, because such events may affect the emergence of novel CoV strains, alter their host range, infection rate, tissue tropism pathogenicity, and their ability to escape vaccination programs. Intratypic recombination among closely related CoVs of the same subgenus has often been reported; however, the patterns and limits of genomic exchange between more distantly related CoV lineages (intertypic recombination) needs further investigation. Here, we report computational/evolutionary analyses that clearly demonstrate a substantial ability for CoVs of different subgenera to recombine. Furthermore, we show that CoVs can obtain-through non-homologous recombination-accessory ORFs from core ORFs, exchange accessory ORFs with different CoV genera, with other viruses (i.e., toroviruses, influenza C/D, reoviruses, rotaviruses, astroviruses) and even with hosts. Intriguingly, most of these radical events result from double-crossovers surrounding the Spike ORF, thus highlighting both the instability and mobile nature of this genomic region. While many such events have often occurred during the evolution of various CoVs, the genomic architecture of the relatively young SARS-CoV/SARS-CoV-2 lineage so far appears to be stable

The Remarkable Evolutionary Plasticity of Coronaviruses by Mutation and Recombination: Insights for the COVID-19 Pandemic and the Future Evolutionary Paths of SARS-CoV-2.

Coronaviruses (CoVs) constitute a large and diverse subfamily of positive-sense single-stranded RNA viruses. They are found in many mammals and birds and have great importance for the health of humans and farm animals. The current SARS-CoV-2 pandemic, as well as many previous epidemics in humans that were of zoonotic origin, highlights the importance of studying the evolution of the entire CoV subfamily in order to understand how novel strains emerge and which molecular processes affect their adaptation, transmissibility, host/tissue tropism, and patho non-homologous genicity. In this review, we focus on studies over the last two years that reveal the impact of point mutations, insertions/deletions, and intratypic/intertypic homologous and non-homologous recombination events on the evolution of CoVs. We discuss whether the next generations of CoV vaccines should be directed against other CoV proteins in addition to or instead of spike. Based on the observed patterns of molecular evolution for the entire subfamily, we discuss five scenarios for the future evolutionary path of SARS-CoV-2 and the COVID-19 pandemic. Finally, within this evolutionary context, we discuss the recently emerged Omicron (B.1.1.529) VoC

Improved Genotyping Vaccine and Wild-Type Poliovirus Strains by Restriction Fragment Length Polymorphism Analysis: Clinical Diagnostic Implications

The combination of preventive vaccination and diagnostic typing of viral isolates from patients with clinical poliomyelitis constitutes our main protective shield against polioviruses. The restriction fragment length polymorphism (RFLP) adaptation of the reverse transcriptase (RT)-PCR methodology has advanced diagnostic genotyping of polioviruses, although further improvements are definitely needed. We report here on an improved RFLP procedure for the genotyping of polioviruses. A highly conserved segment within the 5′ noncoding region of polioviruses was selected for RT-PCR amplification by the UC(53)-UG(52) primer pair with the hope that it would be most resistant to the inescapable genetic alteration-drift experienced by the other segments of the viral genome. Complete inter- and intratypic genotyping of polioviruses by the present RFLP method was accomplished with a minimum set of four restriction endonucleases (HaeIII, DdeI, NcoI, and AvaI). To compensate for potential genetic drift within the recognition sites of HaeIII, DdeI, or NcoI in atypical clinical samples, the RFLP patterns generated with HpaII and StyI as replacements were analyzed. The specificity of the method was also successfully assessed by RFLP analysis of 55 reference nonpoliovirus enterovirus controls. The concerted implementation of these conditional protocols for diagnostic inter- and intratypic genotyping of polioviruses was evaluated with 21 clinical samples with absolute success

Molecular Characterization of Wild-Type Polioviruses Isolated in Greece during the 1996 Outbreak in Albania

During the present study three type 1 poliovirus strains isolated in Greece during the 1996 poliomyelitis outbreak in Albania were retrospectively investigated and determination of their relationship with other epidemic strains isolated in Albania or elsewhere during previous epidemics was attempted. SimPlot analysis revealed that the three Greek strains are the result of a recombination event in the VP2 coding region

Comparative Analysis of SARS-CoV-2 Variants of Concern, Including Omicron, Highlights Their Common and Distinctive Amino Acid Substitution Patterns, Especially at the Spike ORF.

In order to gain a deeper understanding of the recently emerged and highly divergent Omicron variant of concern (VoC), a study of amino acid substitution (AAS) patterns was performed and compared with those of the other four successful variants of concern (Alpha, Beta, Gamma, Delta) and one closely related variant of interest (VoI-Lambda). The Spike ORF consistently emerges as an AAS hotspot in all six lineages, but in Omicron this enrichment is significantly higher. The progenitors of each of these VoC/VoI lineages underwent positive selection in the Spike ORF. However, once they were established, their Spike ORFs have been undergoing purifying selection, despite the application of global vaccination schemes from 2021 onwards. Our analyses reject the hypothesis that the heavily mutated receptor binding domain (RBD) of the Omicron Spike was introduced via recombination from another closely related Sarbecovirus. Thus, successive point mutations appear as the most parsimonious scenario. Intriguingly, in each of the six lineages, we observed a significant number of AAS wherein the new residue is not present at any homologous site among the other known Sarbecoviruses. Such AAS should be further investigated as potential adaptations to the human host. By studying the phylogenetic distribution of AAS shared between the six lineages, we observed that the Omicron (BA.1) lineage had the highest number (8/10) of recurrent mutations