Genetic Adaptation of Coxsackievirus B1 during Persistent Infection in Pancreatic Cells

Abstract

Coxsackie B (CVB) viruses have been associated with type 1 diabetes. We have recently observed that CVB1 was linked to the initiation of the autoimmune process leading to type 1 diabetes in Finnish children. Viral persistency in the pancreas is currently considered as one possible mechanism. In the current study persistent infection was established in pancreatic ductal and beta cell lines (PANC-1 and 1.1B4) using four different CVB1 strains, including the prototype strain and three clinical isolates. We sequenced 5′ untranslated region (UTR) and regions coding for structural and non-structural proteins and the second single open reading frame (ORF) protein of all persisting CVB1 strains using next generation sequencing to identify mutations that are common for all of these strains. One mutation, K257R in VP1, was found from all persisting CVB1 strains. The mutations were mainly accumulated in viral structural proteins, especially at BC, DE, EF loops and C-terminus of viral capsid protein 1 (VP1), the puff region of VP2, the knob region of VP3 and infection-enhancing epitope of VP4. This showed that the capsid region of the viruses sustains various changes during persistency some of which could be hallmark(s) of persistency.

Viruses and Type 1 Diabetes: From Enteroviruses to the Virome

“…Furthermore, long-term passage of CVB1 resulted in the production of smaller viral plaques, highlighting the potential for persistence to reduce the cytopathic effect of EV infection due to changes in translation, adsorption or internalisation. This reinforces the importance of the capsid region during viral persistence, with the potential to identify hallmarks of persistency as an ultimate goal [159].…”

“…Viral adaptation processes such as the establishment of persistence, modulation of viral replication and immune evasion may be facilitated by viral genomic modifications obtained during intra-host evolution. This is highlighted by the divergence of EV genotypes as a direct result of high mutagenic rates of ~4–8 mutations/10 4 nucleotides during viral replication from the lack of proofreading activity in their RNA-dependent RNA polymerase [ 156 , 157 , 158 , 159 ]). Terminal deletions within the 5′ UTR facilitated the establishment of viral persistence in the pancreas of mice by diminishing viral replication and translation, and terminally deleted EV RNA was detected in human heart tissue [ 160 , 161 , 162 , 163 , 164 ].…”

Viruses and Type 1 Diabetes: From Enteroviruses to the Virome

“…Furthermore, long term passage of CVB1 resulted in the production of smaller viral plaques, highlighting the potential for persistence to reduce the cytopathic effect of EV infection due to changes in translation, adsorption or internalisation. This reinforces the importance of the capsid region during viral persistence, with the potential to identify hallmarks of persistency as an ultimate goal [157].…”

“…Viral adaptation processes such as the establishment of persistence, modulation of viral replication and immune evasion may be facilitated by viral genomic modifications obtained during intra-host evolution. This is highlighted by the divergence of EV genotypes as a direct result of high mutagenic rates of ~4-8 mutations/10 4 nucleotides during viral replication from the lack of proofreading activity in their RNA-dependent RNA polymerase (RdRp) [154][155][156][157]). Terminal deletions within the 5' UTR facilitated the establishment of viral persistence in the pancreas of mice by diminishing viral replication and translation, and terminally deleted EV RNA was detected in human heart tissue [158][159][160][161][162].…”

“…Such limitations have driven the demand for more comprehensive, high-throughput and untargeted methods for detection and characterisation of complex viral populations [108]. In the current NGS era, near-complete EV genome sequence can be characterised rapidly through amplicon-based sequencing, enabling in-depth examination of intraand inter-host sequence variations that may contribute importantly to the diabetogenicity of specific EVs [157,197] . Furthermore, recent advances in target enrichment methods have drastically helped to overcome previous bottlenecks of conventional virome sequencing, where an overwhelming abundance of non-viral nucleic acid from the human host, bacteria, fungi and bacteriophage significantly reduced the sensitivity for human viruses [198,199].…”