Molecular and biological characterization of an Asian-American isolate of Chikungunya virus

Autores organización

• 

  Jaime Eduardo Castellanos Parra

  Autor

• 

  Eliana Patricia Calvo Tapiero

  Autor

Autores

• Archila ED
• López LS

Grupos de investigación

• Grupo de Virología

  

  Investigador

  Jaime Eduardo Castellanos Parra

Resumen

Chikungunya virus is an arthropod-transmitted virus that causes chikungunya fever, a disease characterized by severe muscle and joint pain. In 2013, the virus was introduced to the Americas and caused approximately 2.7 million cases of infection during the subsequent two years. The lack of knowledge regarding the biological behavior of the viral strains circulating during the outbreak motivated the characterization of an isolate from the Colombian outbreak, starting from analysis of the complete genome to the biological behavior in vitro. The full genome was retrieved using next-generation sequencing. The infective and replicative capacities were evaluated in HEK293T, Huh-7, and MRC-5 cell lines. The infection rates were determined by flow cytometry, and the cytopathic effect was assessed by a resazurin fluorescent metabolic assay. The viral yield was quantified using the virus plaque formation assay, while the viral proteins and genomic RNA kinetics were subsequently evaluated by western-blot and RT-qPCR. The COL7624 isolate clustered with other American and Caribbean sequences in the Asian American lineage. The T669A substitution in E2 protein distinguished it from other Colombian sequences reported in 2014. After 48 h post infection (hpi), the three cell lines analyzed reached infection percentages exceeding 65%, generating a high load of infectious viral progeny. The infection kinetics indicated that the replication peak of this CHIKV isolate is around 24 hpi, although gRNA is detectable in the culture supernatant from 4 hpi onwards. The infection caused the overexpression of interferon and pro-inflammatory cytokines, such as IL-1ß, TNF-a, and IL-8. The COL7624 CHIKV isolate exhibited a high infective and replicative capacity as well as activation of cellular immune responses, similar to isolates belonging to the other genotypes. Copyright: © 2022 Archila et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Keywords

• Asian Americans; Chikungunya Fever; Chikungunya virus; HEK293 Cells; Humans; Sequence Analysis, DNA; alexa fluor 488; carboxymethylcellulose; caspase 3; cytokine; genomic RNA; interleukin 1beta; interleukin 8; tumor necrosis factor; viral protein; amino acid substitution; antiviral activity; apoptosis; Article; Asian American; cellular immunity; chikungunya; controlled study; fetal bovine serum; flow cytometry; gene overexpression; genome analysis; high throughput sequencing; human; human cell; immunoblotting; metabolic activity assay; mouse; nonhuman; nucleotide sequence; phylogenetic tree; protein expression; quantitative analysis; reverse transcription polymerase chain reaction; RNA extraction; upregulation; virus entry; virus infection; virus load; virus plaque; virus replication; virus transmission; Western blotting; Asian American; chikungunya; Chikungunya virus; DNA sequence; HEK293 cell line