Integrated Functional Characterization of a Panel of Clinical Orthoflavivirus Isolates Reveals Distinct Replication and Innate Immune Response Profiles in Human Keratinocytes.

Authors

Tannya Karen Castro Jiménez, Edwin Antonio Lopez Kelly, Leticia Cedillo-Barrón, Julio García-Cordero, Diego Sait Cruz-Hernández, Nallely Diaz Lima, José Alberto San Juan Luis, Cruz Carlos Castillo Camacho, Eloy Andrés Pérez-Yépez, Cynthia Daniela Ibarra-Moreno, Luis Angel Flores-Mejía, Sergio Roberto Aguilar-Ruíz, Mónica G Mendoza-Rodríguez, Luis I Terrazas, José Bustos-Arriaga

Published in

Viruses. Volume 18. Issue 8. Jul 27, 2026. Epub Jul 27, 2026.

Abstract

Orthoflaviviruses comprise genetically diverse mosquito-borne viruses responsible for a broad spectrum of human diseases. Although naturally circulating clinical isolates exhibit biological variability, the extent to which they generate distinct early epithelial innate immune responses remains incompletely understood. Here, we characterized five clinical orthoflavivirus isolates obtained in Oaxaca, Mexico, using human HaCaT keratinocytes as an in vitro model of early infection. Productive infection was assessed by immunofluorescence microscopy, immunostained focus appearance under isolate-optimized assay conditions, and infectious virus production, whereas host responses were evaluated by transcriptional profiling and quantitative whole-slide single-cell immunofluorescence. All isolates established productive infection and exhibited different viral replication profiles. Temporal transcriptional analyses revealed variable expression of antiviral (IFNβ, Mx1, OAS1, PKR, IFITM3, Viperin, and RANTES) and inflammatory (TNF-α, IL-8, and MCP-1) genes. Quantitative whole-slide analysis provided complementary evidence of variable STAT1 and NF-κB signaling activation across the analyzed isolates. Within this limited panel, viral replication was not consistently aligned with the selected transcriptional and signaling readouts, although the exploratory nature of these comparisons precludes establishing independence between these variables. Together, the virological, transcriptional, and imaging analyses revealed distinct multidimensional functional profiles across the isolate panel. Overall, these findings demonstrate functional heterogeneity among the analyzed clinical orthoflavivirus isolates and highlight integrated functional phenotyping as a useful framework for examining virus-host interactions beyond viral replication alone.

PMID:
42655646
Bibliographic data and abstract were imported from PubMed on 27 Aug 2026.

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