Development of cell combos micromethod to isolate respiratory viruses not detected by molecular techniques
Year: 2025
Type: article
Abstract: . The other approach to detecting new viruses is based on their isolation in cell culture using virus-specific cells and culture conditions. Unfortunately, inoculation on several lines and metagenomics requires a large volume of clinical sample, while the initial sample has already been largely used to carry out multiplexed RT-PCR. Herein, we aimed to develop micro-methods to inoculate combos of cell lines to isolate respiratory viruses missed by molecular techniques. Ten cell lines were selected, and then five cell combos of two cell lines each were developed. Strains of 15 respiratory virus species were inoculated, and viral growth was assessed by cytopathic effect detection and RT-PCR assays. Among these, all grew on at least one combo. The Caco-2/MRC5 combo was the most promising. As proof-of-concept, 859 human respiratory samples found negative by multiplex RT-PCR panels were inoculated into these five cell combos and monitored for the appearance of cytopathic effects. The efficiency of this approach was evidenced by isolating 12 herpes simplex or varicella-zoster viruses not detected by respiratory multiplex PCR assays. In conclusion, this updated approach can detect known but, most importantly, likely emerging respiratory viruses. It could be used to investigate undiagnosed respiratory infection outbreaks especially if optimized with an approach allowing for the detection of viral multiplications with minor or no cytopathic effects.IMPORTANCEThe detection of respiratory viruses relies on a range of laboratory methods each of which has distinct advantages in terms of speed, practicality, and sensitivity. Current molecular methods for respiratory virus detection, such as multiplex PCR, may fail to identify unexpected, genetically divergent, or emerging viruses. This study presents an innovative approach using micromethods inoculating combinations of cell lines (cell combos) to enhance the isolation of a broad panel of respiratory viruses, including those undetected by standard molecular techniques. This strategy revives and modernizes classical virology techniques for use in contemporary diagnostics, particularly during unexplained respiratory outbreaks. It opens up new possibilities for detecting both known and unknown viruses across different sample types.
Source: Microbiology Spectrum
Language: English
Subfield: Epidemiology
Field: Medicine
Domain: Health Sciences
Open Access status: gold
Estimated APC paid: $2,500
Funder: Agence Nationale de la Recherche
OpenAlex ID: W4416124759