Screening human cell lines for viral infections applying RNA-Seq data analysis.

Area of Science:

• Virology
• Bioinformatics
• Cell Biology

Background:

• Viral infections in cell cultures are often overlooked but pose risks to laboratory safety and bioactive agent production.
• Current detection methods like PCR and immunological assays are common but may not capture all infections.
• Cell line contamination can impact experimental results and the safety of derived products.

Purpose of the Study:

• To assess the viral infection status of human cell lines using next-generation sequencing (NGS) data.
• To compare the efficacy of NGS-based methods with traditional virus-specific PCR analyses.
• To evaluate the adequacy of current cell culture testing protocols for virus detection and risk assessment.

Main Methods:

• Screened over 300 Cancer Cell Line Encyclopedia RNA sequencing and 60 whole exome sequencing datasets.
• Applied the Taxonomer bioinformatics tool for viral sequence and nucleotide/protein assessment.
• Compared NGS findings with previous virus-specific PCR results.

Main Results:

• NGS methods (direct alignment and Taxonomer) showed complete concordance with PCR results.
• Twenty cell lines were identified as infected with five distinct virus species.
• A bovine polyomavirus infection in SK-BR-3 cell line was linked to contaminated fetal bovine serum.
• RNA-Seq demonstrated higher sensitivity for virus detection, though low-level contamination can occur.
• No novel, previously unobserved viruses were detected in cell cultures.

Conclusions:

• Next-generation sequencing is a powerful and efficient tool for determining the viral infection status of human cell lines.
• Current cell culture testing methods are sufficient for detecting contamination and assessing risks.
• NGS data analysis requires attention to sequence quality to avoid false positives from nucleotide contamination.