FDA Updates Guidance on Submitting Next-Generation Sequencing Data to the Division of Antivirals
The U.S. Food and Drug Administration (FDA) has updated its technical specifications document “Submitting Next-Generation Sequencing Data to the Division of Antivirals”.
The document, issued by the Center for Drug Evaluation and Research (CDER), was revised in July 2026 and provides the FDA Division of Antivirals’ current thinking on the submission of next-generation sequencing (NGS)protocols, data and analyses in support of resistance assessments for antiviral drug development.
The guidance is nonbinding and describes the information that sponsors should submit when using NGS for sequence-based resistance analysis.
Purpose of the Guidance
The FDA explains that the Division of Antivirals performs independent analyses of NGS data associated with antiviral drugs under development, including small molecule and immunoglobulin-based drugs.
According to the document, comprehensive and accurate resistance information is important to support resistance surveillance, monitor the emergence of viral variants resistant to approved antiviral drugs and protect public health.
The guidance also notes that NGS datasets are complex and that the document aims to address the absence of standardised bioinformatics protocols for large-scale NGS data analysis.
Information Recommended for Submission
The FDA recommends that sponsors provide a detailed NGS protocol, including both wet lab procedures and data analysis methods.
The information to be submitted includes:
NGS protocol;
raw NGS data in FASTQ format;
consensus sequences in FASTA format;
other sequences needed for data analysis, such as reference, primer or adapter sequences;
amino acid frequency tables;
NGS report and summary tables;
conservation analysis of reference sequences, where applicable.
The FDA also recommends that sponsors communicate with the Division early during drug development and provide an NGS protocol before initiating pivotal clinical trials.
NGS Data and Reporting Expectations
The guidance states that sponsors should submit all raw NGS data in FASTQ format and provide consensus sequences in a single FASTA file for each clinical trial.
Sponsors should also provide amino acid frequency tables reporting all amino acid substitutions that differ from the reference sequence at frequencies of 1% or higher. These tables should also include frameshifts, premature stop codons and in-frame insertions and deletions.
The FDA further recommends that the NGS report include a summary of the NGS protocol, resistance testing criteria, mutation frequency cutoffs, baseline polymorphisms, treatment-emergent substitutions and relevant virologic or clinical outcomes.
File Formats and Submission Procedures
The guidance includes recommendations on file formats and submission methods.
The FDA recommends that the NGS protocol, NGS report, reference sequences and primer or adapter sequences be submitted through the FDA Electronic Submissions Gateway.
Raw NGS data, consensus sequences and checksum files should generally be submitted on an external hard drive, while the cover letter and amino acid frequency table should be included in both electronic gateway and hard drive submissions.
The document also provides naming conventions for FASTQ files and consensus sequences in FASTA format.
Impact on Medical Device and IVD Manufacturers
Although this FDA guidance is focused on antiviral drug development, it may be relevant for IVD manufacturers and developers of diagnostic or digital tools that generate, analyse or support NGS-based data used in regulated clinical or regulatory contexts.
Manufacturers and developers should pay particular attention to:
data quality and integrity;
validation of sequencing and bioinformatics methods;
traceability of raw data, reference sequences and analysis outputs;
standardised file formats and naming conventions;
documentation of wet lab and bioinformatics workflows;
management of large-scale sequencing datasets;
early engagement with regulators where NGS data are used to support regulatory decisions.
For IVD manufacturers working with NGS-based assays or companion diagnostic strategies, the document reinforces the importance of structured data submission, validated analytical pipelines and clear reporting of sequence-based results.
