Driving through stop signs: predicting stop codon reassignment improves functional annotation of bacteriophages

Area of Science:

• Virology
• Genomics
• Bioinformatics

Background:

• Bacteriophage diversity is vast, with many uncharacterized lineages.
• Some phage lineages, like Crassvirales, utilize alternate genetic codes by reassigning stop codons.

Purpose of the Study:

• Investigate the prevalence of stop codon reassignment in phage genomes.
• Assess the impact of reassignment on functional annotation quality.
• Develop and evaluate tools for predicting stop codon reassignment before genomic annotation.

Main Methods:

• Predicted stop codon reassignment in 76 INPHARED genomes and 712 Unified Human Gut Virome Catalogue (UHGV) vOTUs.
• Re-annotated sequences using modified Pharokka and Prokka tools (Pharokka-gv and Prokka-gv).
• Compared annotation quality and gene characteristics before and after re-annotation.

Main Results:

• Pharokka-gv and Prokka-gv significantly improved annotation quality.
• Pharokka-gv showed superior performance, increasing median gene length and coding capacity for TAG-to-glutamine reassignment.
• Functional annotation assignment increased, including identification of major capsid proteins.

Conclusions:

• Automatic prediction of stop codon reassignment is beneficial for viral genomic and metagenomic analyses.
• Improved annotation facilitates a deeper understanding of phage biology and diversity.
• The developed tools enhance the accuracy of viral genome interpretation.