PhageTerm: a tool for fast and accurate determination of phage termini and packaging mechanism using next-generation sequencing data

Abstract

The worrying rise of antibiotic resistance in pathogenic bacteria is leading to a renewed interest in bacteriophages as a treatment option. Novel sequencing technologies enable description of an increasing number of phage genomes, a critical piece of information to understand their life cycle, phage-host interactions, and evolution. In this work, we demonstrate how it is possible to recover more information from sequencing data than just the phage genome. We developed a theoretical and statistical framework to determine DNA termini and phage packaging mechanisms using NGS data. Our method relies on the detection of biases in the number of reads, which are observable at natural DNA termini compared with the rest of the phage genome. We implemented our method with the creation of the software PhageTerm and validated it using a set of phages with well-established packaging mechanisms representative of the termini diversity, i.e. 5′cos (Lambda), 3′cos (HK97), pac (P1), headful without a pac site (T4), DTR (T7) and host fragment (Mu). In addition, we determined the termini of nine Clostridium difficile phages and six phages whose sequences were retrieved from the Sequence Read Archive. PhageTerm is freely available (https://sourceforge.net/projects/phageterm), as a Galaxy ToolShed and on a Galaxy-based server (https://galaxy.pasteur.fr).

Genome replication dynamics of a bacteriophage and its satellite reveal strategies for parasitism and viral restriction

“…Intriguingly, the reads peak at near the end of ICP1's annotated genome was prominently visible at 4 minutes post infection, before ICP1 replication has taken place ( Figure 2C). We speculated that this reads peak corresponds with the terminus of infecting ICP1 particles prior to genome circularization as it has previously been established that termini can lead to sequencing biases following DNA library preparation (35). We found that this reads bias was also present in DNA from purified phage particles (Supplementary Figure S4).…”

“…We found that this reads bias was also present in DNA from purified phage particles (Supplementary Figure S4). PhageTerm, which is designed to identify phage termini and packaging methods (35), identified this reads peak as a packaging (pac) site and predicted a headful packaging mechanism for ICP1. The terminus is located in a 1.3kb orfless space between gp1 and gp2 (Supplementary Figure S4A and B).…”

Novel haloarchaeal viruses from Lake Retba infecting Haloferax and Halorubrum species

“…To determine the packaging mechanism employed by HFTV1, we analysed the bias in distribution of the 1,657,094 sequencing reads along the HFTV1 genome using PhageTerm, a tool that relies on the detection of biases in the number of sequencing reads observable at natural DNA termini compared with the rest of the viral genome (Garneau et al ., ). The analysis revealed a pattern of sequencing read coverage consistent with a circularly permuted, terminally redundant genome and headful packaging mechanism initiated from a pac site, similar to that of bacteriophage P1 (Supporting Information Fig.…”

Genomic hypervariability of phage Andromeda is unique among known dsDNA viruses

“…All ORFs are encoded off the same strand. Analysis of the Andromeda genome by both the PhageTerm [28] and Li's methods [29] revealed that this phage is terminally redundant and likely utilises a headful mechanism of packaging similar to that of phage P1 (+ve strand analysis (p = 1.17e-4), -ve strand analysis (p = 2.81e-6)). Based on nucleotide homology, the closest relative to Andromeda is the P. tolaasii phage Bf7 [30] with which it shares 83% identity across 85% of its genome ( Fig.…”