Isolation and Characterization ofPectobacteriumPhage vB_PatM_CB7: New Insights into the GenusCertrevirus

Abstract

To date,Certrevirusis one of two genera of bacteriophage (phage), with phages infectingPectobacterium atrosepticum, an economically important phytopathogen that causes potato blackleg and soft rot disease. This study provides a detailed description ofPectobacteriumphage CB7 (vB_PatM_CB7), which specifically infectsP. atrosepticum. Host range, morphology, latent period, burst size and stability at different conditions of temperature and pH were examined. Analysis of its genome (142.8 kbp) shows that the phage forms a new species ofCertrevirus,sharing sequence similarity with other members, highlighting conservation within the genus. Conserved elements include a putative early promoter like that of theEscherichia colisigma70 promoter, which was found to be shared with other genus members. A number of dissimilarities were observed, relating to DNA methylation and nucleotide metabolism. Some members do not have homologues of a cytosine methylase and anaerobic nucleotide reductase subunits NrdD and NrdG, respectively. Furthermore, the genome of CB7 contains one of the largest numbers of homing endonucleases described in a single phage genome in the literature to date, with a total of 23 belonging to the HNH and LAGLIDADG families. Analysis by RT-PCR of the HNH homing endonuclease residing within introns of genes for the large terminase, DNA polymerase, ribonucleotide reductase subunits NrdA and NrdB show that they are splicing competent. Electrospray ionization-tandem mass spectrometry (ESI-MS/MS) was also performed on the virion of CB7, allowing the identification of 26 structural proteins-20 of which were found to be shared with the type phages of the genera ofVequintavirusandSeunavirus.The results of this study provide greater insights into the phages of theCertrevirusgenus as well as the subfamilyVequintavirinae.