Genome Sequencing, Annotation and Peculiarities of Bacteriophages J1 and PL1

MARIA EUGENIA DIETERLE; CHARLES BOWMAN; DEBORAH JACOBS-SERA; DANIEL A. RUSSELL; GRAHAM HATFULL; MARIANA PIURI

Mar del Plata

Congreso; VIII Congreso de Microbiología General; 2012

SAMIGE

Bacteriophages of acid lactic bacteria are the main cause of fermentation failure in the dairy industry. Phage J1 was originally isolated in 1965 from an abnormal fermentation of Yakult using the strain Lactobacillus casei ?Shirota? as starter. Two years later, phage PL1 was isolated from a strain resistant to J1. Since its isolation, both phages have been well characterized but its genome sequence was not available.We have sequenced and annotated the complete genome of phages J1 and PL1 using 454 Sequencing and DNA Master program. The J1 genome is 40,931bp in length and contains 64 protein-coding genes. The PL1 genome is 38,880 bp in length and contains 59 orfs. Interestingly, the DNA sequences of these genes are very closely related although PL1 has a 1,9 Kb deletion at the genetic switch module. Remarkable features were revealed from the sequences. As consequence of ribosomal frameshifting, two head proteins and also two putative chaperones with the same NH2 termini are coded.  Even J1 was originally described as lytic, a gene that codes for an integrase of the tyrosine-recombinase family and a putative phage attachment site (attP) within an intergenic region were found indicating that this phage could have a lysogenic life cycle in the right host. The presence of turbid plaques in L.casei BL23 and sequence homology between the possible attP region and the bacterial genome (possible attB) support this idea. We have isolated two possible lysogens that do not form plaques in J1 but had an efficiency of plating of 10-4 in PL1.  We are currently studying if the phage integration occurs in the expected place. Surprisingly, phage PL1 showed two different sequences for the same genome region between the positions 13858 and 14221. One is at least two times more represented than the other. The majority sequence is present in L.casei BL23 and BD-II but not in other phages suggesting that during its evolution PL1 has incorporated this region by recombination with the bacterial genome. On the other side, the minority sequence was identical to J1. Both sequences are embedded in putative tail components. By isolation of individual plaques of PL1, we described the presence of a mix population. In some plaques phages carrying one of the sequences were present and in others both sequences could be detected. Further characterization is in progress to understand the maintenance of these two populations in bacteriophage PL-1.