The Linear Plasmid Prophage Vp58.5 of Vibrio parahaemolyticus Is Closely Related to the Integrating Phage VHML and Constitutes a New Incompatibility Group of Telomere Phages

Abstract

Vibrio parahaemolyticus O3:K6 pandemic strains recovered in Chile frequently possess a 42-kb plasmid which is the prophage of a myovirus. We studied the prototype phage VP58.5 and show that it does not integrate into the host cell chromosome but replicates as a linear plasmid (Vp58.5) with covalently closed ends (telomeres). The Vp58.5 replicon coexists with other plasmid prophages (N15, PY54, and ⌽KO2) in the same cell and thus belongs to a new incompatibility group of telomere phages. We determined the complete nucleotide sequence (42,612 nucleotides) of the VP58.5 phage DNA and compared it with that of the plasmid prophage. The two molecules share the same nucleotide sequence but are 35% circularly permuted to each other. In contrast to the hairpin ends of the plasmid, VP58.5 phage DNA contains 5-protruding ends. The VP58.5 sequence is 92% identical to the sequence of phage VHML, which was reported to integrate into the host chromosome. However, the gene order and termini of the phage DNAs are different. The VHML genome exhibits the same gene order as does the Vp58.5 plasmid. VHML phage DNA has been reported to contain terminal inverted repeats. This repetitive sequence is similar to the telomere resolution site (telRL) of VP58.5 which, after processing by the phage protelomerase, forms the hairpin ends of the Vp58.5 prophage. It is discussed why these closely related phages may be so different in terms of their genome ends and their lifestyle.Most temperate bacteriophages integrate into the host chromosome during lysogeny. However, there are some phages (telomere phages) whose prophages are linear plasmids with covalently closed ends. Members of this group of phages are the siphoviruses N15, PY54, and ⌽KO2 isolated from Escherichia coli, Yersinia enterocolitica, and Klebsiella oxytoca, respectively, and the recently described myoviruses ⌽HAP-1 of Halomonas aquamarina and VP882 of Vibrio parahaemolyticus (6,20,23,26,37). Despite their different origins (enterobacteria versus marine bacterium) and morphologies, all known telomere phages share similar genome organizations and some protein similarities. The linear DNA of each phage is a circular permutation of the respective linear plasmid prophage. For the generation of the terminal hairpins of the linear plasmid, the protelomerase (Tel) is essential (8). This enzyme has cleaving/joining activity; its target is a large palindromic DNA sequence called the telomere resolution site (telRL) located upstream of tel on the phage genome. After cleaving telRL by staggered cuts, the resulting self-complementary single-stranded DNA overhangs fold back and are rejoined by the protelomerase (9). Besides tel, all telomere phages possess the gene repA, encoding a multifunctional replication protein. repA of N15 and PY54 was shown to harbor the prophage replication origin and to function as a circular minimal replicon (35,42). Compatibility studies demonstrated that the N15 and ⌽KO2 plasmids belong to the same incompatibility group, whereas the PY54 plasmid is able to coexist...

Identification and characterization of ϕH111-1

“…15 We could not identify any previously characterized phages with ≥ 40% similarity that would belong to the same genus as ϕH111-1. Currently, phages with the most similar proteomes are AcaML1 of Acidithiobacillus caldus (JX507079.1; 28.17% similar) and the "Vhmllikevirus" phages VHML of Vibrio harveyi (NC_004456.1; 28.07% similar) and both [16][17][18][19] Although these phages share subfamily-level similarity, they are very distinct with respect to aspects such as host, gene content, and lifestyle. AcaML1 lysogenizes a thermophilic and acidophilic γ-proteobacterium and has a significantly larger 59 kbp genome with two insertion sequences.…”

“…16 The "Vhmllikevirus" phages VHML, VP58.5, and vB_VpaM_MAR have similar genome sizes to ϕH111-1 (41-43 kbp) but are thought to lysogenize as linear plasmids with telomeres in Vibrio species. [17][18][19] The commonalities among ϕH111-1, AcaML1, and the "Vhmllikevirus" phages are largely restricted to the morphogenesis genes. These phages all have P2-like tail proteins, but encode capsid morphogenesis/DNA packaging and accessory proteins that are unrelated to those of P2 ( Table 2).…”

Two novel temperate bacteriophages co-existing in Aeromonas sp. ARM81 – characterization of their genomes, proteomes and DNA methyltransferases

“…Apart from 'a prototype' -the enterobacteria phage N15, which was isolated in 1964 -only six other phages are known to lysogenize their hosts as linear telomere plasmids (Ravin, 2015). They are the siphoviruses FKO2 of Klebsiella oxytoca (Casjens et al, 2004) and PY54 of Yersinia enterocolitica (Hertwig et al, 2003), and the myoviruses of marine bacteria: VP882, Vp58.5 and vB_VpaM_-MAR of Vibrio parahaemolyticus (Lan et al, 2009;Zabala et al, 2009;Alanis Villa et al, 2012), and FHAP-1 of Halomonas aquamarina (Mobberley et al, 2008). All of them encode a hallmark protein called protelomerase, along with plasmid partitioning proteins ParA and ParB, required for this type of lysogeny.…”

“…All of them encode a hallmark protein called protelomerase, along with plasmid partitioning proteins ParA and ParB, required for this type of lysogeny. Another myovirus, VHML of Vibrio harveyi (Oakey et al, 2002) that is closely related to one of the abovementioned phages, VP58.5 (Zabala et al, 2009) also contains genes encoding homologues of the N15-like protelomerase (Ravin, 2015) and partitioning protein ParA (Oakey et al, 2002). VHML was initially described as a prophage integrated into the host chromosome (Oakey et al, 2002), but later it was suggested that it may be a linear plasmid-like prophage as well (Lima-Mendez et al, 2008).…”

Replication and Maintenance of Linear Phage-Plasmid N15

“…All of them were identified in γ-proteobacteria, and they fall into two groups: the siphoviruses ϕKO2 of Klebsiella oxytoca (15) and PY54 of Yersinia enterocolitica (16), and the myoviruses of marine bacteria: phages VP882, Vp58.5, and vB_VpaM_MAR of Vibrio parahaemolyticus (17)(18)(19) and phage ΦHAP-1 of Halomonas aquamarina (20). All of them are related to N15 and carry a similar genetic determinants responsible for prophage replication and generation of covalently closed telomeres.…”