Complete genome sequence of a giant Vibrio phage ValKK3 infecting Vibrio alginolyticus.

This paper describes the complete sequence of a giant lytic marine myophage, Vibrio phage ValKK3 that is specific to Vibrio alginolyticus ATCC(®) 17749™. Vibrio phage ValKK3 was subjected to whole genome sequencing on MiSeq sequencing platform and annotated using Blast2Go. The complete sequence of ValKK3 genome was deposited in DBBJ/EMBL/GenBank under accession number KP671755.

Direct link to deposited data

www.ncbi.nlm.nih.gov/genome/?term=KP671755

Materials and methods

Vibrio phage ValKK3 was isolated from marine sediment of Kota Kinabalu, Malaysia that was capable to lyse fish and human pathogen, Vibrio alginolyticus ATCC® 17749™ [1]. The phage morphology was determined by a transmission electron microscope. The genome was extracted and purified using DNeasy Blood and Tissue Kit (Qiagen). The genome sequencing and assembly was performed by AITBiotech Pte. Ltd. (Singapore). The DNA library was prepared by Illumina Nextera XT kit. The samples were dual barcoded for multiplexing on the MiSeq instrument. The paired of 250 bp reads (2 × 250,000 samples) were generated and demultiplexed corresponding to the barcodes. The FASQ files were generated on board of MiSeq Instrument. The sequence assembly was performed using de novo assembly using Velvet assembly software version 1.1 (Zerbino, European Bioinformatics Institute, UK). The assembly producing the lowest number of contigs with the largest n50 value was chosen for further assembly with a > 30-fold sequence coverage into single contig. The complete contig sequence was subjected to BLASTn [2] and the sequence was annotated using Blast2GO [3].

Data description

The morphological characteristics of ValKK3 (Fig. 1) showed that it belongs to the family of Myoviridae. To the best of our knowledge, among the giant T4-like phages reported so far (Table 1), the Vibrio phage ValKK3 is the only strain that can infect V. alginolyticus. The genome annotation analysis predicted 390 open reading frames which are numbered consecutively from ORF1 to ORF390 which represented 92.4% of the total genome. The putative genes varied from 111 bp (ORF268) to 4200 bp (ORF349). The protein function analysis showed that some CDSs involve in repression of host gene synthesis (ORF197, ORF202), defense system of own gene synthesis (ORF170, ORF176, ORF207) and quality control (ORF255), DNA synthesis (ORF61, ORF199, ORF367, ORF370, ORF241, ORF256, ORF381), DNA replication and recombination (ORF371, ORF372, ORF320, ORF178, ORF204, ORF106, ORF242, ORF257), structural protein: the head (ORF281–286), tail (ORF287–319) and tail fiber (ORF346–349), DNA packaging protein (ORF289, ORF290) and host lysis protein (ORF304). In this study, the putative functions of ValKK3 genome only represent 15.6% of the genes in the ValKK3. The other 84.4% in the ValKK3 are consisted of the genes which encoded proteins with unknown function or hypothetical proteins. The ValKK3 is similar to the Vibrio phage VH7D, Bacteriophage KVP40, Vibrio phage phi-pp2 and Vibrio phage nt-1 (Table 1). This assigns the ValKK3 to the T4-like phage genus of the Myoviridae family which belongs to the giant vibriophage group [7], [8] with even larger genome size. The blast search revealed that ValKK3 was homologous to vibriophage that is known to infect some members of Harveyi clade bacteria. It is also revealed that T4-like phage is widely spread in the marine environment infecting different Vibrio species. Finally, an additional genome information of ValKK3 is hoped to provide positive contributions to the study of Vibrio bacteriophage.

Fig. 1

Electron micrograph of negative-stained ValKK3. Bars = 100 nm.

Table 1

Genome comparison among giant vibriophages.

Bacteriophage strain	Vibrio phage ValKK3	Vibrio phage VH7D	Bacteriophage KVP40	Vibrio phage phi-pp2	Vibrio phage nt-1
Host species	V. alginolyticus	Vibrio strain 7D	V. parahaemolyticus	V. parahaemolyticus	V. natriegens
GenBank accession no.	KP671755	KC131129	AY283928	JN849462	HQ317393
Head length (nm)	140	155	140	150	120
Head diameter (nm)	79	75	70	90	70
Tail length (nm)	120	Not reported	130	120	110
Genome size (bp)	248,088	246,964	244,834	246,421	247,511
Blastn identity (%)	100	99	81	81	85
ORF number	390	378	386	383	379
G + C level (%)	41.2	41.3	42.6	42.7	41.3
Reference	This study	Luo et al. [4]	Miller et al. [5], Matsuzaki et al. [6]	Lin and Lin [7]	Comeau et al. [8], Zachary [9]

Nucleotide accession number

This complete genome sequence has been deposited at DDBJ/EMBL/GenBank under accession no. KP671755.

Conflict of interest

The authors clarified that this work and writing has no conflict of interest.

Specifications
Organism	Myoviridae
Genus	T4-Like
Sequencer	Illumina (MiSeq)
Data format	Analyzed
Experimental factors	Bacteriophage strain
Experimental features	Whole genome analysis and gene annotation of Vibrio phage ValKK3
Consent	N/A
Sample source location	Marine sediment, Kota Kinabalu, N 06° 02.270′, E 116° 06.710′