Characterization and Genome Sequencing of a Novel Bacteriophage PH101 Infecting Pseudoalteromonas marina BH101 from the Yellow Sea of China

Abstract

A novel Pseudoalteromonas marina bacteriophage, PH101, specifically infecting Pseudoalteromonas BH101 was isolated from the water sample of the Yellow Sea of China using the agar overlay method. 16S rDNA sequence identification was used to identify the host bacteria. Efficiency of infection, multiplicity of infection value, morphological characterization, one-step growth curve, and host range of the bacteriophage were determined. Purified PH101 genomic DNA was extracted and its genome was completely sequenced and analyzed. The phage morphology showed that PH101 belongs to the Myoviridae family with a head of 60 nm in diameter and a tail of 40 nm with a tail fiber of 10-20 nm. Microbiological characterization demonstrated that phage PH101 is stable at a wide range of temperatures (0-70 °C) and showed acid and alkaline resistance (pH 3-12). The one-step growth curve showed a latent period of about 20 min, a rise period of 20 min, and a burst size of about 31.6 virions. The genome sequencing and bioinformatic analysis shows that phage PH101 was a novel bacteriophage which was found to consist of a linear, double-stranded 131,903-bp DNA molecule with a GC content of 37.36 % and 228 putative open reading frames without RNA, which were classified into seven functional groups, including phage structure, adsorption, packaging, gene transfer protease, terminase, DNA binding, and regulation.

Isolation and Complete Genome Sequence of a Novel Pseudoalteromonas Phage PH357 from the Yangtze River Estuary

“…The molecular identification of the bacterial isolate was obtained via 16S rRNA gene sequence analysis [26].…”

“…Phages were purified by picking a single plaque, and suspending the plaque in SM buffer [100 mM NaCl, 8 mM MgSO 4 , 50 mM Tris HCl (pH 7.5)], incubating for 1 h at 37°C, titering and plating by the soft-agar overlay method. The purification step was repeated for three times and the purified phages were amplified and stored at 4°C [7,16,26].…”

“…Stained particles were observed by a JEOL Model JEM-1200EX TEM at 100 kV. Phage dimensions were estimated based on the electron micrographs [16,26].…”

“…The genus Pseudoalteromonas is a marine group of bacteria belonging to class Gammaproteobacteria [26]. Members of the Pseudoalteromonas are motile by a single polar flagellum and have a chemoheterotrophic metabolism [17].…”

“…Members of the Pseudoalteromonas are motile by a single polar flagellum and have a chemoheterotrophic metabolism [17]. Pseudoalteromonas strains are ubiquitous in the ocean including cold environments or other extreme marine habitats, such as deep-sea and polar regions, and many strains have been isolated [21,26,27,31]. Pseudoalteromonas strains play an important role in marine ecosystems due to their ability to form marine biofilms and produce large quantities of bioactive compounds with antifouling, antimicrobial, or algicidal activities [3,20,34].…”