A novel Halomonas ventosae-specific virulent halovirus isolated from the Qiaohou salt mine in Yunnan, Southwest China

Abstract

Although Halomonas phages belonging to the families Myoviridae and Siphoviridae have been reported, no virulent Halomonas siphoviruses are known. In this study, a virulent bacteriophage, QHHSV-1, of the family Siphoviridae that specifically infects H. ventosae QH52-2 was isolated from the Qiaohou salt mine. Restriction analysis indicated that QHHSV-1 is a dsDNA virus with a genome size of 33.5-39.5 kb. Transmission electron microscopy showed that QHHSV-1 is a typical representative of the Siphoviridae, with an icosahedral head (47 nm in diameter) and a non-contractile tail (75 nm in length). We also assessed the adsorption rate of QHHSV-1 for the host bacterium and found significant inhibition after the addition of 10 mM CaCl2. Based on a one-step growth curve, we determined a latent period of 30 min and a burst size of 73 PFU/infected cell. At the optimal pH of 8.0, 25.9 and 15.2 % of the phages survived after a 60-min incubation at 50 and 60 °C, respectively. Phage replication was possible at a wide range of salt concentrations, from 2.0 to 20 % (w/v), with an optimum concentration of 5 %. The survival of QHHSV-1 at different salt concentrations decreased with time and 25 % survival after 25 days at 30 % salt concentration.

Characterization of a novel bacteriophage specific to Exiguobacterium indicum isolated from a plateau eutrophic lake

“…The double‐layer plates were incubated right‐side‐up at 37 °C and examined for the presence of plaques. Individual plaques were selected and purified three times by repeated single‐plaque propagation on strain EI9 or EA31 .…”

“…A one‐step growth experiment was performed according to to determine both the latent period and the phage burst size. In brief, a phage sample was mixed with 500 µl of a fresh EI9 culture to obtain an MOI of 0.1, and the mixture was incubated at 37 °C for 15 min.…”

“…To examine the thermostability of the phage, the phage stock was incubated at 26, 40, 50, 60, and 70 °C for 50 min at pH 7. The survival rate of each treated phage sample was determined using the double‐layer agar technique .…”

“…Different reagent (SDS, ethanol, and proteinase K) were selected to study characteristic of capsid protein. The method reported by Fu et al was used with some modifications. Chloroform, Trition X‐100, sodium dodecyl sulfate (SDS), and absolute ethanol was added to the phage stock to a final concentration of 25% (w/v), 0.3% (w/v), 0.1% (w/v), and 90% (v/v), respectively, and allowed the corresponding mixtures to stand at room temperature for 5 min, 10 min, 30 min, and 60 min, respectively.…”

“…Phage particle purification was performed following the method of Fu et al . Phage DNA was extracted from purified high‐titer lysates using TIANamp Virus DNA/RNA Kit (TIANGEN) according to the manufacturer's instructions.…”