Isolation, Identification and Biological Characteristics Analysis of a New Proteus Phage

To isolate, identify and characterize a new Proteus phage and analyze its genome.

The strains isolated from yak slaughtering in the laboratory were cultured, their DNA was extracted and subjected to 16S rRNA sequencing, and the host bacteria were identified and used for bacteriophage separation from the environmental sewage of a yak slaughterhouse in Chengdu, Sichuan. The bacteriophage was separated and purified by the double-layer agar plate method, and its optimal multiplicity of infection (MOI), one-step growth curve and tolerance were determined. Its morphology was observed by negative staining with phosphoric acid and transmission electron microscopy (TEM). The whole genomic DNA of the phage was extracted using a commercial kit, sequenced on the Illumina MiSeq sequencing platform, assembled, annotated, and genomically analyzed.

Totally 10 Proteus strains were identified from the 67 isolates, and one phage with strong lytic activity, named Proteus phage PV66, was isolated from the sewage. PV66 could lyse Proteus and Enterococcus faecalis. The optimal MOI was 0.1. The results of one-step growth curve showed that the incubation period of bacteria infected with PV66 was about 10 min, the lytic period was about 80 min, and the average lytic amount was 321 PFU/cell. The tolerance pH range was 3-12, and the tolerance temperature was 65 ℃. The bacteriophage had a C3 morphotype under electron microscopy, with a head length of (140± 1) nm, a width of (54± 2) nm, and a tail length of (36± 1) nm. The genome sequencing results showed that the whole sequence length of PV66 was 90492 bp. The phylogenetic tree analysis showed that the phage had high homology with Proteus mirabilis phage and Cronobacter phage and was in the same branch as non-Kuravirus.

The Proteus phage has good application potential. This study provides a basis for future development of new bacteriophages, and enriches the database of Proteus phages.