Host Range, Morphology and Sequence Analysis of Ten Temperate Phages Isolated from Pathogenic Yersinia enterocolitica Strains

Abstract

Yersinia enterocolitica is a heterogeneous species comprising highly pathogenic, weakly pathogenic and non-pathogenic strains. Previous data suggest that gene exchange may occur in Yersinia. Only scarce information exists about temperate phages of Y. enterocolitica, even though many prophage sequences are present in this species. We have examined 102 pathogenic Y. enterocolitica strains for the presence of inducible prophages by mitomycin C treatment. Ten phages were isolated from nine strains belonging to the bio (B)/serotypes (O) B2/O:5,27, B2/O:9 and 1B/O:8. All phages are myoviruses showing lytic activity only at room temperature. Whole-genome sequencing of the phage genomes revealed that they belong to three groups, which, however, are not closely related to known phages. Group 1 is composed of five phages (type phage: vB_YenM_06.16.1) with genome sizes of 43.8 to 44.9 kb, whereas the four group 2 phages (type phage: vB_YenM_06.16.2) possess smaller genomes of 29.5 to 33.2 kb. Group 3 contains only one phage (vB_YenM_42.18) whose genome has a size of 36.5 kb, which is moderately similar to group 2. The host range of the phages differed significantly. While group 1 phages almost exclusively lysed strains of B2/O:5,27, phages of group 2 and 3 were additionally able to lyse B4/O:3, and some of them even B2/O:9 and 1B/O:8 strains.

The Novel Yersinia enterocolitica Telomere Phage vB_YenS_P840 Is Closely Related to PY54, but Reveals Some Striking Differences

“…However, these two Y. enterocolitica phages exhibit completely different host ranges. Unlike PY54, which was isolated from a serotype O:5 strain and can additionally infect the serotype O:5,27, vB_YenS_P840 was shown to lyse exclusively O:3 strains after testing 50 Y. enterocolitica strains, 30 strains belonging to other Yersinia species as well as various other genera of Enterobacterales, e.g., Escherichia , Klebsiella and Salmonella [ 22 , 28 ]. Six out of ten O:3 strains were lysed by the phage (data not available).…”

“…Thereafter, the lysates were centrifuged for 20 min at 10,000× g to remove agar and debris and then filtered (see above). For purification, vB_YenS_P840 particles were concentrated by ultracentrifugation and applied onto a CsCl step gradient as previously described [ 22 ].…”

“… 1 Additional Yersinia sp. isolates were used for host range determination as previously described ([ 22 ]); Abbreviations: Streptomycin (Str R ), Ampicillin (Amp R ), Chloramphenicol (Cat R ) and Kanamycin (Km R ). …”

“…Upon arrival of the cooled tonsil in the lab, five milliliters of SM buffer were added to the sample, which was then incubated on a stirrer at 4 • C overnight. Thereafter, the material was subjected to centrifugation for 20 min at 8000 rpm and 10 • C. The supernatant was passed through a 0.45 µm pore size filter (VWR International, Darmstadt, Germany) and stored until further use at 4 • C. The determination of lytic activity and the host range of the phage were performed by spotting 10 µL of a serial dilution of the sample onto a lawn of Y. enterocolitica indicator strains belonging to various bio-/serotypes [22]. For this, 100 µL of the respective indicator strain was mixed with 6 mL prewarmed NZCYM (VWR International, Darmstadt, Germany) soft agar (0.6%) and poured onto an LB agar plate [23].…”

Characterization of Temperate LPS-Binding Bordetella avium Phages That Lack Superinfection Immunity

“…Phage DNA was extracted from high-titer stocks (approximately 3 × 10 10 PFU mL −1 ) of phages using a phage DNA isolation kit (Norgen Biotek Corp., Thorold, ON, Canada) according to the manufacturer’s instructions. DNA sequencing was performed at the Federal Institute for Risk Assessment (BfR), Berlin, Germany using Next Generation Sequencing (Illumina) as previously described ( 81 ). DNA sequencing libraries were prepared using the Nextera Flex DNA sample preparation kit and subjected to short-read sequencing (2 × 151 cycles) on an NextSeq 500 (Illumina, San Diego, CA, USA).…”