Genome sequence of the phage clP1, which infects the beer spoilage bacterium Pediococcus damnosus

Kelly, David J.; O'Sullivan, Órla; Mills, Susan; McAuliffe, Olivia; Ross, R. Paul; Neve, Horst; Coffey, Aidan

doi:10.1016/j.gene.2012.04.085

Cited by 21 publications

(12 citation statements)

References 72 publications

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“…Their genome sizes render them some of the smallest phages infecting L. plantarum together with phage phiJL-1 and phage ATCC 8014-B1 (accession numbers: NC_006936 and NC_019916), while their G/C content ( 42.5%) is close to that of their host ( 44.5%). In the three 37.9-38.8 kbp, double-stranded (ds) DNA phage genomes, all predicted open reading frames (ORFs) were located at the sense strand, which is in accordance with other phages of L. plantarum 42 and the closely-related P. damnosus 43 .The total number of predicted ORFs was 58-62 for the three phages.…”

Section: Basic Genomic Characteristicssupporting

confidence: 66%

Isolation and characterisation of novel phages infectingLactobacillus plantarumand proposal of a new genus, “Silenusvirus”

Kyrkou

Carstens

Ellegaard-Jensen

et al. 2019

Preprint

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Bacteria of Lactobacillus sp. are very useful to humans. However, the biology and genomic diversity of their (bacterio)phage enemies remains understudied. Knowledge on Lactobacillus phage diversity should broaden to develop efficient phage control strategies. To this end, organic waste samples were screened for phages against two wine-related Lactobacillus plantarum strains. Isolates were shotgun sequenced and compared against the phage database and each other by phylogenetics and comparative genomics. The new isolates had only three distant relatives from the database but displayed a high overall degree of genomic similarity amongst them. The latter allowed for the use of one isolate as a representative to conduct transmission electron microscopy and structural protein sequencing, and to study phage adsorption and growth kinetics. The microscopy and proteomics tests confirmed the observed diversity of the new isolates and supported their classification to the family Siphoviridae and the proposal of the new phage genus "Silenusvirus". Methods Environmental samples, phage assays and bacterial strainsLactobacillus phages were isolated from organic household waste samples. The samples were collected from two different organic waste treatment plants in Denmark (treatment plants A and B). Initially, the samples were split into two subsamples and processed as detailed in Kyrkou et al 12 . The resulting filtrates were screened for phages using the double agar overlay method 16 and a top layer of 0.4% w/v agarose. Specifically, efficiency of plating assays were performed against two indicator strains that had earlier been acquired from private collections, L. plantarum L1 (wine fermentation isolate) and L. plantarum MW-1 (grape isolate). Single plaques were resuspended in 0.7 mL SM buffer and later filtered by 0.45-µm pore size PVDF spin filters (Ciro, Florida, USA). Each purified plaque underwent two further reisolation-filtration cycles to ensure pure stock cultures and was stored at 4°C. For transmission electron microscopy (TEM) and protein sequencing, lysates of 10 10 plaque-forming units (PFUs)/mL) were further purified and concentrated using caesium chloride (CsCl) gradient ultracentrifugation, as described elsewhere 17 . All incubations of phage manipulations were done at 25°C using De Man, Rogosa and Sharpe (MRS) broth and agar media supplemented with 10 mM CaCl 2 (MRSΦ), whereas indicator strains were grown in MRS at 37°C. TEM analysis and structural protein sequencingPhage morphology and structural proteins were determined using the CsCl-purified stocks. Micrographs of phage Silenus were generated as in other studies 12,18 . The mean values and standard deviations of all Silenus virion dimensions were elucidated after inspection of 20-23 phage particles. Structural proteins were sequenced following published protocols 12,18 . Briefly, 100 µL of the CsCl-purified stocks were filtered through an Amicon Ultra filter unit (MWCO 30k Da) and desalted four times. Phage particles (10 µL) were denaturised in 6 M urea, ...

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Section: Basic Genomic Characteristicssupporting

confidence: 66%

Isolation and characterisation of novel phages infectingLactobacillus plantarumand proposal of a new genus, “Silenusvirus”

Kyrkou

Carstens

Ellegaard-Jensen

et al. 2019

Preprint

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“…Interestingly, a high level of identity (97%) with the genome of phage clP1, infecting Pediococcus damnosus, followed by 77% identity with the genome of L. plantarum phage JL-1, was found. Of note, the genome of phage clP1 showed a GC content of 47.6%, which is much higher than those reported for pediococci (37.8 to 41.2%) (35). When each ORF was analyzed, high levels of identity with phage clP1 deduced proteins (65 to 100%) were also observed, while the levels of identity with proteins of phage JL-1 were always lower (29 to 80%) ( Table 2).…”

Section: Methodsmentioning

confidence: 77%

Characterization of Two Virulent Phages of Lactobacillus plantarum

Marcó

Garneau

Tremblay

et al. 2012

Appl Environ Microbiol

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We characterized two Lactobacillus plantarum virulent siphophages, ATCC 8014-B1 (B1) and ATCC 8014-B2 (B2), previously isolated from corn silage and anaerobic sewage sludge, respectively. Phage B2 infected two of the eight L. plantarum strains tested, while phage B1 infected three. Phage adsorption was highly variable depending on the strain used. Phage defense systems were found in at least two L. plantarum strains, LMG9211 and WCSF1. The linear double-stranded DNA genome of the pac-type phage B1 had 38,002 bp, a G؉C content of 47.6%, and 60 open reading frames (ORFs). Surprisingly, the phage B1 genome has 97% identity with that of Pediococcus damnosus phage clP1 and 77% identity with that of L. plantarum phage JL-1; these phages were isolated from sewage and cucumber fermentation, respectively. The double-stranded DNA (dsDNA) genome of the cos-type phage B2 had 80,618 bp, a G؉C content of 36.9%, and 127 ORFs with similarities to those of Bacillus and Lactobacillus strains as well as phages. Some phage B2 genes were similar to ORFs from L. plantarum phage LP65 of the Myoviridae family. Additionally, 6 tRNAs were found in the phage B2 genome. Protein analysis revealed 13 (phage B1) and 9 (phage B2) structural proteins. To our knowledge, this is the first report describing such high identity between phage genomes infecting different genera of lactic acid bacteria.

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“…Analysis of structural proteins was performed as described previously (Kelly et al, 2012) using high titer phage suspensions obtained from ammonium acetate concentration. Samples were mixed with 4X sample loading buffer and heated at 95°C prior to loading in a 12% SDS polyacrylamide gel.…”

Section: Methodsmentioning

confidence: 99%

Phages of non-dairy lactococci: isolation and characterization of ΦL47, a phage infecting the grass isolate Lactococcus lactis ssp. cremoris DPC6860

et al. 2014

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Lactococci isolated from non-dairy sources have been found to possess enhanced metabolic activity when compared to dairy strains. These capabilities may be harnessed through the use of these strains as starter or adjunct cultures to produce more diverse flavor profiles in cheese and other dairy products. To understand the interactions between these organisms and the phages that infect them, a number of phages were isolated against lactococcal strains of non-dairy origin. One such phage, ΦL47, was isolated from a sewage sample using the grass isolate L. lactis ssp. cremoris DPC6860 as a host. Visualization of phage virions by transmission electron microscopy established that this phage belongs to the family Siphoviridae and possesses a long tail fiber, previously unseen in dairy lactococcal phages. Determination of the lytic spectrum revealed a broader than expected host range, with ΦL47 capable of infecting 4 industrial dairy strains, including ML8, HP and 310, and 3 additional non-dairy isolates. Whole genome sequencing of ΦL47 revealed a dsDNA genome of 128, 546 bp, making it the largest sequenced lactococcal phage to date. In total, 190 open reading frames (ORFs) were identified, and comparative analysis revealed that the predicted products of 117 of these ORFs shared greater than 50% amino acid identity with those of L. lactis phage Φ949, a phage isolated from cheese whey. Despite their different ecological niches, the genomic content and organization of ΦL47 and Φ949 are quite similar, with both containing 4 gene clusters oriented in different transcriptional directions. Other features that distinguish ΦL47 from Φ949 and other lactococcal phages, in addition to the presence of the tail fiber and the genome length, include a low GC content (32.5%) and a high number of predicted tRNA genes (8). Comparative genome analysis supports the conclusion that ΦL47 is a new member of the 949 lactococcal phage group which currently includes the dairy Φ949.

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Genome sequence of the phage clP1, which infects the beer spoilage bacterium Pediococcus damnosus

Cited by 21 publications

References 72 publications

Isolation and characterisation of novel phages infectingLactobacillus plantarumand proposal of a new genus, “Silenusvirus”

Isolation and characterisation of novel phages infectingLactobacillus plantarumand proposal of a new genus, “Silenusvirus”

Characterization of Two Virulent Phages of Lactobacillus plantarum

Phages of non-dairy lactococci: isolation and characterization of ΦL47, a phage infecting the grass isolate Lactococcus lactis ssp. cremoris DPC6860

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