An ecological study of lytic bacteriophages of Lactococcus lactis subsp. cremoris isolated in a cheese plant over a five year period

Josephsen, Jytte; Andersen, Nils; Behrndt, H.; Brandsborg, Erik; Christiansen, G.; Hansen, Michael; Hansen, Søren; Nielsen, E. Steemann; Vogensen, Finn K.

doi:10.1016/0958-6946(94)90064-7

Cited by 39 publications

(33 citation statements)

References 24 publications

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“…E. coli and L. lactis plasmid DNA was isolated by using a QIAprep spin plasmid miniprep kit (QIAGEN, Inc., Chatsworth, Calif.) as recommended by the manufacturer, except that L. lactis initially was incubated with lysozyme (20 mg/ml) for 15 min at 37°C. Phage DNA was isolated as described by Josephsen et al (21).…”

Section: Methodsmentioning

confidence: 99%

Identification of the Receptor-Binding Protein in 936-Species Lactococcal Bacteriophages

Dupont

Vogensen

Neve³

et al. 2004

Appl Environ Microbiol

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The aim of this work was to identify genes responsible for host recognition in the lactococcal phages sk1 and bIL170 belonging to species 936. These phages have a high level of DNA identity but different host ranges. Bioinformatic analysis indicated that homologous genes, orf18 in sk1 and orf20 in bIL170, could be the receptor-binding protein (RBP) genes, since the resulting proteins were unrelated in the C-terminal part and showed homology to different groups of proteins hypothetically involved in host recognition. Consequently, chimeric bIL170 phages carrying orf18 from sk1 were generated. The recombinant phages were able to form plaques on the sk1 host Lactococcus lactis MG1614, and recombination was verified by PCR analysis directly with the plaques. A polyclonal antiserum raised against the C-terminal part of phage sk1 ORF18 was used in immunogold electron microscopy to demonstrate that ORF18 is located at the tip of the tail. Sequence analysis of corresponding proteins from other lactococcal phages belonging to species 936 showed that the N-terminal parts of the RBPs were very similar, while the C-terminal parts varied, suggesting that the C-terminal part plays a role in receptor binding. The phages investigated could be grouped into sk1-like phages (p2, fd13, jj50, and 7) and bIL170-like phages (P008, P113G, P272, and bIL66) on the basis of the homology of their RBPs to the C-terminal part of ORF18 in sk1 and ORF20 in bIL170, respectively. Interestingly, sk1-like phages bind to and infect a defined group of L. lactis subsp. cremoris strains, while bIL170-like phages bind to and infect a defined group of L. lactis subsp. lactis strains.Lactococcus lactis is the most important bacterium used for starter cultures by the dairy industry. An important problem in industrial milk fermentation is infection of the starter bacteria by bacteriophages, which leads to bacterial lysis. The consequences of phage infection are fermentation delay, alteration of product quality, and in severe cases loss of the product. All these outcomes result in considerable economic loss to dairies (5,22). Industrial phage ecology is dominated by three phage species: the predominant species 936 and species c2 and P335, which also have considerable importance (5,22).The first step in phage infection is adsorption of the phage to the host cell. Despite the fact that little information concerning the adsorption process of lactococcal phages is available, it seems to be a two-step process that starts with reversible binding to specific carbohydrates exposed on the surface of the cell wall (30, 37, 39), which is followed by, at least in species c2, irreversible binding to a protein in the cell membrane (14,30,38). Information on receptor-binding proteins (RBPs) in phages infecting gram-positive bacteria is sparse compared to the information available for phages infecting gram-negative bacteria. However, recently, Duplessis and Moineau identified the first RBP genes for the Streptococcus thermophilus phages DT1 and MD4 (11). These authors repo...

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Section: Methodsmentioning

confidence: 99%

Identification of the Receptor-Binding Protein in 936-Species Lactococcal Bacteriophages

Dupont

Vogensen

Neve³

et al. 2004

Appl Environ Microbiol

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“…The lack of genome sequences from the less frequently isolated phage species is probably explained by the higher industrial incidences of failed fermentations due to the members of the three above predominant species (37,39).…”

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confidence: 99%

Genome Sequence and Global Gene Expression of Q54, a New Phage Species Linking the 936 and c2 Phage Species ofLactococcus lactis

2006

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The lytic lactococcal phage Q54 was previously isolated from a failed sour cream production. Its complete genomic sequence (26,537 bp) is reported here, and the analysis indicated that it represents a new Lactococcus lactis phage species. A striking feature of phage Q54 is the low level of similarity of its proteome (47 open reading frames) with proteins in databases. A global gene expression study confirmed the presence of two early gene modules in Q54. The unusual configuration of these modules, combined with results of comparative analysis with other lactococcal phage genomes, suggests that one of these modules was acquired through recombination events between c2-and 936-like phages. Proteolytic cleavage and cross-linking of the major capsid protein were demonstrated through structural protein analyses. A programmed translational frameshift between the major tail protein (MTP) and the receptor-binding protein (RBP) was also discovered. A "shifty stop" signal followed by putative secondary structures is likely involved in frameshifting. To our knowledge, this is only the second report of translational frameshifting (؉1) in double-stranded DNA bacteriophages and the first case of translational coupling between an MTP and an RBP. Thus, phage Q54 represents a fascinating member of a new species with unusual characteristics that brings new insights into lactococcal phage evolution.Lactococcus lactis is a low-GϩC gram-positive bacterium extensively used by the dairy industry for its ability to convert sugars into lactic acid, leading to various fermented milk products. However, L. lactis strains are susceptible to attacks by lytic bacteriophages with concomitant low-quality products and economic losses (56). Different strategies have been developed over the past 70 years with the aim of better controlling the indigenous phage population within the dairy environment (54, 71). Nevertheless, the phage population is constantly evolving and new phage isolates are still frequently isolated. Some of these phages are emerging due to current manufacturing practices (49).Classification of the phages is still controversial, and different propositions have been made in recent years (10,65,74). Previous classification studies relied on the comparison of virus morphology and DNA-DNA hybridizations using whole genomes. According to these criteria, 12 lactococcal phage species representing distinct phage groups were proposed (37, 38). This classification of lactococcal phages was recently revisited, and 10 genetically diverse groups of phages, sharing very limited nucleotide similarities, were proposed (26). Among these, two new species were identified, one of which was the type phage Q54.Phage genomics has considerably expanded in the last 5 years, and as of June 2006, 362 complete genomes are available at GenBank. Of those, 14 are from lactococcal phages. Among the available genomes, only members of the three main groups of L. lactis phages are represented, namely, the 936, c2, and P335 species. Ten of the genomes are from phage...

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“…While the 936 species is composed of only lytic phages, P335 species exhibit high levels of DNA homology between temperate and lytic members (17,20). P335 phages have been appearing in cheese plants with increasing frequency in recent years and are now considered members of an important new phage species (1,17,21,28).Workers in our laboratory have previously characterized a number of phages belonging to the P335 species which are virulent for Lactococcus lactis NCK203. In response to the inhibitory pressure of an abortive type of defense (AbiC) (12), phage ul36 can acquire chromosomal sequences from the host in a recombinational process that generates a related but new virulent phage, ul37 (8,29).…”

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confidence: 99%

mentioning

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Genetic Analysis of Chromosomal Regions of Lactococcus lactis Acquired by Recombinant Lytic Phages

Durmaz

Klaenhammer

2000

Appl Environ Microbiol

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Recombinant phages are generated when Lactococcus lactis subsp. lactis harboring plasmids encoding the abortive type (Abi) of phage resistance mechanisms is infected with small isometric phages belonging to the P335 species. These phage variants are likely to be an important source of virulent new phages that appear in dairy fermentations. They are distinguished from their progenitors by resistance to Abi defenses and by altered genome organization, including regions of L. lactis chromosomal DNA. The objective of this study was to characterize four recombinant variants that arose from infection of L. lactis NCK203 (Abi ؉ ) with phage 31. HindIII restriction maps of the variants (31.1, 31.2, 31.7, and 31.8) were generated, and these maps revealed the regions containing recombinant DNA. The recombinant region of phage 31.1, the variant that occurred most frequently, was sequenced and revealed 7. Bacteriophages continue to be a significant economic problem for the dairy industry (9, 30). Starter culture strains which contain naturally occurring phage defense mechanisms can control the problem if they are used in conjunction with sanitation measures. Bacteriophages, nevertheless, have the capacity to evolve and rapidly overcome host defense mechanisms (9,13,22,29). The logical routes of phage evolution include point mutations, deletions, and acquisition of new DNA. Coinfecting phages and the genomic contents of the host cell, which can include functional or defective prophages, are potential sources of new DNA. Identification of the genetic routes by which phages adapt and evolve against industrial starter cultures will be an important part of controlling the appearance of new phages in fermentation environments.Lactococcal phages are classified into 12 species based on morphology and DNA homologies (20). The c2 (prolateheaded) species and the 936 and P335 (small isometricheaded) species are the most important taxa, since these are the major organisms that disrupt dairy fermentations worldwide. While the 936 species is composed of only lytic phages, P335 species exhibit high levels of DNA homology between temperate and lytic members (17,20). P335 phages have been appearing in cheese plants with increasing frequency in recent years and are now considered members of an important new phage species (1,17,21,28).Workers in our laboratory have previously characterized a number of phages belonging to the P335 species which are virulent for Lactococcus lactis NCK203. In response to the inhibitory pressure of an abortive type of defense (AbiC) (12), phage ul36 can acquire chromosomal sequences from the host in a recombinational process that generates a related but new virulent phage, ul37 (8,29). Phage ul37 is not inhibited by AbiC and differs in its base plate and tail morphology. The nature and extent of the sequences and their number or location in the Lactococcus chromosome have not been determined. Knockout insertions directed into the bacterial chromosome in the regions acquired by ul37 eliminated the AbiCinduced metam...

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An ecological study of lytic bacteriophages of Lactococcus lactis subsp. cremoris isolated in a cheese plant over a five year period

Cited by 39 publications

References 24 publications

Identification of the Receptor-Binding Protein in 936-Species Lactococcal Bacteriophages

Identification of the Receptor-Binding Protein in 936-Species Lactococcal Bacteriophages

Genome Sequence and Global Gene Expression of Q54, a New Phage Species Linking the 936 and c2 Phage Species ofLactococcus lactis

Genetic Analysis of Chromosomal Regions of Lactococcus lactis Acquired by Recombinant Lytic Phages

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