Stable expression of the<i>Lactobacillus casei</i>bacteriophage A2 repressor blocks phage propagation during milk fermentation

Álvarez, Miguel A.; Rodrı́guez, Ana; Súarez, Juan E.

doi:10.1046/j.1365-2672.1999.00728.x

Cited by 23 publications

(13 citation statements)

References 18 publications

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“…An A2 repressor gene integrated into the chromosome of Lactobacillus casei rendered the strain completely resistant to phage A2 (2,29). Background expression of the phage adh repressor in Lactobacillus gasseri also conveyed complete resistance to phage adh (13).…”

Section: ϫ8mentioning

confidence: 99%

Lactococcus lactis Lytic Bacteriophages of the P335 Group Are Inhibited by Overexpression of a Truncated CI Repressor

et al. 2002

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Phages of the P335 group have recently emerged as important taxa among lactococcal phages that disrupt dairy fermentations. DNA sequencing has revealed extensive homologies between the lytic and temperate phages of this group. The P335 lytic phage 31 encodes a genetic switch region of cI and cro homologs but lacks the phage attachment site and integrase necessary to establish lysogeny. When the putative cI repressor gene of phage 31 was subcloned into the medium-copy-number vector pAK80, no superinfection immunity was conferred to the host, Lactococcus lactis subsp. lactis NCK203, indicating that the wild-type CI repressor was dysfunctional. Attempts to clone the full-length cI gene in Lactococcus in the high-copy-number shuttle vector pTRKH2 were unsuccessful. The single clone that was recovered harbored an ochre mutation in the cI gene after the first 128 amino acids of the predicted 180-amino-acid protein. Bacteriophages continue to be a significant economic problem for the dairy industry. While naturally occurring defenses, the rotation of starter cultures, and improved sanitation measures have been used to combat the problem, phages continue to evolve to overcome host defense mechanisms. The P335 group, one of three major phage groups that remain problematic for Lactococcus lactis in dairy fermentations, includes both lytic and temperate members (21). Sequence homologies have revealed close links and evidence of DNA exchanges between lytic and temperate P335 phages (3,8,9,12,20,28,33,37,44,46). Furthermore, new, recombinant lytic phages have been recovered after acquisition of chromosomal DNA regions from their lactococcal hosts (3, 12, 33).The lactococcal bacteriophage 31 is a small-isometricheaded, cohesive-ended, lytic phage of the P335 group with a double-stranded DNA genome of 31.9 kb (1). Sequencing over 14.3 kb of phage 31 has defined the following gene clusters: a locus involved in sensitivity to the phage resistance mechanism AbiA (10); a late promoter region and a transcriptional activator of the promoter (11,37,(44)(45)(46); the phage replication module (28, 35); part of the lysis module (28); and a genetic switch region (28). The genetic switch region encodes two divergent promoters, P1 and P2, and homologues to cI and cro genes of temperate phages shown in Fig. 1A. Open reading frame (ORF) 238, downstream of the cro gene, has amino acid-level homology to putative antirepressors from the temperate L. lactis phage TP901-1 and Streptococcus thermophilus phage TP-J34. Lack of an integrase gene and an attachment site explains the lytic life cycle of phage 31 (28). The objective of the present study was to investigate the functionality of the phage 31 CI repressor and determine whether or not the repressor could be constitutively overexpressed in L. lactis to retard infection by phages of the P335 group. Interestingly, the CI repressor of this obligatorily lytic phage failed to provide superinfection immunity when cloned and expressed from a medium-copy-number vector. This finding was unexpected, as ...

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Section: ϫ8mentioning

confidence: 99%

Lactococcus lactis Lytic Bacteriophages of the P335 Group Are Inhibited by Overexpression of a Truncated CI Repressor

et al. 2002

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“…It was soon shown by the same research laboratory that this strain was amenable to genetic transformation by electroporation (Chassy & Flickinger, 1987). From this point in time, L. casei ATCC 393 (pLZ15 2 ) became widely used in genetic and physiological studies on L. casei and it has been used for the characterization of lactose, glucose, xylose and sorbose transport and metabolism (Hemme et al, 1994;Veyrat et al, 1994;Gosalbes et al, 1997Gosalbes et al, , 1999Chaillou et al, 1999;Yebra et al, 2000), the characterization of the role of the general components of the PTS and other elements involved in carbon catabolism repression Gosalbes et al, 1997Gosalbes et al, , 1999Yebra et al, 2000;Viana et al, 2000;Dossonnet et al, 2000), as well as for the design of cloning and integration vectors (Leer et al, 1992;Alvarez et al, 1999;Gosalbes et al, 2000Gosalbes et al, , 2001Pérez-Arellano et al, 2001) and the characterization of protein secretion (Hols et al, 1997;Maassen et al, 1999).…”

Section: Introductionmentioning

confidence: 99%

Significant differences between Lactobacillus casei subsp. casei ATCC 393T and a commonly used plasmid-cured derivative revealed by a polyphasic study

Acedo-Félix

Pérez-Martı́nez

2003

International Journal of Systematic and Evolutionary Microbiology

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INTRODUCTIONStrains belonging to the species Lactobacillus casei can be isolated from very different environments, and some of them are now used as probiotics in commercial products. This species is currently involved in a serious controversy and is the subject of numerous taxonomic studies, because many strains previously characterized as L. casei show genotypic, phenotypic and phylogenetic differences when compared to the standing type strain of the species, L. casei subsp. casei ATCC 393 T (Mills & Lessel, 1973;Dellaglio et al., 1975Dellaglio et al., , 1991Dicks et al., 1996;Collins et al., 1989;Ferrero et al., 1996;Mori et al., 1997;Zhong et al., 1998;Chen et al., 2000;Felis et al., 2001).L. casei has been used in a number of pioneering studies on the physiology and genetics of the genus Lactobacillus. The type strain L. casei subsp. casei (L. casei) ATCC 393 T especially has been used in studies on the fermentation of glucose, lactose, citrate and pyruvate (Hegazi & Abo-Elanga, 1987;Palles et al., 1998), comparative studies on and molecular characterization of the enzyme L-lactate dehydrogenase (Gordon & Doelle, 1976;Hensel et al., 1977;Kim et al., 1991), characterization of an intracellular bglucosidase (Coullon et al., 1998), proteolytic activity (Hegazi & Abo-Elanga, 1987) and studies on the composition of the cell wall, antibiotic resistance and adherence factors (Billot-Klein et al., 1997;Pelletier et al., 1997). Also, the isolation and characterization of extrachromosomal genetic elements in the genus Lactobacillus was reported for the first time in strains of L. casei, such as ATCC 393 T , 61BG, 64H, ATCC 334, ATCC 4646 and ATCC 15820 T (Chassy et al., 1976;Chassy & Giuffrida, 1980;Lee-Wickner & Chassy, 1984, 1985. Flickinger et al. (1986) obtained a number of plasmid-cured derivatives from L. casei strains; among these, an ATCC 393 T variant cured of plasmid pLZ15 [ATCC 393 (pLZ15 2 )], which encodes a lactose permease and b-galactosidase, was found (Chassy & Alpert, 1989). Besides the added value of being a derivative of the standing type strain, this plasmid-cured strain showed a great potential for studies on lactose metabolism, as it still carried a second lactose-specific transport and hydrolysis system, the lactose phosphoenolpyruvate : phosphotransferase system (PTS) and a 6-phospho-b-galactosidase. It was soon shown by the same research laboratory that this strain was amenable to genetic transformation by electroporation (Chassy & Flickinger, 1987). From this point in time, L. casei ATCC 393 (pLZ15 2 ) became widely used in genetic and physiological studies on L. casei and it has been used for the characterization of lactose, glucose, xylose and sorbose transport and metabolism (Hemme et al., 1994; Veyrat et al.,Abbreviations: ARDRA, amplified rDNA restriction analysis; ITS, intergenic spacer sequence; L. casei, L. casei subsp. casei; RAPD analysis, random amplified polymorphic DNA.

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“…In this aspect, it differs from the earlier described superinfection immunity systems, where expression of phage repressor genes from phages of various lactic acid bacterial species (e.g. Streptococcus phage Sfi21, Lactococcus phage TP901-1 or Lactobacillus A2 or Фadh phages) provided immunity against the respective single phage only [85,[87][88][89].…”

Section: Subunit Poisoningmentioning

confidence: 87%

“…Superinfection immunity genes (CI repressors) were also identified in phages of other lactic acid bacteria species (e.g. for Lactococcus phage TP901-1 and Lactobacillus phages A2 and Фadh) [87][88][89]. Their expression in trans was also reported to provide immunity against homologous phage infection.…”

Section: Superinfection Immunity and Exclusionmentioning

confidence: 99%

Lactic Acid Bacteria Resistance to Bacteriophage and Prevention Techniques to Lower Phage Contamination in Dairy Fermentation

Szczepankowska¹,

Górecki²,

Bardowski³

2013

Lactic Acid Bacteria - R &Amp; D for Food, Health and Livestock Purposes

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Stable expression of theLactobacillus caseibacteriophage A2 repressor blocks phage propagation during milk fermentation

Cited by 23 publications

References 18 publications

Lactococcus lactis Lytic Bacteriophages of the P335 Group Are Inhibited by Overexpression of a Truncated CI Repressor

Lactococcus lactis Lytic Bacteriophages of the P335 Group Are Inhibited by Overexpression of a Truncated CI Repressor

Significant differences between Lactobacillus casei subsp. casei ATCC 393T and a commonly used plasmid-cured derivative revealed by a polyphasic study

Lactic Acid Bacteria Resistance to Bacteriophage and Prevention Techniques to Lower Phage Contamination in Dairy Fermentation

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