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At least 19 recordsLinked to original sources

Role of temperate phage in determining lytic phage sensitivity and serotype of Vibrio cholerae.

The effect of lysogenization with five temperate phages from various sources on serotype and lytic phage sensitivity was investigated in six cultures of Vibrio cholerae of both classical and El Tor biotypes. No changes in serotype or in classical phage sensitivity in the classical biotype were observed. Four of the temperate phages were homoimmune and induced resistance to one of the El Tor typing phages, E3, thereby causing a type change in El Tor strains. The sensitivity to the other phages was not changed. In 14 natural isolates too, E3 (group III) phage resistance correlated with the presence of temperate phage. Postadsorption exclusion was found to be the mechanism of resistance involved. The fifth phage, VcA-1, had a unique immunity profile. It could infect the El Tor biotype of V. cholerae but caused no change in the host properties investigated.

Bacteriophages↗

Properties and partial genetic characterization of Nepean phage and other lytic phages of Brucella species.

Nepean (Np), a new brucellaphage, was associated with atypical Brucella abortus strains from Ontario cattle. Carriage of Np was associated with loss of smooth lipopolysaccharide, changes in some protein bands in acrylamide gel electrophoresis profiles, increased susceptibility to colistin, and increased resistance to ultraviolet killing. Nepean (Np) was compared with brucellaphages Tb, Fi, Wb, Iz and R/C. All were morphologically identical, with icosahedral capsids (50-65 nm diameter) and short tails (15-25 nm long), but Np had a more restricted host range, replicating only in smooth strains of B. abortus. All six brucellaphages were generally similar in resistance to chemical and physical agents. Brucellaphage DNA was double stranded and unmethylated; its molecular size was 38 kilobase pairs. The DNAs of Tb, Fi, Wb, Iz and R/C could not be differentiated by restriction endonuclease digest profiles produced by BgII, EcoRI, HindIII or PvuII. Nepean (Np) DNA was very similar to that of the other brucellaphages, but with every enzyme used its profile differed in the number and/or position of at least one fragment. However, there was complete cross-hybridization of Tb and Np DNAs. Hybridization techniques failed to detect Brucella DNA in Dp or Tb phages, or phage DNA in Brucella cells. Extrachromosomal plasmid DNA was not detected.

Animals↗

Structure of colitis phage lytic enzyme gene.

A sequence of 163 amino acids was derived from the nucleotide sequence of a DNA fragment bearing the colitis phage lytic enzyme gene. The molecular weight, basicity and the sequence of the three N-terminal amino acids agreed with those of the phage lytic enzyme. The amino acids at the N-terminal of the colitis phage enzyme are homologous with those of T4, P22 and phi 29 phage lysozymes, to the extent of 18-34%. The transcription initiation site was mapped at 166 nucleotides upstream to the translation initiation codon, ATG. The Shine-Dalgarno sequence, the Pribnow box and the RNA polymerase recognition site were at 5.10 and 35 nucleotdies respectively, upstream to the transcription initiation site. The transcription termination signal, (the inverted repeat) and the termination site (the oligo TS) were at 7 and 28 nucleotides respectively, downstream to the translation termination site, TAA.

Amino Acid Sequence↗

Cloning and sequence determination of a phi AAU2 gene whose product aborts the phage lytic cycle.

This communication describes the cloning of a 1.8-kb fragment from the genome of the corynephage phi AAU2, which aborts the phage lytic cycle when cloned on a high-copy shuttle vector. The associated phenotype, called Apld (aborting phage lytic development), was revealed by noting the reduced plaque size and lower efficiencies of plaquing of phi AAU2 cp, a virulent derivative of phi AAU2, on "Arthrobacter aureus"-C70 Apld+ cells. Adsorption and phage DNA transfection experiments showed evidence that Apld acted once the phage DNA had entered into the cell; apld was confined to a single open reading frame (ORF), encoding a putative 63-aa polypeptide which did not show any homology to proteins contained in the databanks; apld is followed by an ORF the product of which shows homology with a protein expressed by the early region of the Streptomyces phage phi C31.

Amino Acid Sequence↗

Characterization of a temperate Streptococcus thermophilus bacteriophage and its genetic relationship with lytic phages.

The temperate Streptococcus thermophilus bacteriophage phi SFi21 showed an 38-kb-long double-stranded DNA genome with cohesive ends. A single integration site was used in lysogens established in three different S. thermophilus strains. The attP and attB sites were localized on the restriction map of phage DNA and by hybridization on pulsed field separated bacterial DNA. All laboratory-established lysogens showed in addition to integrated prophage DNA unintegrated monomer phage DNA with unligated cos sites. The genetic relatedness of phi SFi21 DNA with DNA from lytic phages was studied in dot blot and Southern blot hybridization by using individual restriction fragments of phiSFi21 DNA as probes. Lytic group I phages hybridized with fragments of the central and the right part of the phiSFi21 genome but failed to hybridize with a fragment joining both parts. Lytic group II phages showed hybridization with the right half of the phiSFi21 genome. In lytic group IV phages, biologically a heterogeneous group, many different combinations of cross hybridization were detected in accordance with the hypothesis of the modular evolution of phage genomes.

Attachment Sites, Microbiological↗

A note on the isolation and propagation of lytic phages from Streptococcus uberis and their potential for strain typing.

Thirty-eight of 98 strains of Streptococcus uberis were shown to be carrying lysogenic phage. Although propagating strains were rare, host modification by field strains sensitive to phage was used to increase the lytic spectra. When 120 nationally-collected strains were challenged with 25 phages, selected on the basis of differing lytic spectra and propagating strains, 30% were susceptible to at least one phage, increasing to 42% when 480 strains from a single farm were considered. A typing system based on susceptibility to lytic phage was considered feasible.

Animals↗

Properties of Brucella-phages lytic for non-smooth Brucella strains.

A series of host-range mutants has been selected for brucella-phage R. Two of these mutants designated R/O and R/C have been used for typing purposes. Phage R/O is lytic for non-smooth strains of Brucella abortus and for B. ovis. It is genetically unstable however and produces mutants lytic for smooth B. obortus and B. suis. Phage R/C is lytic for non-smooth B. abortus and for B. ovis and B. canis. It is much more stable than phages R or R/O and shows little or no lytic activity on smooth Brucella strains. It has been effective in differentiating B. canis from B. suis in tests on a limited number of strains. In their properties, all of the brucella-phages of the R series resemble their parent phage.

Adsorption↗

Characterisation of a new phage lytic for both smooth and non-smooth Brucella species.

A brucella phage of the Izatnagar series, designated Iz1, was lytic for all Brucella species that are normally smooth, although the efficiency of plating varied between biovars and species. The phage was also lytic for rough strains of B melitensis and B suis and, to a lesser extent, B ovis. It displayed negligible lytic activity towards B canis and rough B abortus cultures. In its morphological and serological properties and its stability to inactivating agents, the Iz1 phage resembled other brucella phages.

Bacteriophages↗

Proposals for optimization of the international phage typing system for Listeria monocytogenes: combined analysis of phage lytic spectrum and variability of typing results.

Combined analysis of 5,179 serial phage reactions of 20 Listeria monocytogenes propagating strains over 14 years and phage typing results from 2,659 further L. monocytogenes strains allowed us to estimate lytic spectrum specificity and the variability of the lytic reactions of 35 phages. These included the 26 phages recommended for the international method for phage typing defined in 1985 by Rocourt et al. (J. Rocourt, A. Audurier, A. L. Courtieu, J. Durst, S. Ortel, A. Schrettenbrunner, and A. G. Taylor, Zentralbl. Bakteriol. Abt. 1 Orig. A 259:489-497, 1985). The results are discussed individually for each phage. Proposals for modifying the present system are made with the aim of producing an optimal bacteriophage set for routine use.

Bacteriophage Typing↗

Isolation and properties of a phage lytic for non-smooth Brucella organisms.

A phage for non-smooth cells of Brucella abortus was isolated from a mixture of three brucella-phages incubated with rough brucella cells in the presence of N-methyl-N'-nitro-N-nitrosoguanidine. It did not lyse smooth cells of Br. abortus strains nor those of other Brucella species, but during the course of its replication in rough organisms a small proportion of phage mutants were produced which were similar in properties to smooth-specific phages. The rough-specific phage, R, itself resembled Weybridge phage in its morphological and serological properties. Phage R was found to contain DNA and segregated into high and low density fractions, both with plaque-forming activity, on ultra-centrifugation in CsSO4 gradients. Unlike the smooth-specific phages it attached to heat-labile receptors present on non-smooth brucella cells.

Adsorption↗

Recombinant P4 bacteriophages propagate as viable lytic phages or as autonomous plasmids in Klebsiella pneumoniae.

We demonstrate the use of bacteriophage P4 as a molecular cloning vector in Klebsiella pneumoniae. A hybrid P4 phage, constructed in vitro, that contains a K. pneumoniae hisDG DNA fragment can be propagated either as a lytic viable specialized transducing phage or as an autonomous, self-replicating plasmid. Hybrid P4 genomes existing as plasmids can be readily converted into non-defective P4-hybrid phage particles by superinfection with helper phage P2. Infection of a K. pneumoniae hisD non-P2 lysogen with P4-hisD hybrid phage results in approximately 90% of the infected cells becoming stably transduced to HisD+. Because P4 interferes with P2 growth, high titre stocks of P4 hybrid phages are relatively free (less than or equal to 10(-6) of P2 contamination. The hisG gene product was detected in ultraviolet light irradiated host cells infected by the P4-hisDG hybrid phage. A mutant of P4 (P4sid1) that directs the packaging of P4 DNA into P2 sized capsids should permit the construction of hybrid phages carrying 26 kilobase inserts.

Bacteriophages↗

Development of an expression strategy using a lytic phage to trigger explosive plasmid amplification and gene expression.

A novel plasmid-based expression strategy, exploiting two features of lytic bacteriophages, was developed in Lactococcus lactis. Components of this system include a phage origin of replication and phage expression signals, which were induced to high efficiency upon phage infection of the host. Phage-specific expression signals were cloned from phi 31 in a promoter-screening strategy using the lacZ gene from Streptococcus thermophilus. One clone exhibited a significant induction in beta-galactosidase production and concomitant increase in lacZ mRNA during the phi 31 infection cycle of the host. Molecular characterization of the cloned insert revealed 888 bp positioned near the phi 31 cos site. Primer extension analysis showed that transcription was induced approximately 20 min following phi 31 infection at four points, apparently organized in two sets of tandem promoters on the cloned phage insert. One of these middle phage promoters also showed a basal level of activity prior to phage infection. The phi 31 promoter lacZ cassette was cloned into a low-copy-number vector plasmid containing the phi 31 origin of replication (ori31) and the resulting low-copy-number plasmid exhibited negligible beta-galactosidase production in L. lactis. However, > 2,000 units were detected following a deliberate infection with phi 31. A control expression plasmid without ori31 could only be induced to 85 units. The combination of these phage-inducible expression signals together with ori31 functioned synergistically to drive rapid and high efficiency expression of a heterologous gene in L. lactis.

Amino Acid Sequence↗

Phage HK022 Roi protein inhibits phage lytic growth in Escherichia coli integration host factor mutants.

Temperate coliphage HK022 requires integration host factor (IHF) for lytic growth. The determinant responsible for this requirement was identified as a new gene (roi) located between genes P and Q. This gene encodes a DNA-binding protein (Roi) containing a helix-turn-helix motif. We have shown that Roi binds a site within its own gene that is closely linked to an IHF binding site. By gel retardation experiments, we have found that IHF binding stabilizes the interaction of Roi with its gene. We have isolated three independent phage mutants that are able to grow on an IHF- host. They carry different mutations scattered in the roi gene and specifying single amino-acid changes. The interactions of all three Roi mutant proteins with the Roi binding site differed from that of the wild type. Roi displays strong similarities, in its C-terminal half, to two putative DNA-binding proteins of bacteriophage P1: Ant1 and KilA. The mode of action of the Roi protein and the possibility that IHF is modulating the expression and/or the action of Roi are discussed.

Amino Acid Sequence↗

Genetic analysis of chromosomal regions of Lactococcus lactis acquired by recombinant lytic phages.

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 phi31. HindIII restriction maps of the variants (phi31.1, phi31.2, phi31.7, and phi31.8) were generated, and these maps revealed the regions containing recombinant DNA. The recombinant region of phage phi31.1, the variant that occurred most frequently, was sequenced and revealed 7.8 kb of new DNA compared with the parent phage, phi31. This region contained numerous instances of homology with various lactococcal temperate phages, as well as homologues of the lambda recombination protein BET and Escherichia coli Holliday junction resolvase Rus, factors which may contribute to efficient recombination processes. A sequence analysis and phenotypic tests revealed a new origin of replication in the phi31.1 DNA, which replaced the phi31 origin. Three separate HindIII fragments, accounting for most of the recombinant region of phi31.1, were separately cloned into gram-positive suicide vector pTRK333 and transformed into NCK203. Chromosomal insertions of each plasmid prevented the appearance of different combinations of recombinant phages. The chromosomal insertions did not affect an inducible prophage present in NCK203. Our results demonstrated that recombinant phages can acquire DNA cassettes from different regions of the chromosome in order to overcome Abi defenses. Disruption of these regions by insertion can alter the types and diversity of new phages that appear during phage-host interactions.

Bacteriolysis↗