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H Neve

Publications and source records attributed to H Neve.

15 recordsLinked to original sources
PubMed2000

Gene cloning and expression and secretion of Listeria monocytogenes bacteriophage-lytic enzymes in Lactococcus lactis.

Bacteriophage lysins (Ply), or endolysins, are phage-encoded cell wall lytic enzymes which are synthesized late during virus multiplication and mediate the release of progeny virions. Bacteriophages of the pathogen Listeria monocytogenes encode endolysin enzymes which specifically hydrolyze the cross-linking peptide bridges in Listeria peptidoglycan. Ply118 is a 30.8-kDa L-alanoyl-D-glutamate peptidase and Ply511 (36.5 kDa) acts as N-acetylmuramoyl-L-alanine amidase. In order to establish dairy starter cultures with biopreservation properties against L. monocytogenes contaminations, we have introduced ply118 and ply511 into Lactococcus lactis MG1363 by using a pTRKH2 backbone. The genes were expressed under control of the lactococcal promoter P32, which proved superior to other promoters (P21 and P59) tested in this study. High levels of active enzymes were produced and accumulated in the cytoplasmic cell fractions but were not released from the cells at significant levels. Therefore, ply511 was genetically fused with the (SP)slpA nucleotide sequence encoding the Lactobacillus brevis S-layer protein signal peptide. Expression of (SP)slpA-ply511 from pSL-PL511 resulted in secretion of functional Ply511 enzyme from L. lactis cells. One clone expressed an unusually strong lytic activity, which was found to be due to a 115-bp deletion that occurred within the 3'-end coding sequence of (SP)slpA-ply511, which caused a frameshift mutation and generated a stop codon. Surprisingly, the resulting carboxy-terminal deletion of 80 amino acids in the truncated Ply511 Delta(S262-K341) mutant polypeptide strongly increased its lytic activity. Proteolytic processing of the secretion competent (SP)SlpA-Ply511 propeptide following membrane translocation had no influence on enzyme activity. Immunoblotting experiments using both cytoplasmic and supernatant fractions indicated that the enzyme was quantitatively exported from the cells and secreted into the surrounding medium, where it caused rapid lysis of L. monocytogenes cells. Moreover, transformation of pSL-PL511 delta C into L. lactis Bu2-129, a lactose-utilizing strain that can be employed for fermentation of milk, also resulted in secretion of functional enzyme and showed that the vector is compatible with the native lactococcal plasmids.

Bacteriolysis↗
PubMed1998

Comparison of the lysogeny modules from the temperate Streptococcus thermophilus bacteriophages TP-J34 and Sfi21: implications for the modular theory of phage evolution.

A 7.6-kb DNA segment covering the putative lysogeny module of the pac-site-containing temperate Streptococcus thermophilus bacteriophage TP-J34 was sequenced. Sequence alignment with the lysogeny module from the cos-site-containing S. thermophilus bacteriophage phiSfi21 revealed areas of high sequence conservation (e.g., over the int gene), interspersed with regions of low or no sequence similarity (e.g., over the cro gene). Four of the six sharp transition zones from high to low sequence conservation were found within open reading frames coding for the CI repressor, the Anti-repressor, the Immunity protein, and a protein of unknown function. The transition points in the cI and ant genes appear to separate gene segments coding for distinct functional domains of these proteins. In addition, these two transition points were located at or near the deletion sites observed in spontaneous phage phiSfi21 deletion mutants, thus suggesting these transition points as recombinational hotspots. Furthermore, the sequence at the transition point in the cI gene resembles the attachment site of the phage, suggesting the involvement of the phage integrase in at least some of the exchange reactions. Contrary to the initial formulation of the modular theory of phage evolution the unit of the evolutionary exchange in streptococcal phages is not a group of functional genes, but can be as small as a single gene. Exchange reactions can also occur within genes, possibly between gene segments encoding distinct protein domains.

Amino Acid Sequence↗
PubMed1998

Bacillus subtilis develops competence for uptake of plasmid DNA when growing in milk products.

Transformation with plasmid DNA of naturally competent cells of Bacillus subtilis 168 in milk products was studied. Plasmid pMG36enpr, a broad host-range lactococcal vector carrying an erythromycin resistance and the B. subtilis npr gene encoding neutral protease, was taken up by B. subtilis cells grown in UHT chocolate milk. Under these conditions competence was optimal during transition from exponential to stationary growth phase, resulting in 9 x 10(1) transformants per 0.01 microgram DNA. No manipulation of the cells was necessary for competence to develop. When cells were pregrown in synthetic medium, higher transformation rates were obtained in assays, where the subsequent transformation experiments were either done in chocolate milk diluted 1:1 (v/v) with synthetic growth medium (up to 8 x 10(2) transformants) or in undiluted chocolate milk (1 x 10(2) transformants). The number of transformants was reduced to 4 x 10 (1), when diluted milk or flavored milks were used. No transformants were obtained in diluted yoghurt. Controls, in which both the preculturing and the transformation assays were done in synthetic medium, gave the maximum number of transformants (4 x 10(3) transformants per 0.01 microgram DNA).

Bacillus subtilis↗
PubMed1998

Predicting operative risk for coronary artery surgery in the United Kingdom: a comparison of various risk prediction algorithms.

OBJECTIVE: To compare the ability of four risk models to predict operative mortality after coronary artery bypass graft surgery (CABG) in the United Kingdom. DESIGN: Prospective study. SETTING: Two cardiothoracic centres in the United Kingdom. SUBJECTS: 1774 patients having CABG. MAIN OUTCOME MEASURES: Risk factors were recorded for all patients, along with in-hospital mortality. Predicted mortality was derived from the American Society of Thoracic Surgeons (STS) risk program, Ontario Province risk score (PACCN), Parsonnet score, and the UK Society of Cardiothoracic Surgeons risk algorithm. RESULTS: There were significant differences (p < 0.05) between the British and American populations from which the STS risk algorithm was derived with respect to most variables. The observed mortality in the British population was 3.7% (65 of 1774). The mean predicted mortality by STS score, PACCN, Parsonnet score, and UK algorithms were 1.1%, 1.6%, 4.6%, and 4.7% respectively. The overall predictive ability of the models as measured by the area under the receiver operating characteristic curve were 0.64, 0.60, 0.73, and 0.75, respectively. CONCLUSIONS: There are differences between the British and American populations for CABG and the North American algorithms are not useful for predicting mortality in the United Kingdom. The UK Society of Cardiothoracic Surgeons algorithm is the best of the models tested but still only has limited predictive ability. Great care must be exercised when using methods of this type for comparisons of units and surgeons.

Aged↗
PubMed1995

Virion positions and relationships of lactococcal temperate bacteriophage TP901-1 proteins.

The major proteins of phage TP901-1 virion were characterized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and structural relations were determined using specific antibodies, obtained by affinity purification from a polyclonal serum. A 23-kDa protein was identified as the major tail protein, and a 31-kDa molecule as the major head protein, respectively. Labeling experiments with antibodies against two proteins, with molecular masses of 20 and 19 kDa, indicated that they were baseplate-related components. A 72-kDa protein was found to be part of a neck passage structure, which includes a collar. Evidence for the presence of attached whiskers was also obtained. T7 RNA polymerase-mediated expression of the two major proteins confirmed the gene location of the previously sequenced region of the phage genome. The relation to other lactococcal phages was determined by DNA hybridization and antibody probing, showing that despite low DNA similarity, TP901-1 NPS epitopes were detected in both related and unrelated small isometric-headed phages.

Antibodies, Viral↗
PubMed1993

Direct electron microscopy study on the morphological diversity of bacteriophage populations in lake plusssee.

Direct electron microscopy of bacteriophages adsorbed to a carbon film without prior enrichment by specific host strains or concentration by physical or chemical methods was used to study the morphological diversity of natural bacteriophage assemblages in a North German lake. All samples contained a mixture of morphologically different tailed viruses, which were regarded as bacteriophages. Most of them had isometric heads and long noncontractile tails, belonging to morphotype B1 (Siphoviridae). In addition, members of morphotypes A1 (Myoviridae), B2 (Siphoviridae with elongated heads), and C1 (Podoviridae) were present in lower numbers. Only one cubic virus was detected, while no filamentous or pleomorphic phages were found. Up to 11 different phages per sample, and a total of 39 phages when all samples were considered together, could be distinguished by morphological criteria. The total number of phages was estimated to be on the order of 10/ml.

Journal Article↗
PubMed1992

Sequencing and characterization of pST1, a cryptic plasmid from Streptococcus thermophilus.

Eighty-six strains of S. thermophilus were examined for their plasmid content. Thirteen strains were found to contain one or two plasmids ranging in size from 2.1 to 7.4 kb. DNA-DNA hybridization analysis revealed the presence of five distinct groups of DNA homology. The complete nucleotide sequence of plasmid pST1 (Accession number X65856), which belongs to the major homology group, was determined. It has a molecular size of 2093 bp, a GC content of 35% and contains one major open reading frame of 945 bp (ORF A). The predicted protein, designated Rep A, showed sequence homology with replication proteins from a group of plasmids which are known to replicate via single-stranded DNA intermediates (ssDNA plasmids).

Amino Acid Sequence↗
PubMed1992

Technical Points in Vertical Gastroplasty for Morbid Obesity.

The success of vertical gastroplasty may be jeopardized by gastric leakage or ulceration due to failure of the technique. Reports of band erosion and staple-line leakage have led us to seek technical improvements to reduce technical failures. We describe a modification to the technique of band placement and a manoeuvre to aid the placement of staples when the TA90B staple gun is used.

Journal Article↗
PubMed1991

Bacteriophage receptors of Lactococcus lactis subsp. 'diacetylactis' F7/2 and Lactococcus lactis subsp. cremoris Wg2-1.

Bacteriophage P008 revealed irreversible and uniform adsorption to cell walls of L. lactis subsp. 'diacetylactis' F7/2, whereas phage P127 adsorbed reversibly to a limited number of receptor sites on cell walls of L. lactis subsp. cremoris Wg2-1. Neither extraction of lipids, cell wall- and membrane-teichoic acids nor enzymatic degradation of proteins altered the binding efficiencies of both cell wall fractions. However, phage binding was inhibited, when cell walls were subjected to lysozyme, metaperiodate, or acid treatments. This reflects that a carbohydrate component embedded in the peptidoglycan matrix is part of the phage receptors of strains F7/2 and Wg2-1.

Adsorption↗
PubMed1991

Species and type phages of lactococcal bacteriophages.

Lactococcal phages are classified according to morphology and DNA homology. Phages are differentiated into 12 phage species, and type phages of each species are proposed. Members and possible members of each species are named. Available data on type phages are tabulated including morphology, DNA characteristics and phage protein bands.

Bacteriophages↗
PubMed1988

Plasmid-encoded functions of ropy lactic acid streptococcal strains from Scandinavian fermented milk.

Ropy Streptococcus (Lactococcus) cremoris strains isolated from a ropy Swedish sour milk ("longfil") and a ropy Finnish milk product ("Viili") were screened for their plasmid-encoded functions. Curing experiments strongly indicated that the ropy phenotype was linked to a 17-Md plasmid in the Swedish strains and to a 30 Md plasmid in the Finnish strains. Comparative restriction endonuclease analysis and DNA/DNA-hybridization studies revelated that plasmids from both strain families shared homologous DNA regions. Though be Swedish ropy strains harbored a conjugative 45-Md lactose plasmid, no cotransfer of the 17 Md plasmid occurred.

Animals↗
PubMed1984

Conjugal transfer and characterization of bacteriocin plasmids in group N (lactic acid) streptococci.

Thirteen bacteriocin-producing strains of group N (lactic acid) streptococci were screened for their potential to transfer this property by conjugation to Streptococcus lactis subsp. diacetylactis Bu2-60. Bacteriocin production in three strains was plasmid encoded as shown by conjugal transfer and by analysis of cured, bacteriocin-negative derivatives of the donor strains and the transconjugants. With Streptococcus cremoris strains 9B4 and 4G6 and S. lactis subsp. diacetylactis 6F7 as donors, bacteriocin-producing transconjugants were isolated with frequencies ranging from ca. 2 X 10(-2) to 2 X 10(-1) per recipient cell. Bacteriocin-producing transconjugants had acquired a 39.6-megadalton plasmid from the donor strains 9B4 and 4G6, and a 75-megadalton plasmid from the donor strain 6F7. As shown by restriction endonuclease analysis, the plasmids from strains 9B4 and 4G6 were almost identical. The plasmid from strain 6F7 yielded some additional fragments not present in the two other plasmids. In hybridization experiments any of the three plasmids strongly hybridized with each other and with some other bacteriocin but nontransmissible plasmids from other S. cremoris strains. Homology was also detected to a variety of cryptic plasmids in lactic acid streptococci.

Bacteriocins↗