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Biomedical subjects

D H Steinberg

Publications and source records attributed to D H Steinberg.

14 recordsLinked to original sources
PubMed2003

Pediocin production by recombinant lactic acid bacteria.

Production of the anti-listerial bacteriocin, pediocin, by lactic acid bacteria (LAB) transformed with the cloning vector pPC418 (Ped+, 9.1 kb) was influenced by composition of media and incubation temperature. Maximum pediocin production, tested against Listeria innocua, by electrotransformants of Lactococcus lactis ssp. lactis was measured in tryptone/lactose/yeast extract medium after 24 h growth at 30 degrees C, while incubation at 40 degrees C was optimum for Ped+ transformants of Streptococcus thermophilus and Enterococcus faecalis. The amount of pediocin produced by S. thermophilus in skim milk and cheese whey supplemented with 0.5% yeast extract was estimated as 51,000 units ml(-1) and 25,000 units ml(-1), respectively. Pediocin production remained essentially unchanged in reconstituted skim milk or whey media diluted up to 10-fold. The results demonstrate the capacity of recombinant strains of LAB to produce pediocin in a variety of growth media including skim milk and inexpensive cheese whey-based media, requiring minimum nutritional supplementation.

Animals↗
PubMed1999

Promoter activity of the pER341-borne ST(Phsp) in heterologous gene expression in Escherichia coli and Streptococcus thermophilus(1).

A 138-bp EcoO109/HinfI fragment of Streptococcus thermophilus plasmid pER341 (2798 bp) including the promoter sequence of the heat stress protein gene hsp16.4 was tested in vector constructs for ability to activate the promoterless green fluorescent protein gene (gfp) from a jelly fish in Escherichia coli, S. thermophilus, and lactococci. ST(Phsp) promoted gfp expression in transformed hosts as evidenced by the presence of green fluorescent (GFP(+)) colonies under UV illumination. The results confirmed the potential of ST(Phsp) as a functional promoter in heterologous gene expression in dairy fermentation bacteria.

Base Sequence↗
PubMed1999

Distribution of plasmid-borne stress protein genes in Streptococcus thermophilus and other lactic acid bacteria.

The presence of heat stress protein genes (hsp) was tested by Southern hybridization analysis in total DNA extracts from species of the genus Streptococcus (47 strains), Lactobacillus (34 strains), Lactococcus (24 strains), and Leuconostoc (5 strains). The biotinylated hsp16.4 probe prepared from an ORF2 fragment of pER341 (2.8 kb) tested positively with restricted DNA extracts of seven Streptococcus thermophilus strains and a single strain of Lactococcus lactis subsp. cremoris. In all positive S. thermophilus strains, the hsp was located on plasmids ranging from ca. 2.8 kb to 11 kb in size, while hsp was present in a 7.5-kb plasmid in Lactococcus lactis subsp. cremoris. Southern blots with a rep probe showed that all hsp16.4+ plasmids in S. thermophilus strains also shared homology with the replication function (rep) of pER341, suggesting the common origin of these plasmids.

Bacterial Proteins↗
PubMed1998

Molecular characterization of the erythromycin resistance plasmid pPV142 from Staphylococcus simulans.

The 2.5-kb erythromycin resistance (EmR) plasmid pPV142 of Staphylococcus simulans 13044 was isolated and characterized. Sequence analysis identified ORF1 and ORF2 encoding a 158-residue replication protein (Rep142) and a 244-residue erythromycin resistance protein (Erm, rRNA adenine N-6-methyltransferase), respectively. Structural analysis and Southern hybridization showed that the rep and ermM genes in pPV142 shared homology with the EmR plasmid pPV141 (2.4 kb) of S. chromogenes 3688 and other EmR plasmids known to exist in staphylococci and bacilli. Based on the presence of a 61-bp repeat upstream of the ermM gene, pPV142 is apparently a unique member of the pSN2 family of EmR plasmid able to express erythromycin resistance constitutively.

Amino Acid Sequence↗
PubMed1998

Structural and functional properties of the hsp16.4-bearing plasmid pER341 in Streptococcus thermophilus.

The plasmid pER341 (2798 bp) of Streptococcus thermophilus ST134 was sequenced and its open reading frame (ORF) regions were characterized. Analysis of nucleotide sequences showed the putative translation product of ORF1 (rep) sharing a high level of homology with replication proteins of several small plasmids present in lactic acid bacteria and staphylococci. This and homology of regions of plus-strand (ORI) and minus-strand (ssoA) origin of replication with pC194-class plasmids indicated that pER341 replicates by the rolling-circle mechanism. ORF2 corresponded to a putative hsp gene that apparently encodes Hsp16.4, a 142-amino-acid heat stress protein. Hsp16.4 shared significant identity with other small, 18-kDa-class heat stress proteins from prokaryotic and eukaryotic sources. Hsp16.4 is apparently the first plasmidborne low-molecular-weight heat stress protein reported in dairy fermentation bacteria with a potential role in temperature-regulated functions in S. thermophilus.

Amino Acid Sequence↗
PubMed1998

Permeabilization of Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus with ethanol.

Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus cultures were treated with ethanol and tested for viability and beta-galactosidase activity. Exposure of the biomass of test cultures to 30%-55% ethanol (vol/vol) caused a 100% loss of viability and up to 15-fold increase in measurable beta-galactosidase activity in both streptococci and lactobacilli. Ethanol-treated cell suspensions could be stored for up to 6 months without loss of enzyme activity. The nonviable permeabilized biomass of the more active S. thermophilus was used to achieve up to 80% hydrolysis of lactose in aqueous solutions and non-fat milk.

Anti-Infective Agents, Local↗
PubMed1997

Molecular properties of the erythromycin resistance plasmid pPV141 from Staphylococcus chromogenes.

The 2.3-kb erythromycin resistance (EmR) plasmid pPV141 of Staphylococcus chromogenes 3688 was isolated and characterized. Nucleotide sequence analysis identified ORF1 and ORF2 separated by a 445-bp spacing, encoding a 158-residue replication protein (Rep141) and a 244-residue erythromycin resistance protein (Erm, rRNA adenine N-6-methyltransferase), respectively. Structural analysis and Southern hybridization showed that the rep and ermM genes in pPV141 shared homology with other known EmR plasmids. Based on sequence analysis, pPV141 was classified as a unique member of the pSN2 family of EmR plasmids.

Amino Acid Sequence↗
PubMed1997

Diastolic dysfunction of the left ventricle. A review of the physiology, causes, diagnosis, treatment and implications.

Diastolic dysfunction of the left ventricle frequently occurs in people with left ventricular hypertrophy and coronary artery disease. It is a common cause of congestive heart failure, especially in the elderly. The mechanism of diastolic dysfunction, its causes, diagnosis and treatment, are reviewed. These are important factors to a Medical Director who must assess the results of non-invasive studies. Alerted by the possibility of diastolic dysfunction, the Medical Director can be more sensitive to other signs and symptoms that may represent early signs of congestive heart failure or ischemia.

Aged↗
PubMed1995

Native promoter-plasmid vector system for heterologous cholesterol oxidase synthesis in Streptococcus thermophilus.

The cholesterol oxidase gene (choA) of a streptomycete was used as a model for studying heterologous gene expression in Streptococcus thermophilus, an essential bacterium in dairy food fermentations. The vectors pER82 and pER82P were developed from the 2.2-kb indigenous plasmid (pER8) of S. thermophilus ST108, and sP1, a 51-bp synthetic promoter patterned after a chromosomal sequence of S. thermophilus. The presence of sP1 promoter in pER82PbCOb with the choA insert aligned with the cat gene was essential for the intracellular production of cholesterol oxidase. The pER82PbCOb was apparently stable in S. thermophilus with no detectable evidence of deletion mutational events.

Blotting, Southern↗
PubMed1992

Construction and characterization of shuttle plasmids for lactic acid bacteria and Escherichia coli.

The chimeric plasmid pBN183 was first constructed in Escherichia coli by ligating the BamHI-digested E. coli plasmid pBR322 and a Bg/II-linearized streptococcal plasmid, pNZ18. The pBN183 transformed E. coli to ApR at a frequency of (8.2 +/- 1.2) x 10(5) colony forming units (CFU)/microgram DNA. Electrotransformation of Streptococcus thermophilus with pBN183 yielded CmR, ApS clones at a frequency of (2.6 +/- 0.3) x 10(1) CFU/microgram DNA. Plasmid screening with pBN183-transformed S. thermophilus clones revealed that ca. 70% of these transformants contained deleted plasmids. Plasmid pBN183A, a pBN183 deletion mutant lacking one copy of a tandemly arranged, highly homologous DNA sequence, was isolated for further study. It transformed E. coli to ApR and S. thermophilus to CmR with frequencies of (4.8 +/- 0.1) x 10(5) and (8.1 +/- 0.2) x 10(2) CFU/microgram DNA, respectively. Screening of S. thermophilus transformants did not show the presence of deleted plasmids. Based on the structure of pBN183A, a new shuttle plasmid, pDBN183, was constructed from pBN183 by removal of the small (1.2 kb) Sa/I fragment. Transformation frequencies of pDBN183 were (5.0 +/- 1.3) x 10(5) and (4.6 +/- 0.2) x 10(2) CFU/microgram DNA with E. coli and S. thermophilus, respectively. In contrast to the parent pBN183, only 17% of the pDBN183-transformed S. thermophilus contained deleted plasmids. Plasmid copy numbers of the three vectors in E. coli were estimated at 17-18 per chromosome. The three plasmids conferred ApR and CmR to E. coli, but only CmR to S. thermophilus. The insertion of a Streptomyces cholesterol oxidase gene (choA) into pDBN183 did not affect the plasmid's stability in Lactobacillus casei, but resulted in deletion of the recombinant DNA in S. thermophilus.

Cholesterol Oxidase↗
PubMed1991

Transfer and expression of a Streptomyces cholesterol oxidase gene in Streptococcus thermophilus.

The recombinant plasmid pNCO937 (8.1 kbp) containing a Streptomyces sp. cholesterol oxidase gene was introduced into Streptococcus thermophilus by electrotransformation. Transformation frequency was 7.2 x 10(5) colony forming units/micrograms of DNA. The presence of the cholesterol oxidase gene in S. thermophilus was confirmed with Southern blot analysis using a biotinylated probe. Thin-layer chromatographic analysis showed the expression of the Streptomyces cholesterol oxidase gene resulting in the oxidation of cholesterol to 4-cholesten-3-one. S. thermophilus may be a suitable host for the expression of other genes regulating prokaryotic cholesterol metabolism.

Animals↗
PubMed1988

Genetic transformation of Streptococcus thermophilus by electroporation.

A rapid and convenient electroporation procedure was developed for the genetic transformation of intact cells of Streptococcus thermophilus with various species of plasmid DNA. Transformation frequency was influenced by the capacitance and voltage selected for electric pulsing, the pH and composition of the electroporation medium and the molecular mass of the transforming DNA. Electroporation is a simple and effective technique to introduce plasmid DNA into S. thermophilus and useful in the development of recombinant DNA technology for this important industrial microorganism.

Buffers↗
PubMed1973

Thermal inactivation of staphylococcal enterotoxins B and C.

Thermal inactivation profiles of staphylococcal enterotoxins B (SEB) and C (SEC) at 80, 100, and 121 C showed that SEC is more resistant than SEB to heat. After 24 h of incubation at 25 C, some reactivation (recovery of serological reactivity) occurred in toxins that had been inactivated by heat. If the toxin was stirred during heating, reactivation did not occur. An examination of the reactivation kinetics of heat-treated SEC showed that reactivation was temperature dependent. At 25 C, the incubation temperature of heat-treated crude SEC (80 C for 10 min), 100% reactivation occurred after 24 h, whereas at 4 C only slight reactivation was observed. We and others observed that heat-treated toxins initially lost more serological activity when heated at a low temperature (80 C) than at a higher temperature (100 C); in the present study we demonstrate that this is a reversible phenomenon.

Culture Media↗