Biomedical subjects

L L McKay

Publications and source records attributed to L L McKay.

At least 55 records · Page 3Linked to original sources

PubMed1988

Improvement in symptoms and left ventricular performance after balloon aortic valvuloplasty in patients with aortic stenosis and depressed left ventricular ejection fraction.

The application of balloon aortic valvuloplasty to elderly patients with severe aortic stenosis and a low ventricular ejection fraction is undefined. Balloon aortic valvuloplasty was performed in a subset of 28 patients with low left ventricular ejection fraction (37 +/- 11%), severe aortic stenosis, and a mean age of 79 +/- 5 years. After valvuloplasty, significant increases were seen in aortic valve area (from 0.5 +/- 0.1 to 0.9 +/- 0.2 cm2), aortic systolic pressure (from 120 +/- 12 to 135 +/- 22 mm Hg), and cardiac output (from 4.2 +/- 1.1 to 4.8 +/- 1.6 l/min) (p less than 0.01), and significant decreases were seen in transaortic pressure gradient (from 69 +/- 25 to 35 +/- 15 mm Hg) and pulmonary capillary wedge pressure (from 24 +/- 9 to 20 +/- 7 mm Hg) (p less than 0.01). All patients had symptomatically improved at the time of discharge. Serial radionuclide ventriculography showed an increase in left ventricular ejection fraction from 37 +/- 11% before valvuloplasty to 44 +/- 14% within 48 hours after dilatation and to 49 +/- 13% at 3 months after dilatation. However, there was substantial heterogeneity of response. Thirteen patients (group A) showed progressive increases in left ventricular ejection fraction (from 34 +/- 11% to 49 +/- 15% to 58 +/- 11%, p less than 0.0001), whereas 15 patients (group B) showed no significant change in ejection fraction (from 41 +/- 10% to 40 +/- 13% to 41 +/- 10%, p = NS) over 3 months. There was no difference between these groups with respect to age, extent of coronary artery disease, history of myocardial infarction, and aortic valve area before and after valvuloplasty. However, peak systolic wall stress and left ventricular dimensions were higher in group B compared with group A. In conclusion, balloon aortic valvuloplasty may result in symptomatic improvement in patients with aortic stenosis and depressed left ventricular ejection fraction; some patients develop progressive increases in ejection fraction, whereas others fail to show improvement.

Aged↗

PubMed1988

Plasmid-mediated reduced phage sensitivity in Streptococcus lactis KR5.

The phage insensitivity of Streptococcus lactis KR5 was evaluated for its possible linkage to plasmid DNA. This strain possessed plasmids of 40, 29, 26, 21, 16.5, 10.5, 7.8, and 1.5 Mdal. Plasmid curing using novobiocin resulted in derivatives with increased sensitivity to prolate-headed phage, suggesting the involvement of plasmid DNA in phage insensitivity. Transformation of S. lactis LM0230 protoplasts with the KR5 plasmid DNA pool produced transformants containing a plasmid of about 27 Mdal. These erythromycin-resistant transformants were lactose-positive phage-sensitive or were lactose-negative and exhibited a reduced sensitivity to phage. Agarose gel electrophoresis and restriction endonuclease digestion analysis showed the 27-Mdal plasmid band to be composed of two distinct plasmids of 26 Mdal (pBF61) and 29 Mdal (pBF62), which coded for reduced phage sensitivity and lactose-positive phenotypes, respectively. The mechanisms of reduced phage sensitivity encoded by pBF61 included a restriction/modification system and a mechanism that resulted in reduced plaque size independent of incubation temperature. These results further support the involvement of plasmid DNA in the mechanisms for reduced phage sensitivity in dairy streptococci.

Bacteriophages↗

PubMed1987

Restriction enzyme analysis of lactose and bacteriocin plasmids from Streptococcus lactis subsp. diacetylactis WM4 and cloning of BclI fragments coding for bacteriocin production.

The 131.1-kilobase (kb) bacteriocin production (Bac) plasmid pNP2 and the 63.6-kb lactose metabolism (Lac) plasmid pCS26, from Streptococcus lactis subsp. diacetylactis WM4, as well as pWN8, a 116.7-kb recombinant plasmid from a Lac+ transconjugant, were analyzed with restriction enzymes to determine the origin of pWN8. Plasmid pWN8 conferred a Lac+ Bac- phenotype, contained DNA derived from pCS26 and pNP2, and, like pNP2, exhibited self-transmissibility (Tra+). In cloning attempts, Bac+ transformant S. lactis KSH1 was isolated. The recombinant plasmid, pKSH1, contained three BclI fragments from pNP2. Bac- transformants which individually contained each of the three fragments were also identified. Comparison of restriction maps of pKSH1 and pNP2 revealed an 18.4-kb region common to both plasmids, involving two of the three BclI fragments. S. lactis KSH1 also exhibited greater inhibitory activity against the indicator strain S. diacetylactis 18-16 than did a strain containing the 131.1-kb Bac plasmid.

Bacteriocin Plasmids↗

PubMed1987

Combined percutaneous aortic valvuloplasty and transluminal coronary angioplasty in adult patients with calcific aortic stenosis and coronary artery disease.

Of 120 consecutive balloon aortic valvuloplasty procedures for critical aortic stenosis, valvuloplasty was performed in combination with coronary angioplasty in nine patients (average age 76 years). All nine patients were symptomatic with angina and congestive heart failure before combined procedures. Aortic valvuloplasty was performed with 20 to 23 mm balloon catheter advanced retrogradely from the femoral artery and resulted in an improvement in peak aortic valve gradient (60 +/- 19 to 33 +/- 13 mm Hg; p less than or equal to .01) and calculated aortic valve area (0.7 +/- 0.1 to 1.1 +/- 0.3 cm2; p less than or equal to .01). Single-vessel coronary angioplasty was performed via the femoral approach, with 2.0 to 3.5 mm balloon catheters, and resulted in a mean reduction of a critical coronary stenosis in each patient from 91 +/- 4% to 29 +/- 8%. The site of coronary angioplasty was the left anterior descending artery in three patients, the circumflex artery in three patients, the right coronary artery in two patients, and a bypass graft to the right coronary artery in one patient. Combined procedures were performed with a mean arterial time of 108 min. Complications included groin hematomas (n = 2), transient left bundle branch block (n = 1), and transient atrial fibrillation (n = 1). No patient experienced prolonged chest pain, myocardial infarction, major increase in aortic insufficiency, or embolic phenomena. Eight of the nine patients treated with combined procedures noted significant improvement in symptoms of angina and congestive heart failure and were discharged.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗

PubMed1987

Spontaneous release of temperate phage by relysogenized lactose-positive transductants of Streptococcus lactis C2.

Transduction for lactose-fermenting ability between the lactose-positive lysogen Streptococcus lactis LM0221 and plasmid-cured, prophage-cured, lactose-negative S. lactis LM2301 resulted in the appearance of lactose-positive transductants surrounded by a zone of clearing in the lactose-negative cell lawn. By using plaque assay procedures, the zones were shown to contain bacteriophage particles, and both spontaneous release and UV induction of prophage from these transductants were demonstrated. The DNA hybridization confirmed that LM2301 did not contain the prophage in its chromosome and that the zone-producing transductant KZ1 was relysogenized by the temperate bacteriophage. Further, a 4.35 Kb EcoRI digestion fragment appeared to contain the DNA sequences for integration into the chromosome and may provide a means for stabilizing cloned DNA by effecting chromosomal insertion in LM2301 derivatives. The selection of zone-producing lactose-positive transductants of LM2301 provided a means for detecting strains relysogenized by the temperate phage induced from LM0221.

Bacteriophages↗

PubMed1986

Partial characterization of the genetic basis for sucrose metabolism and nisin production in Streptococcus lactis.

We attempted to identify the genetic loci for sucrose-fermenting ability (Suc+), nisin-producing ability (Nip+), and nisin resistance (Nisr) in certain strains of Streptococcus lactis. To obtain genetic evidence linking the Suc+ Nip+ Nisr phenotype to a distinct plasmid, both conjugal transfer and transformation were attempted. A conjugation procedure modified to protect the recipients against the inhibitory action of nisin allowed the conjugal transfer of the Suc+ Nip+ Nisr marker from three Suc+ Nip+ Nisr donors to various recipients. The frequency of transfer ranged from 1.7 x 10(-4) to 5.6 x 10(-8) per input donor, depending on the mating pair. However, no additional plasmid DNA was apparent in these transconjugants. Transformation of S. lactis LM0230 to the Suc+ Nip+ Nisr phenotype by using the plasmid pool of S. lactis ATCC 11454 was not achieved, even though other plasmids present in the pool were successfully transferred. However, two results imply the involvement of plasmid DNA in coding for the Suc+ Nip+ Nisr phenotype. The Suc+ Nip+ Nisr marker was capable of conjugal transfer to a recipient deficient in host-mediated homologous recombination (Rec-), and the Suc+ Nip+ Nisr marker exhibited bilateral plasmid incompatibility with a number of lactose plasmids found in S. lactis. Although our results indicate that the Suc+ Nip+ Nisr phenotype is plasmid encoded, no physical evidence linking this phenotype to a distinct plasmid was obtained.

Conjugation, Genetic↗

PubMed1986

Cloning and expression of the beta-D-galactosidase gene from Streptococcus thermophilus in Escherichia coli.

The beta-D-galactosidase (beta-gal) gene from Streptococcus thermophilus was cloned to isolate and characterize it for potential use as a selection marker in a food-grade cloning vector. Chromosomal DNA from S. thermophilus 19258 was cleaved with the restriction enzyme PstI and ligated to pBR322 for transformation into Escherichia coli JM108. A beta-galactosidase-positive clone was detected by its blue color on a medium supplemented with 5-bromo-4-chloro-3-indolyl-beta-D-galactoside. This transformant possessed a single plasmid, designated pRH116, which contained, in addition to the vector DNA, a 7.0-kilobase (kb) PstI insertion fragment coding for beta-gal activity. An extract from JM108(pRH116) contained a beta-gal protein with the same electrophoretic mobility as the beta-gal from S. thermophilus 19258. Compared with the beta-gal from E. coli HB101, the S. thermophilus beta-gal was of lower molecular weight. A restriction map of pRH116 was constructed from cleavage of both the plasmid and the purified insert. The construction of deletion derivatives of pRH116 with BglII, BstEII, and HindIII revealed the approximate location of the gene on the 7.0-kb fragment. The beta-gal gene was further localized to a 3.85-kb region.

Chromosome Mapping↗

PubMed1985

Lactate metabolism by pediococci isolated from cheese.

Pediococcus pentosaceus is commonly found among the adventitious microflora of Cheddar cheese. When this organism was incubated with L-(+)-lactate under anaerobic conditions, L-(+)-lactate was rapidly converted to D-(-)-lactate until racemic (DL) lactate was present. Under aerobic conditions this initial reaction was followed by a slower reaction resulting in the use of both lactate isomers and in the production of acetate and CO2. With intact cells the lactate oxidation system had an optimum pH of 5 to 6, depending on the initial lactate concentration. Cells grown anaerobically possessed lactate-oxidizing activity which increased two- to fourfold as sugar was exhausted from the medium. Aerobic growth further increased specific activities. Cheddar cheese was made with the deliberate addition of P. pentosaceus. When the resulting cheese was grated to expose a large surface area to O2, lactate was converted to acetate at a rate which depended on the density of pediococci in the cheese. The lactate oxidation system remained active in cheese which had been ripened for 6 months.

Acetates↗

PubMed1985

Plasmid DNA in Strains of Pediococcus cerevisiae and Pediococcus pentosaceus.

Five parental strains of Pediococcus were examined for plasmid content. Each strain contained three to six resident plasmids, ranging in size from 4.5 to 39.5 megadaltons. A bacteriocin-like substance produced by Pediococcus cerevisiae FBB63 was tentatively linked to a 10.5-megadalton plasmid after being cured with novobiocin.

Journal Article↗

PubMed1985

Isolation and partial characterization of plasmid DNA from Streptococcus thermophilus.

Of 23 strains of Streptococcus thermophilus examined, 5 were found to contain a single small cryptic plasmid, designated pHM1 through pHM5. Through analysis by restriction endonuclease mapping and DNA-DNA hybridization, the five plasmids were found to be closely related. They were present in 4 to 18 copies per cell and ranged in size from 1.4 to 2.2 megadaltons. Plasmids pHM1 and pHM5, as well as pHM2 and pHM4, were found to be identical. Single restriction endonuclease sites were observed on each plasmid for PvuII and MboI. Plasmids pHM2 and pHM3 each had an additional single site for HhaI, and pHM1 had an additional single site for HindIII. The characteristics of these plasmids may make them useful for the development of cloning vectors for use in S. thermophilus.

Base Sequence↗

PubMed1985

Galactose transport in Streptococcus thermophilus.

Although Streptococcus thermophilus accumulated [14C]lactose in the absence of an endogenous energy source, galactose-fermenting (Gal+) cells were unable to accumulate [14C]galactose unless an additional energy source was added to the test system. Both Gal+ and galactose-nonfermenting (Gal-) strains transported galactose when preincubated with sucrose. Accumulation was inhibited 50 or 95% when 10 mM sodium fluoride or 1.0 mM iodoacetic acid, respectively, was added to sucrose-treated cells, indicating that ATP was required for galactose transport activity. Proton-conducting ionophores also inhibited galactose uptake, although N,N'-dicyclohexyl carbodiimide had no effect. The results suggest that galactose transport in S. thermophilus occurs via an ATP-dependent galactose permease and that a proton motive force is involved. The galactose permease in S. thermophilus TS2b (Gal+) had a Km for galactose of 0.25 mM and a Vmax of 195 micromol of galactose accumulated per min per g (dry weight) of cells. Several structurally similar sugars inhibited galactose uptake, indicating that the galactose permease had high affinities for these sugars.

Biological Transport↗

PubMed1985

Galactokinase activity in Streptococcus thermophilus.

ATP-dependent phosphorylation of [14C]galactose by 11 strains of Streptococcus thermophilus indicated that these organisms possessed the Leloir enzyme, galactokinase (galK). Activities were 10 times higher in fully induced, galactose-fermenting (Gal+) strains than in galactose-nonfermenting (Gal-) strains. Lactose-grown, Gal- cells released free galactose into the medium and were unable to utilize residual galactose or to induce galK above basal levels. Gal+ S. thermophilus 19258 also released galactose into the medium, but when lactose was depleted growth on galactose commenced, and galK increased from 0.025 to 0.22 micromol of galactose phosphorylated per min per mg of protein. When lactose was added to galactose-grown cells of S. thermophilus 19258, galK activity rapidly decreased. These results suggest that galK in Gal+ S. thermophilus is subject to an induction-repression mechanism, but that galK cannot be induced in Gal- strains.

Galactokinase↗

PubMed1984

Conjugative 40-megadalton plasmid in Streptococcus lactis subsp. diacetylactis DRC3 is associated with resistance to nisin and bacteriophage.

Streptococcus lactis subsp. diacetylactis DRC3 was examined for plasmid DNA and found to contain a previously unreported plasmid of 40 X 10(6) daltons. This plasmid, designated pNP40, was conjugally transferred to a plasmid-cured derivative of S. lactis C2. Transconjugants containing pNP40 acquired resistance to nisin produced by strains of S. lactis and to commercially available nisin when assay plates were incubated at 21, 32, and 37 degrees C. In addition, c2 phage growth was completely restricted in transconjugants containing pNP40 at 21 and 32 degrees C, but not at 37 degrees C. This result suggests that pNP40 may be coding for a temperature-sensitive enzyme that restricts phage growth at 21 and 32 degrees C, but not at 37 degrees C. Eight consecutive transfers of a transconjugant containing pNP40 in Elliker broth at 37 degrees C resulted in 100% loss of resistance to c2 phage when colonies were tested at 32 degrees C. These phage-sensitive isolates had lost pNP40 and had also become sensitive to nisin. This result suggests that pNP40 may also be thermosensitive in its replication. The finding of a phage resistance determinant located on a conjugative plasmid should prove useful in constructing phage-resistant variants for dairy fermentation processes.

Bacteriophages↗

PubMed1984

In Vivo Cloning of lac Genes in Streptococcus lactis ML3.

The isolation and characterization of a Streptococcus lactis ML3 strain which possessed a recombinant lactose plasmid is described. The recombination events generating this plasmid occurred in vivo in a recombination-deficient strain and appeared to be mediated by transposition events. Restriction mapping revealed that the recombinant plasmid, pDA0307, contained a region of the lactose plasmid, pSK08, linked to another resident plasmid, pSK07. Copy number determinations indicated that the lac genes were present at approximately 20 copies per cell in pDA0307, whereas the lac genes are normally present at approximately 10 copies per cell in pSK08. The strain containing pDA0307 displayed a 21 to 54% increase in the expression of the Lac enzyme phospho-beta-d-galactosidase. However, the strain containing pDA0307 both grew and produced lactic acid in milk at rates identical to that of a strain containing pSK08. This result suggests that lac gene dosage of plasmid-linked lac genes was not limiting the rate at which these derivatives of S. lactis ML3 fermented milk.

Journal Article↗

PubMed1984

Plasmid transformation of Streptococcus lactis protoplasts: optimization and use in molecular cloning.

The parameters affecting polyethylene glycol-induced plasmid transformation of Streptococcus lactis LM0230 protoplasts were examined to increase the transformation frequency. In contrast to spreading protoplasts over the surface of an agar medium, their incorporation into soft agar overlays enhanced regeneration of protoplasts and eliminated variability in transformation frequencies. Polyethylene glycol with a molecular weight of 3,350 at a final concentration of 22.5% yielded optimal transformation. A 20-min polyethylene glycol treatment of protoplasts in the presence of DNA was necessary for maximal transformation. The number of transformants recovered increased as the protoplast and DNA concentration increased over a range of 3.0 X 10(6) to 3.0 X 10(8) protoplasts and 0.25 to 4.0 micrograms of DNA per assay, respectively. With these parameters, transformation was increased to 5 X 10(3) to 4 X 10(4) transformants per microgram of DNA. Linear and recombinant plasmid DNA transformed, but at frequencies 10- to 100-fold lower than that of covalently closed circular DNA. Transformation of recombinant DNA molecules enabled the cloning of restriction endonuclease fragments coding for lactose metabolism into S. lactis LM0230 with the Streptococcus sanguis cloning vector, pGB301. These results demonstrated that the transformation frequency is sufficient to clone plasmid-coded genes which should prove useful for strain improvement of dairy starter cultures.

Base Sequence↗

PubMed1984

Transformation of Streptococcus sanguis Challis with Streptococcus lactis plasmid DNA.

Streptococcus lactis plasmid DNA, which is required for the fermentation of lactose (plasmid pLM2001), and a potential streptococcal cloning vector plasmid (pDB101) which confers resistance to erythromycin were evaluated by transformation into Streptococcus sanguis Challis. Plasmid pLM2001 transformed lactose-negative (Lac-) mutants of S. sanguis with high efficiency and was capable of conferring lactose-metabolizing ability to a mutant deficient in Enzyme IIlac, Factor IIIlac, and phospho-beta-galactosidase of the lactose phosphoenolpyruvate-phosphotransferase system. Plasmid pDB101 was capable of high-efficiency transformation of S. sanguis to antibiotic resistance, and the plasmid could be readily isolated from transformed strains. However, when 20 pLM2001 Lac+ transformants were analyzed by a variety of techniques for the presence of plasmids, none could be detected. In addition, attempts to cure the Lac+ transformants by treatment with acriflavin were unsuccessful. Polyacrylamide gel electrophoresis was used to demonstrate that the transformants had acquired a phospho-beta-galactosidase characteristic of that normally produced by S. lactis and not S. sanguis. It is proposed that the genes required for lactose fermentation may have become stabilized in the transformants due to their integration into the host chromosome. The efficient transformation into and expression of pLM2001 and pDB101 genes in S. sanguis provides a model system which could allow the development of a system for cloning genes from dairy starter cultures into S. sanguis to examine factors affecting their expression and regulation.

Acriflavine↗

PubMed1984

Molecular cloning of the lactose-metabolizing genes from Streptococcus lactis.

Restriction endonucleases and agarose gel electrophoresis were used to analyze plasmid pLM2001, which is required for lactose metabolism by Streptococcus lactis LM0232. The enzymes XhoI, SstI, BamHI, and KpnI each cleaved the plasmid into two fragments, whereas EcoRI and BglII cleaved the plasmid into seven and five fragments, respectively. Sizing of fragments and multiple digestions allowed construction of a composite restriction map. The KpnI fragments of pLM2001 were cloned into the KpnI cleavage site of the vector plasmid pDB101. A recombinant plasmid (pSH3) obtained from a lactose-fermenting, erythromycin-resistant (Lac+ Eryr) transformant of Streptococcus sanguis Challis was analyzed by enzyme digestion and agarose gel electrophoresis. Plasmid pSH3 contained 7 of the 11 KpnI-HindIII fragments from pLM2001 and 5 of the 7 fragments from pDB101. It was determined that a 23-kilobase (kb) KpnI-generated fragment from pLM2001 had been cloned into pDB101 with deletion of part of the vector plasmid. The recombinant plasmid could be transformed with high frequency into several Lac- strains of S. sanguis, conferring the ability to ferment lactose and erythromycin resistance. The presence of pSH3 allowed a strain deficient in Enzyme IIlac, Factor IIIlac, and phospho-beta-galactosidase of the lactose phosphoenolpyruvate-dependent phosphotransferase system to efficiently ferment lactose. Under conditions designed to maximize curing of plasmid DNA with acriflavin, no Lac- derivatives could be isolated from cells transformed with pSH3. Seven of the 40 Lac+ colonies isolated after 10 transfers in acriflavin were shown to be sensitive to erythromycin and did not appear to harbor plasmid DNA.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗

PubMed1984

Genetic and physical characterization of recombinant plasmids associated with cell aggregation and high-frequency conjugal transfer in Streptococcus lactis ML3.

Restriction mapping was employed to characterize the 104-kilobase (kb) cointegrate lactose plasmids from 15 independent transconjugants derived from Streptococcus lactis ML3 as well as the 55-kb lactose plasmid ( pSK08 ) and a previously uncharacterized 48.4-kb plasmid ( pRS01 ) from S. lactis ML3. The data revealed that the 104-kb plasmids were cointegrates of pSK08 and pRS01 and were structurally distinct. The replicon fusion event occurred within adjacent 13.8- or 7.3-kb PvuII fragments of pSK08 and interrupted apparently random regions of pRS01 . Correlation of the transconjugants' clumping and conjugal transfer capabilities with the interrupted region of pRS01 identified pRS01 regions coding for these properties. In the 104-kb plasmids, the pRS01 region was present in both orientations with respect to the pSK08 region. The replicon fusion occurred in recombination-deficient (Rec-) strains and appeared to introduce a 0.8 to 1.0-kb segment of DNA within the junction fragments. The degeneration of the cointegrate plasmids was monitored by examining the lactose plasmids from nonclumping derivatives of clumping transconjugants. These plasmids displayed either precise or imprecise excision of pRS01 sequences or had dramatically reduced copy numbers. Both alterations occurred by rec-independent mechanisms. Alterations of a transconjugant 's clumping phenotype also occurred by rec-independent inversion of a 4.3-kb KpnI-PvuII fragment within the pRS01 sequences of the cointegrate plasmid.

Base Sequence↗