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Biomedical subjects

R A Welch

Publications and source records attributed to R A Welch.

At least 91 records · Page 5Linked to original sources
PubMed1984

Characterization of Escherichia coli hemolysins conferring quantitative differences in virulence.

Recombinant plasmids encoding hemolysins (hly) isolated from four different Escherichia coli strains were found to be very similar by restriction endonuclease fragment analysis within the hemolysin region. Each of the four recombinant plasmids were used to transform a nonhemolytic fecal strain of E. coli. The comparative virulence of the transformants was tested in a rat model of peritonitis. Despite the physical similarity among the hemolysin recombinant plasmids, each conferred different normally avirulent fecal E. coli strain. Reciprocal exchange of similar restriction endonuclease fragments enabled the construction of hybrid hemolysin determinants with two hemolysins of disparate extracellular hemolysin production and relative virulence levels. The hybrid plasmids were introduced into the standard avirulent fecal strain and tested in the rat peritonitis model. The region conferring quantitative differences in extracellular hemolysin production and virulence was found to be within a region of less than 1 kilobase where the hly c cistron is encoded as well as the probable transcription initiation ares for the entire hemolysin operon. A 750-base pair (bp) AvaI fragment from this region was isolated from a virulent hemolysin recombinant and inserted into a common AvaI site present in an avirulent hemolysin plasmid. This insertion resulted in an increase in the amount of extracellular hemolysin activity and the associated virulence when in the fecal E. coli background.

Animals↗
PubMed1983

Molecular cloning and physical characterization of a chromosomal hemolysin from Escherichia coli.

A chromosomal hemolysin determinant was isolated in the form of recombinant plasmid pSF4000 from a human urinary tract isolate. Escherichia coli J96. A restriction endonuclease fragment map of pSF4000 was constructed. The ampicilin resistance transposon Tn/ was used as a site-specific mutagen in conjunction with in vitro derived deletions to localize the hemolysin determinant to a 7.0-kilobase region of pSF4000. Seventeen hemolytic E. coli isolates from human urinary tract infections were found to share similar DNA sequences with J96 hemolysin sequences, using a hemolysin-specific restriction endonuclease fragment as a hybridization probe. In all 17 hemolytic E. coli strains the hemolysin gene sequences were localized to the bacterial chromosome. The hemolysin-specific gene sequences can be found only in hemolytic strains by colony blot hybridization, suggesting that in the evolutionary sense the hemolysin-specific genetic sequences have only recently been introduced into the E. coli chromosome. Evidence is also presented that in the regions neighboring the J96 chromosomal hemolysin there are additional unique DNA sequences not commonly found in other E. coli isolates.

Base Sequence↗
PubMed1983

Induction of lactation in nonpregnant cows by estradiol-17 beta and progesterone from an intravaginal sponge.

An intravaginal sponge impregnated with 500 mg estradiol-17 beta and 1000 mg progesterone to induce lactation in nonpregnant dairy cattle was investigated. Sponge retention (greater than 95%) was achieved by its attachment to a nylon collar. Treatment with the sponge for 10 days resulted in lactogenesis in 25% of treated cows compared with an 89% success rate when an adjunct injection of dexamethasone esters (20 mg intramuscular) was given on day 6 or a 96% success rate when reserpine injections (2.5 mg intramuscular) were given on days 6, 8, and 10 10 after sponge insertion. Milking was begun 12 days after sponge insertion (2 days after removal). Peak milk yield was unaffected by adjunct treatments but was greater in spring (mean 11.5 kg/day) than in autumn treated cows (mean 3 and 6 kg/day) in 2 yr of trials. Concentration of estradiol-17 beta in plasma rose rapidly after sponge insertion to a peak similar to that in cows in late pregnancy. Measurement of estradiol-17 beta concentration in milk indicated less, even at first milking than in commercially distributed pasteurized milk. Milk fat and protein content were higher in milk from induced lactations compared with that from normally calving cows on the same day.

Animals↗
PubMed1982

Novel antibiotic resistance transfer in Bacteroides.

Resistance to tetracycline and lincosamide antibiotics was transferred en bloc from a strain of Bacteroides fragilis (V503) to a plasmidless strain of Bacteroides uniformis (V528) during in vitro filter matings. Resistance transfer was detected at frequencies of 10(-5) to 10(-6) drug-resistant progeny per input donor cell and was dependent on cell-to-cell contact of donors and recipients. Transfer was insensitive to DNase and was not mediated by chloroform- or filter-sterilized donor broth cultures. A determinant for resistance to cefoxitin in V503 was not transferred in this system. V503 contained a 3.7 x 10(6)-dalton plasmid (pVA503). Drug-resistant progeny of V503 x V528 matings usually contained pVA503, but up to 20% of the total progeny of such crosses were plasmid free. Filter blot DNA hybridization studies (Southern method) confirmed that pVA503 was not integrated into the host chromosome of the plasmidless progeny. Drug-resistant progeny from V503 x V528 matings (with or without pVA503) conjugally transferred clindamycin resistance an tetracycline resistance to a suitable recipient strain. None of the drug resistance determinants of V503 were affected by treatment with standard plasmid curing regimens, and methods designed to detect very large plasmid molecules failed to suggest the involvement of extrachromosomal DNA in this resistance transfer system. The well-characterized Bacteroides R plasmid, pBF4 (conferring clindamycin resistance), was found to share hybridizing sequences with bulk cellular V503 DNA when examined by filter blot hybridization. Similarly sized sequences were found in drug-resistant progeny recovered from matings. Neither of the two pBF4 derivatives carrying deletions that abolished clindamycin resistance hybridized with V503 DNA.

Anti-Bacterial Agents↗
PubMed1982

Two independent conjugal transfer systems operating in Bacteroides fragilis V479-1.

Bacteroides fragilis V479-1 (also designated strain 92) has previously been shown to contain a conjugative plasmid, pBF4, that specifies resistance to clindamycin (Cc). A report of inducible tetracycline (Tc) resistance in this strain suggested that this phenotype was also plasmid associated (G. Privitera et al., Nature [London] 278:657-659, 1979) and prompted further investigation. Mating experiments with V469-1 and a Cc-sensitive derivative of V479-1, V598, showed that Tc resistance transfer occurred by a conjugation-like event which was insensitive to DNase, was not mediated by donor culture cell-free filtrates, and required cell-to-cell contact. Results from transfer experiments with V479-1 indicated that Tc and Cc resistance determinants were not linked and segregated independently in matings. Progeny recovered from matings with the V479-1 or V598 donor strain were able to transfer the Tc resistance marker in secondary crosses. Tc resistance transfer from V479-1 or V598 was greatly stimulated by pregrowth in the presence of Tc but not Cc. pBF4-mediated Cc resistance transfer was not affected by pregrowth in the presence of Cc or Tc. Filter blot DNA hybridization studies revealed that pBF4 sequences were not involved in either the Tc resistant phenotype or its associated conjugal transfer properties. The Tc resistance transfer element was not associated with pBF4 or any other extrachromosomal DNA element.

Bacteroides↗
PubMed1981

Dexamethasone concentrations in plasma and milk of cows following the injection of long- and short-acting dexamethasone esters.

A radioimmunoassay has been developed for the measurement of dexamethasone in plasma and milk of cows injected with long- and short-acting dexamethasone esters. Dexamethasone antiserum was produced by injecting cows with a dexamethasone-21-hemisuccinate-human serum albumin complex. The antisera was highly specific for dexamethasone, cross-reacting less than 0.7% for all endogenous steroids tested. Plasma concentrations of dexamethasone in cows injected intramuscularly with either 20 mg dexamethasone-21 trimethyl acetate (n = 2) or the tributyl derivative (n = 2) reached a peak level of 0.6-1.1 ng/ml in 2-6 days then declined to undetectable levels (less than 0.15 ng/ml) and 14 days after injection. In general, dexamethasone concentrations in milk were 0.3-0.5 times the plasma concentrations but showed the same pattern of values. Plasma dexamethasone concentrations were also determined in three lactating dairy cows injected intramuscularly with tritiated dexamethasone-21 trimethyl acetate. In these cows plasma dexamethasone concentrations, as determined by isotopic dilution, reached maximal levels of 1.1-1.6 ng/ml in 1-3 days then declined to levels of around 0.05 ng/ml within 30 days. The concentrations of dexamethasone in milk of two of these cows were, in general, similar to those found in plasma. In three cows injected intramuscularly with 20 mg dexamethasone sodium phosphate the concentrations of dexamethasone in plasma rose sharply to maximum levels of 24-70 ng/ml within 2-20 min after injection and fell to undetectable levels (less than 0.15 ng/ml) after 72 hr.

Animals↗
PubMed1981

Physical characterization of Bacteroides fragilis R plasmid pBF4.

Bacteroides fragilis V479-1 has previously been shown to harbor a self-transmissible 27 X 10(6)-dalton plasmid (pBF4) which confers lincosamide-macrolide resistance. The present study has focused on the physical properties of pBF4. The plasmid was found to be present in 1 to 2 copies per chromosomal equivalent. pBF4 was genetically stable, although spontaneously occurring plasmidless segregants could be detected at low frequency (approximately 1%). This frequency was unaffected by growth of cells in ethidium bromide. About one-third of all spontaneously occurring macrolide-lincosamide-sensitive clones of strain V479-1 were found to contain pBF4 molecules that carried deletions. Ten independently obtained deletion derivatives of pBF4 from lincosamide-macrolide-sensitive strains were compared with the parental pBF4 by restriction endonuclease cleavage analysis. A restriction site map of pBF4 was constructed, and the location of the deletions was approximated. Self-annealed pBF4 molecules, examined by electron microscopy, revealed the presence of two pairs of inverted repeat (IR) sequences on the plasmid. IR-1 was about 400 base pairs in length, and its two component members were separated by an intervening sequence of about 15 kilobases. IR-2 was about 75 base pairs in length, and its component members were separated by 4.2 kilobases. Each of the deletions of pBF4 studied had a terminus at or near the same IR-2 sequence.

Bacteroides fragilis↗
PubMed1981

Transformation of Streptococcus sanguis with monomeric pVA736 plasmid deoxyribonucleic acid.

Monomeric and oligomeric forms of a 5.0 x 10(6)-dalton plasmid (conferring erythromycin resistance) were able to genetically transform naturally competent Streptococcus sanguis. Transformation with electrophoretically purified monomer was a second-order process, whereas transformation with a dye-buoyant density gradient-purified plasmid preparation followed one-hit kinetics.

Centrifugation, Density Gradient↗
PubMed1981

Plasmid- and chromosome-mediated dissimilation of naphthalene and salicylate in Pseudomonas putida PMD-1.

Pseudomonas putida PMD-1 dissimilates naphthalene (Nah), salicylate (Sal), and benzoate (Ben) via catechol which is metabolized through the meta (or alpha-keto acid) pathway. The ability to utilize salicylate but not naphthalene was transferred from P. putida PMD-1 to several Pseudomonas species. Agarose gel electrophoresis of deoxyribonucleic acid (DNA) from PMD-1 and Sal+ exconjugants indicated that a plasmid (pMWD-1) of 110 megadaltons is correlated with the Sal+ phenotype; restriction enzyme analysis of DNA from Sal+ exconjugants indicated that plasmid pMWD-1 was transmitted intact. Enzyme analysis of Sal+ exconjugants demonstrated that the enzymes required to oxidize naphthalene to salicylate are absent, but salicylate hydroxylase and enzymes of the meta pathway are present. Thus, naphthalene conversion to salicylate requires chromosomal genes, whereas salicylate degradation is plasmid encoded. Comparison of restriction digests of plasmid pMWD-1 indicated that it differs considerably from the naphthalene and salicylate degradative plasmids previously described in P. putida.

Catechol 1,2-Dioxygenase↗
PubMed1981

Plasma concentrations of progesterone, oestrogens and prostaglandin F in maternal blood and corticosteroids and oestrogens in foetal blood of cows during dexamethasone-induced deliveries.

Infusing dexamethasone at rates of 0.1, 1.0 or 10 mg/day into 6 foetal calves at day 240 of gestation (2 calves per infusion rate), induced premature calving in a mean 12, 9 and 3 days respectively. Maternal plasma concentrations of progesterone, oestrone, oestradiol-17 beta and prostaglandin F or 13,14-dihydro-15-keto-prostaglandin F, and foetal plasma concentrations of oestrone, oestradiol-17 beta and corticosteroids were monitored until calving. The maternal hormone changes observed for all cows were a decline in progesterone, and a rise in both oestrogen and prostaglandin F concentrations before calving. Dexamethasone infused at a rate of 1.0 and 10.0 mg/day suppressed plasma corticosteroids in the foetus; however at 0.1 mg/day, foetal corticosteroids increased tenfold during the last 3 days before calving. At the lowest rate of infusion, dexamethasone had no effect on foetal oestrogen levels, but at rates of 1.0 and 10.0 mg/day there was a marked rise in foetal plasma levels of oestrogen.

Adrenal Cortex Hormones↗
PubMed1977

Foetal and maternal hormonal changes preceding normal bovine parturition.

Successful chronic cannulation of the foetal posterior vena cava and maternal utero-ovarian and jugular veins in five Jersey cows between days 240 and 260 of gestation enabled changes in plasma hormone levels preceding calving to be monitored. All cows delivered live calves within the expected range of gestation for the breed. Corticosteroids were assayed by competitive protein-binding and prostaglandin F, progesterone, oestrone and oestradiol-17beta by radioimmunoassay. Foetal corticosteroids rose slowly from 5.0 +/- 0.7 ng/ml at 20 days to 9.3 +/- 3.0 ng/ml at 10 days before term, then progressively increased to a mean of 74 ng/ml, though higher concentrations occurred following surgery. Foetal oestrone and oestradiol-17beta concentrations were both less than 50 pg/ml and showed little change toward term. The maternal utero-ovarian oestrogens increased slowly from 20 to 10 days pre-partum and then rose more rapidly reaching peak levels (2.9 +/- 0.6 ng/ml for oestrone and 1.4 +/- 0.3 ng/ml for oestradiol-17beta) 1 to 4 days before delivery. Maternal progesterone concentrations fell towards term, with a rapid decrease over the last 36-48 h before calving when they gradually increased until the last 24 h where was a dramatic rise, reaching peak levels (5.7 +/- 0.6 ng/ml) during labour.

Adrenal Cortex Hormones↗
PubMed1976

Preparturient changes in plasma concentrations of estrone, estradiol-17beta and estradiol-17alpha in the fetal calf: effects of dexamethasone infusion.

The concentrations of total estrogens in fetal calf plasma were determined during a 6-10 day period immediately before delivery. Comparison was made between levels found in untreated calves and calves infused with dexamethasone at the rate of 0.1, 1.0 and 10 mg/24 hours. In untreated calves the plasma estrone, estradiol-17beta and estradio-17alpha levels remained relatively constant at 38 +/- 7 ng ml-1 (mean +/- SEM n = 3), 46 +/- 6 ng ml-1 and 29 +/- 5 ng ml-1 respectively. Infusion with dexamethasone at 0.1 mg/24 hr (3 calves) and 1.0 mg/24 hr (3 calves) was without dramatic effect on plasma estrogen levels. However, in one fetus infused with 10.0 mg/24 hr the dexamethasone treatment may have caused a transitory rise in the levels of all estrogens examined.

Animals↗
PubMed1975

Peripheral plasma progesterone and utero-ovarian prostaglandin F concentrations in the cow around parturition.

The concentration of prostaglandin F in utero-ovarian venous plasma and proseterone in jugular venous plasma were determined by radioimmunoassay in 3 cows over the last 2-3 weeks of gestation. Utero-ovarian prostaglandin F concentrations did not show and consistent pattern in two hours of three cows until 48-72 h before term when the levels rose sharply from 1 ng/ml to maximum 4-9 ng/ml during labour. The concentration of prosterone in jugular venous plasma tended to fall gradually over the last 20 days of gestation with a further fall occurring 48-36 h before delivery.

Animals↗