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D C Coleman

Publications and source records attributed to D C Coleman.

67 records · Page 4Linked to original sources
PubMed1988

Molecular cloning of the plasmid-located determinants for CS1 and CS2 fimbriae of enterotoxigenic Escherichia coli of serotype O6:K15:H16 of human origin.

The plasmid pCS001, isolated from an enterotoxigenic strain of Escherichia coli, mediates expression of the CS1 or CS2 and CS3 fimbrial adhesins in appropriate E. coli hosts. To characterize this further, HindIII-generated DNA fragments of this plasmid were cloned into the vector plasmid pBR322. A chimaera, called pCS200, which mediated expression of the CS1 or CS2 fimbrial antigen but not of CS3 fimbrial antigen in appropriate host strains, was obtained. The DNA inserted into the vector sequences of plasmid pCS200 comprised HindIII fragments of 4.7 kbp and 0.8 kbp. Plasmid pCS200-carrying wild-type E. coli hosts of serotype O6:K15:H16 that expressed the CS1 or CS2 antigen also caused mannose-resistant agglutination of bovine red blood cells, suggesting that functional fimbriae were present on the bacterial surface. As previously observed with strain K12 recipients of CS-fimbriae-associated plasmids mobilized from wild-type enterotoxigenic E. coli, K12 recipients of the chimaeric plasmid pCS200 did not express the CS1 or CS2 fimbrial antigen. An oligonucleotide probe, synthesized on the basis of the published N-terminal amino acid sequence of the CS2 fimbrial subunit, hybridized to plasmid pCS200, indicating that the gene for the structural subunit of this fimbria resided on the plasmid.

Adhesins, Escherichia coli↗
PubMed1987

Molecular cloning and characterization of the genetic determinant encoding CS3 fimbriae of enterotoxigenic Escherichia coli.

The genetic determinant encoding the synthesis and surface expression of CS3 fimbriae of colonization factor antigen II-(CFA/II-) positive enterotoxigenic Escherichia coli was cloned on a 5.1 kb HindIII DNA fragment in pBR322 from the wild-type plasmid pCS001 to yield the CS3+ plasmid pCS100. Subcloning of EcoRI fragments of 1.8 kb and 2.5 kb into vector plasmid pACYC184 and the isolation of a series of pCS100::Tn5 insertion mutants revealed that more than one cistron was involved in the synthesis and expression of CS3 fimbriae. Polypeptides of 94, 26, 24, 17 and 15 kDa were detected in E. coli minicells harbouring pCS100. In Western immunoblotting the 17 kDa and 15 kDa polypeptides reacted with specific anti-CS3 fimbriae serum. The 15 kDa polypeptide comigrated with the structural subunit of CS3 fimbriae. Inhibition of protein processing in minicells by ethanol confirmed that the 17 kDa polypeptide was the precursor form of the 15 kDa structural subunit. A physical map of the cloned DNA was constructed showing the location and direction of transcription of the genes for the 17 and 94 kDa polypeptides. Using the 5.1 kb HindIII fragment of pCS100 as a genetic probe for the CS3 determinant, Southern hybridization analysis of plasmid and total cellular DNA was performed in wild-type enterotoxigenic E. coli strains.

Antigens, Bacterial↗
PubMed1986

Molecular typing of methicillin and gentamicin resistant Staphylococcus aureus in Dublin.

The high incidence of infection in Dublin hospitals caused by non-typable strains of methicillin- and gentamicin-resistant Staphylococcus aureus (MGRSA) has created an important epidemiological problem as conventional methods of sub-dividing these organisms have not been useful. This report describes a novel approach to the typing and analysis of MGRSA strains, particularly non-typable isolates, by comparing restriction endonuclease HindIII digest patterns of total cellular DNA; and by using Southern hybridization analysis to detect size variations or polymorphisms in restriction endonuclease cleavage fragments within small regions of the chromosome. Non-typable MGRSA strains and isolates belonging to two phenotypically related groups of phage-type 77 and 77/84 strains were readily subdivided on the basis of molecular size differences in high molecular weight DNA fragments generated by the enzyme HindIII. Restriction endonuclease fragment size polymorphisms were readily detected in many of the non-typable strains tested in hybridization experiments, and these were used for strain sub-division. Both techniques were useful tools for the separation of closely related MGRSA strains.

DNA Restriction Enzymes↗
PubMed1986

Cloning and expression in Escherichia coli and Staphylococcus aureus of the beta-lysin determinant from Staphylococcus aureus: evidence that bacteriophage conversion of beta-lysin activity is caused by insertional inactivation of the beta-lysin determinant.

The beta-lysin determinant (Hlb) from Staphylococcus aureus CN6708 was cloned in Escherichia coli K-12 using the bacteriophage replacement vector lambda L47.1. The Hlb determinant was localised to a 1250 base pair DNA sequence by cloning fragments from a Hlb+ recombinant phage into the plasmid vectors pACYC184 and pBR322 in E. coli K-12, and by the subsequent construction and analysis of several sub-clones, in vitro deletion and Tn5 insertion mutations. E. coli cells harbouring Hlb+ plasmids expressed readily detectable levels of beta-lysin and sphingomyelinase activity, which were located in the cytoplasm. Two polypeptides of molecular weight 38,000 and 33,000 which were encoded by the Hlb determinant were detected in E. coli minicells, but only the 33,000 dalton protein was detected in immunoblotting experiments with specific anti-beta-lysin serum. Hybridisation analysis with probes made from the cloned Hlb determinant and from DNA of the staphylokinase-converting phage phi 13, indicated that bacteriophage conversion of S. aureus to loss of beta-lysin activity is due to insertion of phi 13 DNA into or adjacent to the beta-lysin determinant. A shuttle plasmid was used to transfer the cloned Hlb determinant into a beta-lysin negative strain of S. aureus where the wild-type chromosomal determinant was inactivated by lysogenic conversion. Beta-lysin activity was readily detected in supernatants of S. aureus harbouring the cloned determinant.

Antimicrobial Cationic Peptides↗
PubMed1985

Susceptibility to antimicrobial agents and analysis of plasmids in gentamicin- and methicillin-resistant Staphylococcus aureus from Dublin hospitals.

Methicillin- and gentamicin-resistant Staphylococcus aureus (MGRSA) strains isolated from Dublin Hospitals were classified into two groups (phenotypes). Phenotype-I strains expressed high level resistance to gentamicin and were susceptible to fusidic acid; strains resistant to tetracycline harboured a 3 X 10(6)-mol. wt plasmid. Strains in phenotype II usually expressed low level resistance to gentamicin, were resistant to fusidic acid and often harboured a (22-24) X 10(6)-mol. wt plasmid that specified resistance to ethidium bromide, tetracycline, kanamycin, neomycin and trimethoprim, or to combinations of these markers. A few phenotype-II strains expressed higher levels of resistance to gentamicin and other aminoglycosides. All MGRSA strains carried a 21 X 10(6)-mol. wt plasmid conferring resistance to penicillin, ethidium bromide, cadmium and mercury. Gentamicin resistance was invariably chromosomal and all strains carried chromosomal resistance to methicillin, erythromycin, streptomycin and spectinomycin. Several methicillin-resistant S. aureus (MRSA) strains isolated before the emergence of gentamicin resistance harboured a 21 X 10(6)-mol. wt penicillinase plasmid with the same restriction endonuclease profile as that from some MGRSA strains. Some MRSA strains carried other plasmids related to those found in MGRSA strains.

Anti-Bacterial Agents↗
PubMed1985

Molecular analysis of multiple-resistance plasmids transferred from gram-negative bacteria isolated in a urological unit.

Forty-one isolates of multiply resistant gram-negative bacteria causing infection in a urological unit of a Dublin hospital were collected during a 6-month period. Twenty-one isolates transferred multiple resistance to an Escherichia coli K-12 recipient in liquid matings. Serratia marcescens, Proteus morganii, Proteus vulgaris, and E. coli isolates harbored similar 120-megadalton IncC plasmids, whereas Enterobacter cloacae strains transferred a 160-megadalton plasmid of a different Inc group. Southern hybridization experiments were performed with purified fragments cloned from one IncC plasmid as probes. They were hybridized to plasmid sequences in total cellular DNA extracts, showing that the IncC plasmids were very closely related. This suggests that the same plasmid has transferred to different bacterial species in the hospital environment.

Bacteriuria↗
PubMed1983

Analysis of tetracycline resistance encoded by transposon Tn10: deletion mapping of tetracycline-sensitive point mutations and identification of two structural genes.

Deletions in the tet genes derived from Tn10 were formed from different tet::Tn5 insertion mutations by removing DNA sequences located between a HindIII site in Tn5 and a HindIII site adjacent to the tet genes. Tetracycline-sensitive point mutations were mapped in recombination tests with the deletions and were thus aligned with the genetic and physical map of the tet region. Plasmids carrying point mutations were tested for complementation with derivatives of pDU938, a plasmid carrying cloned tet genes derived from Tn10 which had been inactivated by Tn5 insertions. Complementation occurred between promoter-proximal tet point mutations and distal tet::Tn5 insertions, suggesting the existence of two structural genes, tetA and tetB. These results, together with the analysis of polypeptides in minicells harboring pDU938tet::Tn5 mutants, suggested that tetA and tetB are expressed coordinately in an operon. The tetB gene encodes the previously characterized 36,000-dalton cytoplasmic membrane TET protein, but the product of tetA was not identified. Point mutations in either tetA or tetB led to the defective expression of the resistance mechanism involving tetracycline efflux. It is suggested that the tetA and tetB products interact cooperatively in the membrane to express resistance.

Biological Transport↗
PubMed1982

Interactions of bacterial and amoebal populations in soil microcosms with fluctuating moisture content.

Sterilized soil samples (20 g of soil per 50-ml flask), amended with 600 mug of glucose-carbon and 60 mug of NH(4)-N . g of dry soil, were inoculated with bacteria (Pseudomonas paucimobilis) alone or with bacteria and amoebae (Acanthamoeba polyphaga). We used wet-dry treatments, which involved air drying the samples to a moisture content of approximately 2% and remoistening the samples three times during the 83-day experiment. Control treatments were kept moist. In the absence of amoebae, bacterial populations were reduced by the first drying to about 60% of the moist control populations, but the third drying had no such effect. With amoebae present, bacterial numbers were not significantly affected by the dryings. Amoebal grazing reduced bacterial populations to 20 to 25% of the ungrazed bacterial populations in both moisture treatments. Encystment was an efficient survival mechanism for amoebae subjected to wet-dry cycles. The amoebal population was entirely encysted in dry soil, but the total number of amoebae was not affected by the three dryings. Growth efficiencies for amoebae feeding on bacteria were 0.33 and 0.39 for wet-dry and constantly moist treatments, respectively, results that compared well with those previously reported for Acanthamoeba spp.

Journal Article↗
PubMed1981

Nitrogen Mineralization by Acanthamoeba polyphaga in Grazed Pseudomonas paucimobilis Populations.

Nitrogen mineralization was studied in a simple grazing system in which the protozoan Acanthamoeba polyphaga was grown with the bacterium Pseudomonas paucimobilis (two soil organisms isolated from the shortgrass prairie in northern Colorado). In different experiments, either carbon or nitrogen was adjusted to be in limiting amounts. When carbon was limiting, grazers were almost entirely responsible for nitrogen mineralization, with bacteria themselves contributing little. When nitrogen was limiting, nitrogen mineralization by grazers permitted continued growth by the grazed bacteria and a greater bacterial biomass production. The increased growth of the grazed bacteria did not result in an increased total amount of carbon used, but the grazed bacteria used carbon more efficiently than the ungrazed bacteria.

Journal Article↗
PubMed1979

Effect of bacteria and amoebae on rhizosphere phosphatase activity.

The contributions of various components of soil microflora and microfauna to rhizosphere phosphatase activity were determined with hydroponic cultures. Three treatments were employed: (i) plants alone (Bouteloua gracilis (H.B.K.) Lag. ex Steud.) (ii) plants plus bacteria (Pseudomonas sp.), and (iii) plants plus bacteria plus amoebae (Acanthamoeba sp.). No alkaline phosphatase was detected, but an appreciable amount of acid phosphatase activity (120 to 500 nmol of p-nitrophenylphosphate hydrolyzed per h per plant) was found in the root culture solutions. The presence of bacteria or bacteria and amoebae increased the amount of acid phosphatase in solution, and properties of additional activity were identical to properties of plant acid phosphatase. The presence of bacteria or bacteria and amoebae increased both solution and root phosphatase activities at most initial phosphate concentrations.

Journal Article↗
PubMedDate not supplied

Multicenter prospective surveillance of oral Candida dubliniensis among adult Brazilian human immunodeficiency virus-positive and AIDS patients.

The incidence of C. dubliniensis in South America has not yet been determined. In the present study, oral swab samples were taken from 108 HIV-infected/AIDS individuals attending 6 separate Brazilian HIV-treatment centers to determine the incidence of C. dubliniensis in this population. Swabs were plated onto CHROMagar Candida medium and 155 isolates, presumptively identified as C. albicans or C. dubliniensis were further investigated. In a preliminary screen for C. dubliniensis, 13 of the 155 isolates showed no or poor growth at 42 degrees C, and all them were subjected to randomly amplified polymorphic DNA (RAPD) and polymerase chain reaction (PCR) analysis using C. dubliniensis-specific primers. We confirmed that 4 out of 13 isolates were C. dubliniensis, representing an incidence rate of 2.8% for the Brazilian HIV-infected population infected with yeasts exhibiting green colonies on CHROMagar Candida. This value is significantly lower than those reported in Ireland and the United States.

AIDS-Related Opportunistic Infections↗