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C J Smyth

Publications and source records attributed to C J Smyth.

At least 19 recordsLinked to original sources
PubMed1992

Identification of erythrocyte-binding antigens in Helicobacter pylori.

The surface antigens of Helicobacter pylori conferring erythrocyte-binding activity were obtained by adsorption onto formaldehyde-treated dog and goat erythrocytes from supernatant fractions of sonicated bacteria and elution using a high concentration of NaCl. The desorbed material was analysed by SDS-PAGE and immunoblotting with anti-whole-cell serum to agar-grown bacteria which had been absorbed with broth-grown, non-haemagglutinating cells (haemagglutination-associated antiserum). Two polypeptides with molecular masses of 25 and 59 kDa were revealed as erythrocyte-binding antigens. Strains which agglutinated both dog and goat erythrocytes possessed both these erythrocyte-binding antigens, whereas an antigenically cross-reactive 24 kDa polypeptide was present in a strain which only agglutinated goat erythrocytes. Haemagglutinin material was extracted from H. pylori using n-octylglucopyranoside and purified by Sepharose chromatography and sucrose density gradient ultracentrifugation. The purified extract directly agglutinated erythrocytes in a neuraminyl-lactose-sensitive and neuraminidase-sensitive manner. The 59 kDa polypeptide was not present in the purified haemagglutinin preparation. The haemagglutination-associated antiserum reacted strongly with the 25 kDa polypeptide band which was the most prominent polypeptide band on analysis of the purified haemagglutinin preparation by SDS-PAGE and silver staining. Thus, H. pylori possesses at least two adhesins, one of which recognises a N-acetylneuraminic acid (alpha 2-3) moiety of receptors, the other being of unknown receptor specificity. Differences in the antigenicity and molecular masses of these adhesins in individual strains may underlie differences in receptor-binding specificities and haemagglutination profiles.

Animals↗
PubMed1990

Legionella in Dublin hospital water supplies.

An environmental survey was carried out which consisted of periodic and random sampling of water tanks and showers in two large Dublin hospitals. Of the samples 5.3% yielded Legionella bacteria. Legionella pneumophila of serogroups 3, 5 and 6 were isolated from these sites with viable counts ranging from 3.0 x 10(2) - 2.5 x 10(3) c.f.u./litre. The implementation of periodic sampling may, however, not be a worthwhile exercise unless an environmental site has been associated with cases of legionellosis. Emphasis should be placed on the prevention of contamination of environmental sites with legionellae and on the development and implementation of protocols and procedures for the isolation of legionellae to gain the necessary expertise should an epidemiological survey be required.

Hospitals↗
PubMed1989

Electron microscopy of adhesive interactions between Gardnerella vaginalis and vaginal epithelial cells, McCoy cells and human red blood cells.

Exfoliated vaginal epithelial cells with attached bacteria, termed 'clue cells', which were procured from a patient with non-specific vaginitis, were stained with ruthenium red and examined by transmission electron microscopy. The attached bacteria appeared to adhere by means of an outer fibrillar coat. An epithelial tissue culture cell line (McCoy) and human red blood cells to which strains of Gardnerella vaginalis attached were similarly examined. The adherence of G. vaginalis to the epithelial cell line appeared to be mediated by an outer fibrillar coat while adherence to red cells appeared to be mediated by fimbriae. Transmission electron microscopy was performed on the Gardnerella strains used. Thin sections of tissue-culture-adherent strains revealed a dense outer fibrillar coat whereas the surface of the haemagglutinating strains showed fine fimbriae. Negative staining of haemagglutinating strains demonstrated fimbriae on a minority of organisms.

Bacterial Adhesion↗
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↗
PubMed1988

Nucleotide sequence of the gene encoding the major subunit of CS3 fimbriae of enterotoxigenic Escherichia coli.

The complete nucleotide sequence of a 612-base-pair DNA fragment containing the gene for the major fimbrial subunit of CS3 of enterotoxigenic Escherichia coli is presented. A possible promoter region, a ribosome-binding site, and two potential signal peptidase cleavage sites are indicated. Unlike the best-studied fimbrial proteins, the predicted CS3 sequence has no Cys residues.

Amino Acid Sequence↗
PubMed1987

Carriage rates of enterococci in the dental plaque of haemodialysis patients in Dublin.

The incidence of carriage of enterococci in dental plaque was determined in haemodialysis patients attending three clinics in the Dublin area either as outpatients or as hospitalised patients. Their carriage rates were compared with a University group comprising normal healthy students, academic staff, technicians and ancillary personnel and with a cohort of otherwise healthy toothache patients. The carriage rates among the staffs of the dialysis units also were examined. The overall carriage rates of enterococci of the University group, the toothache patient group, the haemodialysis patients and the dialysis unit staffs did not differ significantly from each other, ranging from 5%-20%. However, the dental plaque of a mainly hospitalised group of haemodialysis patients and their attendant staff at one clinic was colonised to a statistically significant higher degree with enterococci than that of the haemodialysis patients and the staff at the outpatient clinics, both separately and as combined patient and staff groups. Age, sex, a history of recent antibiotic therapy, and elapsed time since the last dental visit did not affect isolation rates to a significant extent. The commonest enterococcus isolated from subjects was Streptococcus faecalis, followed by its variety liquefaciens. Only one subject harboured Streptococcus durans in dental plaque. Ten of the 21 subjects yielding enterococci harboured two different enterococci in their plaque. The isolation of S. faecalis var liquefaciens alone or in combination with S. faecalis did not correlate with subject-history parameters. The findings obtained imply that antibiotic prophylaxis specifically against enterococci may be necessary only for a small number of haemodialysis patients in whom oral carriage of enterococci has been demonstrated bacteriologically.

Adolescent↗
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↗
PubMed1987

Haemagglutination and tissue culture adhesion of Gardnerella vaginalis.

Six strains of Gardnerella vaginalis were studied to examine the adhesin-receptor mechanism involved in their attachment to human red blood cells and an epithelial tissue culture cell line (McCoy). The adhesins involved in the attachment of the bacteria to each of these cells were proteinaceous but showed marked differences after various chemical or physical treatments, indicating that separate adhesins were present. Haemagglutinating strains were more hydrophobic than tissue-culture-adherent strains. Haemagglutination of human red blood cells by strains of G. vaginalis was inhibited by galactose, lactose, N-acetylneuraminic acid and phosphatidylserine. In contrast, the tissue-culture adherence of strains was not inhibited by these substances.

Bacterial Adhesion↗
PubMed1987

Biotyping, serotyping and phage typing of Streptococcus faecalis isolated from dental plaque in the human mouth.

Thirty Streptococcus faecalis isolates from mixed dental plaque samples were classified into four groups on the basis of biotype, tetracycline susceptibility, phage type and serotype combinations. The organisms were from patients on haemodialysis, from staff of the dialysis unit, and from controls. Three biotypes were distinguished by seven biochemical tests: production of acid from inositol, sucrose and xylose; rapid or delayed production of acid from sorbitol; gelatin liquefaction; and production of alkaline phosphatase and beta-galactosidase. With a set of eight typing antisera for S. faecalis, 15 strains were non-typable, 12 were serotype 1 and three were serotype 19. With a set of 17 bacteriophages specific for S. faecalis, all of the oral isolates were typable; 40% were lysotype I1 and the remainder lysotype V6b. On the basis of biotype-serotype-phage-type combinations, indications of possible spread of strains between haemodialysis patients and dialysis unit staff were obtained. Biotyping and serotyping of 13 German isolates of S. faecalis of phage type I1 from four clinical sources and tripartite typing of three control strains provided additional evidence for the potential of biotyping in distinguishing between strains of identical serotype and phage type. One oral isolate of S. faecium was of phage type XX. None of the oral isolates of S. faecalis, of which 14 exhibited delayed sorbitol fermentation, reacted with group-G streptococcal grouping reagents or antiserum. Slow sorbitol fermentation does not appear to be a definitive phenotypic marker for S. faecalis strains possessing antigens that react with both group-D and group-G grouping reagents.

Adolescent↗
PubMed1987

In vitro adhesiveness and biotype of Gardnerella vaginalis strains in relation to the occurrence of clue cells in vaginal discharges.

Haemagglutination and tissue culture adherence tests using a McCoy cell line were used to examine the adherence characteristics of 105 strains of Gardnerella vaginalis. Each strain represented one isolate per patient. For each patient, a direct smear of vaginal discharge was examined for clue cells. The relation between in vitro adherence and the presence of clue cells was examined. There seemed to be no appreciable relation between the presence of clue cells in smears and the haemagglutinating activity of strains. In contrast, adherence as judged by the McCoy tissue culture system showed a significant relation to the presence of clue cells (p less than 0.001). Though both adhesive characteristics were not inhibited by mannose, the mechanism of haemagglutination of human red cells appeared to differ from that of adherence of tissue culture cells. The findings imply that the clue cell phenomenon is due to attachment of adherent strains of G vaginalis to epithelial cells. Adherent strains of G vaginalis may play a part in the pathogenesis of bacterial vaginosis.

Bacterial Adhesion↗
PubMed1986

Development of intestinal antibodies against Escherichia coli antigens in piglets with experimental neonatal E. coli diarrhoea.

Intestinal immune responses to Escherichia coli antigens were studied in conventionally reared piglets orally infected on the first day of life with a virulent enterotoxigenic E. coli (O149: K88). During the first week of life intestinal antibodies were produced against the homologous lipopolysaccharide (LPS) as well as against the K88 antigen and the heat-labile enterotoxin (LT). On Day 7, anti-LPS antibodies of the IgA and IgG classes were detected in most piglets, whereas anti-K88 antibodies of the IgG and IgM classes predominated; antibodies against the enterotoxin were usually of the IgG class. In 21-day-old piglets antibodies of all immunoglobulin classes had usually been produced. In most cases, the levels of intestinal antibodies were substantially higher on Day 21 compared to Day 7, but the levels varied considerably both between and within litters. The intestinal immune responses did not correlate with the severity of clinical symptoms. One-, 7- and 21-day-old piglets reared in a specific-pathogen-free (SPF) herd lacked significant intestinal antibodies to the antigens examined. The oral challenge did not stimulate systemic immune responses. After colostral intake, all piglets had high antibody levels in the circulation. These levels decreased continuously during the 3-week study period. The possibility that high amounts of antibodies in colostrum could interfere with this early intestinal antibody formation should be considered when planning vaccination programmes against E. coli diarrhoea in piglets.

Animals↗
PubMed1986

Plasmids coding for colonization factor antigens I and II, heat-labile enterotoxin, and heat-stable enterotoxin A2 in Escherichia coli.

Colonization factor antigens I and II (CFA/I and CFA/II) are important in the pathogenesis of diarrhea in humans caused by some enterotoxigenic Escherichia coli (ETEC). Plasmid DNA from 16 CFA/I+ and five CFA/II+ ETEC were examined by Southern blot analysis with enterotoxin gene probes and were compared with plasmid DNA from derivatives of the same ETEC that had lost the ability to produce these colonization factors. Among the 16 CFA/I+ ETEC strains, the loss of CFA/I was accompanied by the loss of a plasmid of between 34 and 68 megadaltons (MDa) coding for heat-stable enterotoxin A2 (ST-A2) in 12 strains, by the loss of a 60-MDa plasmid coding for heat-labile enterotoxin (LT) and ST-A2 in one strain, or by deletions of a segment of DNA encoding for ST-A2 in three strains. Among five CFA/II+ ETEC strains, the loss of CFA/II was associated with the loss of a plasmid of 75 MDa coding for LT and ST-A2 in three strains, with the loss of genes coding for LT and ST-A2 from a 68-MDa plasmid in one strain, or with no discernible loss of a plasmid or DNA sequences coding for enterotoxins in the remaining strain. The loss of CFA/I and CFA/II production was associated with the loss of DNA sequences encoding for ST-A2 in 20 of 21 ETEC examined.

Antigens, Bacterial↗
PubMed1982

Effect of parenteral vaccination of dams on intestinal Escherichia coli in piglets with diarrhea.

To evaluate the effect of widely used parenteral vaccination of dams against neonatal colibacillosis, the virulence factors of the intestinal Escherichia coli flora, namely, O serogroup, enterotoxin(s) produced (heat labile, porcine heat stable, and murine heat stable) and adhesins (K88, K99, and 987P antigens) of 149 piglets from different herds in Sweden were investigated. Three categories were investigated: healthy piglets, diarrheal piglets born to unvaccinated dams, and diarrheal piglets born to dams vaccinated with a polyvalent Formalin-killed whole-cell vaccine containing K88 antigen (Porcovac; Hoechst Pharmaceuticals, Hounslow, England). Piglets less than 1 week old and those 1 to 8 weeks old were evaluated separately. Diarrheal piglets less than 1 week old from vaccinated dams yielded a higher incidence of K99 antigen-positive E. coli of the murine heat-stable enterotoxigenicity type compared with piglets of the same age group from unvaccinated dams. The percentage of diarrheal cases from which E. coli lacking recognized virulence attributes were isolated was also higher in the former compared with the latter group. In the 1- to 8-week-old diarrheal piglets of vaccinated dams, the overall incidence, enterotoxigenicity type, and serotype of the E. coli isolates resembled those of diarrheal piglets less than 1 week of age from unvaccinated herds. Enterotoxigenic E. coli bearing 987P antigen detectable in vitro was rare. Most of the enterotoxigenic isolates lacking K88, K99, and 987P antigens produced only ST. The investigation pinpoints some of the inadequacies of vaccines of the type studied under field conditions.

Animals↗
PubMed1981

K99 antigen-positive enterotoxigenic Escherichia coli from piglets with diarrhea in Sweden.

K88 antigen-negative enterotoxigenic Escherichia coli and non-enterotoxigenic strains isolated from piglets with diarrhea were examined for K99 antigen by agglutination tests after growth on Minca-IsoVitaleX (BBL Microbiology Systems, Cockeysville, Md.) agar medium. Of 64 K88-negative enterotoxigenic strains from as many piglets, 17 were found to be K99 positive. Of these, 10 were Swedish and 7 were of Norwegian origin. All 17 produced heat-stable enterotoxin detectable in the infant mouse assay, but only 2 gave positive ligated loop tests in 3- to 7-week-old piglets. Ligated loop tests in 5- to 12-day-old piglets were positive for each of the 15 K99-positive strains tested. Each of the Swedish K99-positive isolates was from a piglet of lesser than or equal to 7 days of age. One piglet harboring a K99-positive strain also harbored an O141:K88 enterotoxigenic strain producing only heat-stable enterotoxin. Five of the Swedish piglets yielding K99-positive isolates were from dams vaccinated wih a multicomponent bacterial vaccine containing K88 antigen. The K99 strains were O:K:H serotyped. The O serogroups represented were O8, O9, O64, O101, and O140. None of the 101 non-enterotoxigenic porcine isolates, representing 42 serogroups and non-O-groupable and rough strains, was found to be K99 positive. The findings indicate that so-called class 2 or atypical porcine enterotoxigenic E. coli should be routinely examined for the presence of K99 antigen.

Animals↗
PubMed1978

Inhibition of noradrenaline-stimulated lipolysis and cyclic AMP accumulation in isolated rat adipocytes by purified phospholipase C and theta-toxin from Clostridium perfringens.

Purified phospholipae C (phosphatidylcholine cholinephosphohydrolase, EC 3.1.4.3) and theta-toxin from Clostridium perfringens both inhibited noradrenaline-stimulated lipolysis and cyclic AMP accumulation in isolated rat adipocytes in a dose-dependent manner. The action of phospholipase C was gradual in onset, while the effect of theta-toxin was almost immediate. Phospholipase C, but not theta-toxin, hydrolyzed membrane phospholipids and inhibited adenylate cyclase (EC 4.6.1.1) in a crude membrane fraction from fat cells. The inhibitory effects of phospholipase C were associated with morphological alterations detectable by electron microscopy, whereas effects of theta-toxin were observed at a time when no clearcut morphological alterations could be observed. It is concluded that the two purified principles from C. perfringens, which are both present in commercial preparations of phospholipase C, antagonize noradrenaline-stimulated cyclic AMP accumulation and lipolysis. Although their exact mechanisms of action have not been elucidated, phospholipase C and theta-toxin have different modes of attack.

Adenylyl Cyclases↗