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J F Torres

Publications and source records attributed to J F Torres.

15 recordsLinked to original sources
PubMed1996

Antigenicity of amino-acid sequences from Clostridium difficile toxin B.

Clostridium difficile toxins A and B cause antibiotic-associated colitis. Whereas antigenic determinants specifying neutralisation of toxin A have been partially elucidated, those of toxin B remain unknown. To define antigenic determinants of toxin B, synthetic peptides were prepared for five linear sequences selected by computer analysis for putative T and B epitopes. Peptides spanning the carboxy terminal region (aa 2155-2283) were also selected because this region contains repetitive units thought to bind the toxin to cell receptors. Multiple antigenic peptides were synthesised by linking four peptide copies to a core of four lysine residues (tetraMAP). Outbred mice were given four doses of each tetraMAP by intraperitoneal injection and specific immunoglobulins G and A were measured by enzyme-linked immunosorbent assay (ELISA) in serum, ascitic fluid and faeces. All 14 MAPs induced strong IgG responses against the homologous peptide; peptides representing aa 2155-2179 and 2246-2270 induced the strongest responses, of 592 and 493 ELISA units, respectively - although, to a lower extent, all 14 MAPs induced serum and faecal IgA responses against the homologous peptide. All MAPs induced IgG1 and IgG2b subclasses, documenting their capacity to elicit Th2-dependent mucosal immunity. IgG anti-MAPs were assayed for reaction with native toxins A and B; most anti-MAPs recognised the toxins only weakly or did not recognise them. Antibodies against peptide representing aa 2168-2192 recognised both native toxin B (19 ELISA units) and toxin A (2 ELISA units). None of the antibodies neutralised cytotoxicity of either toxin in cell culture. In contrast, four MAPs (aa 2080-2095, 2168-2192, 2220-2244 and 2233-2257) inhibited cytotoxicity when mixed with toxin B before addition to cells; inhibition was mediated by a direct interaction with toxin B.

Amino Acid Sequence↗
PubMed1995

Evaluation of formalin-inactivated Clostridium difficile vaccines administered by parenteral and mucosal routes of immunization in hamsters.

Clostridium difficile produces toxins that cause inflammation, necrosis, and fluid in the intestine and is the most important cause of nosocomial antibiotic-associated diarrhea and colitis. We evaluated C. difficile antigens as vaccines to protect against systemic and intestinal disease in a hamster model of clindamycin colitis. Formalin-inactivated culture filtrates from a highly toxigenic strain were administered by mucosal routes (intranasal, intragastric, and rectal) with cholera toxin as a mucosal adjuvant. A preparation of culture filtrate and killed whole cells was also tested rectally. The toxoid was also tested parenterally (subcutaneously and intraperitoneally) and by a combination of three intranasal immunizations followed by a combined intranasal-intraperitoneal boost. Serum antibodies against toxins A and B and whole-cell antigen were measured by enzyme-linked immunosorbent assay, neutralization of cytotoxic activity, and bacterial agglutination. The two rectal immunization regimens induced low antibody responses and protected only 20% of hamsters against death and 0% against diarrhea. The intragastric regimen induced high antibody responses but low protection, 40% against death and 0% against diarrhea. Hamsters immunized by the intranasal, intraperitoneal, and subcutaneous routes were 100% protected against death and partially protected (40, 40, and 20%, respectively) against diarrhea. Among the latter groups, intraperitoneally immunized animals had the highest serum anticytotoxic activity and the highest agglutinating antibody responses. Hamsters immunized intranasally and revaccinated intraperitoneally were 100% protected against both death and diarrhea. Protection against death and diarrhea correlated with antibody responses to all antigens tested. The results indicate that optimal protection against C. difficile disease can be achieved with combined parenteral and mucosal immunization.

Administration, Intranasal↗
PubMed1995

Indirect immunofluorescent assay (IFA) in buffy coat as a rapid diagnostic test for invasive candidiasis.

In order to have a diagnostic test for invasive candidiasis, an immunofluorescent assay (IFA) in buffy coat was developed and evaluated. This test was compared to buffy coat culture and blood culture. The study was divided into two parts: evaluation of the diagnostic test in a rabbit model, and for diagnosis in patients at high risk for invasive candidiasis. Part I: Rabbits were experimentally infected. Of 13 rabbits with disseminated candidiasis blood was drawn from central venous catheter and from peripheral veins. From catheters, Candida was detected in 13/13, 10/13 and 8/13, by buffy coat culture, IFA, and blood culture, respectively. From peripheral veins, Candida was detected in 10/10 by buffy coat cultures, in 6/10 by IFA and in 3/10 by blood cultures. The gold standard was histopathological diagnosis; sensitivity for IFA was 60% with peripheral blood and 76% with catheter blood samples. Part II: IFA and blood cultures were compared in clinical samples from hospitalized children. Forty patients were included. Candida was isolated from blood cultures in four patients whereas IFA detected Candida in 22 patients. Differences were statistically significant (p < 0.05). We conclude that IFA in buffy coat smears is a reliable test to detect candidemia, and would be a useful tool to help physicians with treatment decisions with amphotericin B.

Animals↗
PubMed1991

Purification and characterisation of toxin B from a strain of Clostridium difficile that does not produce toxin A.

Most toxigenic strains of Clostridium difficile produce both toxin A and toxin B. The toxin produced by C. difficile strain 8864 was characterised and compared with those produced by C. difficile strain 10463. Toxin A was not detected by immunoassay in cultures from strain 8864 and all the cytotoxic activity produced by this strain was neutralised by antiserum to toxin B. Toxin B from strain 8864 was purified and compared with toxin B from strain 10463. The size of the purified subunits of toxin B from strain 8864 differed slightly from those of strain 10463 and there were small immunological differences. The effect on fibroblast cells was more like that of C. sordellii cytotoxin than of toxin B from strain 10463. These results suggest that C. difficile strain 8864 produces a modified toxin B and does not produce toxin A.

Bacterial Proteins↗
PubMed1989

Production, purification and characterization of Clostridium difficile toxic proteins different from toxin A and from toxin B.

The purification and characterization of three new proteins called C1, C2, and C3 from Clostridium difficile are described. Their estimated molecular mass were about 350 (C1), 270 (C2) and 140 (C3) kDa, consisting of subunits of 39 (C1), 43 (C2) and 41 (C3) kDa, respectively. Immunodiffusion revealed that the three proteins contained similar but not identical antigenic determinants to toxin A. Each protein induced a cytotonic effect on hamster ovaric cells; the combined proteins, had a specific activity on cells 5-times higher than that of toxin A. In rat intestinal loops, they induced a clear fluid secretion, while toxin A elicited a haemorrhagic fluid response. The cytotonic activities of all three proteins were abolished by antiserum against toxin A, while antiserum against toxin B inhibited only the activity of the 270 kDa protein. In contrast to toxin A, the cytotoxicity of the three proteins was inactivated by trypsin. Thus, the chemical, antigenic and biological properties of these proteins differed from those of toxin A and toxin B.

Bacterial Toxins↗
PubMed1988

Purification and characterisation of two forms of toxin B produced by Clostridium difficile.

Toxin B from Clostridium difficile was purified to homogeneity by gel filtration and high resolution ion exchange chromatography. Two forms of toxin B were found. Form 1 which seemed to consist of two identical subunits of 220-300 kDa; femtogram amounts of this toxin induced rounding of fibroblast cells. Form 2 contained subunits of 43 kDa and 105 kDa; the stoichiometric ratio probably being 4:1; picogram amounts were needed to induce rounding of fibroblast cells. Immunological studies suggested that both subunit types were antigenic and had epitopes which were identical with those of form 1.

Bacterial Proteins↗
PubMed1987

Epidemiological aspects of Clostridium difficile in a pediatric hospital and its role in diarrheal disease.

The influence of antibiotics on the frequency of colonization by Clostridium difficile and the presence of its cytotoxin in infants and older children was examined to determine its role in diarrheal disease. Cytotoxin was more closely associated with cases of diarrhea, both in infants and in children than the microorganism, although not significantly. The isolates were typed by means of sensitivity to bacteriophages and bacteriocins and their cytotoxigenic potential was also determined. Less than 30% of the colonized patients had toxigenic strains. A study of strain variability over a four-year period in the same hospital showed that two bacteriophage-bacteriocin types and non-toxigenic strains predominated. The common presence of non-toxigenic strains could explain in part the lack of correlation between isolation of Clostridium difficile and diarrhea. Most of the non-toxigenic strains showed moderate resistance to tetracycline, a property which might explain their ability to persist for long periods in the hospital.

Adolescent↗
PubMed1984

Prevalence of Clostridium difficile and its cytotoxin in infants in Mexico.

The incidence of Clostridium difficile and its cytotoxic activity were determined in the feces of 122 children under 1 year of age. Samples were obtained from children receiving antibiotics and with (52 cases) or without (26 cases) diarrhea, from children with diarrhea who did not receive antibiotics (22 cases), and from healthy children (22 cases). Isolation of C. difficile in feces from children in all groups was similar (mean 23.4%) except for the group with non-antibiotic-associated diarrhea (4.5%). In both groups of children receiving antibiotics, with or without diarrhea, the cytotoxin was detected in 7.6% of the cases. In the group with non-antibiotic-associated diarrhea, none of the samples was positive for cytotoxicity. In healthy children, cytotoxin was positive in 4.5% of the cases.

Anti-Bacterial Agents↗
PubMedDate not supplied

Toxigenicity and adherence in Clostridium difficile strains isolated from patients with and without diarrhoea.

The mechanisms by which Clostridium difficile causes diarrhoea are unknown. The expression of putative virulence factors by 44 Clostridium difficile strains isolated from patients with and without diarrhoea was studied. Toxins A and B were tested in CHO and MRC-5 cells, respectively; adherence was measured in two substrates: HEp-2 cells and polystyrene plates. The in vitro expression of toxins A and B by strains isolated from patients with diarrhoea was not significantly different from that by strains isolated from patients without diarrhoea. The ability of adherence to both HEp-2 cells and polystyrene by strains isolated from patients with diarrhoea was not significantly different when compared with strains isolated from patients without diarrhoea; however, strains isolated from adults with diarrhoea seem to adhere to a greater extent to both substrates than strains isolated from adults without diarrhoea. Twenty three strains which did not produce toxins A and B were tested for enterotoxicity in rat small intestine. Eight such strains induced fluid accumulation and seven of them were isolated from children. Adherence to cells and to polystyrene might be an important virulence factor in strains causing diarrhoea in adults; whereas the production of toxins other than A and B might be an important pathogenic mechanism in strains causing diarrhoea in children.

Adult↗