Cyclic enterobacterial common antigens from Escherichia coli O157 as microbe-associated molecular patterns.

In a previous study, we described 2 forms of cyclic enterobacterial common antigen (ECACYC), a tetramer and a pentamer, from Escherichia coli O157. ECACYC is present in several representatives of the Enterobacteriaceae. To date, functional studies on ECACYC are sparse. Cyclic oligosaccharides in other bacteria, like the cyclic ß-glucans in Rhizobiaceae, represent microbe-associated molecular patterns involved in host-bacteria interaction. This observation determined the aim of the present study: to test whether the tetrameric and pentameric ECACYC from E. coli O157 can be recognised by host humoral and cellular mechanisms. ELISA tests designed to compare the 2 ECACYC with the O157 lipopolysaccharide showed that both ECACYC were not recognised by polyclonal anti-O157 serum but were good ligands for mannan-binding lectin. The lectin had a higher affinity for the tetramer than the pentamer. ECACYC deposited more C3b than did the lipopolysaccharide. To examine the interactions with human circulating neutrophils, the antigens were loaded onto fluorescent latex beads and applied in a phagocytosis experiment. Spheres coated with the 2 ECACYC occasionally adhered to phagocyte surfaces but, unlike O157-loaded spheres, failed to induce free-radical release. The results show that the 2 ECACYC represent microbe-associated molecular patterns recognised by host humoral non-self-recognition mechanisms.

Released products of pathogenic bacteria stimulate biofilm formation by Escherichia coli K-12 strains.

It has recently been shown that pathogens with a limited capacity for sessile growth (like some Escherichia coli O157 strains) can benefit from the presence of other bacteria and form mixed biofilms with companion strains. This study addresses the question whether pathogens may influence attached growth of E. coli non-pathogenic strains via secreted factors. We compared the biofilm-modulating effects of sterile stationary-phase culture media of a biofilm non-producing strain of E. coli O157:H, a laboratory biofilm-producing E. coli K-12 strain and a biofilm-forming strain of the pathogen Yersina enterocolitica O:3. Sessile growth was monitored as biomass (crystal violet assay), exopolysaccharide (ELLA) and morphology (scanning electron and confocal laser microscopy). With two of the E. coli K-12 strains stimulation of biofilm formation by all supernatants was achieved, but only the pathogens' secreted products induced biomass increase in some 'biofilm-deficient' K-12 strains. Lectin-peroxidase labeling indicated changes in colanic acid and poly-N-acetylglucosamine amounts in extracellular matrices. The contribution of indole, protein and polysaccharide to the biofilm-modulating activities of the supernatants was compared. Indole, in concentrations equal to those established in the supernatants, suppressed sessile growth in one K-12 strain. Proteinase K significantly reduced the stimulatory effects of all supernatants, indicating a prominent role of protein/peptide factor(s) in biofilm promotion. The amount of released polysaccharides (rPS) in the supernatants was quantitated then comparable quantities of isolated rPS were applied during biofilm growth. The three rPS had notable strain-specific effects with regard to both the strain-source of the rPS and the E. coli K-12 target strain.

Triglyceride levels and apolipoprotein E polymorphism in patients with acute pancreatitis.

BACKGROUND: Hypertriglyceridemia is an unusual cause of acute pancreatitis and sometimes considered to be an epiphenomenon. This study aimed to investigate the clinical and analytical features and the APOE genotypes in patients with acute pancreatitis and severe hypertriglyceridemia. METHODS: We undertook a one-year, prospective study of patients with acute pancreatitis whose first laboratory analysis on admission to the emergency department included measurement of serum triglycerides. The APOE genotype was determined and the patients answered an established questionnaire within the first 24 hours concerning their alcohol consumption, the presence of co-morbidities and any medications being taken. The patients' progression, etiological diagnosis, hospital stay and clinical and radiological severity were all recorded. RESULTS: Hypertriglyceridemia was responsible for 7 of 133 cases of pancreatitis (5%); the remaining cases were of biliary (53%), idiopathic (26%), alcoholic (11%) or other (5%) origin. Compared with these remaining cases, the patients with hypertriglyceridemia were significantly younger, had more relapses, and more often had diabetes mellitus. They usually consumed alcohol or consumed it excessively on the days before admission. Also, the ε4 allele of the APOE gene was more common in this group (P<0.05). CONCLUSION: One of 20 episodes of acute pancreatitis is caused by hypertriglyceridemia and it is linked to genetic (ε4 allele) and comorbid factors such as diabetes and, especially, alcohol consumption.

[Left-sided endocarditis due to gram-negative bacilli: epidemiology and clinical characteristics]. / Endocarditis sobre válvulas izquierdas por bacilos gran negativos: epidemiología y características clínicas.

INTRODUCTION: The aim of this study is to describe the epidemiological, clinical characteristics, and outcome of patients with left-side endocarditis caused by gram-negative bacteria. METHOD: Prospective multicenter study of left-sided infective endocarditis reported in the Andalusian Cohort for the Study of Cardiovascular Infections between 1984 and 2008. RESULTS: Among the 961 endocarditis, 24 (2.5%) were caused by gram-negative bacilli. The most common pathogens were Escherichia coli, Pseudomonas aeruginosa and Salmonella enterica. Native valves (85.7%) were mainly affected, most of them with previous valve damage (57%). Comorbidity was greater (90% vs 39%; P=.05) than in endocarditis due to other microorganism, the most frequent being, diabetes, hepatic cirrhosis and neoplasm. A previous manipulation was found in 47.6% of the cases, and 37% were considered hospital-acquired. Renal failure (41%), central nervous system involvement (33%) and ventricular dysfunction (45%) were the most frequent complications. Five cases (21%) required cardiac surgery, mostly due to ventricular dysfunction. More than 50% of cases were treated with aminoglycosides, but this did not lead to a better outcome or prognosis. Mortality (10 patients) was higher than that reported with other microorganisms (41% vs 35%; P=.05). CONCLUSIONS: Left-sided endocarditis due to gram-negative bacilli is a rare disease, which affects patients with major morbidities and often with a previous history of hospital manipulations. Cardiac, neurological and renal complications are frequent and associated with a high mortality. The association of aminoglycosides in the antimicrobial treatment did not involve a better outcome or prognosis.

Lead, cadmium, zinc, and copper bioavailability in the soil-plant-animal system in a polluted area.

A comparative research study on the bioavailability of Pb, Cd, Zn, and Cu in the soil-plant-animal-system was carried out. The connection between the total quantity and the mobile forms of Pb, Cd, Zn, and Cu in soils with different levels of contamination; the transition of these metals into rapeseed; and their assimilation by rabbits fed with a food that consisted mainly of rapeseed was studied. It was established that the absorption of heavy metals by the rapeseed definitely has a selective character, as the affinity towards Zn is most strongly expressed. The accumulation of Pb, Cd, Zn, and Cu in the organs of the rapeseed occurs in the following order: inflorescences > leaves > stems. A direct connection between the quantity of the mobile forms and their accumulation in the plants was not found. The environmental contamination has a significant effect on heavy metal levels and distribution, as the largest quantity of all four elements is accumulated in the kidneys and liver. A well-expressed impact of the level of Cd contamination on the absorption of essential trace metals (Zn and Cu) and their accumulation into some of the organs of the animals was found.

Cross-reactive monoclonal antibodies raised against the lipopolysaccharide antigen of Salmonella minnesota Re chemotype: diagnostic relevance.

Three cross-reactive monoclonal antibodies (MAbs) of IgM and IgG2b isotype were generated in two separate fusions after immunization of BALB/c mice with heat killed Salmonella minnesota R595 of Re chemotype and acid-treated bacteria, coated with Re lipopolysaccharide (LPS) antigen. The specificity of the MAbs was demonstrated as the Re LPS antigen. The activity and cross-reactivity against purified elementary bodies of Chlamydia trachomatis and various S- and R-LPS antigens of other Gram-negative bacteria were characterized in enzyme-linked immunosorbent assay, passive hemolysis assay, immunoblot and immunofluorescence with the available chlamydial strains. The results demonstrated cross-reaction between the Re LPS antigen, the genus-specific chlamydial LPS and the LPS antigens of Escherichia coli O119 and Acinetobacter baumannii, suggesting the presence of identical or similar epitopes in the lipopolysaccharide antigens. The findings are implying the necessity of novel approaches, improving the specificity of serologic assays in the laboratory diagnosis of chlamydial infections.

Monoclonal antibody against O:5 Salmonella antigen cross-reacts with unidentified lipopolysaccharide epitope of Salmonella serogroup O:8 (C(2)-C(3)).

Monoclonal antibody (MAb) 8aC10 against Salmonella O:5 antigen was obtained after immunization of BALB/c mice with live attenuated mutant of Salmonella typhimurium. Antigen specificity of the MAb was characterized by ELISA, immunoblotting, passive hemagglutination (PHA), passive hemolysis and agglutination tests. In ELISA, PHA and immunoblotting the MAb reacted only with lipopolysaccharides (LPS) of Salmonella strains from group O:4 (B), expressing O:5 antigen. The MAb agglutinated in addition Salmonella strains with O:8 antigen from group C(2)-C(3) but did not react with purified LPS. These results demonstrate O:5 specificity of MAb 8aC10. Cross-agglutination with group C(2)-C(3) suggests the presence of similar but not identical epitope in O:8 expressing strains, which is possibly localized onto O-acetyl-abequose and abequose residues bound with a alpha-1-->3 linkage to the basic polysaccharide backbone of Salmonella LPS with O:5 and O:8 antigen respectively.

Production and characterization of monoclonal immunoglobulin A antibodies directed against Salmonella H:g,m flagellar antigen.

Hybridomas were generated after intragastral immunization of BALB/c mice with live Salmonella suberu and subsequent fusion between isolated spleen lymphoblasts and myeloma cells. Three monoclonal antibodies (MAbs) of immunoglobulin A (IgA) isotype were selected and characterized. All of them were found to recognize the H:g epitope in enzyme-linked immunosorbent assay and immunoblotting but did not react with all H:g-expressing strains in slide agglutination test. All MAbs strongly agglutinated Salmonella enteritidis type strain and a large number of S. enteritidis clinical isolates. They were not bactericidal in the presence of complement. All hybridoma clones produced secretory IgA forms, which were found in the gastrointestinal tract of mice bearing hybridoma as a subcutaneous 'backpack' tumor or after intravenous application of purified MAbs. The IgA MAbs stability demonstrated in different tests together with their antigen specificity and strong agglutination ability make them a useful diagnostic tool for serotyping of Salmonella strains.

Welfare Homes for Patients Living with HIV/AIDS in Andalusia.

AIMS: To study the AIDS welfare homes (AWHs) in Andalusia, assess their resources and the services provided, and describe the characteristics of their residents. PATIENTS AND METHODS: Cross-sectional, observational study; an interview questionnaire technique was used with the managers and the residents of the AWHs. RESULTS: A total of 7 AWHs and 96 residents were included; 32% of the staff were health care workers and 45.5% volunteers. The occupancy rate was 86% (2007) and 96% (2008). Residents' characteristics: mean age 45.6 years, 73% male, 92% with at least 1 AIDS-defining disease, median Karnofsky index 60 (50-80), and median Barthel index 80 (40-100). Half the residents had physical sequelae and 31% mental sequelae. CONCLUSION: The AWHs perform an important role in the care of certain types of patients with HIV infection. They require human and material resources to be able to tackle the immense difficulties associated with this group of patients.

Occurrence and structure of cyclic Enterobacterial Common Antigen in Escherichia coli O157:H⁻.

Two cyclic forms of the Enterobacterial Common Antigen were isolated from Escherichia coli O157:H(-). These antigenic determinants were purified from the biomass through extensive chemical, enzymatic and chromatographic procedures whereas MALDI MS spectrometry indicated their cyclic nature with a polymerization degree of 4 or 5. The two species, denoted as ECA(CYC-4) and ECA(CYC-5), were assigned by NMR and showed no further substitution with other appendages such as acetyl groups as usually described for similar cyclic antigens from other Enterobacteriaceae.

Lectin-binding epitopes at the surface of Escherichia coli K-12: examination by electron microscopy, with special reference to the presence of a colanic acid-like polymer.

The presence and distribution of lectin-binding epitopes at the surface of Escherichia coli K-12, strain W1655, was studied by electron microscopy after lectin-gold labeling and negative staining. A comparison was made between the lectin-binding capacity of cells cultivated at 20 degrees C and 37 degrees C (in broth or on agar). A variety of pre-treatment protocols were applied prior to labeling. The gold-conjugated lectins used were wheat germ agglutinin (WGA), soybean agglutinin (SBA) and Ulex europaeus lectin (UEA-I). For all culture conditions, the bacteria had moderate exposure of WGA-binding sites, and this was not changed after pre-treatment. Cells cultivated at 37 degrees C had exposed SBA- and UEA-I-binding epitopes apparently associated with the cell surface. These significantly increased in number after boiling the cells for 10 min. With bacteria cultivated at 20 degrees C these two lectins recognized sites situated on exopolysaccharide filaments. Affino dot-blot experiments with isolated polysaccharides of the strain identified the K-12 lipooligosaccharide as the source of WGA-binding epitopes, and the exopolysaccharide, colanic acid (CA) as the source of SBA- and UEA-I-binding sites. The interaction with these two lectins of bacteria cultivated at 37 degrees C could be due to altered translocation of CA from the cytoplasm to the environment. This suggestion was supported by the demonstration by electron microscopy of SBA and UEA-I binding at the surface of hot phenol-water extracted cell walls.