Immunoglobulin G antibody response to infection with coccoid forms of Helicobacter pylori.

An increasing number of studies support a potential role for coccoid forms in Helicobacter pylori infection. Evidence for this was obtained through scanning microscopy, genetic analysis for virulence traits, examination of the presence and activity of key enzymes, and other methods. We studied the serum immunoglobulin G responses to coccoid H. pylori forms by enzyme-linked immunosorbent assay (ELISA) and immunoblotting and compared them with those of bacillary cells. Sera from a total of 295 infected individuals were studied; these included sera from 100 patients with duodenal ulcers, 98 patients with nonulcer dyspepsia, 11 patients with gastroduodenal cancer, and 86 asymptomatic individuals. Initially, we characterized and selected coccoid and bacillary antigenic preparations by one-dimensional (1-D) and 2-D gel electrophoresis and immunoblotting. Data showed that coccoid and bacillary preparations with comparable protein contents have similar patterns in 1-D and 2-D electrophoresis gels and antigenic recognition at blotting. These results revealed that coccoid and spiral antigens in ELISA can equally recognize specific antibodies to H. pylori in sera from infected individuals. The analysis of the spiral and coccoid preparations by Western blotting showed no major differences in antigen recognition. No specific bands or profiles associated with a single gastric condition were identified.

Portación de Staphylococcus aureus enterotoxigénicos en manipuladores de alimentos / Carriage of enterotoxigenic Staphylococcus aureus in food handlers

BACKGROUND: One of the most common pathogens causing alimentary toxi-infections is Staphylococcus aureus (S aureus). The presence of S aureus in food, indicates flaws during food manipulations. For this reason most sanitary norms require the detection of S aureus carriers. AIM: To determine the carriage rate of enterotoxin producing S aureus strains in food handlers, and to evaluate the antibiotic susceptibility to six antimicrobial agents. MATERIALS AND METHODS: A total of 102 food handlers from 19 restaurants in Santiago, were analyzed. Samples for microbiological analysis were obtained with a swab from the retropharynx. RESULTS: S aureus grew in 35 out of the 102 samples obtained (34 per cent). Further analysis revealed that 19/35 (54 per cent) strains were able to produce enterotoxins. Therefore the corrected carriage rate was 19 per cent (19/102). The most frequently detected enterotoxin was the type A (12/19). All S aureus isolates were resistant to penicillin and susceptible to oxacillin, clindamycin, kanamycin, vancomycin and linezolid. CONCLUSIONS: The carriage rate of S aureus in food handlers in similar to the rate reported in the general population in our country. These results confirm the need for education and training programs in food safety, directed to food handlers.

Prevalence of serum antibodies to Helicobacter pylori VacA and CagA and gastric diseases in Chile.

The objective of this study was to evaluate the prevalence of antibodies to Helicobacter pylori CagA and VacA proteins and correlate this prevalence with gastric diseases in colonised Chileans. The study was performed in 418 adults colonised with H. pylori: 316 with gastroduodenal pathology (152 duodenal ulcer, 14 gastric cancer and 150 gastritis patients) and 102 asymptomatic subjects. Serum IgG antibodies to H. pylori were determined by enzyme immunoassay (EIA). Antibodies to VacA and CagA proteins were detected by Western blotting. In a subgroup of the patients, the vacuolating activity was determined by HeLa cell assay and the CagA product was confirmed by PCR assay. IgG antibodies to both VacA and CagA proteins of H. pylori were found in 270 (85%) of 316 colonised gastric patients and in 72 (71%) of 102 asymptomatic subjects. Colonisation with virulent strains was significantly higher among duodenal ulcer and gastric cancer patients than in gastritis patients or asymptomatic subjects. Infections with VacA+/ CagA+ H. pylori strains is common in Chile but, in contrast to some Asian countries, this phenotype was more prevalent in isolates from patients with more severe gastric pathologies.

Restricted growth of ent(-) and tonB mutants of Salmonella enterica serovar Typhi in human Mono Mac 6 monocytic cells.

Monocytes and macrophages are an important host defense in humans infected with Salmonella enterica serovar Typhi. Bacterial ability to survive in these cells is therefore a crucial virulence characteristic of this pathogen. In this study, we demonstrate that growth of a Salmonella enterica serovar Typhi enterochelin synthesis mutant and a tonB mutant in the human monocyte cell line Mono Mac 6 is restricted compared to that of the parental wild-type Ty2 strain. These results suggest that enterochelin- and TonB-mediated iron uptake plays a role in S. enterica serovar Typhi pathogenesis, and also suggest that mutations in iron uptake may attenuate S. enterica serovar Typhi strains for human beings.

Disfunción endotelial como alteración primaria en las patologías vasculares / Endothelial dysfunction as a primary alteration in vascular diseases

Endothelium controls vascular smooth muscle tone by secreting relaxing and contracting factors. There is a constant release of endothelium derived relaxing factors, mainly nitric oxide, a potent vasodilator, inhibitor of platelet aggregation, monocyte adhesion and smooth muscle proliferation. In addition, the endothelium may increase the release of NO in response to humoral stimulation by vasoactive substances such as acetylcholine, bradikinin or substance P. Although the endothelium releases a number of products, no single blood test has yet proved useful to determine normal endothelial function or as early abnormalities. The most useful test of endothelial function relies on the meassurement of endothelium-dependent dilatation in response to pharmacological or physiologic stimuli. The alteration of this response is known as endothelial dysfunction and has been observed in a variety of circumstances related to cardiovascular risk. This review summarizes the evidence that sustains this association and emphasizes the clinical utility of assessing endothelial function presenting two clinical cases of hypercholesterolemia in which a high-resolution vascular ultrasound in the braquial artery was used

Lipopolysaccharide-independent radioimmunoprecipitation and identification of structural and in vivo induced immunogenic surface proteins of Salmonella typhi in typhoid fever.

The humoral response to Salmonella typhi is important for protective immunity against typhoid fever, as indicated by the protection obtained with killed cell vaccines and component vaccines (outer membrane proteins, Vi antigen) in animals and human beings. Nonetheless, analysis and interpretation of host humoral immune response to S. typhi surface antigens have been difficult because of the complex structure of the S. typhi envelope and the lack of purified reagents for detection of immune response to individual surface components. Normal and convalescent human sera from typhoid fever patients were absorbed with S. typhi lipopolysaccharide. These sera were used in radioimmunoprecipitation assays of whole S. typhi cells and S. typhi membranes labelled with either 125I or 35S-methionine. This strategy has permitted the unequivocal identification of a humoral immune response to structural and in vivo induced outer membrane proteins of S. typhi. In this manner, we have identified the porins, lipoprotein, the iron-starvation-induced proteins, and three proteins of 30, 18.5 and 15 kDa as surface-exposed immunogens of S. typhi in patients with typhoid fever. These studies suggest that further experimental work is needed to characterize the relevance of both anti-S. typhi outer membrane protein and antilipopolysaccharide antibodies in recovery from S. typhi infections and protective immunity.

Absence of a role for DNA polymerase II in SOS-induced translesion bypass of phi X174.

In order to examine the possible role of Escherichia coli DNA polymerase II in SOS-induced translesion bypass, Weigle reactivation and mutation induction were measured with single-stranded phi X174 transfecting DNA containing individual lesions. No decrease in bypass of thymine glycol or cyclobutane pyrimidine dimers in the absence of DNA polymerase II was observed. Furthermore, DNA polymerase II did not affect bypass of abasic sites when either survival or mutagenesis was the endpoint. Lastly, repair of gapped DNA molecules, intermediates in methyl-directed mismatch repair, was also unaffected by the presence or absence of DNA polymerase II.

Processing of model single-strand breaks in phi X-174 RF transfecting DNA by Escherichia coli.

The inactivation efficiency and repair of single-strand breaks was investigated using model strand breaks created by endonucleolytic incision of damaged DNA. Phi X-174 duplex transfecting DNA containing either thymine glycols, urea residues, or abasic (AP) sites was incubated with AP endonucleases that produce breaks on the 3' side, the 5' side, or both sides of the lesion. For each lesion, incubation with Escherichia coli endonuclease III results in a single-strand break containing a 3' alpha, beta-unsaturated aldehyde (4-hydroxy-2-pentenal), while treatment of AP- or urea-containing DNA with E. coli endonuclease IV results in a single-strand break containing a 5' deoxyribose or a 5' deoxyribosylurea moiety, respectively. Incubation of lesion-containing DNA with both enzymes results in a base gap. Ligatable nicks containing 3' hydroxyl and 5' phosphate moieties were produced by subjecting undamaged DNA to DNase I. When the biological activity of these DNAs was assessed in wild-type cells, ligatable nicks were not lethal, but each of the other strand breaks tested was lethal, having inactivation efficiencies between 0.12 and 0.14. These inactivation efficiencies are similar to those of the base lesions from which the strand breaks were derived. In keeping with the current model of base excision repair, when phi X duplex DNA containing strand breaks with a blocked 3' terminus was transfected into an E. coli double mutant lacking the major 5' cellular AP endonucleases, a greater than twofold decrease in survival was observed. Moreover, when this DNA was treated with a 5' AP endonuclease prior to transfection, the survival returned to that of wild type. As expected, when DNA containing strand breaks with a 5' blocked terminus or DNA containing base gaps was transfected into the double mutant lacking 5' AP endonucleases, the survival was the same as in wild-type cells. The decreased survival of transfecting DNA containing thymine glycols, urea, or AP sites observed in appropriate base excision repair-defective mutants was also obviated if the DNA was incubated with the homologous enzyme prior to transfection. Thus, in every case, with both base lesions and single-strand breaks, the lesion was repaired in the cell by the enzyme that recognizes it in vitro. Furthermore, the repair step in the cell could be eliminated if the appropriate enzyme was added in vitro prior to transfection.(ABSTRACT TRUNCATED AT 400 WORDS)

Choleriform syndrome and production of labile enterotoxin (CT/LT1)-like antigen by species of Salmonella infantis and Salmonella haardt isolated from the same patient.

Isolates of Salmonella infantis from the stool and blood and an isolate of Salmonella haardt from the stool of a patient with choleriform diarrhea produced labile enterotoxin (CT/LT1)-like antigen. Genetic and molecular experiments indicated that the production of CT/LT1-like antigen was chromosomally encoded.

[Evaluation of an immunoenzyme technique (ELISA) to diagnose typhoid fever]. / Evaluación de un ensayo inmunoenzimático (ELISA) para el diagnóstico de la fiebre tifoídea.

The efficiency of an ELISA method, designed to detect polyvalent IgG and IgM antibodies to Salmonella typhi polysaccharide was evaluated in patients admitted or convalescing from typhoid fever and in control subjects. Polyvalent antibodies to S typhi were demonstrated in 28/30 (93%) typhoid patients, 0/11 bacteremic patients (E coli or S paratyphi A) and 0/15 asymptomatic individuals. Widal test showed significant anti-0 agglutinin values (> = 1: 160) in only 12/30 (40%), 1/11 (9%) and 0/15 subjects from each group respectively. Typhoid patients tested on admission or at discharge showed similar high reactivity rates to ELISA. On the contrary, the Widal test detected only 2/15 (13%) patients on admission (p < 0.02) and 10/15 (67%) at discharge. These results and additional immunoblotting tests suggest that ELISA developed to detect polyvalent anti-LPS antibodies could represent a highly specific diagnostic tool to confirm typhoid fever in endemic areas.

Chromosomal DNA, iron-transport systems, outer membrane proteins, and enterotoxin (heat labile) production in Salmonella typhi strains.

We examined a representative collection of Salmonella typhi strains from Chile, Peru, Mexico, India, and England for the presence of several properties. All strains had a conserved pattern of outer membrane proteins, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The electrophoresis profiles of chromosomal DNA digested with EcoRI and PstI restriction enzymes were similar for all the strains. A conserved pattern of hybridization was observed when digested chromosomal DNA was hybridized with DNA probes for the 36-kilodalton porin, enterobactin synthesis, and enterobactin receptor genes. All the strains produced enterobactin but not aerobactin in bioassays. None of the strains produced heat-labile toxin, as measured by an enzyme-linked immunosorbent assay. Colony and Southern hybridizations with DNA probes for aerobactin synthesis and its receptor and heat-labile toxin genes were negative. These results indicate that S. typhi strains from different origins have similar phenotypic and genetic properties and, as has been suggested, constitute a clone.

Characterization of enteroinvasive Escherichia coli strains isolated from children with diarrhea in Chile.

Analysis of stool samples from 912 cases of diarrhea among Chilean infants and 1,112 controls resulted in the isolation of 17 enteroinvasive Escherichia coli (EIEC) strains from diarrhea cases (1.9%) and 3 EIEC from the asymptomatic controls (0.3%). Biochemical analysis of the 20 isolates showed variability among them. However, the majority were lysine decarboxylase negative and nonmotile and utilized sodium acetate. The strains belonged to the O groups 28ac, 124, 143, or 144 or were untypable with the antisera used. Most of them had conjugative plasmids which mediated multiple antibiotic resistance. There was a strong correlation in this group of strains between a positive Sereny test, the presence of a plasmid of 120 megadaltons, and hybridization with the invasiveness probe, an HindIII fragment derived from the plasmid pPS15A. The isolates had a wide range of plasmid profiles. Bioassays and colony and Southern hybridization tests with iron uptake DNA probes indicated that 80% of the EIEC strains produced aerobactin and expressed its receptor, the genes for which are known to be chromosomally located.

Role of the inner protein capsid on in vitro human rotavirus transcription.

The inner protein shell of human rotavirus consists of a single polypeptide called VP6 which was removed from the single-shelled virus by treatment with CaCl2, leaving the viral core. The core thus obtained was unable to transcribe. However, the addition of a supernatant containing VP6 in the absence of Ca2+ restored the transcriptional activity. VP6 obtained from different electropherotypes and serotypes was able to restore transcriptional activity to homologous and heterologous cores. Viral cores obtained after incubation with purified VP6 had electron microscopic characteristics, polypeptide compositions, and transcription products similar to those of the single-shelled virus. The results suggested the successful in vitro reconstitution of the single-shelled virus.

In vitro transcription of human pararotavirus.

Purified human pararotavirus obtained from stool samples from a 6-month-old infant was characterized. Electron microscopy of the viral particles subjected to different treatments suggested that the protein shells differed from those described for rotavirus. Treatment with both EDTA or ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid in the presence or absence of Mg2+ seemed to convert the virions into core particles by removal of both the outer and inner shells, and no particles equivalent to single-shelled rotavirus were observed. Different procedures were used to activate the human pararotavirus-associated RNA-dependent RNA polymerase. The enzyme was not activated by chelating agents or by thermal shock as in rotavirus. Activation by thermal shock occurred only in the presence of the four ribonucleoside triphosphates and Mg2+. However, the polymerase of pararotavirus was found to be similar to those described for rotaviruses. When in vitro transcripts were analyzed, 11 RNA species having a migration pattern similar to that of the original genomic RNA were detected.