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Biomedical subjects

L R Trabulsi

Publications and source records attributed to L R Trabulsi.

At least 19 recordsLinked to original sources

Inhibition of enteropathogenic Escherichia coli adherence to HeLa cells by immune rabbit sera.

We have studied the effect of immune rabbit sera on the localized (LA) and diffuse (DA) adherence to HeLa cells of 10 enteropathogenic Escherichia coli (EPEC) strains belonging to serogroups O55, O86, O111, O119, and O142. Anti-La1 serum, obtained by rabbit immunization with an E. coli strain harboring a cloned DNA fragment from an EPEC LA plasmid, strongly inhibited the adherence of all serogroups but one (O142). Similar results were obtained with anti-LA2 serum, which is anti-O111 serum absorbed with a non-adherent O111:H- EPEC strain. In contrast, non-absorbed anti-O55 and anti-O111 sera showed an inhibitory effect mainly on the adherence of homologous strains. Except for one experiment diffuse adherence was not inhibited by any antiserum used. The inhibitory effect of immune sera on localized adherence does not seem to be correlated with plasmid curing since adherence plasmid pMS49 proved to be stable after treatment with anti-O55 and anti-O111 sera. The cross-inhibition of adherence by anti-LA sera suggests that localized adherence-related adhesins of the O55, O86, O111, and O119 strains share similar antigens.

Animals

Enteropathogens associated with acute diarrheal disease in urban infants in São Paulo, Brazil.

To determine the prevalence and epidemiology of enteropathogens in acute infantile diarrhea, 500 infants less than or equal to 12 months of age with diarrhea and 500 age-matched control subjects coming to a São Paulo emergency room were studied. Enteropathogens were identified in 55% of case infants and 10% of controls; enteropathogenic Escherichia coli (EPEC) of classic EPEC serotypes producing EPEC adherence factor (EAF) (26% of case infants), rotavirus (14%), Salmonella species (8%), enterotoxigenic E. coli (7%), and Shigella species (5%) were associated with diarrhea. Isolation of EAF+ classic EPEC decreased with increasing age of cases and peaked in spring, whereas rotavirus was least common in early infancy and peaked in fall and winter. Bloody stool had a 36% positive predictive value for Shigella infection, EAF+ classic EPEC were highly resistant to antimicrobial drugs. Among poor São Paulo infants, EAF+ classic EPEC equaled or exceeded rotavirus throughout the year as a cause of diarrhea bringing children to medical attention.

Acute Disease

Use of ampicillin plus ranitidine for Helicobacter pylori gastritis.

1. Helicobacter pylori (formerly Campylobacter pylori) is now recognized as an etiological factor in gastritis and duodenal ulcers and probably also gastric ulcers. Eradication of the bacteria is fundamental to avoid ulcer relapse. Although bismuth salts have been shown to be effective for treatment, they are not commercially available in Brazil. 2. We report an attempt to treat patients with Helicobacter pylori-associated gastritis with ampicillin (1000 mg twice daily for one month) and compare the results with the conventional treatment used in Brazil (ranitidine, 300 mg daily for one month) and with a combination of the two drugs. We studied 44 patients with histologically confirmed gastritis and with Helicobacter pylori, who were examined at the beginning and after one month of treatment. 3. Ampicillin associated with ranitidine was better than ampicillin or ranitidine alone for the treatment of gastritis. Although ampicillin may be more efficient in patients with lower acid output we did not find a statistically significant difference between these two groups (ampicillin vs drug combination), perhaps owing to the small number of patients studied. When ampicillin was combined with ranitidine there was 25% normalization of the histological picture of the gastric mucosa. 4. We conclude that ampicillin in combination with ranitidine may be a useful treatment for Helicobacter pylori-associated gastritis.

Ampicillin

New invasive serotype of Escherichia coli.

A new invasive Escherichia coli strain which presented delayed or variable reactions in lactose, mucate, and acetate tests is described. It did not agglutinate with antisera for all known E. coli and Shigella groups A, B, C, D, and provisional Shigella serovar. We propose the designation of E. coli BH until the situation of its 0 antigen is settled.

Antigens, Bacterial

Production of Shiga-like toxin among Escherichia coli strains and other bacteria isolated from diarrhea in São Paulo, Brazil.

An elevated level of Shiga-like toxin I (SLT-I) production was found in 1 of 466 Escherichia coli strains studied. Among the 34 sonic lysates obtained from classical enteropathogenic E. coli, 5 produced SLT-I. The Aeromonas, Citrobacter, Edwardsiella, Enterobacter, Klebsiella, Proteus, Providencia, Pseudomonas, Salmonella, Serratia, Shigella, Yersinia, and Vibrio strains also studied were not SLT producers, except for a Shigella dysenteriae type 1 strain. Although SLT-I-producing E. coli strains were isolated from diarrhea, they seem to be an uncommon cause of disease in children less than 1 year old in our community.

Bacterial Toxins

A study of the gram-negative bacterial flora in patients with ankylosing spondylitis.

1. Twenty-eight patients with active definite primary ankylosing spondylitis and fifty-four healthy control subjects were studied. 2. The HLA-B27 antigen was found in 75% of patients and 3.7% of controls. 3. Fecal samples from these subjects were cultured for gram-negative enteric bacteria on two occasions within one month. Positive cultures for Klebsiella sp were found in 32.1% of patients and in 22.2% of healthy controls, but this difference was not statistically significant. All other microorganisms detected were qualitatively and quantitatively similar in both groups. 4. Significantly increased mean values of serum IgA levels were found in the patient group when compared with the control group (P less than 0.01). The mean serum IgG and IgM levels did not differ statistically between the two groups. There was no correlation between any laboratory or clinical parameter and presence of Klebsiella sp carriage in ankylosing spondylitis patients. 5. These data are consistent with the view that a long time elapses between exposure to a trigger factor and clinical manifestations of the disease.

Adolescent

Serotype-specific prevalence of Escherichia coli strains with EPEC adherence factor genes in infants with and without diarrhea in São Paulo, Brazil.

To examine interrelationships of classic enteropathogenic Escherichia coli (EPEC) serotypes, EPEC adherence factor (EAF) genes, and diarrheal disease, E. coli were studied from stools of 500 infants less than 1 y of age with acute diarrhea and 500 age-matched controls. EAF-containing (EAF+) E. coli of three common classic EPEC serotypes (O111:H-, odds ratio [OR] 36.0; O111:H2, OR 55.0; O119:H6, OR 3.7) were individually strongly associated with diarrhea, as were EAF+ strains of less common classic serotypes combined (OR 5.3). Among EPEC serogroups, neither EAF+ strains of nonclassic serotypes (OR 1.8) nor EAF-strains of classic (OR 2.2) or nonclassic (OR 1.4) serotypes were significantly associated with diarrhea. At least one EAF+ non-EPEC serogroup serotype (O88:H25) may represent an unrecognized EPEC serotype. Serotype-specific variation in the association of EAF+ E. coli with diarrhea suggests that other factors are also important in determining virulence; thus, both EAF detection and E. coli serotyping are desirable in studying the etiology of diarrheal disease.

Acute Disease

Virulence factors and biochemical characteristics of serotypes of Escherichia coli serogroup O29.

Escherichia coli strains belonging to serogroup O29 were studied. Invasiveness was the most common virulence factor described in this serogroup, but a few papers also reported production of heat-stable (ST) enterotoxin. In the present study invasive ability was found in O29:H- strains, whereas production of ST-I enterotoxin was observed only in serotype O29:H21 strains, showing that virulence was a characteristic of specific serotypes or bioserotypes within the O29 serogroup. Different serotypes were found among strains that were neither invasive nor toxigenic. Invasive strains were biochemically less active than the toxigenic ones and presented the invasiveness plasmid (pINV) of about 120 to 140 megadaltons, whereas hybridization tests showed that ST-I production was related to a plasmid of about 90 megadaltons. A diffuse adherence pattern to HeLa cells was observed in all ST-I isolates, but the role of this adherence in the pathogenicity of these strains was not determined. Thus, a unique biochemical pattern and plasmid profile may be useful characteristics to distinguish between pathogenic (toxigenic or invasive) and nonpathogenic O29 strains.

Antigens, Bacterial

Prevalence of Escherichia coli strains with localized, diffuse, and aggregative adherence to HeLa cells in infants with diarrhea and matched controls.

To determine the possible role of Escherichia coli strains with three different patterns of adherence to HeLa cells in causing diarrhea in infants in São Paulo, Brazil, we studied stool specimens from 100 infants up to 1 year of age with acute diarrheal illnesses and 100 age-matched control infants without recent diarrhea. E. coli with localized adherence to HeLa cells was much more common in patients (23%) than in controls (2%) (P less than 0.0001) and was detected more frequently than rotavirus (19%) was in patients, even though the study was conducted during the coldest months of the year. Most (80%) of the E. coli colonies with localized adherence were of traditional enteropathogenic E. coli serotypes. Little difference was found between patients and controls in the rate of isolation of E. coli with diffuse adherence (31 and 32%, respectively) or aggregative adherence (10 and 8%, respectively). A genetic probe used to detect a plasmid-mediated adhesin which confers expression of localized adherence proved to be 100% sensitive and 99.9% specific in detecting E. coli with localized adherence to HeLa cells. Although E. coli strains with localized adherence have now been shown to be enteric pathogens in several parts of the world, the role of strains showing diffuse adherence and aggregative adherence is still uncertain.

Adhesins, Escherichia coli

Aggregation of membrane-associated actin filaments following localized adherence of enteropathogenic Escherichia coli to HeLa cells.

We have previously observed that enteropathogenic Escherichia coli (EPEC) adhere to HeLa cells in a localized manner, which we designated localized adherence as opposed to the diffuse pattern of adhesion. In this paper we have examined the effects of localized adherence of EPEC on the actin microfilament system of host HeLa cells. Centrifugation of bacteria onto HeLa cells improved the localized adherence and rapid rearrangements of actin filaments were detected by immunofluorescence and electron microscopy. Aggregation of microfilaments is consistently observed at the sites of localized adherence, and is abolished by cytochalasin D and low temperatures. Scanning electron microscopy indicates that these aggregates are surface microvilli entangled with attached EPEC.

Actin Cytoskeleton

Protracted diarrhea: the importance of the enteropathogenic E. coli (EPEC) strains and Salmonella in its genesis.

The etiology of the protracted diarrhea is ill-defined, but in the underdeveloped countries acute gastroenteritis might be the most common triggering factor, especially due to certain enteropathogenic bacteria, such as enteropathogenic E. coli (EPEC) and Salmonella. We investigated the role of these agents in the genesis of protracted diarrhea in 29 infants with a mean age of 4.6 months. The patients underwent the following tests: stool culture, culture of the jejunal secretion, and small bowel and rectal biopsies. The stool culture was positive for some enteropathogenic bacteria in 17 (58.6%) patients: EPEC serotypes 0126, 0125, 055, 026, 0111, 0127, 0114, 0158, and 0119 and Salmonella were identified. The jejunal secretion culture revealed bacterial proliferation in 15 (51.7%) patients, and the following bacteria were isolated: EPEC 0142, Proteus, Klebsiella, Enterobacter, EPEC 0114, Pseudomonas, EPEC 0111, Salmonella, and EPEC 0119. The small bowel biopsy showed subtotal villous atrophy in 13 (44.8%) patients, and the rectal biopsy revealed colitis in 13 (44.8%) patients. These findings stress the importance of those enteropathogenic bacteria in the genesis of protracted diarrhea in underdeveloped countries mainly due to food intolerance leading to aggravation of the nutritional status.

Diarrhea, Infantile

Isolation and characterization of the localized adherence factor of enteropathogenic Escherichia coli.

The binding factor of enteropathogenic Escherichia coli O111:H- responsible for localized adherence (LA) on HeLa cells was investigated. Inhibition of LA by carbohydrates and lectins showed that the reactive epitope on HeLa cells contains N-acetylgalactosamine units. Treatment of bacteria with EDTA for extraction of lipopolysaccharides eliminated these polymers as binding factors. Such treatment also caused a marked increase in adhesion suggesting steric hindrance by lipopolysaccharides of the LA factor binding capacity. Immunoblotting with rabbit antibodies showed a strong reaction with two components with approximate molecular sizes of 29 and 32 kilodaltons (kDa) present in the outer membrane preparations of bacteria. Both the absorbed rabbit immune serum and the outer membrane preparation of the bacteria inhibited bacterial adhesion by 100%. Outer membrane components were isolated from an N-acetylgalactosamine-agarose column by elution with KSCN, labeled with 125I, and immunoprecipitated with absorbed rabbit hyperimmune antiserum. The only component precipitated was the protein doublet at 29 to 32 kDa corresponding to the components detected by immunoblotting. The predominant component was always the 32-kDa polypeptide. We conclude that this component of the outer membrane is the best candidate for the LA factor in enteropathogenic E. coli.

Acetylgalactosamine

DNA probes for identification of enteroinvasive Escherichia coli.

Eighty-one Escherichia coli strains belonging to all known invasive O serogroups were tested with two distinct invasiveness probes (pMR17 and pSF55). All 54 Sereny test-positive strains and 5 strains that lost Sereny positivity during storage hybridized with both probes. Probe-positive strains carried a 120- to 140-megadalton plasmid, did not produce lysine decarboxylase, and, with the exception of certain serotypes, were nonmotile. Motile strains of serotype O144:H25 were for the first time characterized as invasive by hybridization with the probes.

DNA, Bacterial

Plasmids coding for drug resistance and localized adherence to HeLa cells in enteropathogenic Escherichia coli O55:H- and O55:H6.

Plasmids coding for drug resistance and localized adherence (LA) to HeLa cells were found in two enteropathogenic Escherichia coli strains belonging to serotypes O55:H- and O55:H6. Strain 49-81 HSJ (O55:H-) carries two plasmids, one coding for both ampicillin resistance (Apr) and LA (pMS49). Strain 71-82 HSJ (O55:H6) harbors only one plasmid, coding for resistance to sulfadiazine, chloramphenicol, kanamycin, ampicillin, and LA (pMS71). Plasmids pMS49 and pMS71 were transferred to E. coli K-12 711 and from this strain to E. coli K-12 J53. Curing with acridine orange of an Apr LA+ transconjugant showed that both characteristics were lost simultaneously. The plasmids have a molecular weight of approximately 55 X 10(6) and are the first naturally recombinant plasmids coding for adherence and drug resistance described in enteropathogenic E. coli.

Adhesiveness

Variation in chemical properties and antigenic determinants among type II heat-labile enterotoxins of Escherichia coli.

Type II heat-labile enterotoxin (LT-II) from Escherichia coli 41 was purified and compared with prototype LT-II encoded by genes from E. coli SA53. Both toxins were oligomeric proteins consisting of polypeptides A (Mr, 28,000) and B (Mr, 11,800). The A polypeptides were cleaved by trypsin into fragments A1 (Mr, 21,000) and A2 (Mr, about 7,000). These two toxins were shown to belong to two different subclasses of LT-II. We propose to designate the prototype toxin LT-IIa and the new variant LT-IIb. The pI of LT-IIb was between 5.2 and 5.6, significantly lower than the pI of 6.8 for LT-IIa, and the behavior of LT-IIb during purification differed significantly from that of LT-IIa. The toxic dose of unnicked LT-IIb in the Y1 adrenal-cell assay was 94 pg, but trypsin-treated, nicked LT-IIb was toxic at about 3 pg. In contrast, the toxic dose of LT-IIa was previously shown to be 0.5 to 1 pg for several preparations that varied from unnicked to partially nicked, and treatment with trypsin was not required for full toxicity. The titer of LT-II antiserum in neutralization tests was 100-fold greater against LT-IIa than against LT-IIb. In immunodiffusion tests, LT-IIa and LT-IIb gave a reaction of partial identity. In a radioimmunobinding assay, the titer of LT-IIa antiserum against homologous LT-IIa was approximately 10-fold greater than against LT-IIb. The cholera-E. coli family of heat-labile enterotoxins has been divided into serogroup I, which includes cholera toxin and the antigenic variants of E. coli heat-labile toxin designated LTh-I and LTp-I, and serogroup II, which includes LT-IIa and LT-IIb. The type I and type II toxins do not cross-react in neutralization or immunodiffusion tests. By using very sensitive radioimmunobinding assays, it was possible to demonstrate common antigenic determinants between the type I and type II toxins. However, the titers of antibodies in hyperimmune sera that recognized these common determinants were very low.

Antigen-Antibody Complex

Production of type II heat-labile enterotoxin by Escherichia coli isolated from food and human feces.

Escherichia coli strains isolated in Sao Paulo, Brazil, from feces of patients with diarrhea and from food samples produced toxin(s) that was shown to be related both immunologically and genetically to the recently characterized type II heat-labile enterotoxin of E. coli. The new isolates of type II heat-labile enterotoxin-producing E. coli belonged to five different serotypes and did not represent a single clone.

Bacterial Toxins