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Claudia Toma

Publications and source records attributed to Claudia Toma.

At least 19 recordsLinked to original sources
PubMed2006

Detection and characterization of Shiga toxin-producing Escherichia coli in captive non-domestic mammals.

Shiga toxin producing-Escherichia coli (STEC) is an important emerging pathogen, and ruminants are recognized as their main natural reservoir. The aim of this work was to establish the frequency of STEC in non-domestic mammals of the Zoo and Botanical Garden of La Plata City, Argentina, and to pheno-genotypically characterize STEC isolates. By polymerase chain reaction (PCR), Shiga toxin (stx) gene sequences were detected in 50.8% of 65 fecal samples. Twenty-five STEC strains were isolated from 38.5% of the Zoo's animals. Ten species of order Cetartiodactyla and one species of order Rodentia were recognized as new STEC carriers. STEC strains belonged to 7 different serotypes including new serotypes O12:H25 and O13:H6. Serotype O146:H28, previously associated with human infections, represented 24% of STEC isolates. The most frequent Shiga toxin identified were type 1c and type 2c. Nineteen strains were positive for iha gene, 8 strains were positive for ehxA gene. Moreover, all strains were positive for lpfAO113 and negative for rfbO157, eae, saa, lpfAO157/OI-141, lpfAO157/OI-154, efa1, and toxB genes. Results obtained by XbaI-pulsed-field gel electrophoresis (XbaI-PFGE) confirmed the transmission of STEC strains among different animal species and suborders. In addition, we observed a potential association between STEC-harboring animal and factors such as belonging to order Cetartiodactyla, living in a pit, and belonging to a non-autochthonous species. This is the first work developed with zoological mammals and STEC in Argentina.

Animals↗
PubMed2006

Development of a rapid immunochromatographic test to identify enteropathogenic and enterohemorrhagic Escherichia coli by detecting EspB.

A rapid immunochromatographic (IC) test to identify enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC, respectively) was developed to detect EspB secreted by the type III secretion system of these bacteria. The detection limit of the test system was 4 ng/mL. All 33 of 34 strains harboring the eae gene encoding intimin were positive in the IC test, and all 40 of the eae-negative strains were negative. The results showed that the sensitivity was 96.9% and specificity was 100%. The IC test also detected EspB in a stool sample artificially supplemented with 60 ng EspB/mL. The IC test for the detection of EspB may be a practical method to define EPEC or EHEC both in clinical laboratories and the field.

Adhesins, Bacterial↗
PubMed2006

A Vibrio cholerae protease needed for killing of Caenorhabditis elegans has a role in protection from natural predator grazing.

Vibrio cholerae is the causal bacterium of the diarrheal disease cholera, and its growth and survival are thought to be curtailed by bacteriovorous predators, e.g., ciliates and flagellates. We explored Caenorhabditis elegans as a test organism after finding that V. cholerae can cause lethal infection of this nematode. By reverse genetics we identified an extracellular protease, the previously uncharacterized PrtV protein, as being necessary for killing. The killing effect is associated with the colonization of bacteria within the Caenorhabditis elegans intestine. We also show that PrtV is essential for V. cholerae in the bacterial survival from grazing by the flagellate Cafeteria roenbergensis and the ciliate Tetrahymena pyriformis. The PrtV protein appears to have an indirect role in the interaction of V. cholerae with mammalian host cells as judged from tests with tight monolayers of human intestinal epithelial cells. Our results demonstrate a key role for PrtV in V. cholerae interaction with grazing predators, and we establish Caenorhabditis elegans as a convenient organism for identification of V. cholerae factors involved in host interactions and environmental persistence.

Animals↗
PubMed2005

The long polar fimbriae genes identified in Shiga toxin-producing Escherichia coli are present in other diarrheagenic E. coli and in the standard E. coli collection of reference (ECOR) strains.

Long polar fimbriae (LPF) are related to type I fimbriae in genetic organization and were first identified in Salmonella enterica serovar Typhimurium. Four lpfA genetic variants designated lpfA(O157/OI-141), lpfA(O157/OI-154), lpfA(O26) and lpfA(O113) have been identified in Shiga toxin-producing Escherichia coli (STEC). In this study, PCR was employed to determine the distribution of STEC-lpfAs in enteropathogenic, enteroaggregative, enterotoxigenic and enteroinvasive E. coli (EPEC, EAEC, ETEC and EIEC) and in the standard E. coli collection of reference (ECOR). Among the 97 diarrheagenic strains from our collection, only 2 EPEC strains of serotypes O55:H7 and O119:NM were positive for both lpfA(O157/OI-141) and lpfA(O157/OI-154). lpfA(O157/OI-141) was also positive in 1 of 25 ETEC strains. lpfA(O113) was present in 51 of 97 strains and lpfA(O26) in 13 of 97 strains of diverse diarrheagenic categories. STEC-lpfAs were also present in non-pathogenic ECOR strains of all phylogenetic groups. This study showed that the lpfA genes identified in the genome of STEC strains are not specific to this category. Our results suggest that there is a relationship between the lpfA variant and the phylogenetic group.

Adhesins, Bacterial↗
PubMed2005

Oxalate-degrading Providencia rettgeri isolated from human stools.

BACKGROUND: Oxalate-degrading bacteria are thought to metabolize intestinal oxalate and thus decrease the urinary excretion of oxalate by reducing its intestinal absorption. METHODS: We have isolated several novel oxalate-degrading bacteria from human stools. Oxalate degrading bacteria were investigated to characterize their protein profiles with antibodies against oxalyl-coenzyme A decarboxylase (65 kDa) and formyl-coenzyme A transferase (48 kDa) purified from Oxalobacter formigenes. RESULTS: One of these isolates was identified as Providencia rettgeri, which showed two proteins (65 kDa and 48 kDa) on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) that were not found in non-oxalate-degrading P. rettgeri. Antibodies reacted with the 65 and 48 kDa proteins from the P. rettgeri strain on Western blotting. An Oxalobacter formigenes formyl-coenzyme A transferase gene probe reacted with chromosomal DNA from P. rettgeri on Southern blotting under high stringency conditions, while an Oxalobacter formigenes oxalyl-coenzyme A decarboxylase gene probe did not react under the same conditions. CONCLUSIONS: The mechamism of oxalate degradation by P. rettgeri appears to be similar to that of Oxalobacter formigenes. This is the first report of a facultative oxalate-degrading organism that is one of the Enterobacteriaceae.

Blotting, Southern↗
PubMed2005

The relationship between O-antigens and pathogenic genes of diarrhea-associated Escherichia coli.

To evaluate the serogrouping-based diagnosis of diarrheagenic Escherichia coli, a total of 1,130 strains of E. coli isolated in several countries were studied. The strains were regarded as enterovirulent on the basis of their O-antigens determined using a commercially available kit containing 43 antisera, and the presence of diarrhea-associated genes (eae, stx, aggR, est, elt, ipaH) was evaluated by PCR. Two hundred sixty-three strains of 1,130 (23.3%) were identified as diarrheagenic based on the presence of at least one pathogenic gene. The probability that E. coli identified as diarrheagenic on the basis of serogrouping actually possessed some pathogenic gene was highest for serogroup O119 (78.4%); other serogroups with a positive rate for pathogenic genes higher than 60% were O111 and O126. No target genes were detected among the strains belonging to serogroups O1, O29, O112ac, O143, O158 and O168. Our results suggest that, in practice, serogrouping is useful for the identification of diarrheagenic E. coli in a very limited number of serogroups.

Antigens, Bacterial↗
PubMed2004

Purification and characterization of a novel metalloprotease isolated from Aeromonas caviae.

A novel protease produced by Aeromonas caviae was purified and characterized. The molecular weight of the protease (AP19) was estimated as 19 kDa on SDS-polyacrylamide gel electrophoresis. The protease activity was not inhibited completely by heating at 100 degrees C for 15 min. The proteolytic activities were inhibited by metalloprotease inhibitor. The N-terminal amino acid sequence of AP19 was VTASVSFSGRCTN. AP19 did not activate Aeromonas proaerolysin, and did not show fluid accumulation in the rabbit intestinal loop test. A high concentration of the protease showed cytotoxic activity against Vero cells.

Aeromonas↗
PubMed2004

Aerolysin is activated by metalloprotease in Aeromonas veronii biovar sobria.

Aeromonas veronii is an opportunistic human pathogen that causes diarrhoea and extraintestinal infections, such as wound infection and septicaemia. An A. veronii protease (AVP) from a biovar sobria strain, AeG1, was partially purified and characterized. Mature AVP hydrolysed casein but not elastin, and protease activity was inhibited by metalloprotease inhibitors. Nucleotide sequence analysis showed that AVP belongs to the thermolysin family of proteases. An AVP-deficient mutant was constructed to investigate the role of AVP in aerolysin activation. Western blot analysis using anti-aerolysin antisera revealed that proaerolysin (52 kDa) in the AVP-deficient mutant was not completely activated to mature aerolysin (47 kDa) as seen in the wild-type strain. The AVP-deficient mutant showed lower cytotoxic and haemolytic activities than wild type. AVP and proaerolysin had no haemolytic activity; however, activity appeared after incubating both proteins. Taken together, these results suggested that AVP plays an indirect role in virulence through activating aerolysin, which is an essential step for cytotoxic activity.

Aeromonas↗
PubMed2004

Antibiotic resistance conferred by a class I integron and SXT constin in Vibrio cholerae O1 strains isolated in Laos.

Changes in the drug susceptibility pattern were observed in Vibrio cholerae O1 isolated in the Lao People's Democratic Republic during 1993 to 2000. In this study, 50 V. cholerae O1 strains were selected during this period for studying the presence of class I integron and SXT constin. Twenty-four streptomycin-resistant strains out of 26 isolated before 1997 contained a class I integron harboring the aadA1 gene cassette. Twenty-four strains isolated after 1997 contained an SXT constin (a large conjugative element). Twenty of the strains were resistant to chloramphenicol, tetracycline, streptomycin, and trimethoprim-sulfamethoxazole, while four strains were susceptible to the antibiotic tested. The resistance genes included in the SXT constins were floR, tetA, strAB, and sulII, which encode resistance to chloramphenicol, tetracycline, streptomycin, and sulfamethoxazole, respectively. The antibiotic resistance gene cluster was found to be deleted in the four susceptible strains. SXT(LAOS) did not contain dfrA1 or dfr18, which confer resistance to trimethoprim in SXT(ET) and SXT(MO10), respectively. A hot spot region of SXT(LAOS) was sequenced, and we identified two novel open reading frames showing homology to sO24 (exonuclease) and sO23 (helicase) of the genomic island associated with the multidrug resistance region of Salmonella enterica serovar Typhimurium DT104. Analysis of SXT(LAOS) showed that there is a continuous flux of genes among V. cholerae SXT constins which should be carefully monitored.

Bacterial Proteins↗
PubMed2004

Distribution of putative adhesins in different seropathotypes of Shiga toxin-producing Escherichia coli.

The distribution of eight putative adhesins that are not encoded in the locus for enterocyte effacement (LEE) in 139 Shiga toxin-producing Escherichia coli (STEC) of different serotypes was investigated by PCR. Five of the adhesins (Iha, Efa1, LPF(O157/OI-141), LPF(O157/OI-154), and LPF(O113)) are encoded in regions corresponding to genomic O islands of E. coli EDL933, while the other three adhesins have been reported to be encoded in the STEC megaplasmid of various serotypes (ToxB [O157:H7], Saa [O113:H21], and Sfp [O157:NM]). STEC strains were isolated from humans (n = 54), animals (n = 52), and food (n = 33). They were classified into five seropathotypes (A through E) based on the reported occurrence of STEC serotypes in human disease, in outbreaks, and in the hemolytic-uremic syndrome (M. A. Karmali, M. Mascarenhas, S. Shen, K. Ziebell, S. Johnson, R. Reid-Smith, J. Isaac-Renton, C. Clark, K. Rahn, and J. B. Kaper, J. Clin. Microbiol. 41:4930-4940, 2003). The most prevalent adhesin was that encoded by the iha gene (91%; 127 of 139 strains), which was distributed in all seropathotypes. toxB and efa1 were present mainly in strains of seropathotypes A and B, which were LEE positive. saa was present only in strains of seropathotypes C, D, and E, which were LEE negative. Two fimbrial genes, lpfA(O157/OI-141) and lpfA(O157/OI-154), were strongly associated with seropathotype A. The fimbrial gene lpfA(O113) was present in all seropathotypes except for seropathotype A, while sfpA was not present in any of the strains studied. The distribution of STEC adhesins depends mainly on serotypes and not on the source of isolation. Seropathotype A, which is associated with severe disease and frequently is involved in outbreaks, possesses a unique adhesin profile which is not present in the other seropathotypes. The wide distribution of iha in STEC strains suggested that it could be a candidate for vaccine development.

Adhesins, Escherichia coli↗
PubMed2004

Comparative chracterization of Staphylococcus aureus isolates from throats and noses of healthy volunteers.

Staphylococcus aureus isolates in 2001 from the nose and the throat of an adult population were characterized for their incidence and type. The incidence was 51%, present in 80 out of 157 individuals examined, consisting of 34 nasal carriers, 24 throat carriers, and 22 who carried the isolates in both the nose and throat. Among these isolates, 2 and 5 from the nose and the throat, respectively, were identified as methicillin-resistant S. aureus. S. aureus from the nose and throat of the same individuals were characterized for identification. Examination of their phenotypes revealed that in 11 individuals the clone of S. aureus in the throat was different from the nasal clone. These results suggested that staphylococcal flora in the nose and the throat were independently formed, and that attention should also be directed to the carriers of S. aureus in the throat for the control of nosocomial infection.

Adult↗
PubMed2003

Molecular analysis of VcfQ protein involved in Vibrio cholerae type IV pilus biogenesis.

The nucleotide sequence of an ORF (vcfQ) within the type IV pilus gene cluster of Vibrio cholerae O34 strain NAGV14 was determined, thereby completing the sequence analysis of the structural operon. The vcfQ gene showed homology to the mshQ gene of the mannose-sensitive haemagglutinin pilus gene cluster. The vcfQ was 651 bp larger than mshQ, and the G+C content of the extra 651 bp portion (35.6 mol%) was lower than that of the overall vcfQ gene (42.5 mol%). Except for the first 270 aa residues, the deduced amino acid sequence of VcfQ showed high homology to the MshQ protein. There was immunological cross-reaction between VcfQ and MshQ by Western blotting. Cell fractionation studies showed that VcfQ is located in both the inner and the outer membranes. Mutational analysis showed that vcfQ-deficient mutant expressed detectable levels of major pilin (VcfA), but failed to assemble them into pili, indicating that VcfQ is essential for pilus assembly. Colony-blotting analyses showed that the N-terminal region of vcfQ is variable in V. cholerae strains.

Amino Acid Sequence↗
PubMed2003

Multiplex PCR assay for identification of human diarrheagenic Escherichia coli.

A multiplex PCR assay for the identification of human diarrheagenic Escherichia coli was developed. The targets selected for each category were eae for enteropathogenic E. coli, stx for Shiga toxin-producing E. coli, elt and est for enterotoxigenic E. coli, ipaH for enteroinvasive E. coli, and aggR for enteroaggregative E. coli. This assay allowed the categorization of a diarrheagenic E. coli strain in a single reaction tube.

Diarrhea↗
PubMed2003

The incidence of Escherichia coli having pathogenic genes for diarrhea: a study in the People's Democratic Republic of Lao.

The incidence of Escherichia coli having pathogenic genes for diarrhea was studied in Laos in 2002. A total of 525 E. coli strains from 278 patients (basically, two E. coli isolates from each patient) were examined by PCR to detect the known pathogenic genes (stx, eae, elt, est, ipaH, and aggR). These genes were detected in 23 strains from 16 patients (16/278: 5.8%). In 10 cases of the 16, one of the two isolates from each individual was negative for the gene, and in the other six cases, both isolates had the gene (same gene in four cases). E. coli having eae but no stx (enteropathogenic E. coli [EPEC]) was found in two cases out of 278 (0.7%). Nevertheless, Class I classical EPEC (serogroup-based) was found in 77 cases (28%). Enterotoxigenic E. coli, enteroaggregative E. coli, and enterohemorrhagic E. coli were found in 9, 4, and 1 cases, respectively. Enteroinvasive E. coli was not detected. This study suggested that the incidence of diarrhea due to E. coli is not as high as has been previously thought.

Adolescent↗
PubMed2002

Minor pilin subunits are conserved in Vibrio cholerae type IV pili.

The nucleotide sequences of five open reading frames within the Vibrio cholerae NAGV14 type IV pilus gene cluster were determined. The genes showed high homology to the mannose-sensitive hemagglutinin (MSHA) pilus genes mshB, mshC, mshD, mshO and mshP. PCR analysis showed that a MSHA-like gene cluster is highly conserved among different V. cholerae strains, with the exception of the previously reported major pilin subunit. Recombinant MshB and MshO proteins were purified and specific antiserum was raised to each of them. Western blotting analyses showed that these antisera reacted with purified NAGV14 and MSHA pili. The results suggested that MshB and MshO are minor components of the pilus fiber. Although there was no cross-reaction between the major pilin subunits of NAGV14 and MSHA pili, minor components seemed to be highly homologous and immunologically cross-reactive.

Antigens, Bacterial↗