Ana Coelho | UFRJ - Academia.edu (original) (raw)

Papers by Ana Coelho

Memórias do Instituto Oswaldo Cruz, 1998

The pathogenic O1 Amazonia variant of Vibrio cholerae has been shown previously to have a cytotox... more The pathogenic O1 Amazonia variant of Vibrio cholerae has been shown previously to have a cytotoxin acting on cultured Vero and Y-1 cells, and to lack important virulence factors such as the cholera toxin (Coelho et al. 1995a). This study extends the molecular analysis of the Amazonia strains, detecting the presence of the toxR gene, with a very similar sequence to that of the El Tor and classical biotypes. The outer membrane proteins are analyzed, detecting a variation among the group of Amazonia strains, with three different patterns found. As a by-product of this work a polymerase chain reaction fragment was sequenced, reading part of the sequence of the Lon protease of the Amazonia strains. This gene was not previously described in V. cholerae, but its sequence is present in the TIGR database specific for this species.

PROTEOMICS, 2004

A proteome reference map has been constructed for Vibrio cholerae El Tor, in the pI range of 4.0 ... more A proteome reference map has been constructed for Vibrio cholerae El Tor, in the pI range of 4.0 to 7.0. The map is based on two-dimensional gels (2-D) and the identification, by peptide mass fingerprint, of proteins in 94 spots, corresponding to 80 abundant proteins. Two strains are compared, strain N16961 and a Latin American El Tor strain C3294. The consensus map contains 340 spots consistently seen with both strains grown in Luria-Bertani broth (LB) or minimal M9 medium. The results were obtained from nine gels run with 18 cm immobilized pH gradient strips and precast gels. The 2-D gels were anchored to real N16961 proteins identified by mass spectrometry. Various energy metabolism components and periplasmic ATP-binding cassette (ABC) transporter proteins were identified among the abundant proteins. Two isoforms of OmpU were found. Five operons are proposed and seven hypothetical proteins were experimentally confirmed. Comparisons are made with protein 2-D gels for a classical strain and to microarray analysis available for the N16961 El Tor strain. New results were obtained from the proteome analysis, indicating an abundance of periplasmic ABC transporter proteins not found in microarray studies.

PROTEOMICS, 2008

This is the first broad proteomic description of Gluconacetobacter diazotrophicus, an endophytic ... more This is the first broad proteomic description of Gluconacetobacter diazotrophicus, an endophytic bacterium, responsible for the major fraction of the atmospheric nitrogen fixed in sugarcane in tropical regions. Proteomic coverage of G. diazotrophicus PAL5 was obtained by two independent approaches: 2-DE followed by MALDI-TOF or TOF-TOF MS and 1-DE followed by chromatography in a C18 column online coupled to an ESI-Q-TOF or ESI-IT mass spectrometer. The 583 identified proteins were sorted into functional categories and used to describe potential metabolic pathways for nucleotides, amino acids, carbohydrates, lipids, cofactors and energy production, according to the Enzyme Commission of Enzyme Nomenclature (EC) and Kyoto Encyclopedia of genes and genomes (KEGG) databases. The identification of such proteins and their possible insertion in conserved biochemical routes will allow comparisons between G. diazotrophicus and other bacterial species. Furthermore, the 88 proteins classified as conserved unknown or unknown constitute a potential target for functional genomic studies, aiming at the understanding of protein function and regulation of gene expression. The knowledge of metabolic fundamentals and coordination of these actions are crucial for the rational, safe and sustainable interference on crops. The entire dataset, including peptide sequence information, is available as Supporting Information and is the major contribution of this work.

Photochemistry and Photobiology, 1996

The results of this work show that the resistance of Escherichk coli cells to the photodynamic ac... more The results of this work show that the resistance of Escherichk coli cells to the photodynamic action of methylene blue is increased by the addition of glucose to the media in which they are grown. It is postulated that the increased resistance may be due to lowered retention of the dye by cells grown in the presence of glucose, leading to the diminution in DNA damage revealed in the alkaline sucrose gradients. The role of cyclic adenosine-monophosphate in the protective action of glucose is discussed.

Oral Surgery, Oral Medicine, Oral Pathology, Oral Radiology, and Endodontology, 2001

The "red complex," composed of Bacteroides forsythus, Porphyromonas gingivalis,... more The "red complex," composed of Bacteroides forsythus, Porphyromonas gingivalis, and Treponema denticola, is implicated in severe forms of periodontal diseases. The purpose of this study was to assess the occurrence of the red complex in root canal infections through the use of a sensitive technique-the 16S rDNA-directed polymerase chain reaction (PCR). Samples were obtained from 50 necrotic pulps with periradicular pathosis. Ten cases were diagnosed as acute periradicular abscesses. DNA was extracted from the samples and analyzed with a PCR-based identification assay. At least 1 member of the red complex was found in 33 of 50 cases. T denticola, P gingivalis, and B forsythus were detected in 44%, 30%, and 26% of the cases, respectively. The red complex was found in 4 of 50 cases. No particular signs or symptoms were associated with the presence of these bacterial species. Despite what is indicated in reports with respect to marginal periodontitis, red complex bacteria-either singularly or collectively-was not associated with any particular pattern of clinical symptoms. However, because the bacterial species from the red complex are recognized oral pathogens, their occurrence in root canal infections suggests that they may play a role in the pathogenesis of periradicular diseases.

Memórias do Instituto Oswaldo Cruz, 1994

Memórias do Instituto Oswaldo Cruz, 2005

The neuraminidase gene, nanH, is present in the O1, non-toxigenic Vibrio cholerae Amazonia strain... more The neuraminidase gene, nanH, is present in the O1, non-toxigenic Vibrio cholerae Amazonia strain. Its location has been assigned to a 150 kb NotI DNA fragment, with the use of pulsed-field gel electrophoresis and DNA hybridization. This NotI fragment is positioned inside 630 kb SfiI and 1900 kb I-CeuI fragments of chromosome 1. Association of the pathogenicity island VPI-2, carrying nanH and other genes, with toxigenic strains has been described by other authors. The presence of nanH in a non-toxigenic strain is an exception to this rule. The Amazonia strain nanH was sequenced (Genbank accession No. AY825932) and compared to available V. cholerae sequences. The sequence is different from those of pandemic strains, with 72 nucleotide substitutions. This is the first description of an O1 strain with a different nanH allele. The most variable domain of the Amazonia NanH is the second lectin wing, comprising 13 out of 17 amino acid substitutions. Based on the presence of nanH in the same region of the genome, and similarity of the adjacent sequences to VPI-2 sequences, it is proposed that the pathogenicity island VPI-2 is present in this strain.

Journal of Applied Microbiology, 2009

Aims: We performed the first characterization of the microbiota associated with the reef coral Mu... more Aims: We performed the first characterization of the microbiota associated with the reef coral Mussismilia braziliensis by means of a culture-independent approach. Methods and Results: The main groups were Proteobacteria, Cyanobacteria and unclassified bacteria according to the 16S rDNA libraries. Most of the sequences of the mucus of healthy and diseased M. braziliensis did not find close matches in GenBank (i.e. >97% 16S rDNA similarity). Most of the sequences of seawater and mucus of healthy coral fell into tight clusters (17 and 15 clusters respectively). In contrast, most of the sequences of mucus of diseased coral did not form clusters. The rarefaction curves indicate saturation in the recovery of higher taxa (approximately 40 phyla). However, the number of species in the coral mucus (n = 130-170) and seawater (n = 170) did not reach a plateau. Conclusions: The coral microbiota encompasses several potentially novel species and higher taxa. The microbiota of M. braziliensis appears to be species-specific. Diseased coral may have provided a suitable place for colonization by opportunistic bacteria, resulting in a greater bacterial diversity. Significance and Impact of the Study: The first study on the diversity of the microbiota of the endemic and endangered of extinction coral M. braziliensis.

FEMS Microbiology Letters, 1981

FEMS Microbiology Letters, 1999

Polymerase chain reaction has been used to detect the presence of the virulence associated gene, ... more Polymerase chain reaction has been used to detect the presence of the virulence associated gene, tcpA and part of the promoter distal region of the toxin-co-regulated pilus cluster in non-O1, non-toxigenic, Vibrio cholerae. The amplified regions were characterised by restriction fragment length polymorphism and heteroduplex motility assay. We describe the nucleotide sequence of the tcpA gene fragment from non-toxigenic vibrios from clinical and environmental sources. The present study shows that there are at least three types of the tcpA gene among V. cholerae and the primers specific for the classical tcpA gene, amplify all biotypes. A sequence similarity in other regions of the toxin-co-regulated pilus cluster is suggested. The evidences for the presence of this cluster among non-toxigenic vibrios is, to our knowledge, reported for the first time. The use of restriction fragment length polymorphism for typing the tcpA and studying the alleles distribution is proposed.

Cellular Microbiology, 2002

Several strains of Vibrio cholerae secrete a haemolytic toxin of 63 kDa, termed V. cholerae cytol... more Several strains of Vibrio cholerae secrete a haemolytic toxin of 63 kDa, termed V. cholerae cytolysin (VCC). This toxin causes extensive vacuolation and death of cells in culture and forms an anion-selective channel in planar lipid bilayers and in cells. Here, we identify inhibitors of the VCC anion channel and show that the formation of the anion channel is necessary for the development of the vacuoles and for the cell death induced by this toxin. Using markers of cell organelles, we show that vacuoles derive from different intracellular compartments and we identify the contribution of late endosomes and of the trans-Golgi network in vacuole biogenesis.

BMC Genomics, 2009

Background Gluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that liv... more Background Gluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that lives in association with sugarcane plants. It has important biotechnological features such as nitrogen fixation, plant growth promotion, sugar metabolism pathways, secretion of organic acids, synthesis of auxin and the occurrence of bacteriocins. Results Gluconacetobacter diazotrophicus Pal5 is the third diazotrophic endophytic bacterium to be completely sequenced. Its genome is composed of a 3.9 Mb chromosome and 2 plasmids of 16.6 and 38.8 kb, respectively. We annotated 3,938 coding sequences which reveal several characteristics related to the endophytic lifestyle such as nitrogen fixation, plant growth promotion, sugar metabolism, transport systems, synthesis of auxin and the occurrence of bacteriocins. Genomic analysis identified a core component of 894 genes shared with phylogenetically related bacteria. Gene clusters for gum-like polysaccharide biosynthesis, tad pilus, quorum sensing...

Memórias do Instituto …, 2005

Memórias do Instituto …, 1998

BMC …, 2009

Background: Gluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that li... more Background: Gluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that lives in association with sugarcane plants. It has important biotechnological features such as nitrogen fixation, plant growth promotion, sugar metabolism pathways, secretion of organic acids, synthesis of auxin and the occurrence of bacteriocins. Results: Gluconacetobacter diazotrophicus Pal5 is the third diazotrophic endophytic bacterium to be completely sequenced. Its genome is composed of a 3.9 Mb chromosome and 2 plasmids of 16.6 and 38.8 kb, respectively. We annotated 3,938 coding sequences which reveal several characteristics related to the endophytic lifestyle such as nitrogen fixation, plant growth promotion, sugar metabolism, transport systems, synthesis of auxin and the occurrence of bacteriocins. Genomic analysis identified a core component of 894 genes shared with phylogenetically related bacteria. Gene clusters for gum-like polysaccharide biosynthesis, tad pilus, quorum sensing, for modulation of plant growth by indole acetic acid and mechanisms involved in tolerance to acidic conditions were identified and may be related to the sugarcane endophytic and plant-growth promoting traits of G. diazotrophicus. An accessory component of at least 851 genes distributed in genome islands was identified, and was most likely acquired by horizontal gene transfer. This portion of the genome has likely contributed to adaptation to the plant habitat. Conclusion: The genome data offer an important resource of information that can be used to manipulate plant/bacterium interactions with the aim of improving sugarcane crop production and other biotechnological applications.

Infection and …, 2000

A Vibrio cholerae cytotoxin, designated VcVac, was found to cause vacuolation in Vero cells. It w... more A Vibrio cholerae cytotoxin, designated VcVac, was found to cause vacuolation in Vero cells. It was originally detected in the pathogenic O1 Amazonia variant of V. cholerae and later shown to be produced in environmental strains and some El Tor strains. Comparison of VcVac ...

Journal of Clinical Microbiology, 1995

A survey of pathogenic Vibrio cholerae O1 strains from the north of Brazil by using arbitrarily p... more A survey of pathogenic Vibrio cholerae O1 strains from the north of Brazil by using arbitrarily primed PCR fingerprints revealed a group of strains with similar fingerprint patterns that are distinct from those of the current El Tor epidemic strain. These strains have been analyzed by in vivo and in vitro techniques and the group has been denominated the Amazonia variant of V. cholerae O1. Vibrio cholerae is causing a severe epidemic in Latin America after being absent from the continent for about 100 years (30). The taxonomy of this species has been the object of our interest, and we recently developed a method for distinguishing pathogenic groups by using arbitrarily primed PCR (AP-PCR) fingerprints (5, 6) on the basis of the general methodology of AP-PCR (31, 32). By this technique a single oligonucleotide with an arbitrary sequence is used in a PCR with the DNA of the strain under analysis. Low-stringency conditions for hybridization are used, and the oligonucleotide can find regions of pairing, leading to the amplification of various genome fragments. Our original study (6) involved four groups of pathogenic V. cholerae: classical, El Tor, Gulf of Mexico endemic El Tor, and Bengal, all of which are distinguishable with the fingerprints. When applied to strains causing the Latin American epidemics the results were similar to those for other Old World El Tor strains. However, when this method was used to study a group of strains from patients with diarrheal disease in the westernmost part of the Brazilian Amazon, a quite distinct fingerprint pattern emerged for some of these strains. In the work described here we further extended this observation by using other in vitro and in vivo techniques to evaluate the degree of relatedness between this group and the other pathogenic strains. MATERIALS AND METHODS Strains of V. cholerae. The strains used in the present study are listed in Table 1. AP-PCR. The AP-PCR mixtures consisted of 10 mM Tris-HCl (pH 8.3), 50 mM KCl, 4 mM MgCl 2 , 100 M (each) deoxynucleoside triphosphate (dNTP), 30 pmol of one of the oligonucleotides, and 100 ng of DNA in a total volume of 25 l. The mixture was overlaid with oil, and 1.5 U of Taq DNA polymerase was added. The program consisted of 45 cycles, and an annealing temperature of 32ЊC was used (6). Two sets of fingerprints were done, one of them with oligonucleotide 1 (5Ј-GGTGCGGGAA) and the other with oligonucleotide 3 (5Ј-CCAGATGCAC) (6). Analysis of the amplified fragments was done on 1.4% agarose gels (GIBCO-Bethesda Research Laboratories) in Tris-borate buffer (TBE) (27) running at 130 V for 2.5 h. Multilocus enzyme electrophoresis. The multilocus enzyme electrophoresis method used in the present study was the same as that described previously (24). Ribotyping. Chromosomal DNA was prepared from 5 ml of an overnight culture in alkaline peptone water (6, 28). A total of 15 g of chromosomal DNA from each of the strains was cut with 30 U of BglI for a period of 8 h. DNA fragments were separated in 0.7%, 22-cm agarose gels in TBE at 50 V for 15 h.