RESUMEN

Background: Virulence factors (VF) are bacteria-associated molecules that assist to colonize the host at the cellular level. Bacterial virulence is highly dynamic and specific pathogens have a broad array of VFs. The genus Helicobacter is gram-negative, microaerobic, flagellated, and mucus-inhabiting bacteria associated with gastrointestinal inflammation. To investigate about their pathogenicity, several Helicobacter species have been characterized and sequenced. Since the variability and possible origin of VF in the genus are not clear, our goal was to perform a comparative analysis of Helicobacter species in order to investigate VF variability and their evolutionary origin. Methods: The complete genomes of 22 Helicobacter species available in NCBI were analyzed, using computational tools. We identifyed gain and loss events in VF genes, which were categorized in seven functional groups to determine their most parsimonious evolutionary origin. After verifying the annotation of all VF genes, a phylogeny from conserved VF organized by Helicobacter species according to gastric Helicobacter species (GHS) or enterohepatic (EHS) classification was obtained. Results: Gain and loss analysis of VF orthologous in Helicobacter ssp revealed the most possible evolutionary origin for each gene set. Microevolutionary events in urease and flagella genes were detected during the evolution of the genus. Our results pointed that acquisition of ureases and adherence genes and deletion of cytotoxins in some lineages, as well as variation in VF genes copy number, would be related to host adaptation during evolution of the Helicobacter genus. Our findings provided new insights about the genetic differences between GHS and EHS and their relationship with pathogenicity.

Asunto(s)

Helicobacter , Helicobacter/genética , Virulencia/genética , Factores de Virulencia/genética , Filogenia , Genoma , Ureasa/genética

RESUMEN

An unusual biotype of KPC-2-producing Klebsiella pneumoniae (KPC-Kpn) isolates was detected in Corrientes, Argentina, which, to their isolation date, had been free of KPC-Kpn outbreaks. Our aim was to describe the clinical epidemiology focused on genomic characterization of atypical urease-negative KPC-Kpn clinical isolates belonging to the high-risk hospital-associated clonal lineage ST340/CC258. Thirteen isolates were recovered, all of them from inpatients with KPC-Kpn infection (August 2015 to January 2016). These isolates displayed identical enterobacterial repetitive intergenic consensus-PCR electropherotype belonging to a single clonal sequence type ST340. Whole genome sequencing was performed on two KPC-Kpn and the resistome analyses revealed the following acquired resistance genes: blaKPC-2, blaCTX-M-15, blaOXA-1, blaSHV-11, aac(3)-IId, aph(3')-Ia, aac(6')-Ib-cr, sul1, dfrA14, catB3, fosA, and arr-3. Mutations in GyrA (S83I) and ParC (S80I) were also identified. Among the virulence determinants, yersiniabactin was detected in both strains, specifically the ybt9 locus located in ICEKp3. Five plasmid incompatibility groups were observed in this clone and an unusual IncP6 plasmid bearing blaKPC-2 gene (named pKpn3KP) was fully characterized. In this study, we present the first draft genome sequences of two clinical isolates of KPC-2/CTX-M-15-producing K. pneumoniae belonging to the high-risk clonal lineage ST340/CC258 associated with nosocomial outbreaks in Argentina.

Asunto(s)

Infecciones por Klebsiella , Klebsiella pneumoniae , Humanos , Infecciones por Klebsiella/epidemiología , Infecciones por Klebsiella/microbiología , beta-Lactamasas/genética , Ureasa/genética , Antibacterianos/farmacología , Plásmidos/genética , Tipificación de Secuencias Multilocus

RESUMEN

BACKGROUND: Brucella canis is the etiological agent of canine brucellosis, a worldwide neglected zoonosis that constitutes one of the major infectious causes of infertility and reproductive failure in dogs. Although genomic information available for this pathogen has increased in recent years, here we report the first genome sequencing of a B. canis strain in Chile, and the differences in virulence genes with other B. canis strains. RESULTS: Genome assembly produced a total length of 3,289,216 bp, N50 of 95,163 and GC% of 57.27, organized in 54 contigs in chromosome I, and 21 contigs in chromosome II. The genome annotation identified a total of 1981 CDS, 3 rRNA and 36 tRNA in chromosome I, and 1113 CDS and 10 tRNA in chromosome II. There is little variation between the different strains and the SCL isolate. Phylogenetic analysis showed that the Chilean SCL strain is closely related to B. canis and B. suis strains. Small differences were found when compared to the Serbian isolate, but all strains shared the same recent common ancestor. Finally, changes in the sequence of some virulence factors showed that the SCL strain is similar to other South American B. canis strains. CONCLUSIONS: This work sequenced and characterized the complete genome of B. canis strain SCL, evidencing the complete presence of all the genes of the virB operon, and minor changes in outer membrane proteins and in the urease operon. Our data suggest that B. canis was introduced from North America and then spread throughout the South American continent.

Asunto(s)

Animales , Perros , Brucelosis/epidemiología , Brucella canis/genética , Brucella canis/patogenicidad , Ureasa/genética , Brucelosis/transmisión , Zoonosis , Chile , Genoma

RESUMEN

Staphylococcus saprophyticus is an important pathogen responsible for community urinary tract infections (UTI). Besides composing the human microbiota, this species is widely distributed in the environment and the origins of this organism for human infection is not fully characterized. Although some virulence determinants are known, such as d-serine deaminase (DsdA), urease and cell-wall associated proteins, few studies investigated the distribution of virulence-associated genes and analyzed the pathogenic potential of S. saprophyticus strains from different sources. The aim of the present study was to detect the presence of S. saprophyticus genes encoding surface proteins UafA, Aas, Ssp, SdrI, SssF as well as the DsdA and urease enzymes. A total of 142 S. saprophyticus strains were obtained from four sources: UTI, colonization, water and food. It was found, in every tested strain, the presence of genes encoding the surface proteins UafA, Aas, Ssp and SssF and the DsdA and urease enzymes. In contrast, the gene encoding SdrI surface protein was not detected in any of the strains of S. saprophyticus. These results provide a better understanding of the characteristics of S. saprophyticus strains and suggest that isolates from non-human sources have a potential to colonize the urinary tract.

Asunto(s)

Proteínas Bacterianas/genética , Genes Bacterianos/genética , Infecciones Estafilocócicas/microbiología , Staphylococcus saprophyticus/genética , Staphylococcus saprophyticus/aislamiento & purificación , Factores de Virulencia/genética , Brasil , ADN Bacteriano/aislamiento & purificación , Femenino , Humanos , Hidroliasas/genética , Proteínas de la Membrana/genética , Staphylococcus saprophyticus/enzimología , Staphylococcus saprophyticus/patogenicidad , Ureasa/genética , Sistema Urinario/microbiología , Infecciones Urinarias/microbiología , Virulencia/genética

RESUMEN

Whole genome sequencing and analyses of Ureaplasma diversum ATCC 49782 was undertaken as a step towards understanding U. diversum biology and pathogenicity. The complete genome showed 973,501 bp in a single circular chromosome, with 28.2% of G+C content. A total of 782 coding DNA sequences (CDSs), and 6 rRNA and 32 tRNA genes were predicted and annotated. The metabolic pathways are identical to other human ureaplasmas, including the production of ATP via hydrolysis of the urea. Genes related to pathogenicity, such as urease, phospholipase, hemolysin, and a Mycoplasma Ig binding protein (MIB)-Mycoplasma Ig protease (MIP) system were identified. More interestingly, a large number of genes (n = 40) encoding surface molecules were annotated in the genome (lipoproteins, multiple-banded antigen like protein, membrane nuclease lipoprotein and variable surface antigens lipoprotein). In addition, a gene encoding glycosyltransferase was also found. This enzyme has been associated with the production of capsule in mycoplasmas and ureaplasma. We then sought to detect the presence of a capsule in this organism. A polysaccharide capsule from 11 to 17 nm of U. diversum was observed trough electron microscopy and using specific dyes. This structure contained arabinose, xylose, mannose, galactose and glucose. In order to understand the inflammatory response against these surface molecules, we evaluated the response of murine macrophages J774 against viable and non-viable U. diversum. As with viable bacteria, non-viable bacteria were capable of promoting a significant inflammatory response by activation of Toll like receptor 2 (TLR2), indicating that surface molecules are important for the activation of inflammatory response. Furthermore, a cascade of genes related to the inflammasome pathway of macrophages was also up-regulated during infection with viable organisms when compared to non-infected cells. In conclusion, U. diversum has a typical ureaplasma genome and metabolism, and its surface molecules, including the identified capsular material, represent major components of the organism immunopathogenesis.

Asunto(s)

Genoma Bacteriano/genética , Interacciones Huésped-Patógeno/genética , Infecciones por Ureaplasma/genética , Ureaplasma/genética , Composición de Base/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Inflamasomas/genética , Lipoproteínas/genética , Redes y Vías Metabólicas/genética , Anotación de Secuencia Molecular , Mycoplasma/genética , Mycoplasma/patogenicidad , Fosfolipasas/genética , Receptores Toll-Like/genética , Ureaplasma/patogenicidad , Infecciones por Ureaplasma/microbiología , Infecciones por Ureaplasma/patología , Ureasa/genética

RESUMEN

Ureases (EC 3.5.1.5) are Ni(2+) -dependent metalloenzymes produced by plants, fungi and bacteria that hydrolyze urea to produce ammonia and CO2 . The insertion of nickel atoms into the apo-urease is better characterized in bacteria, and requires at least three accessory proteins: UreD, UreF, and UreG. Our group has demonstrated that ureases possess ureolytic activity-independent biological properties that could contribute to the pathogenicity of urease-producing microorganisms. The presence of urease in pathogenic bacteria strongly correlates with pathogenesis in some human diseases. Some medically important fungi also produce urease, including Cryptococcus neoformans and Cryptococcus gattii. C. gattii is an etiological agent of cryptococcosis, most often affecting immunocompetent individuals. The cryptococcal urease might play an important role in pathogenesis. It has been proposed that ammonia produced via urease action might damage the host endothelium, which would enable yeast transmigration towards the central nervous system. To analyze the role of urease as a virulence factor in C. gattii, we constructed knockout mutants for the structural urease-coding gene URE1 and for genes that code the accessory proteins Ure4 and Ure6. All knockout mutants showed reduced multiplication within macrophages. In intranasally infected mice, the ure1Δ (lacking urease protein) and ure4Δ (enzymatically inactive apo-urease) mutants caused reduced blood burdens and a delayed time of death, whereas the ure6Δ (enzymatically inactive apo-urease) mutant showed time and dose dependency with regard to fungal burden. Our results suggest that C. gattii urease plays an important role in virulence, in part possibly through enzyme activity-independent mechanism(s).

Asunto(s)

Criptococosis/microbiología , Cryptococcus gattii/enzimología , Cryptococcus gattii/patogenicidad , Ureasa/metabolismo , Factores de Virulencia/metabolismo , Virulencia , Animales , Southern Blotting , Western Blotting , Células Cultivadas , Criptococosis/metabolismo , Criptococosis/mortalidad , Criptococosis/patología , Femenino , Humanos , Ratones , Ratones Endogámicos BALB C , Fagocitosis , Filogenia , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tasa de Supervivencia , Ureasa/genética , Factores de Virulencia/genética

RESUMEN

Ureases catalyze the hydrolysis of urea into NH3 and CO2. They are synthesized by plants, fungi and bacteria but not by animals. Ureases display biological activities unrelated to their enzymatic activity, i.e., platelet and neutrophil activation, fungus inhibition and insecticidal effect. Urease from Canavalia ensiformis (jack bean) is toxic to several hemipteran and coleopteran insects. Jaburetox is an insecticidal fragment derived from jack bean urease. Among other effects, Jaburetox has been shown to interact with lipid vesicles. In this work, the ion channel activity of C. ensiformis urease, Jaburetox and three deletion mutants of Jaburetox (one lacking the N-terminal region, one lacking the C-terminal region and one missing the central ß-hairpin) were tested on planar lipid bilayers. All proteins formed well resolved, highly cation-selective channels exhibiting two conducting states whose conductance ranges were 7-18pS and 32-79pS, respectively. Urease and the N-terminal mutant of Jaburetox were more active at negative potentials, while the channels of the other peptides did not display voltage-dependence. This is the first direct demonstration of the capacity of C. ensiformis urease and Jaburetox to permeabilize membranes through an ion channel-based mechanism, which may be a crucial step of their diverse biological activities, including host defense.

Asunto(s)

Canavalia/metabolismo , Insecticidas/metabolismo , Canales Iónicos/metabolismo , Membrana Dobles de Lípidos/metabolismo , Péptidos/metabolismo , Proteínas de Plantas/metabolismo , Ureasa/metabolismo , Secuencia de Aminoácidos , Canavalia/química , Canavalia/genética , Permeabilidad de la Membrana Celular , Insecticidas/química , Datos de Secuencia Molecular , Péptidos/química , Péptidos/genética , Proteínas de Plantas/química , Proteínas de Plantas/genética , Eliminación de Secuencia , Ureasa/química , Ureasa/genética

RESUMEN

Ureases are nickel-dependent enzymes which catalyze the hydrolysis of urea to ammonia and carbamate. Despite the apparent wealth of data on ureases, many crucial aspects regarding these enzymes are still unknown, or constitute matter for ongoing debates. One of these is most certainly their structural organization: ureases from plants and fungi have a single unit, while bacterial and archaean ones have three-chained structures. However, the primitive state of these proteins--single- or three-chained--is yet unknown, despite many efforts in the field. Through phylogenetic inference using three different datasets and two different algorithms, we were able to observe chain number transitions displayed in a 3-to-1 fashion. Our results imply that the ancestral state for ureases is the three-chained organization, with single-chained ureases deriving from them. The two-chained variants are not evolutionary intermediates. A fusion process, different from those already studied, may explain this structural transition.

Asunto(s)

Modelos Moleculares , Ureasa/química , Archaea/clasificación , Archaea/enzimología , Bacterias/clasificación , Bacterias/enzimología , Hongos/clasificación , Hongos/enzimología , Filogenia , Plantas/clasificación , Plantas/enzimología , Estructura Terciaria de Proteína , Ureasa/genética

RESUMEN

Ureases, nickel-dependent enzymes that catalyze the hydrolysis of urea into ammonia and bicarbonate, are widespread in plants, bacteria, and fungi. Previously, we cloned a cDNA encoding a Canavalia ensiformis urease isoform named JBURE-II, corresponding to a putative smaller urease protein (78kDa) when compared to other plant ureases. Aiming to produce the recombinant protein, we obtained jbure-IIb, with different 3' and 5' ends, encoding a 90kDa urease. Three peptides unique to the JBURE-II/-IIb protein were detected by mass spectrometry in seed extracts, indicating that jbure-II/-IIb is a functional gene. Comparative modeling indicates that JBURE-IIb urease has an overall shape almost identical to C. ensiformis major urease JBURE-I with all residues critical for urease activity. The cDNA was cloned into the pET101 vector and the recombinant protein was produced in Escherichia coli. The JBURE-IIb protein, although enzymatically inactive presumably due to the absence of Ni atoms in its active site, impaired the growth of a phytopathogenic fungus and showed entomotoxic properties, inhibiting diuresis of Rhodnius prolixus isolated Malpighian tubules, in concentrations similar to those reported for JBURE-I and canatoxin. The antifungal and entomotoxic properties of the recombinant JBURE-IIb apourease are consistent with a protective role of ureases in plants.

Asunto(s)

Canavalia/enzimología , Canavalia/genética , Ureasa/genética , Secuencia de Aminoácidos , Antifúngicos/química , Antifúngicos/aislamiento & purificación , Antifúngicos/metabolismo , Antifúngicos/farmacología , Secuencia de Bases , Canavalia/química , Clonación Molecular , Isoenzimas/genética , Isoenzimas/aislamiento & purificación , Pruebas de Sensibilidad Microbiana , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/aislamiento & purificación , Proteínas de Plantas/metabolismo , Proteínas de Plantas/farmacología , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacología , Homología de Secuencia de Ácido Nucleico , Ureasa/aislamiento & purificación , Ureasa/metabolismo , Ureasa/farmacología

RESUMEN

A novel reverse primer (GLM MR1) was designed for detection of the glmM gene in Helicobacter pylori by PCR. The percentage of amplification in clinical isolates using GLM MR1 was 100% for detection of the glmM gene and 86.36% for the ureA gene. The primer designed is useful for the identification of H. pylori.

Asunto(s)

Técnicas Bacteriológicas/métodos , Cartilla de ADN/genética , Infecciones por Helicobacter/diagnóstico , Helicobacter pylori/enzimología , Fosfoglucomutasa/genética , Reacción en Cadena de la Polimerasa/métodos , Adolescente , Niño , Preescolar , ADN Bacteriano/genética , Helicobacter pylori/genética , Humanos , Lactante , Sensibilidad y Especificidad , Ureasa/genética

RESUMEN

Jaburetox-2Ec, a recombinant peptide derived from an urease isoform (JBURE-II), displays high insecticidal activity against important pests such as Spodoptera frugiperda and Dysdercus peruvianus. Although the molecular mechanism of action of ureases-derived peptides remains unclear, previous ab initio data suggest the presence of structural motifs in Jaburetox-2Ec with characteristics similar to those found in a class of pore-forming peptides. Here, we investigated the molecular aspects of the interaction between Jaburetox-2Ec and large unilamellar vesicles. Jaburetox-2Ec displays membrane-disruptive ability on acidic lipid bilayers and this effect is greatly influenced by peptide aggregation. Corroborating with this finding, molecular modeling studies revealed that Jaburetox-2Ec might adopt a well-defined beta-hairpin conformation similar to those found in antimicrobial peptides with membrane disruption properties. In addition, molecular dynamics simulations suggest that the protein is able to anchor at a polar/non-polar interface. In the light of these findings, for the first time it was possible to point out some evidence that the peptide Jaburetox-2Ec interacting with lipid vesicles promotes membrane permeabilization.

Asunto(s)

Insecticidas/química , Insecticidas/farmacología , Ureasa/química , Ureasa/farmacología , Secuencia de Aminoácidos , Animales , Canavalia/enzimología , Canavalia/genética , Heterópteros , Membrana Dobles de Lípidos , Modelos Moleculares , Datos de Secuencia Molecular , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/farmacología , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacología , Homología de Secuencia de Aminoácido , Spodoptera , Liposomas Unilamelares , Ureasa/genética

RESUMEN

The aim of the present work is to identify the presence of Helicobacter pylori bacterium in samples of gastric mucosa fragments, obtained by gastric biopsy, from Brazilian patients with peptic ulcer and chronic gastritis and also to determine differences among the prevalent strains in these two diseases by urease C and urease B genes amplification utilizing nested polymerase chain reaction (PCR) and PCR. We encountered 17 genotyping patterns for urease C and 7 for urease B and, although no significant differences were found among the patterns encountered for both diseases, we found predominant groups for each disease. Typing methods of the products obtained by nested PCR and PCR show a functional scheme and are of great importance for epidemiologic studies and H. pylori strain characterization, in addition to allowing correlation among the several strains and their role in the diseases caused by this microorganism.

Asunto(s)

Mucosa Gástrica/microbiología , Gastritis/microbiología , Infecciones por Helicobacter/complicaciones , Helicobacter pylori/genética , Úlcera Péptica/microbiología , Polimorfismo de Longitud del Fragmento de Restricción , Adulto , Proteínas Bacterianas , Técnicas de Tipificación Bacteriana , Brasil , Proteínas Portadoras , Enfermedad Crónica , Electroforesis en Gel de Agar , Femenino , Genes Bacterianos , Genotipo , Helicobacter pylori/clasificación , Helicobacter pylori/aislamiento & purificación , Humanos , Masculino , Persona de Mediana Edad , Ureasa/genética

RESUMEN

BACKGROUND: Recent studies showed that Helicobacter pylori existed in the New World prior to the arrival of Columbus. The purpose of the present study was to detect the presence of Helicobacter pylori in pre-Columbian mummies from Northern Mexico. METHODS: Six samples were studied (four samples of gastric remains, tongue-soft palate, and brain remained as negative controls) from two of the six naturally mummified corpses studied (adult male and infant male). Samples were taken from tissues suitable for DNA amplification by Polymerase chain reaction (PCR). DNA was extracted and H. pylori detection was carried out by PCR and hybridized with the pHp probe from 16S rRNA gene. The purified PCR products were cloned and sequenced in both directions. DNA sequences were analyzed with ALIGN and BLAST software. A second amplification was performed using ureB gene by real-time PCR. RESULTS: From four samples of gastric remnant, only two were H. pylori-positive for amplification of a 109 bp DNA fragment; the remaining two were negative, as were the tongue-soft palate and the brain biopsies as well. These PCR products were hybridized with a pHp probe. Nucleotide sequence analysis showed homology with H. pylori in 98 of 99% when compared with the gene bank nucleotide sequence. Only one sample of gastric remnant H. pylori-positive with 16S rRNA gene was also positive for ureB gene from H. pylori. CONCLUSION: This data supported infection with H. pylori in Mexican pre-Columbian mummies dating from approximately 1,350 AC.

Asunto(s)

Infecciones por Helicobacter/historia , Helicobacter pylori/genética , Helicobacter pylori/aislamiento & purificación , Momias/microbiología , Adulto , Antropología , ADN Bacteriano/análisis , ADN Bacteriano/aislamiento & purificación , Infecciones por Helicobacter/microbiología , Historia Medieval , Humanos , Recién Nacido , Masculino , México , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S/genética , Estómago/microbiología , Ureasa/genética

RESUMEN

Canatoxin, a urease isoform from Canavalia ensiformis seeds, shows insecticidal activity against different insect species. Its toxicity relies on an internal 10 kDa peptide (pepcanatox), released by hydrolysis of Canatoxin by cathepsins in the digestive system of susceptible insects. In the present work, based on the N-terminal sequence of pepcanatox, we have designed primers to amplify by PCR a 270-bp fragment corresponding to pepcanatox using JBURE-II cDNA (one of the urease isoforms cloned from C. ensiformis, with high identity to JBURE-I, the classical urease) as a template. This amplicon named jaburetox-2 was cloned into pET 101 vector to obtain heterologous expression in Escherichia coli of the recombinant protein in C-terminal fusion with V-5 epitope and 6-His tag. Jaburetox-2Ec was purified on Nickel-NTA resin and bioassayed in insect models. Dysdercus peruvianus larvae were fed on cotton seed meal diets containing 0.01% (w/w) Jaburetox-2Ec and, after 11 days, all individuals were dead. Jaburetox-2Ec was also tested against Spodoptera frugiperda larvae and caused 100% mortality. In contrast, high doses of Jaburetox-2Ec were innocuous when injected or ingested by mice and neonate rats. Modeling of Jaburetox-2Ec, in comparison with other peptide structures, revealed a prominent beta-hairpin motif consistent with an insecticidal activity based on either neurotoxicity or cell permeation.

Asunto(s)

Canavalia/enzimología , Insecticidas/aislamiento & purificación , Ureasa/química , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Cartilla de ADN , Electroforesis en Gel de Poliacrilamida , Insecticidas/química , Insecticidas/toxicidad , Datos de Secuencia Molecular , Proteínas de Plantas , Reacción en Cadena de la Polimerasa , Homología de Secuencia de Aminoácido , Ureasa/genética , Ureasa/aislamiento & purificación , Ureasa/toxicidad

RESUMEN

Canavalia ensiformis (jackbean) seeds contain the proteins urease and canatoxin, a variant form of the jackbean urease. Here we have cloned a cDNA encoding another isoform of urease, called JBURE-II. This cDNA was obtained by RT-PCR using as template total RNA extracted from C. ensiformis tissues. Nucleotide sequence analysis showed that JBURE-II clones share 86% similarity with known jackbean urease. The presence in C. ensiformis of a family of urease-related genes with at least three members was demonstrated by Southern blot analysis. In order to understand the pattern of expression of the JBURE-II gene, we collected tissue samples from different stages of flower and embryo development. The results of RT-PCR show that JBURE-II is expressed from flower buds throughout seed maturation. Semi-quantitative RT-PCR indicates that expression of urease and JBURE-II genes is induced in seedlings and in leaves treated with abscisic acid, a phytohormone involved in seed maturation and wound response. This work constitutes the first report on the presence of a family of urease genes in jackbean, and provides characterization of a cDNA encoding a new member of this gene family.

Asunto(s)

Canavalia/genética , Proteínas de Plantas/genética , Ureasa/genética , Ácido Abscísico/farmacología , Secuencia de Aminoácidos , Secuencia de Bases , Canavalia/enzimología , Clonación Molecular , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Hibridación Genética , Datos de Secuencia Molecular , Familia de Multigenes/genética , Hojas de la Planta/efectos de los fármacos , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Proteínas de Plantas/metabolismo , ARN Mensajero/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Semillas/efectos de los fármacos , Semillas/genética , Semillas/metabolismo , Homología de Secuencia de Aminoácido , Ureasa/metabolismo

RESUMEN

The proteobacterial genus Helicobacter is composed of gastric species, all of them urease-positive, and enteric species (gastrointestinal, intestinal, hepatic, biliary), some of them urease-positive, others not. Here, we point out that the gastric species are divided in at least two phylogenetic groups, one is homogeneous, clearly separated from the enteric species, and another is forming a tight cluster within the enteric species. This feature is apparent in the phylogeny of the genus as inferred from both the 16S rRNA gene and the alpha-subunit of the urease. Our observation shows that the ability to colonize the gastric mucosa appeared more than once in the history of the genus, and suggests that acquiring this ability may be a relatively simple and punctual process, involving a limited number of genes. Such a process may be the lateral transfer acquisition of a functional copy of the gene ureI which encodes a urea channel activated at acidic pH that is essential for gastric colonization by Helicobacter pylori.

Asunto(s)

Helicobacter/genética , Estómago/microbiología , Evolución Biológica , Helicobacter/patogenicidad , Humanos , Filogenia , ARN Ribosómico 16S , Ureasa/genética

RESUMEN

Molecular methods for detection of Helicobacter pylori infection have been shown to be highly sensitive in gastric biopsies and cultures. The objective of this work was to compare PCR detection of H. pylori DNA in string-absorbed gastric juice and in gastric biopsies. The study was performed in 47 dyspeptic adult patients undergoing endoscopy, and infection was detected by amplification of a segment of H. pylori ureA gene. Of the 29 patients positive in biopsy analysis, 23 (79%) were also positive in the gastric string. PCR analysis of gastric strings is a sensitive and safe procedure to detect H. pylori when endoscopy is not indicated, and may be of great clinical and epidemiological usefulness in determining effectiveness of eradication therapies, typing virulence genes and detecting antibiotic resistance mutations.

Asunto(s)

Jugo Gástrico/microbiología , Infecciones por Helicobacter/diagnóstico , Helicobacter pylori/aislamiento & purificación , Reacción en Cadena de la Polimerasa , Adulto , Biopsia , Helicobacter pylori/genética , Humanos , Antro Pilórico/microbiología , Sensibilidad y Especificidad , Manejo de Especímenes , Ureasa/genética