RESUMEN
The development of novel nanoenergetic materials with enhanced bulk properties requires an understanding of the intermolecular interactions occurring between molecular components. We investigate the surface interactions between 1,3,5-trinitro-1,3,5-triazacyclohexane (RDX) and polyethylene (PE) crystals on the basis of combined use of molecular dynamics (MD) simulations and force-distance spectroscopy, in conjunction with Lifshitz macroscopic theory of van der Waals forces between continuous materials. The binding energy in the RDX-PE system depends both on the degree of PE crystallinity and on the RDX crystal face. Our MD simulations yield binding energies of approximately 132 and 120 mJ/m(2) for 100% amorphous and 100% crystalline PE on RDX (210), respectively. The average value is about 36% greater than our experimental value of 81 ± 15 mJ/m(2) for PE (â¼48% amorphous) on RDX (210). By comparison, Liftshitz theory predicts a value of about 79 mJ/m(2) for PE interacting with RDX. Our MD simulations also predict larger binding energies for both amorphous and crystalline PE on RDX (210) compared to the RDX (001) surface. Analysis of the interaction potential indicates that about 60% of the binding energy in the PE-RDX system is due to attractive interactions between HPE-ORDX and CPE-NRDX pairs of atoms. Further, amorphous PE shows a much longer interaction distance than crystalline PE with the (210) and (001) RDX surfaces due to the possibility of larger polymer elongations in the case of amorphous PE as strain is applied. Also, we report estimates of the binding energies of energetic materials RDX and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) with PE, propylene, polystyrene, and several fluorine-containing polymers using Lifshitz theory and compare these with reported MD calculations.
RESUMEN
PURPOSE: Gram-positive (GP) organisms are among the most common cause of infections in early postsurgical and immunocompromised populations. Patients recovering from lung transplantation (LT) are particularly susceptible owing to the physiologic stress imposed by surgery and induction with intense immunosuppression. Sites, types, and timing of GP infections following LT are not well documented. This report describes the clinical spectrum of GP infections and their effects on surgical airway complications (SAC) and bronchiolitis obliterans syndrome (BOS) following LT. METHODS AND MATERIALS: Data were collected from 202 patients undergoing 208 LT procedures at a single institution between November 1990 and November 2005. Data were retrospectively analyzed according to timing, location, and causative pathogen. RESULTS: In the median follow-up period of 2.7 years (range, 0-13.6 years), 137 GP infections were confirmed in 72 patients. Sites of infection included respiratory tract (42%), blood (27%), skin, wound and catheter (21%), and other (10%). GP pathogens identified were Staphylococcus species (77%), Enterococcus species (12%), Streptococcus species (6%), Pneumococcus (4%), and Eubacterium lentum (1%). The likelihood of SAC and BOS was increased in lung allograft recipients with GP pneumonia as compared with those without (hazard ratio 2.1; 95% confidence interval=1.5-3.1). CONCLUSIONS: GP organisms were responsible for infections in 40% of lung allograft recipients and most commonly isolated from the respiratory tract and blood stream. Staphylococcal species were most frequently identified, 42% of which were methicillin-resistant Staphylococcus aureus (MRSA). Given the strong association of respiratory tract infections with the development of SAC and BOS, empiric antimicrobial strategies after LT should include agents directed against GP organisms, especially MRSA.
Asunto(s)
Bronquiolitis Obliterante , Bacterias Grampositivas , Infecciones por Bacterias Grampositivas/fisiopatología , Trasplante de Pulmón/efectos adversos , Infección de la Herida Quirúrgica , Adolescente , Adulto , Anciano , Bacteriemia/microbiología , Bronquiolitis Obliterante/microbiología , Bronquiolitis Obliterante/fisiopatología , Niño , Femenino , Bacterias Grampositivas/clasificación , Bacterias Grampositivas/aislamiento & purificación , Bacterias Grampositivas/patogenicidad , Infecciones por Bacterias Grampositivas/microbiología , Humanos , Masculino , Staphylococcus aureus Resistente a Meticilina/aislamiento & purificación , Persona de Mediana Edad , Infecciones del Sistema Respiratorio/microbiología , Infecciones del Sistema Respiratorio/fisiopatología , Infecciones Estafilocócicas/microbiología , Infecciones Estafilocócicas/fisiopatología , Staphylococcus/clasificación , Staphylococcus/aislamiento & purificación , Infección de la Herida Quirúrgica/microbiología , Infección de la Herida Quirúrgica/fisiopatología , Síndrome , Adulto JovenRESUMEN
PURPOSE: Clostridium difficile colitis (CDC) is the most common nosocomial infection of the gastrointestinal tract in patients with recent antibiotic use or hospitalization. Lung transplant recipients receive aggressive antimicrobial therapy postoperatively for treatment and prophylaxis of respiratory infections. This report describes the epidemiology of CDC in lung recipients from a single center and explores possible associations with bronchiolitis obliterans syndrome (BOS), a surrogate marker of chronic rejection. METHODS: Patients were divided into those with confirmed disease (CDC+) and those without disease (CDC-) based on positive C. difficile toxin assay. Because of a bimodal distribution in the time to develop CDC, the early postoperative CDC+ group was analyzed separately from the late postoperative CDC+ cohort with respect to BOS development. RESULTS: Between 1990 and 2005, 202 consecutive patients underwent 208 lung transplantation procedures. Of these, 15 lung recipients developed 23 episodes of CDC with a median follow-up period of 2.7 years (range, 0-13.6). All patients with confirmed disease had at least 1 of the following 3 risk factors: recent antibiotic use, recent hospitalization, or augmentation of steroid dosage. Of the early CDC+ patients, 100% developed BOS, but only 52% of the late CDC+ patients developed BOS, either preceding or following infection. CONCLUSION: CDC developed in 7.4% of lung transplant patients with identified risk factors, yielding a cumulative incidence of 14.7%. The statistical association of BOS development in early CDC+ patients suggests a relationship between early infections and future chronic lung rejection.
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Clostridioides difficile , Enterocolitis Seudomembranosa/epidemiología , Trasplante de Pulmón/efectos adversos , Adolescente , Adulto , Anciano , Bronquiolitis Obliterante/epidemiología , Bronquiolitis Obliterante/etiología , Niño , Enterocolitis Seudomembranosa/microbiología , Femenino , Rechazo de Injerto , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Factores de RiesgoRESUMEN
Salmonella typhi, the causative agent of typhoid fever, annually infects 16 million people and kills 600 000 world wide. Plasmid-encoded multiple drug resistance in S. typhi is always encoded by plasmids of incompatibility group H (IncH). The complete DNA sequence of the large temperature-sensitive conjugative plasmid R27, the prototype for the IncHI1 family of plasmids, has been compiled and analyzed. This 180 kb plasmid contains 210 open reading frames (ORFs), of which 14 have been previously identified and 56 exhibit similarity to other plasmid and prokaryotic ORFs. A number of insertion elements were found, including the full Tn 10 transposon, which carries tetracycline resistance genes. Two transfer regions, Tra1 and Tra2, are present, which are separated by a minimum of 64 kb. Homologs of the DNA-binding proteins TlpA and H-NS that act as temperature-regulated repressors in other systems have been located in R27. Sequence analysis of transfer and replication regions supports a mosaic-like structure for R27. The genes responsible for conjugation and plasmid maintenance have been identified and mechanisms responsible for thermosensitive transfer are discussed.
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Resistencia a Múltiples Medicamentos/genética , Factores R/química , Salmonella typhi/genética , Secuencia de Aminoácidos , Secuencia de Bases , Conjugación Genética , ADN Nucleotidiltransferasas/química , ADN Nucleotidiltransferasas/genética , Desoxirribonucleasa I/química , Desoxirribonucleasa I/genética , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Alineación de Secuencia , Homología de Secuencia de Aminoácido , TemperaturaRESUMEN
Tellurium compounds are used in several industrial processes, although they are relatively rare in the environment. Genes associated with tellurite resistance (TeR) are found in many pathogenic bacteria. Tellurite can be detoxified through interactions with cellular thiols, such as glutathione, or a methyltransferase-catalyzed reaction, although neither process appears involved in plasmid-mediated TeR.
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Antibacterianos/farmacología , Bacterias/efectos de los fármacos , Telurio/farmacología , Antibacterianos/toxicidad , Bacterias/genética , Farmacorresistencia Microbiana/genética , Humanos , Telurio/toxicidadRESUMEN
The availability of two complete Helicobacter pylori genome sequences and recent studies of its population genetics have provided a detailed picture of genetic diversity in this important human gastric pathogen. It is believed that, in addition to genetic recombination, de novo mutation could have a role in generating the high level of genetic variation in H. pylori.
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Variación Genética/genética , Genoma Bacteriano , Helicobacter pylori/genética , Daño del ADN , Reparación del ADN , Humanos , MutaciónRESUMEN
The magnitude and mathematical nature of short-term variations of pulse rate and mean arterial blood pressure has been studied in 27 post-operative cardiac surgical patients over a continuous period of 200 min in each case. Similar variability was observed in all patients. Short-term variations were composed predominantly of a series of rhythmic changes ranging from one synchronous with respiration to others between 2 and 5 min in cycle length. There was consistent variance of the beat-by-beat values for both variables about a continuously updating 5 min mean. The average standard deviation was 3.75 beats/min for pulse rate and 0.48 kPa (3.64 mm Hg) for mean arterial blood pressure. For both variables the distribution about the 5-min mean did not differ significantly from a normal distribution in 50 out of the 54 records. These findings have implications for the reproducibility of current methods of estimating mean pulse rate and blood pressure, and the change necessary before two estimates may be regarded as significantly different. The results are applicable both to the interpretation of Ward Charts by medical staff and to automated monitoring systems.
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Presión Sanguínea , Procedimientos Quirúrgicos Cardíacos , Pulso Arterial , Adolescente , Adulto , Análisis de Varianza , Niño , Femenino , Humanos , Masculino , Persona de Mediana Edad , Factores de TiempoRESUMEN
A chloramphenicol-resistance determinant (CmR), originally cloned from Campylobacter coli plasmid pNR9589 in Japan, was isolated and the nucleotide sequence determined, which contained an open reading frame of 621 bp. The gene product was identified as Cm acetyltransferase (CAT), which had a putative amino acid sequence that showed 43% to 57% identity with other CAT proteins of both Gram+ and Gram- origin. Although expression of the cat gene was constitutive in both C. coli and Escherichia coli, results of primer extension experiments indicated that transcription was initiated at different sites in these two species. A kanamycin-resistance determinant, identified as the aphA-3 gene, was located downstream from the cat gene. The codon usage of the cat gene is very different from that used in E. coli, however, the CAT polypeptide was synthesized in large amounts in E. coli maxicells. Therefore, the codon usage bias is not one of the obstacles which affects Campylobacter spp. gene expression in E. coli. New Campylobacter cloning vectors were constructed in this study.
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Campylobacter/genética , Cloranfenicol O-Acetiltransferasa/genética , Resistencia al Cloranfenicol/genética , Secuencia de Aminoácidos , Secuencia de Bases , Campylobacter/enzimología , Clonación Molecular/métodos , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Escherichia coli/genética , Genes Bacterianos , Vectores Genéticos , Datos de Secuencia Molecular , Plásmidos , Mapeo Restrictivo , Homología de Secuencia de Ácido NucleicoRESUMEN
Agarose-immobilized chromosomal DNA from the nalidixic-acid-resistant Campylobacter coli strain UA417 and its streptomycin-resistant (StrR) derivative, UA417R, were digested with the restriction enzymes SalI (GTCGAC) and SmaI (CCCGGG). The sizes of the resulting fragments were determined using pulsed-field gel electrophoresis. The two genomes showed similar restriction patterns of seven and 13 fragments for the two respective enzymes and the total genome size was determined to be approx. 1.7 Mb. Analysis of partial digestion fragments, as well as Southern-blot hybridization, were used to construct a physical map of the C. coli UA417R genome. Natural transformation studies using DNA fragments extracted from UA417R, as well as the erythromycin-resistant (EryR) C. coli strain UA585, were used to locate the StrR and EryR resistance markers on the genomic map.
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Campylobacter/genética , Genes Bacterianos , Southern Blotting , Mapeo Cromosómico , Farmacorresistencia Microbiana/genética , Electroforesis en Gel de Poliacrilamida , Ácido Nalidíxico , Polimorfismo de Longitud del Fragmento de Restricción , EstreptomicinaRESUMEN
Resistance to tetracycline in the microaerophilic Gram-negative bacterium Campylobacter jejuni is plasmid-mediated. A 6.9-kb HindIII DNA fragment containing the tetracycline-resistance (TcR) gene (designated tetO) from the C. jejuni conjugative plasmid pUA466 was cloned into pUC8, and the resultant plasmid pUOA1 was used to transform Escherichia coli to Tc resistance. The tetO gene was localized at a 2.0-kb region comprising 0.2-kb and 1.8-kb HincII fragments, and the nucleotide sequences were determined. The protein coding region of tetO contained a 1911-bp open reading frame which corresponded to a 72.3-kDa protein. Upstream from the start codon were hexanucleotides that resembled the canonical sequences found at the -10 region, -35 region and the ribosome-binding site of the prokaryotic promoter. The tetO gene product was expressed utilizing an E. coli-derived in vitro transcription/translation system. The polypeptide had an apparent Mr of 68,000. Comparison of the amino acid sequences of TetO to those of TetM (derived from the Gram-positive Streptococcus pneumoniae) revealed 76% homology. Hydrophilicity plot analyses of TetO and TetM proteins provided almost identical profiles. These results clearly support our earlier [Taylor et al., J. Bacteriol. 169 (1987) 2984-2989] suggestion that TcR determinants found in Gram-positive bacteria and in C. jejuni may have a common ancestry.
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Proteínas Bacterianas/genética , Campylobacter fetus/genética , Proteínas Portadoras , Secuencia de Aminoácidos , Secuencia de Bases , Elementos Transponibles de ADN , Datos de Secuencia Molecular , Resistencia a la TetraciclinaRESUMEN
The transcription and translation of the tellurite-resistance (TeR) genes of the HII incompatibility group plasmid, pHH1508a, were studied. The nucleotide (nt) sequence of the TeR region was determined and two possible open reading frames, tehA and tehB, were identified. The direction of transcription and translation of these genes was confirmed through the preparation of lacZ and phoA (encoding alkaline phosphatase) fusions. The transcription start point was identified in the sequence using RNA primer extension. The tehA gene codes for a 36-kDa polypeptide which is highly hydrophobic. The TehA protein appears to be located in the inner membrane of the bacterial cell since tehA fusions with both phoA and lacZ were obtained and expressed. The tehB gene codes for a 23-kDa polypeptide which appears to be relatively hydrophilic and is probably located in the cytoplasm. Both proteins were overproduced using a T7 RNA polymerase/promoter system. No nt or amino acid sequence homology could be found between this TeR determinant and the TeR genes from the IncHI-2 plasmid, pMER610, and the IncP alpha plasmid, RK2.
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Resistencia a Medicamentos/genética , Plásmidos/genética , Telurio , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Electroforesis en Gel de Poliacrilamida , Expresión Génica , Marcadores Genéticos , Datos de Secuencia Molecular , Mutagénesis Insercional , Biosíntesis de Proteínas , Mapeo Restrictivo , Transcripción GenéticaRESUMEN
In this study, we cloned and sequenced the Helicobacter pylori genes encoding fumarate reductase (FRD). H. pylori frdA, frdB and frdC specify polypeptides of 715, 245 and 254 aa, respectively. The deduced aa sequences of FrdA and FrdB are highly homologous to those of the corresponding subunits of Wolinella succinogenes FRD and also exhibit a significant sequence identity with other bacterial FRD and succinate dehydrogenase subunits A and B. However, H. pylori FrdC shares a striking degree of sequence identity only with W. succinogenes FrdC, which is a cytochrome b with two haem groups. The products encoded by H. pylori frdA, frdB and frdC were overproduced in maxicells and H. pylori FrdA was characterized using an anti-E. coli FrdA serum. H. pylori FRD activity, which was measured as fumarate-dependent benzyl viologen oxidation, is membrane-associated. Inactivation of frdA led to the loss of such activity and the mutant H. pylori cells were delayed (approx. 10-20 h behind their parent cells) in entering the mid-log phase, suggesting that FRD-driven metabolism plays an active but non-essential role for growth of H. pylori cells in vitro. H. pylori FRD contains three subunits, of which FrdA and FrdB appear to form the catalytic dimer, whereas FrdC serves as a membrane anchor.
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Genes Bacterianos , Helicobacter pylori/enzimología , Operón , Succinato Deshidrogenasa/genética , Secuencia de Aminoácidos , Anticuerpos Antibacterianos/inmunología , Clonación Molecular , Escherichia coli/enzimología , Escherichia coli/inmunología , Datos de Secuencia Molecular , Mutagénesis , Homología de Secuencia de Aminoácido , Succinato Deshidrogenasa/metabolismoRESUMEN
A comparative study has been carried out on the biotolerance to subcutaneous, intramuscular and intraperitoneal implants of silicone rubber, microporous PTFE and polyacrylamide-agarose gel for 6 wk in the rat. Assessment was quantitative histology for capsule thickness, fibroblast density, mononuclear cell density, multinucleated giant cell density and also collagen packing density. The trial polymers were compared to historical data on healing in dummy procedures and on two hydrophilic graft copolymers of high biotolerance namely polyethylene-acrylic acid and microporous polypropylene-acrylic acid. The rank ordering of the trial polymers for all criteria except collagen packing, was for the least response to be to polyacrylamide-agarose and the most to silicone rubber, with the differences being significant (p less than 0.05) for capsule thickness and mononuclear cell density. Polyacrylamide-agarose was similar in response to the two hydrophilic copolymers; microporous PTFE was slightly more reactive and silicone rubber was significantly more reactive (p less than 0.05).
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Resinas Acrílicas , Politetrafluoroetileno , Prótesis e Implantes , Sefarosa , Elastómeros de Silicona , Animales , Materiales Biocompatibles , Colágeno/fisiología , Geles , Ratas , Propiedades de Superficie , Cicatrización de HeridasRESUMEN
Gastric biopsy specimens were examined microbiologically and histologically for the presence of Campylobacter pyloridis. Of 51 randomly selected patients, 22 (43%) were found to harbor C. pyloridis in the gastric mucosa. The histologic demonstration of spiral organisms observed by staining with hematoxylin and eosin correlated well with microbiologic isolation of the organisms. There was a strong association (95.5%) between C. pyloridis in the gastric mucosa and histologically defined gastritis. However, there was no obvious association between C. pyloridis and ulcers. All C. pyloridis strains isolated exhibited uniform biochemical characteristics and had almost identical protein profiles, which indicated that they belong to a relatively homogeneous group distinct from other Campylobacter species. All C. pyloridis isolates were uniformly susceptible to ampicillin, amoxicillin, cephalothin, gentamicin, kanamycin, tetracycline, coumermycin, ciprofloxacin, novobiocin, clorobiocin, and nitrofurantoin. They were moderately resistant to nalidixic acid.
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Campylobacter/aislamiento & purificación , Mucosa Gástrica/microbiología , Gastritis/microbiología , Gastropatías/microbiología , Campylobacter/efectos de los fármacos , Femenino , Gastritis/patología , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Microscopía Electrónica , Coloración y Etiquetado , Gastropatías/patologíaRESUMEN
Prolonged hyperoxia causes lung injury and respiratory failure secondary to oxidative tissue damage mediated, in part, by the superoxide anion. We hypothesized that aerosol treatment with recombinant human manganese superoxide dismutase (rhMnSOD) would attenuate hyperoxic lung damage in primates. Adult baboons were anesthetized and ventilated with 100% oxygen for 96 h or until death. Six animals were treated with aerosolized rhMnSOD (3 mg . kg-1 . day-1 in divided doses), and six control animals did not receive enzyme therapy. Physiological variables were recorded every 12 h, and ventilation-perfusion ratio relationships were evaluated by using the multiple inert-gas elimination technique. After the experiments, surfactant composition and lung edema were measured. We found that rhMnSOD significantly decreased pulmonary shunt fraction (P < 0.01) and preserved arterial oxygenation (P < 0.01) during hyperoxia. The rhMnSOD increased lung phospholipids, phosphatidylcholine and disaturated phosphatidylcholine, and decreased lung edema in this model. Testing of higher and lower doses of MnSOD (1 and 10 mg . kg-1 . day-1) in two other groups of baboons produced variable physiological protection, suggesting a "window" of effective dosage. We conclude that aerosolized MnSOD (3 mg . kg-1 . day-1) affords significant preservation of pulmonary gas exchange during hyperoxic lung injury.
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Hiperoxia/patología , Pulmón/patología , Superóxido Dismutasa/farmacología , Aerosoles , Animales , Hemodinámica , Humanos , Hiperoxia/metabolismo , Hiperoxia/fisiopatología , Pulmón/metabolismo , Pulmón/fisiopatología , Masculino , Papio , Edema Pulmonar/etiología , Intercambio Gaseoso Pulmonar , Surfactantes Pulmonares/química , Proteínas Recombinantes , Respiración , Análisis de Supervivencia , Relación Ventilacion-PerfusiónRESUMEN
Two virus isolates, 1 each from Aedes campestris and Psorophora signipennis mosquitoes collected in south central New Mexico in August 1985, were shown by neutralization tests to be identical to each other, but not to any of more than 250 arthropod-borne and other viruses. Electron microscopy of 1 isolate (85-488NM, chosen as the prototype) indicated that this strain shares morphologic characteristics with viruses of the family Rhabdoviridae. Indirect fluorescent antibody tests indicated that this virus is a member of the genus Vesiculovirus, but is not closely related to any of the North American or other rhabdoviruses with which it was tested, including vesicular stomatitis (Indiana) and vesicular stomatitis (New Jersey) viruses. The name Malpais Spring virus is proposed for this newly recognized vesiculovirus. A serologic survey indicated that Malpais Spring virus infects indigenous (mule deer and pronghorn) and exotic (gemsbok) ungulates at and near the sites where the mosquitoes from which the virus strains were isolated were collected. Antibody prevalence in wild animals indicates that the pronghorn and gemsbok may play roles as hosts for Malpais Spring, epizootic hemorrhagic disease (New Jersey), and bluetongue viruses in this area.
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Aedes/microbiología , Animales Salvajes , Rhabdoviridae/aislamiento & purificación , Rumiantes , Virosis/veterinaria , Animales , Pruebas de Fijación del Complemento , Reacciones Cruzadas , Efecto Citopatogénico Viral , Ciervos , Ensayo de Inmunoadsorción Enzimática , Femenino , Técnica del Anticuerpo Fluorescente , Ratones , Microscopía Electrónica , Pruebas de Neutralización , New Mexico , Rhabdoviridae/ultraestructura , Células Vero , Virosis/microbiologíaRESUMEN
Streptococcus pneumoniae is a Gram-positive bacterium which is naturally resistant to tellurite. In this study, we cloned and sequenced a homologue of the Escherichia coli tellurite resistance gene tehB from S. pneumoniae. It encoded a protein of 284 amino acids which is 86 residues longer than E. coli TehB, but similar in size to Haemophilus influenzae TehB and Eikenella corrodens hemagglutinin (Hag1) as well as homologues from Actinobacillus actinomycetemcomitans, Neisseria gonorrhoeae and Neisseria meningitidis. The S. pneumoniae TehB displayed 46-58% identity (52-68% similarity) to these proteins. The results in this study showed that the S. pneumoniae tehB alone not only conferred on E. coli high level resistance to tellurite, but also caused filamentous morphology in E. coli.
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Proteínas Bacterianas/genética , Farmacorresistencia Microbiana/genética , Proteínas de Escherichia coli , Genes Bacterianos , Streptococcus pneumoniae/efectos de los fármacos , Telurio/farmacología , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Escherichia coli/fisiología , Pruebas de Sensibilidad Microbiana , Datos de Secuencia Molecular , Análisis de Secuencia de ADN , Streptococcus pneumoniae/genéticaRESUMEN
We previously reported that a gene hpcopA isolated from Helicobacter pylori is associated with copper transport. In the present study, the DNA upstream of the hpcopA was cloned and the nucleotide sequence analyzed. An open reading frame coding for 124 amino acids was predicted, which was connected in frame to the hpcopA. The deduced protein sequence exhibits striking homology with known copper-transporting P-type ATPases. Disruption of this ORF renders H. pylori hypersensitive to copper present in growth media, indicating that it encodes the N-terminal amino acid sequence of the hpCopA protein. Measurement of copper content in the wild-type and hpcopA-disrupted cells showed that the copper content was increased in the mutant cells, further supporting that the previous proposal that the gene hpcopA is involved in copper export. In addition, the cop operon consisting of the genes hpcopA and hpcopP was identified by primer extension and reverse transcription-polymerase chain reaction. Our results indicate that the genes for copper import and export are located in separate operons within H. pylori.
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Adenosina Trifosfatasas/genética , Proteínas Portadoras/genética , Proteínas de Transporte de Catión , Cobre/metabolismo , Helicobacter pylori/genética , Secuencia de Bases , ATPasas Transportadoras de Cobre , Genes Bacterianos/fisiología , Helicobacter pylori/enzimología , Datos de Secuencia Molecular , Mutagénesis/fisiología , Conformación de Ácido Nucleico , Sistemas de Lectura Abierta , Operón/genética , Reacción en Cadena de la Polimerasa , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Transcripción Genética/genéticaRESUMEN
The genomic map of Campylobacter jejuni UA580 was expanded and more precisely constructed using I-CeuI, Sal/I and SmaI restriction endonucleases in conjunction with pulsed-field gel electrophoresis (PFGE). The presence of three fragments after digestion with I-CeuI confirmed the presence of three copies of the 23S ribosomal rRNA (rrl) gene. The genome size of Campylobacter jejuni UA580 was determined to be 1725 +/- 5.9 kbp by I-CeuI with fragment sizes of 1053 +/- 4.4, 361 +/- 2.7 and 311 +/- 3.6 kbp. Analysis of a PCR product from C. jejuni UA580 23S rRNA gene showed that I-CeuI did cut within the gene. The precise locations of the three genes were determined using I-CeuI with two copies of the 23S and 5S rRNA genes located separately from the 16S rRNA gene whereas the third copy of the 23S and 5S rRNA genes had a closer linkage to a 16S rRNA gene copy. Homologous gene probes were used to map additional genes and allowed the realignment of a few previously mapped genes on the chromosome. Other strains of C. jejuni were also cut into three fragments with I-CeuI, which generated variable PFGE patterns.
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Campylobacter jejuni/genética , Mapeo Cromosómico , Endodesoxirribonucleasas/metabolismo , ARN Ribosómico 23S/genética , Sondas de ADN/genética , Electroforesis en Gel de Campo Pulsado , Genoma , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S/genética , ARN Ribosómico 5S/genética , Mapeo RestrictivoRESUMEN
Genomic DNA from 15 strains of Helicobacter mustelae was subjected to pulsed-field gel electrophoresis (PFGE) after digestion with PacI and SfiI. H. mustelae genome DNA appeared very similar in all strains examined, whether isolated from ferrets or mink or from animals bred in either the USA or in the UK. The H. mustelae genome size was estimated to be 1.7 Mb, similar in size to that of H. pylori. A minor difference in PacI PFGE pattern and genome size was observed between rifampicin-resistant and rifampicin-susceptible derivatives of H. mustelae F251. Another minor difference in genome pattern based on PFGE with SfiI was observed between an H. mustelae strain used to experimentally infect four ferrets which resulted in loss of an SfiI site in strains obtained from the newly infected ferrets. Thus, although minor differences in PFGE pattern were noted, H. mustelae lacks the genomic diversity observed in H. pylori.