RESUMEN
Increasingly, positron emission tomography/computed tomography (PET/CT) is being used as a tumor surveillance modality for multiple tumor types. A 73-year-old postmenopausal female with stage IV nonsmall cell lung cancer presented after a PET/CT demonstrated focal uptake in the superior and lateral aspects of the uterus. The patient reported a history of intermittent postmenopausal bleeding and an endometrial biopsy documented uterine carcinosarcoma. Postoperative pathologic review and immunohistochemical staining with thyroid transcription factor-1 revealed metastatic adenocarcinoma consistent with her lung primary in her uterus and adnexa. Our case represents a rare occurrence in which lung cancer has metastasized to multiple female pelvic organs. Increasing use of PET/CT may lead to the discovery of occult metastases masquerading as a second primary malignancy.
Asunto(s)
Carcinoma/patología , Carcinosarcoma/diagnóstico por imagen , Carcinosarcoma/secundario , Neoplasias Pulmonares/patología , Neoplasias Primarias Desconocidas/diagnóstico por imagen , Neoplasias Uterinas/diagnóstico por imagen , Neoplasias Uterinas/secundario , Anciano , Femenino , Humanos , Tomografía de Emisión de Positrones , Tomografía Computarizada por Rayos XRESUMEN
Bacillus subtilis and related species (Bacillus licheniformis, Bacillus pumilus, Bacillus amyloliquefaciens, and Bacillus mojavensis) represent a group of bacteria largely studied and widely employed by industry. Small rolling circle replicating plasmids of this group of bacteria have been intensively studied as they represent a convenient model for genetic research and for the construction of molecular tools for the genetic modification of their hosts. Through the computational analysis of the available plasmid sequences to date, the first part of this review focuses on the main stages that the present model for rolling circle replication involves, citing the research data which helped to elucidate the mechanism by which these molecules replicate. Analysis of the distribution and phylogeny of the small RC plasmids inside the Bacillus genus is then considered, emphasizing the low level of diversity observed among these plasmids through the in silico analysis of their organization and the sequence divergence of their replication module. Finally, the parasitic vs. mutualistic nature of small rolling circle plasmids is briefly discussed.
Asunto(s)
Bacillus subtilis/fisiología , ADN Bacteriano/genética , ADN Circular/genética , Plásmidos , Bacillus subtilis/genética , Especificidad de la EspecieRESUMEN
The degree of biodiversity among Bacillus licheniformis plasmids and their relation to other Bacillus subtilis group plasmids has been evaluated. To attain this goal we surveyed the diversity and linkage of replication modules in a collection of 21 naturally occurring plasmids of B. licheniformis strains, isolated from different geographical areas. On the basis of rep gene sequence analysis it was possible to group the B. licheniformis plasmids rep genes in two main cluster. Comparison with known rep genes from Bacillus rolling-circle-replicating (RCR) plasmids revealed the presence in B. licheniformis plasmids of replication genes with a DNA sequence peculiar to B. licheniformis species together with rep genes with a very high sequence similarity to B. subtilis plasmids. Furthermore, the molecular organization of an atypical replicon, pBL63.1, was shown. This plasmid did not display any significant similarity with known Bacillus RCR plasmids. The complete nucleotide sequence evidenced a replication module with an unexpected similarity with Rep proteins from RCR plasmids of bacterial species phylogenetically distantly related to Bacillus. pBL63.1 represents an exception to the low-level diversity hypothesis among Bacillus RC replicons.
Asunto(s)
Bacillus/genética , Variación Genética , Plásmidos/genética , Replicón , Secuencia de Bases , Southern Blotting , Replicación del ADN , Datos de Secuencia Molecular , Filogenia , Análisis de Secuencia , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido NucleicoRESUMEN
The complete nucleotide sequences of two plasmids, pFL5 and pFL7, isolated from soil bacteria, Bacillus licheniformis FL5 and FL7, have been determined. The plasmids pFL5 and pFL7 were analyzed and found to be 9150 and 7853 bp in size with a G+C content of 41.0 and 43.6 mol%, respectively. Computer assisted analysis of sequence data revealed 11 possible ORFs in pFL5, four of which could be assigned no function from homology searches. Instead, eight putative ORFs were identified in pFL7, two of which appeared to have no biological function. All the ORFs were preceded by a ribosome binding site. The ORFs 9.5 and 6.7, each of 340 amino acids, were postulated to encode a replication protein similar to known replication proteins of rolling circle replicons, particularly those of the pC194 family. The structural organization of the two pFL plasmids is similar to the pTA plasmids family, with only a few putative coding regions that cannot be attributed to these plasmid backbone genes. In contrast to pTA plasmids, the majority of the genes have an orientation of transcription opposite to the direction of replication. The identified probable sso sequences seem to belong to a different group of those found in Bacillus plasmids; in fact, a significant level of homology was found with ssoA group sequences. These plasmids seem to be related to plasmids identified within the Bacillus subtilis group, confirming the low-level diversity among these replicons.
Asunto(s)
Bacillus/genética , Plásmidos/genética , Microbiología del Suelo , Composición de Base , Secuencia de Bases , Orden Génico , Datos de Secuencia Molecular , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de SecuenciaRESUMEN
AIMS: The milk acidification rate of Streptococcus thermophilus strains can be affected by several factors, one of which is the hydrolysis of urea by the urease complex. To evaluate the technological suitability of S. thermophilus strains deprived of urease activity in milk fermentation, the genetic cluster related to urease enzymatic activity has been characterized in the type strain DSM 20167T. METHODS AND RESULTS: Amplification of the urease genes of S. thermophilus DSM 20167T was developed on the basis of the urease gene cluster of the phylogenetically related S. salivarius. Nucleotide sequencing revealed the presence of eight open reading frames, which were most homologous to ureABC (structural genes) and ureI, ureEFGD (accessory genes) of S. salivarius and other ureolytic bacteria. Reverse transcriptase PCR experiments were in agreement with an operon organization for the eight genes (ureIABCEFGD). A food grade mutant A16 (DeltaureC3) with a 693 bp in-frame deletion in ureC gene exhibited a urease negative (Ure-) phenotype. Unlike the wild-type strain, the acidification rate of the mutant in reconstituted skimmed milk was not affected by the presence of urea or nickel ions. A small-scale yoghurt fermentation trials were carried out using the wild-type or the Ure- mutant A16 (DeltaureC3) in co-culture with Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 in presence of urea. The result obtained underlines that when the Ure- mutant was used as a co-starter the acidification rate was higher than that obtained using the wild-type strain. CONCLUSIONS: The study provides the first genetic characterization and the technological implication of S. thermophilus DSM 20617T urease activity. SIGNIFICANCE AND IMPACT OF THE STUDY: The detrimental effect of ureolytic activity on the rate of milk acidification was evaluated and superseded using a food-grade Ure- recombinant strain. Small-scale yoghurt production trials highlighted the positive role of a Ure-S. thermophilus mutant as a co-starter in milk fermentations. Moreover, the vector pMI108 developed for the construction of the Ure- strain, should be considered as a potential tool for the generation of Ure- dairy S. thermophilus strains selected for other relevant technological properties but characterized by the undesirable ureolytic phenotype.
Asunto(s)
Genes Bacterianos , Leche/microbiología , Familia de Multigenes , Streptococcus/genética , Ureasa/genética , Animales , ADN Bacteriano/genética , Microbiología de Alimentos , Humanos , Concentración de Iones de Hidrógeno , Mutagénesis Sitio-Dirigida , Fenotipo , Análisis de Secuencia de ADN/métodos , Streptococcus/enzimología , Streptococcus/crecimiento & desarrollo , Yogur/microbiologíaRESUMEN
AIMS: To evaluate the genetic diversity and the technological properties of 44 strains of Streptococcus thermophilus isolated from dairy products. Methods METHODS AND RESULTS: Strains were analysed for some relevant technological properties, i.e. exopolysaccharide (EPS) production, growth kinetic in skim milk medium, urease activity and galactose fermentation. The EPS production, determined by evaluating the colour of the colonies grown in ruthenium red milk agar, was observed in 50% of the analysed strains. Urease activity, determined by colorimetric and conductimetric methods, showed that 91% of the isolates, all except four, could hydrolyse urea. A conductimetric approach was also used for the evaluation of the overall metabolic behaviour in milk of Strep. thermophilus strains and the differences observed allowed grouping of the strains in seven different clusters. A total of 11 strains were able to produce acid in presence of galactose. Genetic diversity of Streptococcus thermophilus strains, evaluated by Random Amplified Polymorphic DNA fingerprinting (RAPD) and amplified epsC-D restriction analysis, allowed the identification of 21 different genotypes. CONCLUSIONS: Comparison between the genotypic and phenotypic data highlights an interesting correlation between some important technological properties and well-defined genotypes. SIGNIFICANCE AND IMPACT OF THE STUDY: The genetic and technological characterization carried out on several Strep. thermophilus strains of dairy origin should expand the knowledge on this important lactic acid bacteria species and lead to a simple, rapid, and reliable identification of strains on the basis of well-defined biotechnological properties.
Asunto(s)
Productos Lácteos/microbiología , Variación Genética , Streptococcus/genética , Animales , Genotipo , Lactosa/metabolismo , Fenotipo , Polisacáridos Bacterianos/biosíntesis , Técnica del ADN Polimorfo Amplificado Aleatorio , Streptococcus/clasificación , Streptococcus/metabolismo , Ureasa/metabolismoRESUMEN
Two species of dairy streptococci, Streptococcus waius and Streptococcus macedonicus, were originally characterized by 16S-23S intergenic spacer sequence analysis, random amplified polymorphic DNA fingerprinting, PFGE analysis and DNA-DNA reassociation experiments. All genetic data suggested that S. waius strains belong to the previously described species S. macedonicus. Likewise, the phenotypic characterization showed that strains of S. macedonicus and S. waius were highly related and easily differentiated from the closest phylogenetic neighbour, Streptococcus bovis, principally by their failure to produce a blackening reaction in medium containing aesculin. The utilization of maltose and cellobiose by S. macedonicus/S. waius strains allowed their differentiation from the most studied dairy species, Streptococcus thermophilus. On the basis of genetic and phenotypic data S. macedonicus and S. waius species should be considered synonyms and S. macedonicus has the priority.
Asunto(s)
Productos Lácteos/microbiología , Streptococcus/clasificación , Técnicas de Tipificación Bacteriana , Composición de Base , Electroforesis en Gel de Campo Pulsado , Genotipo , Hibridación de Ácido Nucleico , Fenotipo , Filogenia , Reacción en Cadena de la Polimerasa , Técnica del ADN Polimorfo Amplificado Aleatorio , Streptococcus/genética , Streptococcus/fisiologíaRESUMEN
The vacuolar proton pump (V-ATPase) located on the plasma membrane of the osteoclast is a potential molecular target for the discovery of novel bone antiresorptive agents useful for the treatment of osteoporosis. In order to design novel compounds able to selectively inhibit the osteoclast V-ATPase we firstly identified the minimal structural requirements of bafilomycin A1, a macrolide antibiotic which potently inhibits all V-ATPases. This information allowed the design of 2-(indole)pentadienamide derivatives whose optimization led to a novel class of potent inhibitors that demonstrated a high degree of selectivity for the osteoclast V-ATPase. The most interesting derivative, SB-242784, was able to inhibit bone resorption by human osteoclasts in vitro and to completely prevent ovariectomy-induced bone loss in rats when administered orally at 10 mg kg(-1) day(-1). Structure activity relationships of this class of compounds were investigated further by replacing the 2,4-pentadienoyl chain with suitable spacers able to maintain the correct orientation and distance between the indole ring and the amide moiety.
Asunto(s)
Resorción Ósea/prevención & control , Inhibidores Enzimáticos/farmacología , Osteoclastos/efectos de los fármacos , ATPasas de Translocación de Protón/antagonistas & inhibidores , ATPasas de Translocación de Protón Vacuolares , Animales , Humanos , Osteoclastos/enzimología , Osteoporosis/prevención & control , Relación Estructura-ActividadRESUMEN
In the last decade a number of selective and potent non-peptidic agents became available to explore the usefulness of the delta-opioid receptor in modulation of pain of different origins. As a continuing effort in this field, potent and selective delta-opioid agonists based on the pyrrolomorphinan framework have been designed, synthesised and characterised biologically in our laboratories. In animal models, a selected compound of interest, SB 235863, has proved the concept that selective delta-opioid agonists may have great potential as pain relief agents in inflammatory and neuropathic pain conditions. Importantly, such a compound was free of the unwanted side effects usually associated with narcotic analgesics such as morphine.
Asunto(s)
Analgésicos Opioides/farmacología , Receptores Opioides delta/agonistas , Animales , Diseño de Fármacos , Humanos , Inflamación/tratamiento farmacológico , Dolor/tratamiento farmacológico , Relación Estructura-ActividadRESUMEN
A stepwise chemical modification from human neurokinin-3 receptor (hNK-3R)-selective antagonists to potent and combined hNK-3R and hNK-2R antagonists using the same 2-phenylquinoline template is described. Docking studies with 3-D models of the hNK-3 and hNK-2 receptors were used to drive the chemical design and speed up the identification of potent and combined antagonsits at both receptors. (S)-(+)-N-(1-Cyclohexylethyl)-3-[(4-morpholin-4-yl)piperidin-1-yl]methyl-2-phenylquinoline-4-carboxamide (compound 25, SB-400238: hNK-3R binding affinity, K(i) = 0.8 nM; hNK-2R binding affinity, K(i) = 0.8 nM) emerged as the best example in this approach. Further studies led to the identification of (S)-(+)-N-(1,2,2-trimethylpropyl)-3-[(4-piperidin-1-yl)piperidin-1-yl]methyl-2-phenylquinoline-4-carboxamide (compound 28, SB-414240: hNK-3R binding affinity, K(i) = 193 nM; hNK-2R binding affinity, K(i) = 1.0 nM) as the first hNK-2R-selective antagonist belonging to the 2-phenylquinoline chemical class. Since some members of this chemical series showed a significant binding affinity for the human mu-opioid receptor (hMOR), docking studies were also conducted on a 3-D model of the hMOR, resulting in the identification of a viable chemical strategy to avoid any significant micro-opioid component. Compounds 25 and 28 are therefore suitable pharmacological tools in the tachykinin area to elucidate further the pathophysiological role of NK-3 and NK-2 receptors and the therapeutic potential of selective NK-2 (28) or combined NK-3 and NK-2 (25) receptor antagonists.
Asunto(s)
Morfolinas/síntesis química , Piperidinas/síntesis química , Quinolinas/síntesis química , Receptores de Neuroquinina-2/antagonistas & inhibidores , Receptores de Neuroquinina-3/antagonistas & inhibidores , Secuencia de Aminoácidos , Animales , Células CHO , Clonación Molecular , Cricetinae , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Morfolinas/química , Morfolinas/metabolismo , Piperidinas/química , Piperidinas/metabolismo , Quinolinas/química , Quinolinas/metabolismo , Ensayo de Unión Radioligante , Receptores de Neuroquinina-2/química , Receptores de Neuroquinina-2/metabolismo , Receptores de Neuroquinina-3/química , Receptores de Neuroquinina-3/metabolismo , Receptores Opioides mu/química , Receptores Opioides mu/metabolismo , Relación Estructura-ActividadRESUMEN
Specific regions in three genes coding for aminopeptidases C and N, and a trypsin-like serine protease were selected as species-specific primer sequences for rapid and reliable identification of Lactobacillus helveticus strains. The PCR procedures carried out gave specific 524-, 726- and 918-bp amplificates, with DNA isolated from L. helveticus. No PCR product was generated for closely related bacteria. The amplification products were also screened for their species specificity in dot blot hybridization with representatives of the most closely related genera and species and a number of other bacterial species.
Asunto(s)
Aminopeptidasas , Proteínas Bacterianas/genética , Proteínas de Choque Térmico , Lactobacillus/clasificación , Lactobacillus/genética , Proteínas Periplasmáticas , Reacción en Cadena de la Polimerasa , Serina Endopeptidasas/genética , Cartilla de ADN , Sondas de ADN , ADN Bacteriano/genética , Genes Bacterianos , Especificidad de la EspecieRESUMEN
A polyphasic taxonomic study was performed on the type strain of Bacillus thermosphaericus DSM 10633T and three related soil isolates. On the basis of phenotypic characteristics, chemotaxonomic profiles and phylogenetic data a new genus, Ureibacillus gen. nov., is proposed for the strains in the Bacillus thermosphaericus cluster. Strains of this cluster fall into two DNA-DNA similarity groups: while one group contains the type strain of Ureibacillus thermosphaericus comb. nov. and a single soil isolate, the other contains two soil isolates. The two groups differed in the composition of isoprenoid quinones and some phenotypic properties. These data support the description of a novel species of Ureibacillus for which the name Ureibacillus terrenus is proposed. The type strain of this new species is TH9AT (= DSM 12654T = LMG 19470T).
Asunto(s)
Bacillus/clasificación , Microbiología del Suelo , Bacillus/citología , Técnicas de Tipificación Bacteriana , Composición de Base , ADN Ribosómico , Ácidos Grasos/análisis , Datos de Secuencia Molecular , Fenotipo , ARN Ribosómico 16S , Homología de Secuencia de Ácido Nucleico , Terminología como Asunto , Vitamina K/análisisRESUMEN
A polyphasic study was performed on five thermophilic strains belonging to the genus Bacillus, isolated from soil of different geographical areas. 16S rRNA gene sequence analysis placed these isolates in RNA group 5, with Saccharococcus caldoxylosilyticus and [Bacillus] thermoglucosidasius being the closest phylogenetic neighbours. The type species of Saccharococcus, Saccharococcus thermophilus, was only moderately related to these two species and the novel isolates. DNA-DNA hybridization studies and comparison of morphological, chemotaxonomic and phenotypic features supported the close relationship between the novel isolates and Saccharococcus caldoxylosilyticus. These data justify the reclassification of Saccharococcus caldoxylosilyticus. Following the transfer of the validly described Bacillus species of group 5 into the genus Geobacillus, the reclassification of Saccharococcus caldoxylosilyticus as Geobacillus caldoxylosilyticus comb. nov. is proposed. This species can be distinguished genomically from Geobacillus thermoglucosidasius, Geobacillus stearothermophilus, Geobacillus thermodenitrificans and Saccharococcus thermophilus by a specific PCR-RFLP assay targeting the 16S rDNA.
Asunto(s)
Bacillaceae/clasificación , Bacillaceae/genética , Calor , Microbiología del Suelo , Bacillaceae/química , Bacillaceae/fisiología , Composición de Base , ADN Ribosómico , Ácidos Grasos/análisis , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Fenotipo , Filogenia , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S , Mapeo RestrictivoRESUMEN
A RAPD analysis performed using a single primer targeted to the pediocin AcH/PA-1 gene was carried out on several P. acidilactici strains and on some related species of lactic acid bacteria. The high degree of genetic variability detected in P. acidilactici strains did not allow the selection of a common RAPD fragment that could be chosen as a potential species-specific DNA marker. Nevertheless a 700 bp fragment, that was found to be peculiar of all potential pediocin producer strains analyzed, was cloned and sequenced with the aim to develop a species specific PCR marker. Sequence analysis of the cloned 700 bp fragment showed one putative small open reading frame (ORF1), with no significant homology with known genes, and a partial putative second coding region (ORF2) with a high degree of similarity with several methionyl tRNA synthesis (metS) genes. The two coding regions were separated by a short spacer region. Primers targeted to ORF2 plus part of the spacer region and primers designed for the amplification of the entire cloned RAPD fragment were found to be species-specific for the detection of P. acidilactici strains. Furthermore primers designed on the ORF1 sequence allowed the amplification of a 439 bp fragment only in some P. acidilactici strains, including pediocin producing strains.
Asunto(s)
Bacteriocinas/genética , Pediococcus/clasificación , Técnica del ADN Polimorfo Amplificado Aleatorio/métodos , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Marcadores Genéticos , Datos de Secuencia Molecular , Pediocinas , Pediococcus/genética , Análisis de Secuencia de ADN , Homología de Secuencia de AminoácidoRESUMEN
A polyphasic study was performed on 10 soil isolates of thermophilic denitrifying Bacillus strains from different geographical areas. The presence of two main characteristic bands following amplification of the internal transcribed spacer (ITS) region of rrn operons suggests a close relatedness to 'Bacillus thermodenitrificans'. The isolates cluster around two strains of 'B. thermodenitrificans' in riboprint and fatty acid analyses, though differences occur at the strain level. Subsequent DNA-DNA reassociation studies including the 10 isolates, 'B. thermodenitrificans' DSM 465T and DSM 466, and Bacillus stearothermophilus ATCC 12980T and Bacillus thermoleovorans ATCC 43513T revealed such a high level of genomic relatedness between the isolates and the DSM strains (> 73% similarity) that they must be considered strains of the same taxon. The degree of DNA-DNA similarity between the 12 strains of 'B. thermodenitrificans' and the type strains of the other two phylogenetically neighbouring Bacillus species was significantly lower (21-43% similarity). Based upon phylogenetic, chemotaxonomic and phenotypic evidence, the designation of B. thermodenitrificans sp. nov., nom. rev. is proposed. The type strain of B. thermodenitrificans is DSM 465T.
Asunto(s)
Bacillus/clasificación , Nitratos/metabolismo , Microbiología del Suelo , Bacillus/química , Bacillus/aislamiento & purificación , Bacillus/fisiología , Composición de Base , Dermatoglifia del ADN , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Ribosómico/química , ADN Ribosómico/genética , Ácidos Grasos/análisis , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Fenotipo , ARN Ribosómico 16S/genética , ARN Ribosómico 23S/genética , Mapeo Restrictivo , Temperatura , Operón de ARNr/genéticaRESUMEN
Pediococcus acidilactici, Pediococcus parvulus and Lactobacillus plantarum pediocin AcH/PA-1 produces strains were studied with the aim to investigate their common genetic pediocin determinant using pedA, pedB, pedC and pedD gene-targeted PCR assay. Single Strand Conformation Polymorphism and restriction analysis of pedA and pedC amplified fragments from the three different species did not show any differences in sequence while these analysis carried out on pedB and pedD amplified fragments highlighted differences related to the three species analyzed harboring these plasmid encoded genes. Furthermore different multiplex PCR assay using IdhD, pedA and pedD as target genes were developed to clearly identify the pediocin AcH/PA-1 producer strains and to obtain the simultaneous identification of the P. acidilactici strains.
Asunto(s)
Bacteriocinas/genética , Genes Bacterianos , Lactobacillus/genética , Pediococcus/genética , Bacteriocinas/biosíntesis , Southern Blotting , Amplificación de Genes , Lactobacillus/clasificación , Lactobacillus/metabolismo , Operón/genética , Pediocinas , Pediococcus/clasificación , Pediococcus/crecimiento & desarrollo , Pediococcus/metabolismo , Plásmidos/genética , Reacción en Cadena de la Polimerasa , Polimorfismo Conformacional Retorcido-Simple , Mapeo Restrictivo , Análisis de Secuencia de ADNRESUMEN
The vacuolar H+-ATPase (V-ATPase), located on the ruffled border of the osteoclast, is a proton pump which is responsible for secreting the massive amounts of protons that are required for the bone resorption process. With the aim to identify new agents which are able to prevent the excessive bone resorption associated with osteoporosis, we have designed a novel class of potent and selective inhibitors of the osteoclast proton pump, starting from the structure of the specific V-ATPase inhibitor bafilomycin A1. Compounds 3a-d potently inhibited the V-ATPase in chicken osteoclast membranes (IC50 = 60-180 nM) and were able to prevent bone resorption by human osteoclasts in vitro at low-nanomolar concentrations. Notably, the EC50 of compound 3c in this assay was 45-fold lower than the concentration required for half-maximal inhibition of the V-ATPase from human kidney cortex. These results support the validity of the osteoclast proton pump as a useful molecular target to produce novel inhibitors of bone resorption, potentially useful as antiosteporotic agents.
Asunto(s)
Resorción Ósea/prevención & control , Compuestos Bicíclicos Heterocíclicos con Puentes , Inhibidores Enzimáticos , Indoles , Osteoclastos/efectos de los fármacos , ATPasas de Translocación de Protón/antagonistas & inhibidores , Pirimidinas , Vacuolas/efectos de los fármacos , Animales , Compuestos Bicíclicos Heterocíclicos con Puentes/síntesis química , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Membrana Celular , Pollos , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Humanos , Indoles/síntesis química , Indoles/farmacología , Corteza Renal/enzimología , Osteoclastos/enzimología , Osteoclastos/ultraestructura , Pirimidinas/síntesis química , Pirimidinas/farmacología , Células Tumorales Cultivadas , Vacuolas/enzimologíaRESUMEN
A genotypic study using amplified ribosomal DNA restriction analysis (ARDRA), random amplified polymorphic DNA fingerprinting (RAPD) and ribosomal spacer analysis (RSA) in comparison with DNA-DNA reassociation experiments was carried out with 85 thermophilic Bacillus isolates from uncultivated soil of 14 different geographical areas and seventeen reference strains representing defined thermophilic Bacillus species. This approach permitted the attribution of 51% of the new isolates to the Bacillus thermoleovorans group and the identification of 40% of the new isolates as B. "thermodenitrificans". Moreover, 2 strains were assigned to B. pallidus species and 1 isolate to B. thermosphaericus species. The remaining 6% of our thermophilic isolates from soil, constituting 2 DNA-DNA homology groups, are still unidentified. A detailed genotypic characterization of the heterogeneous species of B. thermoleovorans and B. stearothermophilus was also presented.
Asunto(s)
Bacillus/clasificación , Bacillus/genética , Técnicas de Tipificación Bacteriana , Microbiología del Suelo , Bacillus/aislamiento & purificación , Dermatoglifia del ADN , ADN Bacteriano/análisis , ADN Ribosómico/análisis , Calor , Hibridación de Ácido Nucleico , Técnica del ADN Polimorfo Amplificado Aleatorio , Mapeo RestrictivoRESUMEN
A study of phenotypic and genotypic characteristics was carried out on 182 strains isolated from soil of different geographical areas; the type strains were B. licheniformis, B. subtilis, B. pumilus, B. cereus and B. coagulans. The results showed, primarily on the basis of phenotypic features, that all the isolates belonged to the B. licheniformis species, however DNA relatedness studies revealed only 161 to be genetically related to B. licheniformis, the DNA relatedness levels ranging from 66 to 100%. The other 21 isolates appeared to be genetically distinct not only from B. licheniformis but also from B. subtilis and B. pumilus, where there were low levels of DNA relatedness (from 4 to 37%). Nevertheless the ARDRA results indicate that the 21 atypical isolates were phylogenetically related to B. licheniformis. Our data and the phenotypic homogeneity found suggest the presence of three different genomovars.
Asunto(s)
Bacillus/genética , Microbiología del Suelo , Bacillus/clasificación , Bacillus/fisiología , Bacillus cereus/clasificación , Bacillus cereus/genética , Bacillus cereus/fisiología , Bacillus subtilis/clasificación , Bacillus subtilis/genética , Bacillus subtilis/fisiología , Metabolismo de los Hidratos de Carbono , ADN Bacteriano/genética , Genotipo , Fenotipo , Plásmidos/genética , Reacción en Cadena de la Polimerasa , ARN Bacteriano/análisis , ARN Ribosómico 16S/análisis , Especificidad de la EspecieRESUMEN
Optimisation of a novel series of osteoclast ATPase inhibitors led to (2Z,4E)-5-(5,6-dichloro-2-indolyl)-2-methoxy-N-(1,2,2,6,6- pentamethylpiperidin-4-yl)-2,4-pentadienamide (1) that was the most potent compound in an in vitro osteoclast ATPase assay and in human bone resorption assays. Two of the possible geometric isomers have also been prepared and shown to be significantly less potent than 1.