RESUMEN
In the present study, 16 women with recurrent vulvovaginal candidiasis (RVVC) due to Candida albicans and Candida (Torulopsis) glabrata were followed for a period of 4 to 12 months, and 36 vaginal isolates were evaluated by pulsed-field gel electrophoresis (PFGE). Eleven women were infected by C. albicans and 5 by C. glabrata. Three electrophoretic karyotypes of C. albicans and 3 of C. glabrata were identified throughout the follow-up. All patients but one was infected with the same karyotype of C. albicans or C. glabrata during the follow-up period. Two different karyotypes of C. glabrata were identified in one patient in the course of 12 months. The results confirmed the diversity of the karyotypes of C. albicans and C. glabrata causing vulvovaginitis, and demonstrated the persistence of colonization with the same strain over different periods of time despite therapy (15/16 women).
Asunto(s)
Candida albicans/clasificación , Candida albicans/genética , Candida/clasificación , Candida/genética , Candidiasis Vulvovaginal/microbiología , Candida/aislamiento & purificación , Candida albicans/aislamiento & purificación , Cromosomas Fúngicos , Electroforesis en Gel de Campo Pulsado , Femenino , Humanos , Cariotipificación , Técnicas de Tipificación Micológica , Recurrencia , Vagina/microbiologíaRESUMEN
The antimicrobial effects of 30 trifluoromethyl ketones [1-30] were studied on various representative bacteria. Of the ketones, 4,4,4-trifluoro-1-phenyl-1,3-butanedione [10], 1,1,1-trifluoro-3-(4,5-dimethyloxazol-2-yl)-2-propanone [11] and 1-(2-benzoxazolyl)-3,3,3-trifluoro-2-propanone [18] were found to exhibit potent antibacterial activity against the Gram-positive Bacillus megaterium and Corynebacterium michiganese, but not against Gram-negative bacteria such as Pseudomonas aeruginosa and Serratia marcescens. Compounds 11 and 18 inhibited the Escherichia coli. Compound 18 was also effective against yeasts. The combination of promethazine with 18 was significantly synergistic against E. coli strains, especially the proton pump deficient mutant. The results suggest that membrane transporters are the target of trifluoromethyl ketones. The inhibition was more marked in the proton pump deficient E. coli mutant than in the wild type, which suggested that the antibacterial effect of trifluoromethyl ketones is partly prevented by the proton pump system.
Asunto(s)
Antiinfecciosos/farmacología , Bacterias/efectos de los fármacos , Hongos/efectos de los fármacos , Cetonas/química , Cetonas/farmacología , Prometazina/farmacología , Antibacterianos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , VIH-1/efectos de los fármacos , Humanos , Pruebas de Sensibilidad Microbiana/métodosRESUMEN
Fungal isolates belonging to four Rhizopus species were screened for the presence of double-stranded RNA (dsRNA) molecules. Five (two R. stolonifer, two R. microsporus, and one R. oryzae) of the 27 isolates examined harboured such genetic elements. Electrophoresis of the nucleic acids revealed five RNA patterns, with 1-5 discrete dsRNA bands. The molecular sizes corresponding to these bands were 2.2-14.8 kb. Gel electrophoresis of purified virus-like particles (VLPs) indicated only one capsid of similar size in all virus-harbouring strains; when investigated by electron microscopy, they were found to be polyhedral VLPs 40 nm in diameter. In one of the R. microsporus isolates an uncapsidated large dsRNA segment (14.8 kb) was observed. No phenotypic differences were observed between uninfected and virus-harbouring Rhizopus isolates.
Asunto(s)
ARN Bicatenario/aislamiento & purificación , ARN Viral/aislamiento & purificación , Rhizopus/virología , Virus/ultraestructuraRESUMEN
Random amplified polymorphic DNA (RAPD) and isoenzyme polymorphisms among 16 isolates of the postharvest pathogen Gilbertella persicaria were examined. Six different 10-bp primers were used to determine the extent of intraspecific genetic variability. Nine composite amplification types were identified. RAPD markers were obtained which correlated with the mating types of the G. persicaria isolates. The variability of the isoenzyme patterns was very low and no correlation was found between the isoenzyme markers and the mating abilities. When 80 single carbon substrates were tested in utilization assays, most of them were utilized uniformly by the 16 G. persicaria strains. However, some compounds elicited differences between the isolates representing the two mating types. Beta-alanine (0.2%) has little effect on the germination of the sporangiospores of the (+) isolates, but inhibited the germination of (-) sporangiospores. Glycerol-1-monoacetate supported the growth of both mating types, but at concentrations higher than 4% this was accompanied with a compact (colonial) growth for plus mating type isolates only.
Asunto(s)
Mucorales/genética , Carbono/metabolismo , ADN de Hongos/genética , ADN de Hongos/aislamiento & purificación , Variación Genética , Isoenzimas/genética , Mucorales/enzimología , Mucorales/crecimiento & desarrollo , Mucorales/patogenicidad , Plantas Comestibles/microbiología , Polimorfismo Genético , Técnica del ADN Polimorfo Amplificado AleatorioRESUMEN
Contour clamped homogeneous electric field (CHEF) gel electrophoresis was used to obtain electrophoretic karyotypes from nine Mucor strains representing five different species (M. bainieri, M. circinelloides, M. mucedo, M. plumbeus and M. racemosus). The chromosomal banding patterns revealed high variability among the isolates. The sizes of the DNA in the Mucor chromosomes were estimated to be between 2.5 and 8.7 Mb. The total genome sizes were calculated to be between 30.0 and 44.7 Mb. The applicability of these electrophoretic karyotypes for the investigation of genome structure, for strain identification and for species delimitation is considered.
Asunto(s)
Cromosomas Fúngicos , ADN de Hongos/aislamiento & purificación , Mucor/genética , Electroforesis en Gel de Campo Pulsado , Genoma Fúngico , Cariotipificación , Especificidad de la EspecieRESUMEN
For interpretation of intraspecific polymorphism and the considerable differences in the size of mtDNAs among three groups of A. carbonarius, restriction maps were constructed from several enzymes. Functional maps were also developed to compare genome organisations and gene content. The appearance of various mtDNAs of A. carbonarius strains are different in size, but their gene content is almost identical. The 1.1 kb size difference between two closely related subgroups (1a, 1b) can be attributed to the presence or absence of an intron in cox2 gene. This phenomenon demonstrates that the migration of introns is possibly responsible for the development of variable mitochondrial genomes in nature. The striking differences in size and restriction patterns between two main mtDNA groups might derive from both the intronal variations and the altered intergenic organisation.
Asunto(s)
Aspergillus/genética , ADN Mitocondrial/genética , Variación Genética , Aspergillus/clasificación , Secuencia de Bases , Clonación Molecular , Sondas de ADN , ADN de Hongos/genética , Intrones/genética , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Mapeo Físico de Cromosoma , Mapeo Restrictivo , Análisis de Secuencia de ADNRESUMEN
DNA was purified from the isolated mitochondria of a Mucor piriformis wild-type strain (NRRL 26211). A circular restriction map of the mitochondrial DNA was established on the basis of single and double digests with several restriction endonucleases. The average mitochondrial DNA size calculated from these data was 33.53 kbp; this is in good agreement with the contour length size (33.62 kbp) of the open circular molecules detected by electron microscopy. Heterologous hybridizations with cloned Aspergillus nidulans mitochondrial genes were used to locate some coding regions on the map.
Asunto(s)
ADN de Hongos/genética , ADN Mitocondrial/genética , Mitocondrias/genética , Mucor/genética , ADN de Hongos/aislamiento & purificación , ADN de Hongos/ultraestructura , ADN Mitocondrial/aislamiento & purificación , ADN Mitocondrial/ultraestructura , Genoma , Mapeo RestrictivoRESUMEN
Sexual activity was induced in the basidiomyceteous Phaffia rhodozyma (Xanthophyllomyces dendrorhous) by depletion of nitrogen from the culture medium. This activity involved both mating between two yeast cells and the formation of basidiospores. Mating is possibly started by a G1 phase arrest of the cell cycle, as in other yeasts. The life cycle exhibited homothallic features. Crosses between genetically marked strains, and pulse-field gel electrophoresis of the chromosomal DNA of cells derived from individual spores revealed evidence of karyogamy, meiosis and even recombination. The segregation ratio in tetrads pointed to diploid vegetative cells, which formed tetraploid zygotes and the immediate meiosis then gave rise to diploid progenies again. Apart from the type strain Phaffia rhodozyma CBS 5905, all the examined strains were able to sporulate.
Asunto(s)
Basidiomycota/crecimiento & desarrollo , Basidiomycota/genética , Basidiomycota/fisiología , Ciclo Celular , Cruzamientos Genéticos , Medios de Cultivo , ADN de Hongos , Electroforesis en Gel de Campo Pulsado , Fase G1 , Meiosis , Nitrógeno/farmacología , Esporas Fúngicas/aislamiento & purificación , Esporas Fúngicas/fisiologíaRESUMEN
The presence of double-stranded RNA elements was examined in 123 strains representing 18 Mucor species. These genetic elements were found to be present in 6 strains: 1 M. aligarensis, 1 M. hiemalis, 2 M. corticolus, 1 M. mucedo and 1 M. ramannianus. Electrophoretic separation of the nucleic acids revealed 4 different RNA patterns, with 1 to 5 discrete dsRNA bands. The molecular weights corresponding to these bands were 1.42-4.15 x 10(6) D. Using electronmicroscopy, for the first time the presence of virus like particles in Mucor species has been revealed.
Asunto(s)
Mucor/genética , Mucor/virología , Virus ARN/ultraestructura , ARN Bicatenario/aislamiento & purificación , Electroforesis en Gel de Agar , Microscopía Electrónica , Mucor/aislamiento & purificación , Virus ARN/genética , Virus ARN/aislamiento & purificación , ARN Bicatenario/genéticaRESUMEN
Previous mitochondrial transmission experiments between oligomycin-resistant and oligomycin-sensitive incompatible strains of the A. niger aggregate bearing various mtDNA RFLP profiles resulted in a great variety of mitochondrial recombinants under selection pressure. Apart from the recombinant mtDNAs, resistant clones harbouring unchanged RFLP profiles of resistant donor mtDNAs with the recipient nuclear backgrounds were rarely isolated. These strains were anastomosed with nuclearly isogenic oligomycin-sensitive recipient partners and the mitochondria of the resulting progeny were examined under non-selective conditions. These experiments provide insights into events which are possibly similar to those occurring in nature. The heterokaryons obtained formed both oligomycin-resistant and -sensitive sectors, most of which were found to be homoplasmons. Progenies harbouring oligomycin-resistant and -sensitive mtDNAs may originate either from individual recombination events or be due to parental segregation. MtDNA recombination might take place in the heterokaryons without selection by oligomycin. The most frequent recombinant types of mtDNA RFLP profiles were indistinguishable from those recombinant mtDNAs which were frequently obtained under selection pressure from directed transfer experiments between incompatible strains. We present evidence that mixed mitochondrial populations may influence the compatibility reactions in the presence of an isogenic nuclear background, that recombination may take place without selection pressure, and that the process does not require specific nuclear sequences of both parental strains.
Asunto(s)
Aspergillus niger/genética , ADN Mitocondrial/genética , Recombinación Genética/genética , Antibacterianos/farmacología , Aspergillus niger/efectos de los fármacos , Núcleo Celular , Cruzamientos Genéticos , ADN de Hongos/genética , Farmacorresistencia Microbiana/genética , Marcadores Genéticos , Oligomicinas/farmacología , Polimorfismo de Longitud del Fragmento de Restricción , Técnica del ADN Polimorfo Amplificado Aleatorio , Selección GenéticaRESUMEN
Protoplasts of Saccharomyces cerevisiae were inactivated by treatment with different concentrations of antifungal compounds for various periods. Of the 14 compounds tested, N-ethylmaleimide proved to be the most efficient. The inactivation effect was fully reproducible. The inactivated protoplasts could be reactivated and still function as fusion partners. They were fused with untreated protoplasts by polyethylene glycol treatment and produced viable hybrid cells. Nuclear and extrachromosomal genetic analysis and chromosome separation of the fusion products from fusion experiments involving inactivated and non-inactivated protoplasts revealed that N-ethylmaleimide did not affect either of the genomes and hence it was perfectly suited for the hybridization of any type yeast cells without genetic markers.
Asunto(s)
Fusión Celular , Polietilenglicoles/farmacología , Protoplastos/citología , Saccharomyces/citología , Tensoactivos/farmacología , Cruzamientos Genéticos , ADN Mitocondrial/análisis , Electroforesis en Gel de Campo Pulsado , Activación Enzimática , Etilmaleimida/farmacología , Marcadores Genéticos , Hibridación Genética , Cariotipificación , Plásmidos/análisis , Protoplastos/efectos de los fármacos , Protoplastos/enzimología , Saccharomyces/efectos de los fármacos , Saccharomyces/enzimología , Saccharomyces/genética , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/enzimología , Saccharomyces cerevisiae/genética , Especificidad de la EspecieRESUMEN
Electrophoretic karyotypes of 80 auxotrophic and morphological mutants obtained from two Phaffia rhodozyma strains (ATCC 24203 and ATCC 24229) by gamma-radiation were investigated. Contour-clamped homogeneous gel electrophoresis separation of the chromosomal size DNAs revealed 29 new chromosomal patterns after mutagen treatment. No correlation was found between a given type of chromosomal aberration and any phenotypic character. However, analysis of the chromosomal rearrangements proved to be useful for a more exact determination of chromosome number and genome size. The total genome size of ATCC 24229 was found to be 19.3 Mb, with nine chromosomes, while analysis of the mutant derivatives of ATCC 24203 suggested the presence of 11 chromosomes, with an estimated total genome size of 22.2 Mb. The advantages of the analysis of mutant electrophoretic karyotypes for genome characterization are discussed.
Asunto(s)
Cromosomas Fúngicos/genética , Rayos gamma , Cariotipificación/métodos , Hongos Mitospóricos/genética , Cromosomas Fúngicos/química , Cromosomas Fúngicos/efectos de la radiación , Electroforesis en Gel de Campo Pulsado/métodos , Peso MolecularRESUMEN
The present survey demonstrates polymorphism in both the length and the number of double-stranded RNAs (dsRNAs) among six Phaffia rhodozyma strains. Strains with one-, three- and four-types of dsRNA molecules were found, while two strains proved to be dsRNA-free. Elongated icosahedral virus-like particles (VLPs) 34x26 nm in size were detected in strains carrying four- or three-types of dsRNAs. One 3.7-kb dsRNA molecule was found not to form part of the VLP genome. Transmission of the VLPs of strain ATCC 24203 was followed through the basidiospores during the sexual cycle. Cytoplasmic inheritance was observed.
Asunto(s)
Basidiomycota/genética , Cuerpos de Inclusión Viral/genética , ARN Bicatenario/genética , Basidiomycota/fisiología , Electroforesis , Herencia Extracromosómica , Cuerpos de Inclusión Viral/química , Cuerpos de Inclusión Viral/ultraestructura , Microscopía Electrónica , Fenotipo , Polimorfismo Genético , ARN Bicatenario/efectos de los fármacos , ARN de Hongos/genética , ARN Viral/genética , Esporas Fúngicas/genéticaRESUMEN
Sixty-one isolates and collection strains of Aspergillus fumigatus were compared for their phenotypic (morphological features and isoenzyme profiles) and genotypic (restriction enzyme-generated mitochondrial DNA and ribosomal DNA profiles and random amplified polymorphic DNA patterns) features. The examined strains exhibited highly variable colony morphologies and growth rates at different temperatures, but their micromorphologies and conidial diameters were characteristic of the species. Of the isoenzymes studied, the beta-arylesterase and phosphatase patterns were the most divergent, and the 61 strains could be classified into seven groups. The glucose 6-phosphate dehydrogenase and catalase isoenzyme patterns displayed only a limited variability, while the profiles of superoxide dismutase, lactate dehydrogenase, and glutamate dehydrogenase were highly conserved. The HaeIII-generated mitochondrial DNA patterns and SmaI-digested repetitive DNA and ribosomal DNA hybridization patterns of almost all strains were also invariable. The level of variation was much higher when random amplified polymorphic DNA analysis was applied. Although the patterns of the strains were very similar with most of the primers, the application of some primers made it possible to cluster the A. fumigatus isolates into several groups. The results indicate that the random amplified polymorphic DNA technique could be used more efficiently than isoenzyme analysis for typing A. fumigatus isolates. A good correlation was found between the dendrograms obtained from the isoenzyme and random amplified polymorphic DNA data, but the isoenzyme and amplified DNA patterns did not correlate with the pathogenicity, pigment production, or geographical origin of the strains. One "A. fumigatus" strain (strain FRR 1266) exhibited unique isoenzyme, mitochondrial DNA, ribosomal DNA, and random amplified polymorphic DNA patterns; it is proposed that this strain represents a new species of the section Fumigati.
Asunto(s)
Aspergillus fumigatus/clasificación , Variación Genética , Aspergillus fumigatus/citología , Aspergillus fumigatus/enzimología , Aspergillus fumigatus/genética , Secuencia de Bases , Genotipo , Isoenzimas , Datos de Secuencia Molecular , Fenotipo , Polimorfismo de Longitud del Fragmento de Restricción , Técnica del ADN Polimorfo Amplificado AleatorioRESUMEN
Chromosome-substituted haploid segregants of an A. nidulans x A. tetrazonus somatic hybrid were used to allocate several random amplified polymorphic DNA and isoenzyme markers to parental chromosomes. Twenty-six amplified DNA fragments, and nine isoenzyme activities, including lactate dehydrogenase, superoxide dismutase, and arylesterase isoenzymes were assigned to chromosomes. Chromosomes-specific markers were found for each A. nidulans and A. tetrazonus chromosome. These markers could be used to saturate the genetic map of A. nidulans. The formation of two secondary metabolites was also assigned to chromosomes III and VIII. Attempts were made to allocate extracellular enzyme activities to parental chromosomes, mostly without success, possibly because multiple enzyme forms located on different chromosomes could be responsible for the production of an enzyme activity.
Asunto(s)
Aspergillus nidulans/genética , Aspergillus/genética , Mapeo Cromosómico , Cromosomas Fúngicos , Técnica del ADN Polimorfo Amplificado Aleatorio , Aspergillus/enzimología , Aspergillus nidulans/enzimología , Secuencia de Bases , Proteínas Fúngicas/genética , Marcadores Genéticos , Isoenzimas/genética , Datos de Secuencia Molecular , Especificidad de la EspecieRESUMEN
The validity of the species concept was examined with strains of Phaffia rhodozyma by comparing the isoenzyme profiles, ribosomal DNAs, and random amplified polymorphic DNA (RAPD) patterns of the strains. The isoenzyme profiles appeared to be more stable than the RAPD patterns or the electrophoretic karyotypes determined previously (A. Nagy, N. Garamszegi, C. Vágvölgyi, and L. Ferenczy, FEMS Microbiol. Lett., in press). The ribosomal DNA patterns revealed only a limited degree of variability, while RAPD analysis proved to be the most useful method for differentiating the strains studied. Strain CBS 5905T (T = type strain) produced characteristic RAPD patterns, which were different from those produced by the other strains. However, despite the high degree of variability observed, the isoenzyme data and the slightly variable ribosomal DNA hybridization profiles confirmed that the strains which we examined belong to one species.
Asunto(s)
ADN de Hongos/análisis , Amplificación de Genes , Isoenzimas/análisis , Polimorfismo de Longitud del Fragmento de Restricción , Levaduras/clasificación , Secuencia de Bases , Datos de Secuencia Molecular , Levaduras/enzimología , Levaduras/genéticaRESUMEN
Contour-clamped homogeneous electric field (CHEF) gel electrophoresis was used to separate chromosomal size DNA molecules of two Mucor circinelloides strains. Electrophoretic karyotypes revealed the presence of eight distinct bands for the M. circinelloides f. lusitanicus strain, and four, presumably multiple, bands for the M. circinelloides f. gryseo-cyanus strain. The approximate sizes of the resolved chromosomal DNA bands ranged from 2.3 to 8.1 Mb, giving estimated genome sizes of 38.7 and 32.6 Mb, respectively. Hybridisation techniques were used to assign the leuA gene to a chromosome.
Asunto(s)
ADN de Hongos/genética , Electroforesis en Gel de Campo Pulsado , Cariotipificación , Mucor/genética , Cromosomas Fúngicos , Genes FúngicosRESUMEN
The abilities of 14 phenothiazines, 8 benzo[a]phenothiazines and 12 benz[c]acridines to induce an antibacterial effect against Escherichia coli K12 were compared. Several phenothiazines, which showed antiplasmid activity, displayed the most potent antibacterial activity. All benz[c]acridine derivatives were moderately antibacterial, whereas benzo[a]phenothiazines were inactive. The active phenothiazine derivatives had more potent inhibitory activity against fungi, including phytopathogen filamentous, human pathogen filamentous fungi and yeasts, than against gram-positive and -negative bacteria. Taken together with previously reported data, the induction mechanism of antimicrobial and antiplasmid activity by these compounds seems to be different from that of antitumor, differentiation-inducing and carcinogenic activity.
Asunto(s)
Acridinas/farmacología , Escherichia coli/efectos de los fármacos , Hongos/efectos de los fármacos , Fenotiazinas/farmacología , Acridinas/química , Pruebas de Sensibilidad Microbiana , Fenotiazinas/química , Relación Estructura-ActividadRESUMEN
Nuclei were isolated from protoplasts of a haploid auxotrophic Aspergillus nidulans strain. Transformation of protoplasts prepared from a complementary haploid auxotrophic strain with these purified nuclei resulted in both heterokaryotic and diploid colonies. The nutritionally-complementing colonies appeared at a frequency of 5 x 10(-7) to 10(-8).