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At least 937 records · Page 52Linked to original sources
PubMed2004

Identification of genes preferentially expressed in the pathogenic yeast phase of Paracoccidioides brasiliensis, using suppression subtraction hybridization and differential macroarray analysis.

Paracoccidioides brasiliensis, a thermodimorphic fungus, is the causative agent of paracoccidioidomycosis (PCM), the most prevalent systemic mycosis in Latin America. Pathogenicity appears to be intimately related to the dimorphic transition from the hyphal to the yeast form, which is induced by a shift from environmental temperature to the temperature of the mammalian host. Little information is available on the P. brasiliensis genes that are necessary during the pathogenic phase. We have therefore undertaken Suppression Subtraction Hybridization (SSH) and macroarray analyses with the aim of identifying genes that are preferentially expressed in the yeast phase. Genes identified by both procedures as being more highly expressed in the yeast phase are involved in basic metabolism, signal transduction, growth and morphogenesis, and sulfur metabolism. In order to test whether the observed changes in gene expression reflect the differences between the growth conditions used to obtain the two morphological forms rather than differences intrinsic to the cell types, we performed real-time RT-PCR experiments using RNAs derived from both yeast cells and mycelia that had been cultured at 37 degrees C and 26 degrees C in either complete medium (YPD or Sabouraud) or minimal medium. Twenty genes, including AGS1 (alpha-1,3-glucan synthase) and TSA1 (thiol-specific antioxidant), were shown to be more highly expressed in the yeast cells than in the hyphae. Although their levels of expression could be different in rich and minimal media, there was a general tendency for these genes to be more highly expressed in the yeast cells.

DNA, Complementary↗
PubMed2001

Molecular characterization of field isolates of human pathogenic trypanosomes.

The accurate identification of each of the three subspecies of Trypanosoma brucei remains a challenging problem in the epidemiology of sleeping sickness. Advances in molecular characterization have revealed a much greater degree of heterogeneity within the species than previously supposed. Only group 1 T. b. gambiense stands out as a separate entity, defined by several molecular markers. T. b. rhodesiense is generally too similar to sympatric T. b. brucei strains to be distinguished from them by any particular molecular markers. Nevertheless, characterization of trypanosome isolates from humans and other animals has allowed the identification of potential reservoir hosts of T. b. rhodesiense. The recent discovery of a gene for human serum resistance may provide a useful marker for T. b. rhodesiense in the future. There have been few attempts to find associations between genetic markers and other biological characters, except human infectivity. However, virulence or fly transmissibility have been correlated with molecular markers in some instances.

Africa↗
PubMed2005

Rapid identification and strain-typing of respiratory pathogens for epidemic surveillance.

Epidemic respiratory infections are responsible for extensive morbidity and mortality within both military and civilian populations. We describe a high-throughput method to simultaneously identify and genotype species of bacteria from complex mixtures in respiratory samples. The process uses electrospray ionization mass spectrometry and base composition analysis of PCR amplification products from highly conserved genomic regions to identify and determine the relative quantity of pathogenic bacteria present in the sample. High-resolution genotyping of specific species is achieved by using additional primers targeted to highly variable regions of specific bacterial genomes. This method was used to examine samples taken from military recruits during respiratory disease outbreaks and for follow up surveillance at several military training facilities. Analysis of respiratory samples revealed high concentrations of pathogenic respiratory species, including Haemophilus influenzae, Neisseria meningitidis, and Streptococcus pyogenes. When S. pyogenes was identified in samples from the epidemic site, the identical genotype was found in almost all recruits. This analysis method will provide information fundamental to understanding the polymicrobial nature of explosive epidemics of respiratory disease.

Bacteria↗
PubMed1992

Identification of multiple structural domains regulating viroid pathogenicity.

To investigate the role of individual structural domains in viroid pathogenicity and replication, a series of interspecific chimeras was constructed by exchanging the terminal left (TL) and/or pathogenicity (P) domains between tomato apical stunt (TASVd) and citrus exocortis (CEVd) viroids. All six chimeras tested were replicated stably in tomato, and the symptoms exhibited by infected plants were intermediate between those induced by the parental viroids. Quantitative comparisons of symptom development and progeny accumulation revealed that: (i) the TL domain of TASVd contains a determinant required for appearance of severe veinal necrosis in tomato, (ii) the severe epinasty and stunting characteristic of TASVd requires the presence of its TL and P domains, and (iii) the variable (V) and terminal right (TR) domains comprising the right side of the native structure also play an important role in viroid pathogenicity. Chimeras containing the right side of TASVd accumulated to higher levels early in infection, and infected plants developed more severe symptoms than those whose right halves were derived from CEVd. Although the individual contributions of the TL and P domains to symptom induction could not be completely separated from that of viroid titer, the TL domain appears to exert a greater effect upon symptom severity than does the P domain. The TL, P, V, and TR domains of TASVd and CEVd contain three discrete regions of sequence and/or structural variability that may correspond to the pathogenicity determinants uncovered by our genetic analysis.

Base Sequence↗
PubMed2001

Identification of DNA sequences from a second pathogenicity island of uropathogenic Escherichia coli CFT073: probes specific for uropathogenic populations.

Uropathogenic Escherichia coli is the leading cause of urinary tract infection and hospital visits in North America. Cystitis and acute pyelonephritis, infection of the bladder and kidney, respectively, are the two most common syndromes encountered in patients with urinary tract infection. We sequenced and annotated 71,684 bases of a previously unidentified pathogenicity-associated island (PAI) from E. coli strain CFT073. This PAI contained 89 open-reading frames encoding a pap operon, iron-regulated genes, mobile genetic elements, and a large proportion of unknown or unidentified open-reading frames. Dot blot analysis with 11 DNA sequences from this PAI demonstrated that 7 sequences were more prevalent among uropathogens: 2 probes were more prevalent among cystitis and pyelonephritis isolates, 2 among pyelonephritis isolates only, and 3 among cystitis isolates only than among fecal isolates. These data suggest that groups of uropathogens have genetic differences that may be responsible for the different clinical outcomes.

Bacterial Proteins↗
PubMed2001

Identification of virulence mutants of the fungal pathogen Cryptococcus neoformans using signature-tagged mutagenesis.

Cryptococcus neoformans var. neoformans is an important opportunistic fungal pathogen of patients whose immune system has been compromised due to viral infection, antineoplastic chemotherapy, or tissue transplantation. As many as 13% of all AIDS patients suffer a life-threatening cryptococcal infection at some time during the course of their HIV disease. To begin to understand the molecular basis for virulence in Cryptococcus neoformans var. neoformans serotype A, we have employed signature-tagged mutagenesis (STM) to identify mutants with altered virulence in a mouse model. The critical parameters of signature-tagged mutagenesis in C. neoformans are explored. Data are presented showing that at least 100 different strains can be mixed together in a single animal with each participating in the infection and that there is no apparent interaction between a virulent strain and an avirulent strain in our animal model. Using signature-tagged mutagenesis, we identified 39 mutants with significantly altered growth in a competitive assay. Molecular analyses of these mutants indicated that 19 (49%) contained an insertion in the actin promoter by homologous recombination from a single crossover event, creating a duplication of the actin promoter and the integration of single or multiple copies of the vector. Analysis of the chromosomal insertion sites of those mutants that did not have an integration event in the actin promoter revealed an approximately random distribution among the chromosomes. Individual challenge of the putative mutants in a mouse model revealed five hypovirulent mutants and one hypervirulent mutant.

Actins↗
PubMed1995

Identification and characterization of specific sequences encoding pathogenicity associated proteins in the genome of commensal Neisseria species.

The distribution of distinct sequences in pathogenic and commensal Neisseria species was investigated systematically by dot blot analysis. Probes representing the genes of Rmp, pilin and IgA1 protease were found to hybridize exclusively to the chromosomal DNA of the pathogenic species, Neisseria gonorrhoeae and/or Neisseria meningitidis. In contrast, specific sequences for the genes of the porin protein Por and the opacity protein (Opa) were also detected in a panel of commensal Neisseria species such as N. lactamica, N. subflava, N. flava, N. mucosa and N. sicca. Using opa-specific oligonucleotides as probes in chromosomal blots, the genomes of the commensal Neisseria species show a totally reduced repertoire of cross-hybridizing loci compared to the complex opa gene family of N. gonorrhoeae. DNA sequence analysis of one opa-related gene derived from N. flava and N. sicca, respectively, revealed a large degree of homology with previously described gonococcal and meningococcal genes, e.g., a typical repetitive sequence in the leader peptide and the distribution of the hypervariable and conserved regions. This observation, together with the finding, that the gene is constitutively transcribed, leads to the assumption that some of the commensal Neisseria species may have the potential for the expression of a protein harboring similar functions as the Opa proteins in pathogenic Neisseriae.

Amino Acid Sequence↗
PubMed2004

Simultaneous detection and identification of common cell culture contaminant and pathogenic mollicutes strains by reverse line blot hybridization.

We have developed a reverse line blot (RLB) hybridization assay to detect and identify the commonest mollicutes causing cell line contamination (Mycoplasma arginini, Mycoplasma fermentans, Mycoplasma hyorhinis, Mycoplasma orale, and Acholeplasma laidlawii) and human infection (Mycoplasma pneumoniae, Mycoplasma hominis, Mycoplasma genitalium, Ureaplasma parvum, and Ureaplasma urealyticum). We developed a nested PCR assay with "universal" primers targeting the mollicute 16S-23S rRNA intergenic spacer region. Amplified biotin-labeled PCR products were hybridized to membrane-bound species-specific oligonucleotide probes. The assay correctly identified reference strains of 10 mollicute species. Cell cultures submitted for detection of mollicute contamination, clinical specimens, and clinical isolates were initially tested by PCR assay targeting a presumed mollicute-specific sequence of the 16S rRNA gene. Any that were positive were assessed by the RLB assay, with species-specific PCR assay as the reference method. Initially, 100 clinical and 88 of 92 cell culture specimens gave concordant results, including 18 in which two or more mollicute species were detected by both methods. PCR and sequencing of the 16S-23S rRNA intergenic spacer region and subsequent retesting by species-specific PCR assay of the four cell culture specimens for which results were initially discrepant confirmed the original RLB results. Sequencing of amplicons from 12 cell culture specimens that were positive in the 16S rRNA PCR assay but negative by both the RLB and species-specific PCR assays failed to identify any mollicute species. The RLB hybridization assay is sensitive and specific and able to rapidly detect and identify mollicute species from clinical and cell line specimens.

Acholeplasma↗
PubMed1973

Potential pathogens in the environment: isolation, enumeration, and identification of seven genera of intestinal bacteria associated with small green pet turtles.

Bacteriological analyses were performed on fecal swabs and the aquarium water of 27 individually purchased specimens of the small green pet turtle, Pseudemys scripta elegans. Representatives of Aeromonas, Citrobacter, Enterobacter, Klebsiella, Proteus, Salmonella, and Serratia were isolated. Enterobacter, Klebsiella, and Salmonella were encountered in 20% or more of the specimens, whereas Aeromonas was isolated from 63%. Klebsiella pneumoniae counts ranged from 10(3) to 10(4) per milliliter of aquarium water, whereas Aeromonas routinely exceeded 10(4) per milliliter. Aeromonas cultures from turtles were identical to 7 human isolates in some 29 biochemical tests. On the basis of our findings, we question whether the Salmonella-free certification program alone is sufficient to render these reptiles as safe pets.

Aeromonas↗
PubMed1995

Identification of a new intestinal spirochete with pathogenicity for chickens.

Two intestinal spirochete isolates obtained from chickens with diarrhea were examined by electron microscopy, biochemical tests, rRNA gene restriction pattern analysis, and multilocus enzyme electrophoresis. One isolate (strain 91-1207/C1) was pathogenicity tested in vivo in chickens. The chicken spirochetes were morphologically indistinguishable from Serpulina innocens and Serpulina hyodysenteriae and phenotypically similar to S. innocens. However, the chicken spirochetes could be distinguished from S. innocens, S. hyodysenteriae, and other swine intestinal spirochetes by rRNA gene restriction pattern analysis and multilocus enzyme electrophoresis. In pathogenicity tests in 1-day-old chicks and 14-month-old hens, chicken spirochete 91-1207/C1 produced pale-yellow, watery cecal contents and mild lymphocytic typhlitis. These findings support the conclusion that avian intestinal spirochetes can be pathogenic to commercial poultry and that the microorganisms are different from intestinal spirochetes that infect pigs.

Animals↗
PubMed1988

New plate medium for screening and presumptive identification of gram-negative urinary tract pathogens.

A new selective, differential plating medium to screen the common gram-negative urinary tract pathogens is described. The medium combines adonitol fermentation, phenylalanine deaminase, and beta-glucuronidase tests and allows the indole and cytochrome oxidase tests to be performed directly from the plates. High-level agreement with individual conventional tests was recorded in comparative studies with 504 cultures of gram-negative rods. There was 100% agreement, except for the Providencia spp. indole spot test (61.6% agreement). Adonitol fermentation by Providencia species could not be determined. Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, and Pseudomonas aeruginosa were identified with a high efficiency (100, 85.7, 83.5, and 100% agreement, respectively) without further testing. There was 96% overall agreement for the 267 infected urine samples tested.

Bacteriuria↗
PubMed1998

Rapid identification of Candida albicans and other human pathogenic yeasts by using short oligonucleotides in a PCR.

A PCR system that can quickly and accurately identify 14 species of human pathogenic yeasts was developed. The procedure distinguished between nine species of a closely related clade, Lodderomyces elongisporus, Candida parapsilosis, a new Candida sp., C. sojae, C. tropicalis, C. maltosa, C. viswanathii, C. albicans, and C. dubliniensis and between another five more divergent species, Pichia guilliermondii, C. glabrata, C. zeylanoides, C. haemulonii, and C. haemulonii type II. A rapid DNA extraction procedure that yields purified DNA in about 1 h is also described. The system uses uniform conditions with four primers for each reaction, two 40- to 50-mer universal primers that serve as a positive control and two 23- to 30-mer species-specific primers. Species-specific primers were derived from a 600-nucleotide variable region (D1/D2) at the 5' end of the large-subunit (26S) ribosomal DNA gene and were generally designed to use mismatches at the 3' end. Universal primers were developed from conserved nucleotide sequences in the small-subunit (18S) rRNA gene. In this system, a control 1,200- to 1,300-base DNA fragment was produced in all reactions and a smaller 114- to 336-base DNA fragment was produced if the chromosomal DNA from the target species was present. The PCR procedure is rapid and easy to interpret and may be used with mixed cultures.

Candida↗
PubMedDate not supplied

Molecular identification of the turf grass rapid blight pathogen.

Rapid blight is a newly described disease on turf grasses, primarily found on golf courses using suboptimal water for irrigation purposes. On the basis of shared morphological characteristics, it has been proposed that the rapid blight pathogen belongs to a genus of stramenopiles, Labyrinthula, which had been known to cause disease of marine plants only. We have collected 10 isolates from four species of turf grass in five states and sequenced portions of the SSU (18S) rDNA gene from each to provide a definitive taxonomic placement for rapid blight pathogens. We also included sequences from Labyrinthuloides yorkensis, Schizochytrium aggregatum, Aplanochytrium sp., Thraustochytrium striatum, Achlya bisexualis and several nonturf-grass isolates of Labyrinthula. We found that rapid blight isolates indeed are placed firmly within the genus Labyrinthula and that they lack detectable genetic diversity in the 18S rDNA region. We propose that the rapid blight pathogens share a recent common ancestor and might have originated from a single, infected population.

Genes, rRNA↗