Search results for "Real-Time Polymerase Chain Reaction"
showing 10 items of 385 documents
Identification and Detection of Phoma tracheiphila, Causal Agent of Citrus Mal Secco Disease, by Real-Time Polymerase Chain Reaction.
2006
Phoma tracheiphila is the causal agent of a tracheomycotic disease of citrus called mal secco causing the dieback of twigs and branches. This pathogen is of quarantine concern; therefore, fast and reliable protocols are required to detect it promptly. A specific primer pair and a dual-labeled fluorogenic probe were used in a real-time polymerase chain reaction (PCR) with the Cepheid Smart Cycler II System (Transportable Device TD configuration) to detect this fungus in citrus samples. Real-time PCR assay was compared to modified conventional PCR assay. The sensitivity of the former was evaluated by testing P. tracheiphila DNA dilutions, and the minimum amount detectable was about 500 fg, wh…
Phenotypic and genotypic evaluation of slime production by conventional and molecular microbiological techniques.
2009
Twenty-nine staphylococcal isolates from different clinical samples were tested for slime production: phenotypic characterization was carried out using Christensen test (tube test) and Congo red agar plate test (CRA plate test), while the presence and expression of icaA and icaD genes were evaluated by real-time PCR. In 79.3% of studied strains there was a concordance between slime production and presence of icaA and icaD genes, and between lack of slime production and absence of both or only one of the tested genes. In four of five strains where positive phenotype was not associated with the presence of ica genes, gene co-expression (evaluated by mRNA determination) was lacking, while in o…
Comparative evaluation of molecular methods for the quantitative measure of torquetenovirus (TTV) viremia, the new surrogate marker of immune compete…
2019
Background Torquetenovirus (TTV) viremia is emerging as a promising tool to assess functional immune competence, to predict posttransplant immune-related complications, and eventually to customize immunosuppression. Methods In this study, 327 blood samples were tested using two real-time PCR (rtPCR) assays both targeted to the untranslated region of the TTV genome. The first assay was an in-house rtPCR developed by our group, the second one was the recently marketed TTV R-GENE assay. Results In the validation study, the TTV R-GENE showed good performances in precision and reproducibility, and sensitivity as low as 12 TTV DNA copies/mL, like previously reported for the in-house rtPCR. The Bl…
Differential expression of Cryptosporidium parvum genes encoding sporozoite surface antigens in infected HCT-8 host cells.
2006
Intracellular replication of Cryptosporidium parvum (Apicomplexa) involves the generation of several asexual and sexual forms of the parasite. During the stage conversions, complex mechanisms lead to differential structural and functional properties of the parasite. These require a well tuned gene transcription machinery. For the first time the gene expression of four surface proteins of C. parvum sporozoites, CP15, CP17, P23, and GP900 were analysed in parallel by reverse transcription polymerase chain reaction. In addition, CP17 and P23 antigens were detected in infected host cells by immunofluorescence using antisera raised against recombinant forms of the proteins. The results show that…
Dynamic changes in the subcellular distribution of the tobacco ROS-producing enzyme RBOHD in response to the oomycete elicitor cryptogein.
2014
Highlight text The oomycete elicitor cryptogein triggers the relocation of RBOHD from intracellular compartments to the plasma membrane in tobacco cells. This suggests that intracellular trafficking is a potential determinant of RBOHD activity.
Aneuploid IMR90 cells induced by depletion of pRB, DNMT1 and MAD2 show a common gene expression signature
2019
Chromosome segregation defects lead to aneuploidy which is a major feature of solid tumors. How diploid cells face chromosome mis-segregation and how aneuploidy is tolerated in tumor cells are not completely defined yet. Thus, an important goal of cancer genetics is to identify gene networks that underlie aneuploidy and are involved in its tolerance. To this aim, we induced aneuploidy in IMR90 human primary cells by depleting pRB, DNMT1 and MAD2 and analyzed their gene expression profiles by microarray analysis. Bioinformatic analysis revealed a common gene expression profile of IMR90 cells that became aneuploid. Gene Set Enrichment Analysis (GSEA) also revealed gene-sets/pathways that are …
dltA overexpression: A strain-independent keystone of daptomycin resistance in methicillin-resistant Staphylococcus aureus
2013
The mechanisms leading to reduced susceptibility to daptomycin (DAP) are multifactorial and have not been fully elucidated. We analysed, by sequencing and expression studies, the role of the major molecular targets (cell-envelope charge genes, dltA, mprF, cls2; cell-wall turnover and autolysis genes, sceD, atl) involved in the emergence of DAP resistance in three series of isogenic clinical methicillin-resistant Staphylococcus aureus (MRSA) in which DAP resistance emerged after a heterogeneous glycopeptide-intermediate S. aureus (hGISA) step under teicoplanin and DAP therapy. All of the isolates had different genotypes and were delta-haemolysin negative, reflecting a strain proclivity to ac…
Quantification of a novel group of nitrate-reducing bacteria in the environment by real-time PCR
2004
Abstract Nitrate reduction is performed by phylogenetically diverse bacteria. Analysis of narG (alpha subunit of the membrane bound nitrate reductase) trees constructed using environmental sequences revealed a new cluster that is not related to narG gene from known nitrate-reducing bacteria. In this study, primers targeting this as yet uncultivated nitrate-reducing group were designed and used to develop a real-time SYBR® Green PCR assay. The assay was tested with clones from distinct nitrate-reducing groups and applied to various environmental samples. narG copy number was high ranging between 5.08×108 and 1.12×1011 copies per gram of dry weight of environmental sample. Environmental real-…
pilF polymorphism-based real-time PCR to distinguish Vibrio vulnificus strains of human health relevance
2012
The Gram-negative bacterium Vibrio vulnificus is a common inhabitant of estuarine environments. Globally, V. vulnificus is a significant foodborne pathogen capable of causing necrotizing wound infections and primary septicemia, and is a leading cause of seafood-related mortality. Unfortunately, molecular methods for the detection and enumeration of pathogenic V. vulnificus are hampered by the genetically diverse nature of this pathogen, the range of different biotypes capable of infecting humans and aquatic animals, and the fact that V. vulnificus contains pathogenic as well as non-pathogenic variants. Here we report an alternative approach utilizing the development of a real-time PCR assay…
A TaqMan-based real-time PCR assay for the specific detection and quantification ofLeuconostoc mesenteroidesin meat products
2007
A new real-time PCR procedure was developed for the specific detection and quantification of Leuconostoc mesenteroides in meat products. It is a TaqMan assay based on 23S rRNA gene targeted primers and probe. Specificity was evaluated using purified DNA from 132 strains: 102 lactic acid bacteria (LAB), including 57 reference strains and 46 food isolates, belonging to genus Leuconostoc and related genera, and 30 non-LAB strains. Quantification was linear over at least 5 log units using both serial dilutions of purified DNA and calibrated cell suspensions from Leuconostoc mesenteroides ssp. dextranicum CECT 912T. This assay was able to detect at least five genomic equivalents, using purified …