Search results for "Chain reaction"
showing 10 items of 1862 documents
Comparison of different primer sets for use in Automated Ribosomal Intergenic Spacer Analysis of complex bacterial communities.
2004
ABSTRACT ITSF and ITSReub, constituting a new primer set designed for the amplification of the 16S-23S rRNA intergenic transcribed spacers, have been compared with primer sets consisting of 1406F and 23Sr (M. M. Fisher and E. W. Triplett, Appl. Environ. Microbiol. 65:4630-4636, 1999) and S-D-Bact-1522-b-S-20 and L-D-Bact-132-a-A-18 (L. Ranjard et al., Appl. Environ. Microbiol. 67:4479-4487, 2001), previously proposed for automated ribosomal intergenic spacer analysis (ARISA) of complex bacterial communities. An agricultural soil and a polluted soil, maize silage, goat milk, a small marble sample from the façade of the Certosa of Pavia (Pavia, Italy), and brine from a deep hypersaline anoxi…
Shifts in diversity and microscale distribution of the adapted bacterial phenotypes due to Hg(II) spiking in soil.
2003
In a previous experiment [Ranjard et al. (2000) FEMS Microbiol Ecol 31:107–115], the spatial heterogeneity of a mercury impact on soil bacterial community was revealed by an increase of mercury-resistant (HgR) bacterial numbers in the outer fraction and the sand fractions when compared to those in the silt fractions. The objectives of the present study were (i) to investigate whether mercury exposure affects the diversity and the distribution within the various fractions of the HgR populations and (ii) to evaluate the contribution of the HgR populations to the overall community adaptation. A total of 236 strains isolated before (104 isolates) and 30 days (132 isolates) after spiking were ch…
Characterisation of rpsL, rrs and embB mutations associated with streptomycin and ethambutol resistance in Mycobacterium tuberculosis.
2003
In order to characterise molecular mechanisms of first-line drug resistance in Mycobacterium tuberculosis and to evaluate the use of molecular markers of resistance (gene point mutations), we analysed 66 multi-drug-resistant (MDR) isolates from Latvian tuberculosis patients. They were all resistant to rifampin (RIF), isoniazid (INH) and streptomycin (SM), and 33 were resistant to ethambutol (EMB). Enzymatic digestion by MboII and nucleotide sequencing of the rpsL gene fragment detected a single nucleotide substitution K43R in 40 (61%) of the 66 SM-resistant M. tuberculosis isolates. Of the other 26 SM-resistant isolates, 16 (24%) had mutations at positions 513A--C and 516C--T of the rrs gen…
Use of nodulation pattern, stress tolerance, nodC gene amplification, RAPD-PCR and RFLP-16S rDNA analysis to discriminate genotypes of Rhizobium legu…
2005
Twenty-seven new Rhizobium isolates were obtained from root nodules of wild and crop legumes belonging to the genera Vicia, Lathyrus and Pisum from different agroecological areas in central and southern Italy. A polyphasic approach including phenotypic and genotypic techniques was used to study their diversity and their relationships with other biovars and species of rhizobia. Analysis of symbiotic properties and stress tolerance tests revealed that wild isolates, showed a wide spectrum of nodulation and a marked variation in stress tolerance compared with reference strains tested in this study. All rhizobial isolates (except for the isolate CG4 from Galega officinalis) were presumptively i…
Polyphasic taxonomic characterization of Lactobacillus rossiae isolates from Belgian and Italian sourdoughs reveals intraspecific heterogeneity.
2009
Abstract (GTG) 5 -PCR fingerprinting and pheS sequence analysis of 18 Lactobacillus rossiae isolates, mainly originating from Belgian and Italian artisan sourdoughs, revealed intraspecies grouping as evidenced by the delineation of three and two subgroups, respectively. On the other hand, 16S rRNA and rpoA gene sequence analysis and DNA–DNA hybridizations supported the accommodation of all isolates in a single species. No correlation between genetic and phenotypic heterogeneity was observed. Collectively, these data do not warrant taxonomic division of L. rossiae . On the other hand, the considerable differences in intraspecies sequence variation of L. rossiae isolates displayed by the pheS…
A multiplex RTi-PCR reaction for simultaneous detection of Escherichia coli O157:H7, Salmonella spp. and Staphylococcus aureus on fresh, minimally pr…
2007
In this work, a new multiplex single-tube real-time PCR approach is presented for the detection of Escherichia coli O157:H7, Salmonella spp. and Staphylococcus aureus, three of the more frequent food-borne bacterial pathogens that are usually investigated in a variety of food matrices. The study includes the design and specificity testing, of a new primer and probe specific for Salmonella spp. Reaction conditions were adjusted for the simultaneous amplification and detection of specific fragments in the beta-glucuronidase (uidA, E. coli) and Thermonulease (nuc, Sta. aureus) genes, and in the replication origin sequence (oriC, Salmonella spp.). Melting-curve analysis using a SYBR Green I RTi…
Use of a species-specific multiplex PCR for the identification of pediococci.
2008
In this study, the 23S rRNA genes of nine different Pediococcus type strains were sequenced. By using a multiple sequence alignment with 23S rDNA sequences of related lactic acid bacteria two primer pairs were constructed, one for the general identification of the genus Pediococcus and one for the identification of the atypical species, P. dextrinicus. Furthermore, a primer set for a rapid multiplex PCR identification of the eight typical Pediococcus species was developed. With this technique, the species P. damnosus, P. parvulus, P. inopinatus, P. cellicola, P. pentosaceus, P. acidilactici, P. claussenii, and P. stilesii could be discriminated simultaneously in a single PCR. Experiments wi…
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…
Development of a real-time PCR assay for detection and quantification of enterotoxigenic members of Bacillus cereus group in food samples
2009
A highly sensitive real-time PCR (qPCR) procedure, targeting the phosphatidylcholine-specific phospholipase C gene (pc-plc), was developed for specific detection and quantification of strains belonging to Bacillus cereus group. The target region was selected based on the enterotoxigenic profiles of 75 Bacillus strains. The inclusivity and exclusivity of the RTi-PCR assay were assessed with 59 isolates of the B. cereus group, 16 other Bacillus spp., and 4 non-Bacillus strains. The assay was also used to construct calibration curves for different food matrices, and it had a wide quantification range of 6 log units using both serial dilutions of purified DNA and calibrated cell suspensions of …
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 …