Detection ofBacillus sphaericusmosquitocidal toxin genes by multiplex colony PCR

2009 ◽  
Vol 55 (2) ◽  
pp. 207-209 ◽  
Author(s):  
Santosh C. Jagtap ◽  
Chandrakant B. Jagtap ◽  
Pradeep Kumar ◽  
R. B. Srivastava
Keyword(s):  
Bacillus Sphaericus ◽  
Pcr Assay ◽  
Single Tube ◽  
Colony Pcr ◽  
Toxin Genes ◽  
Specific Pcr ◽  
Genes Encoding ◽  
Single Tube Reaction

A multiplex colony PCR assay was developed for the detection of 5 genes encoding Bacillus sphaericus mosquito larvicidal toxins, namely binA, binB, mtx1, mtx2, and mtx3. Primers designed for these 5 genes yielded specific PCR amplicons of the expected size from type cultures of B. sphaericus. This method of detecting multiple toxin genes by colony PCR in a single tube reaction is a simple, rapid, and economical technique for identification of highly toxic environmental B. sphaericus isolates.

scholarly journals A PCR Assay To Discriminate Human and Ruminant Feces on the Basis of Host Differences in Bacteroides-Prevotella Genes Encoding 16S rRNA

2000 ◽  
Vol 66 (10) ◽  
pp. 4571-4574 ◽  
Author(s):  
Anne E. Bernhard ◽  
Katharine G. Field
Keyword(s):  
Natural Waters ◽  
Fragment Length Polymorphism ◽  
Sequence Data ◽  
Pcr Primers ◽  
Pcr Markers ◽  
Human Feces ◽  
Inexpensive Method ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Genes Encoding

ABSTRACT Our purpose was to develop a rapid, inexpensive method of diagnosing the source of fecal pollution in water. In previous research, we identified Bacteroides-Prevotella ribosomal DNA (rDNA) PCR markers based on analysis. These markers length heterogeneity PCR and terminal restriction fragment length polymorphism distinguish cow from human feces. Here, we recovered 16S rDNA clones from natural waters that were close phylogenetic relatives of the markers. From the sequence data, we designed specific PCR primers that discriminate human and ruminant sources of fecal contamination.

scholarly journals Detection of an Allele Conferring Resistance to Bacillus sphaericus Binary Toxin in Culex quinquefasciatus Populations by Molecular Screening

2008 ◽  
Vol 75 (4) ◽  
pp. 1044-1049 ◽  
Author(s):  
Karlos Diogo de Melo Chalegre ◽  
Tatiany Patr�cia Rom�o ◽  
Liliane Barbosa Amorim ◽  
Daniela Bandeira Anastacio ◽  
Rosineide Arruda de Barros ◽  
...  
Keyword(s):  
Culex Quinquefasciatus ◽  
Specific Binding ◽  
Bacillus Sphaericus ◽  
Allelic Frequency ◽  
Binary Toxin ◽  
Pcr Assay ◽  
Diagnostic Pcr ◽  
Specific Pcr ◽  
Specific Pcr Assay ◽  
High Level

ABSTRACT The activity of the Bacillus sphaericus binary (Bin) toxin on Culex quinquefasciatus larvae depends on its specific binding to the Cqm1 receptor, a midgut membrane-bound α-glucosidase. A 19-nucleotide deletion in the cqm1 gene (cqm1 REC ) mediates high-level resistance to Bin toxin. Here, resistance in nontreated and B. sphaericus-treated field populations of C. quinquefasciatus was assessed through bioassays as well as a specific PCR assay designed to detect the cqm1 REC allele in individual larvae. Resistance ratios at 90% lethal concentration, gathered through bioassays, were close to 1 and indicate that the selected populations had similar levels of susceptibility to B. sphaericus, comparable to that of a laboratory colony. A diagnostic PCR assay detected the cqm1 REC allele in all populations investigated, and its frequency in two nontreated areas was 0.006 and 0.003, while the frequency in the B. sphaericus-treated population was significantly higher. Values of 0.053 and 0.055 were detected for two distinct sets of samples, and homozygote resistant larvae were found. Evaluation of Cqm1 expression in individual larvae through α-glucosidase assays corroborated the allelic frequency revealed by PCR. The data from this study indicate that the cqm1 REC allele was present at a detectable frequency in nontreated populations, while the higher frequency in samples from the treated area is, perhaps, correlated with the exposure to B. sphaericus. This is the first report of the molecular detection of a biolarvicide resistance allele in mosquito populations, and it confirms that the PCR-based approach is suitable to track such alleles in target populations.

scholarly journals Real-Time PCR Assay for Detection and Enumeration of Dekkera bruxellensis in Wine

2003 ◽  
Vol 69 (12) ◽  
pp. 7430-7434 ◽  
Author(s):  
Trevor G. Phister ◽  
David A. Mills
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Pcr Primers ◽  
Rrna Gene ◽  
Dekkera Bruxellensis ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Spoilage Yeast ◽  
Pcr Method ◽  
26S Rrna

ABSTRACT Traditional methods to detect the spoilage yeast Dekkera bruxellensis from wine involve lengthy enrichments. To overcome this difficulty, we developed a quantitative real-time PCR method to directly detect and enumerate D. bruxellensis in wine. Specific PCR primers to D. bruxellensis were designed to the 26S rRNA gene, and nontarget yeast and bacteria common to the winery environment were not amplified. The assay was linear over a range of cell concentrations (6 log units) and could detect as little as 1 cell per ml in wine. The addition of large amounts of nontarget yeasts did not impact the efficiency of the assay. This method will be helpful to identify possible routes of D. bruxellensis infection in winery environments. Moreover, the time involved in performing the assay (3 h) should enable winemakers to more quickly make wine processing decisions in order to reduce the threat of spoilage by D. bruxellensis.

scholarly journals Shiga Toxins, and the Genes Encoding Them, in Fecal Samples from Native Idaho Ungulates

2008 ◽  
Vol 75 (3) ◽  
pp. 862-865 ◽  
Author(s):  
Jeremy J. Gilbreath ◽  
Malcolm S. Shields ◽  
Rebekah L. Smith ◽  
Larry D. Farrell ◽  
Peter P. Sheridan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Wild Ungulates ◽  
Fecal Samples ◽  
Shiga Toxins ◽  
Toxin Genes ◽  
Genes Encoding ◽  
Shiga Toxin Genes

ABSTRACT Cattle are a known reservoir of Shiga toxin-producing Escherichia coli. The prevalence and stability of Shiga toxin and/or Shiga toxin genes among native wild ungulates in Idaho were investigated. The frequency of both Shiga genes and toxin was similar to that reported for Idaho cattle (∼19%).

Individual single tube genotyping and DNA pooling by allele-specific PCR to uncover associations of polymorphisms with complex diseases

2007 ◽  
Vol 376 (1-2) ◽  
pp. 155-162 ◽  
Author(s):  
Antonio Casado-Díaz ◽  
Rafael Cuenca-Acevedo ◽  
José Manuel Quesada ◽  
Gabriel Dorado
Keyword(s):  
Complex Diseases ◽  
Dna Pooling ◽  
Single Tube ◽  
Specific Pcr ◽  
Allele Specific ◽  
Allele Specific Pcr

Identification of Klebsiella oxytoca using a specific PCR assay targeting the polygalacturonase pehX gene

2003 ◽  
Vol 154 (8) ◽  
pp. 587-592 ◽  
Author(s):  
Gennadiy Kovtunovych ◽  
Tetyana Lytvynenko ◽  
Valentyna Negrutska ◽  
Olena Lar ◽  
Sylvain Brisse ◽  
...  
Keyword(s):  
Klebsiella Oxytoca ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Specific Pcr Assay
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