+ Site Statistics
+ Search Articles
+ Subscribe to Site Feeds
EurekaMag Most Shared ContentMost Shared
EurekaMag PDF Full Text ContentPDF Full Text
+ PDF Full Text
Request PDF Full TextRequest PDF Full Text
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter
Follow on LinkedInFollow on LinkedIn

+ Translate

Multicenter evaluation of PCR reagents for Bacillus anthracis

Multicenter evaluation of PCR reagents for Bacillus anthracis

Abstracts of the General Meeting of the American Society for Microbiology 102: 145-146

We have developed and optimized PCR reagents for five gene targets of Bacillus anthracis. The genes targeted are protective antigen, lethal factor, edema factor (all px01 genes), CAP A (px02 gene) and a chromosomal marker. Optimization for each of the reagents as wet chemistry resulted in assays that detect 100% of samples containing 100 fg of target DNA (approximately 20 gene copies). Although the assay can detect DNA concentrations down to 10 fg, the rate of detection varies depending on the assay at the lower concentrations. Lyophilization of PCR chemistry offers advantages over use of wet chemistry. It standardizes the reaction mixture over a large set of reagent tubes, extends shelf life, allows single use reagent sets, simplifies PCR set up, reduces contamination of reagent from external sources and provides a highly reproducible reaction. We conducted a multicenter study of three reagents as wet chemistry and dry chemistry for B. anthracis to determine the reproducibility of the assays in other laboratories. Five laboratories participated in the study. AFIP prepared the test panels and wet reagents. The dried reagents were manufactured by Idaho Technologies. Ct values were reproducible with both wet and dried chemistry. Each reagent was tested against a panel of 80 organisms consisting of B. anthracis and other Bacillus species in addition to limiting dilutions of B. anthracis DNA. Reproducibility was very good at high concentrations of DNA and was good at low concentrations of DNA. As expected, the lower the concentration, the greater the variation of the standard deviation of the Ct value.

(PDF 0-2 workdays service: $29.90)

Accession: 035349766

Download citation: RISBibTeXText

Related references

Mechanism of killing of spores of Bacillus anthracis in a high-temperature gas environment, and analysis of DNA damage generated by various decontamination treatments of spores of Bacillus anthracis, Bacillus subtilis and Bacillus thuringiensis. Journal of Applied Microbiology 116(4): 805-814, 2014

Cloning and expression of determinants of Bacillus anthracis protective antigen in Escherichia coli, Bacillus subtilis, and Bacillus anthracis cells. Molekuliarnaia Genetika, Mikrobiologiia i Virusologiia: 13-16, 1993

Cloning and expression of a determinant for a protective antigen of Bacillus anthracis in the cells of Escherichia coli, Bacillus subtilis, Bacillus anthracis. Molekulyarnaya Genetika Mikrobiologiya i Virusologiya 0(2): 13-16, 1993

The induction of motility in Bacillus anthracis by means of bacteriophage lysates: significance for the relationship of Bacillus anthracis to Bacillus cereus. Journal of Bacteriology 69(5): 590-602, 1955

Etest for antibiotic susceptibility testing of Bacillus anthracis, Bacillus cereus and Bacillus thuringiensis: evaluation of a French collection. International Journal of Antimicrobial Agents 31(5): 490-492, 2008

Characterization of Bacillus cereus isolates associated with fatal pneumonias: strains are closely related to Bacillus anthracis and harbor B. anthracis virulence genes. Journal of Clinical Microbiology 44(9): 3352-3360, 2006

Differential analysis of Bacillus anthracis after pX01 plasmid curing and comprehensive data on Bacillus anthracis infection in macrophages and glial cells. Proteomics 7(20): 3743-3758, 2007

The chemical basis of the virulence of Bacillus anthracis. VIII. Fractionation of the intracellular material of Bacillus anthracis. British Journal of Experimental Pathology 37(4): 361-371, 1956

Evaluation of different methods to discriminate Bacillus anthracis from other bacteria of the Bacillus cereus group. Journal of Applied Microbiology 100(4): 673-681, 2006

Evaluation of Bacillus strains as model systems for the work on Bacillus anthracis spores. International Journal of Food Microbiology 145 Suppl 1: S129-S136, 2011

Evaluation of a microfluorometer in immunofluorescence assays of individual spores of Bacillus anthracis and Bacillus cereus. Journal of Immunological Methods 49(3): 271-282, 1982

Evaluation of Bacillus subtilis strain IS53 for the production of Bacillus anthracis protective antigen. Letters in Applied Microbiology 19(4): 225-227, 1994

Characterisation and evaluation of synthetic antimicrobial peptides against Bacillus globigii, Bacillus anthracis and Burkholderia thailandensis. International Journal of Antimicrobial Agents 36(4): 359-363, 2011

I. Soil as a reservoir of Bacillus anthracis. II. Cultural, morphological and virulent properties of Bacillus anthracis strains recovered from soil and from infected and dead animals. Gigiena i veterinarno sanitarnye trebovaniya k promyshlennym zhivotnovodcheskim kompleksam Trudy VNIIVS: 95-105, 1979

Susceptibility of Bacillus anthracis, Bacillus cereus, Bacillus mycoides, Bacillus pseudomycoides and Bacillus thuringiensis to 24 antimicrobials using Sensititre automated microbroth dilution and Etest agar gradient diffusion methods. Journal of Antimicrobial ChemoTherapy 60(3): 555-567, 2007