A multiplex real-time PCR for identifying and differentiating B. anthracis virulent types.
Bacillus anthracis is closely related to the endospore forming bacteria Bacillus cereus and Bacillus thuringiensis. For accurate detection of the life threatening pathogen B. anthracis, it is essential to distinguish between these three species. Here we present a novel multiplex real-time PCR for simultaneous specific identification of B. anthracis and discrimination of different B. anthracis virulence types. Specific B. anthracis markers were selected by whole genome comparison and different sets of primers and probes with optimal characteristic for multiplex detection of the B. anthracis chromosome, the B. anthracis pXO1 and pXO2 plasmids and an internal control (IC) were designed. The primer sets were evaluated using a panel of B. anthracis strains and exclusivity was tested using genetically closely related B. cereus strains. The robustness of final primer design was evaluated by laboratories in three different countries using five different real-time PCR thermocyclers. Testing of a panel of more than 20 anthrax strains originating from different locations around the globe, including the recent Swedish anthrax outbreak strain, showed that all strains were detected correctly.
Wielinga
Peter R
PR
National Institute for Public Health and the Environment, Centre for infectious Disease Control, Laboratory for Zoonoses and Environmental Microbiology, Antonie van Leeuwenhoeklaan 9, PO Box 1, Bilthoven, The Netherlands. peter.wielinga@rivm.nl
Hamidjaja
Raditijo A
RA
Agren
Joakim
J
Knutsson
Rickard
R
Segerman
Bo
B
Fricker
Martina
M
Ehling-Schulz
Monika
M
de Groot
Astrid
A
Burton
Jane
J
Brooks
Tim
T
Janse
Ingmar
I
van Rotterdam
Bart
B
D001408:Bacillus anthracis / Q000145:classification* / Q000302:isolation & purification / Q000472:pathogenicity; D001409:Bacillus cereus / Q000145:classification / Q000235:genetics; D001413:Bacillus thuringiensis / Q000145:classification / Q000235:genetics; D017931:DNA Primers; D016133:Polymerase Chain Reaction / Q000379:methods*; D014774:Virulence / Q000235:genetics
D017931:DNA Primers