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Molecular characterization of Bacillus anthracis using multiplex PCR, ERIC-PCR and RAPD
Y H Shangkuan1, Y H Chang, J F Yang
1Division of Bacteriology, Institute of Preventive Medicine, National Defense Medical Center, Taipei, Taiwan, ROC. yhshang@tpts6.seed.net.tw
Letters in Applied Microbiology
|March 27, 2001
Summary
Molecular characterization of Bacillus anthracis strains was achieved using multiplex PCR, ERIC-PCR, and RAPD. These methods effectively differentiated B. anthracis strains and identified virulence factors.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Bacillus anthracis is a significant bacterial pathogen.
- Accurate strain differentiation is crucial for epidemiological studies and outbreak investigations.
- Virulence factors and plasmid content are key characteristics for understanding B. anthracis pathogenicity.
Purpose of the Study:
- To molecularly characterize Bacillus anthracis strains.
- To evaluate the utility of multiplex PCR, ERIC-PCR, and RAPD for strain differentiation.
- To identify virulence factors and assess plasmid-associated genetic elements.
Main Methods:
- Multiplex PCR was employed to detect specific virulence genes (cya, cap, cereolysinAB).
- Enterobacterial Repetitive Intergenic Consensus-PCR (ERIC-PCR) and Random Amplification of Polymorphic DNA (RAPD) were performed for genetic fingerprinting.
- Southern hybridization was used to confirm the origin of amplified DNA fragments, with probes derived from ERIC-PCR and RAPD.
Main Results:
- Multiplex PCR successfully identified B. anthracis virulence factors.
- ERIC-PCR and RAPD generated distinct fragment patterns, dividing B. anthracis into two groups.
- Sequence analysis revealed that the cloned fragments originated from the pXO1 plasmid of B. anthracis.
- Hybridization confirmed that probes targeted cya+ strains.
Conclusions:
- Multiplex PCR enables simultaneous identification of Bacillus cereus group isolates and B. anthracis virulence factors.
- ERIC-PCR and RAPD, coupled with Southern hybridization, effectively differentiate B. anthracis strains from B. cereus.
- These molecular techniques can distinguish virulent from avirulent B. anthracis strains and detect amplification of large plasmids.