In silico and in vitro evaluation of PCR-based assays for the detection of Bacillus anthracis chromosomal signature sequences

• Bacillus anthracis, the causative agent of anthrax, is a zoonotic pathogen that is relatively common throughout the world and may cause life threatening diseases in animals and humans. There are many PCR-based assays in use for the detection of B. anthracis. While most of the developed assays rely on unique markers present on virulence plasmids pXO1 and pXO2, relatively few assays incorporate chromosomal DNA markers due to the close relatedness of B. anthracis to the B. cereus group strains. For the detection of chromosomal DNA, different genes have been used, such as BA813, rpoB, gyrA, plcR, S-layer, and prophage-lambda. Following a review of the literature, an in silico analysis of all signature sequences reported for identification of B. anthracis was conducted. Published primer and probe sequences were compared for specificity against 134 available Bacillus spp. genomes. Although many of the chromosomal targets evaluated are claimed to be specific to B. anthracis, cross-reactions with closely related B. cereus and B. thuringiensis strains were often observed. Of the 35 investigated PCR assays, only 4 were 100% specific for the B. anthracis chromosome. An interlaboratory ring trial among five European laboratories was then performed to evaluate six assays, including the WHO recommended procedures, using a collection of 90 Bacillus strains. Three assays performed adequately, yielding no false positive or negative results. All three assays target chromosomal markers located within the lambdaBa03 prophage region (PL3, BA5345, and BA5357). Detection limit was further assessed for one of these highly specific assays. © 2013 Landes Bioscience.

Ämnesord

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinska och farmaceutiska grundvetenskaper -- Mikrobiologi inom det medicinska området (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Basic Medicine -- Microbiology in the medical area (hsv//eng)

NATURVETENSKAP  -- Biologi -- Immunologi (hsv//swe)

NATURAL SCIENCES  -- Biological Sciences -- Immunology (hsv//eng)

Nyckelord

Bacillus anthracis

Chromosomal marker

Detection

Diagnostic sensitivity

In silico analysis

Inter-laboratory trial

qPCR

Specificity

anthrax

Bacillus cereus

bacterial strain

chromosome banding pattern

computer model

controlled study

cross reaction

DNA extraction

fluorescence resonance energy transfer

fluorometry

limit of detection

nonhuman

point mutation

real time polymerase chain reaction

review

sensitivity and specificity

single nucleotide polymorphism

world health organization

animal

article

Bacillus thuringiensis

bacterial chromosome

biology

chromosome marker

comparative study

evaluation study

genetics

human

isolation and purification

methodology

molecular diagnosis

polymerase chain reaction

Animalia

Bacillus cereus group

bacterial DNA

primer DNA

Animals

Chromosomes

Bacterial

Computational Biology

DNA Primers

DNA

Bacterial

Humans

Molecular Diagnostic Techniques

Publikations- och innehållstyp

ref (ämneskategori)

art (ämneskategori)