Evaluation of a microarray-hybridization based method applicable for discovery of single nucleotide polymorphisms (SNPs) in the Pseudomonas aeruginosa genome.
BACKGROUND
Whole genome sequencing techniques have added a new dimension to studies on bacterial adaptation, evolution and diversity in chronic infections. By using this powerful approach it was demonstrated that Pseudomonas aeruginosa undergoes intense genetic adaptation processes, crucial in the development of persistent disease. The challenge ahead is to identify universal infection relevant adaptive bacterial traits as potential targets for the development of alternative treatment strategies.


RESULTS
We developed a microarray-based method applicable for discovery of single nucleotide polymorphisms (SNPs) in P. aeruginosa as an easy and economical alternative to whole genome sequencing. About 50% of all SNPs theoretically covered by the array could be detected in a comparative hybridization of PAO1 and PA14 genomes at high specificity (> 0.996). Variations larger than SNPs were detected at much higher sensitivities, reaching nearly 100% for genetic differences affecting multiple consecutive probe oligonucleotides. The detailed comparison of the in silico alignment with experimental hybridization data lead to the identification of various factors influencing sensitivity and specificity in SNP detection and to the identification of strain specific features such as a large deletion within the PA4684 and PA4685 genes in the Washington Genome Center PAO1.


CONCLUSIONS
The application of the genome array as a tool to identify adaptive mutations, to depict genome organizations, and to identify global regulons by the "ChIP-on-chip" technique will expand our knowledge on P. aeruginosa adaptation, evolution and regulatory mechanisms of persistence on a global scale and thus advance the development of effective therapies to overcome persistent disease.
Dötsch
Andreas
A
Helmholtz Centre for Infection Research, Braunschweig, Germany. andreas.doetsch@helmholtz-hzi.de
Pommerenke
Claudia
C
Bredenbruch
Florian
F
Geffers
Robert
R
Häussler
Susanne
S
D000220:Adaptation, Biological / Q000235:genetics; D055028:Comparative Genomic Hybridization; D004269:DNA, Bacterial / Q000235:genetics; D016680:Genome, Bacterial; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D011550:Pseudomonas aeruginosa / Q000235:genetics*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA / Q000379:methods
D004269:DNA, Bacterial