Genotyping of measles virus in clinical specimens on the basis of oligonucleotide microarray hybridization patterns.
An oligonucleotide microarray hybridization method for identification of most known measles virus (MV) genotypes was developed. Like the conventional genotyping method, the microarray relied on detecting sequence differences in the 450-nucleotide region coding for the COOH-terminal 150 amino acids of the nucleoprotein (N). This region was amplified using PCR primers binding to all known MV genotypes. The microarray included 71 pairs of oligonucleotide probes (oligoprobes) immobilized on glass slides. Each pair consisted of a genotype-specific oligoprobe, which matched the sequence of only one target genotype, and a control oligoprobe, which contained mismatches at the nucleotide positions unique to this genotype. A pattern recognition algorithm based on cluster analysis of the ratios of hybridization signals from specific and control oligoprobes was used to identify the specific MV genotype. Following the initial validation, the method was used for rapid genotyping of two panels of coded samples. The results of this study showed good sensitivity (90.7%), specificity (100%), and genotype agreement (91.8%) for the new method compared to the results of genotyping conducted using phylogenetic analysis of viral sequences of the C terminus of the N gene. In addition, the microarray demonstrated the ability to identify potential new genotypes of MV based on the similarity of their hybridization patterns with those of known MV genotypes.
Neverov
Alexander A
AA
Center for Biologics Evaluation and Research, Food and Drug Administration, Rockville, MD 20852-1448, USA. alexander.neverov@fda.hhs.gov
Riddell
Michaela A
MA
Moss
William J
WJ
Volokhov
Dmitriy V
DV
Rota
Paul A
PA
Lowe
Luis E
LE
Chibo
Doris
D
Smit
Sheilagh B
SB
Griffin
Diane E
DE
Chumakov
Konstantin M
KM
Chizhikov
Vladimir E
VE
D005838:Genotype; D008459:Measles virus / Q000235:genetics* / Q000302:isolation & purification; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010802:Phylogeny; D015203:Reproducibility of Results