Identification of upper respiratory tract pathogens using electrochemical detection on an oligonucleotide microarray.
Bacterial and viral upper respiratory infections (URI) produce highly variable clinical symptoms that cannot be used to identify the etiologic agent. Proper treatment, however, depends on correct identification of the pathogen involved as antibiotics provide little or no benefit with viral infections. Here we describe a rapid and sensitive genotyping assay and microarray for URI identification using standard amplification and hybridization techniques, with electrochemical detection (ECD) on a semiconductor-based oligonucleotide microarray. The assay was developed to detect four bacterial pathogens (Bordetella pertussis, Streptococcus pyogenes, Chlamydia pneumoniae and Mycoplasma pneumoniae) and 9 viral pathogens (adenovirus 4, coronavirus OC43, 229E and HK, influenza A and B, parainfluenza types 1, 2, and 3 and respiratory syncytial virus. This new platform forms the basis for a fully automated diagnostics system that is very flexible and can be customized to suit different or additional pathogens. Multiple probes on a flexible platform allow one to test probes empirically and then select highly reactive probes for further iterative evaluation. Because ECD uses an enzymatic reaction to create electrical signals that can be read directly from the array, there is no need for image analysis or for expensive and delicate optical scanning equipment. We show assay sensitivity and specificity that are excellent for a multiplexed format.
Lodes
Michael J
MJ
CombiMatrix Corporation, Mukilteo, Washington, USA. mlodes@combimatrix.com
Suciu
Dominic
D
Wilmoth
Jodi L
JL
Ross
Marty
M
Munro
Sandra
S
Dix
Kim
K
Bernards
Karen
K
Stöver
Axel G
AG
Quintana
Miguel
M
Iihoshi
Naomi
N
Lyon
Wanda J
WJ
Danley
David L
DL
McShea
Andrew
A
D000256:Adenoviridae / Q000235:genetics / Q000302:isolation & purification; D001424:Bacterial Infections / Q000175:diagnosis / Q000382:microbiology; D001886:Bordetella pertussis / Q000235:genetics / Q000302:isolation & purification; D016993:Chlamydophila pneumoniae / Q000235:genetics / Q000302:isolation & purification; D028941:Coronavirus 229E, Human / Q000235:genetics / Q000302:isolation & purification; D028962:Coronavirus OC43, Human / Q000235:genetics / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D004563:Electrochemistry / Q000379:methods*; D006801:Humans; D009980:Influenza A virus / Q000235:genetics / Q000302:isolation & purification; D009981:Influenza B virus / Q000235:genetics / Q000302:isolation & purification; D009177:Mycoplasma pneumoniae / Q000235:genetics / Q000302:isolation & purification; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010222:Parainfluenza Virus 1, Human / Q000235:genetics / Q000302:isolation & purification; D010223:Parainfluenza Virus 2, Human / Q000235:genetics / Q000302:isolation & purification; D010224:Parainfluenza Virus 3, Human / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D012136:Respiratory Syncytial Viruses / Q000235:genetics / Q000302:isolation & purification; D012137:Respiratory System / Q000382:microbiology* / Q000821:virology*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D013297:Streptococcus pyogenes / Q000235:genetics / Q000302:isolation & purification; D014777:Virus Diseases / Q000175:diagnosis / Q000821:virology
D004269:DNA, Bacterial; D004279:DNA, Viral