Abstract
A common technique used for sensitive and specific diagnostic virus detection in clinical samples is PCR that can identify one or several viruses in one assay. However, a diagnostic microarray containing probes for all human pathogens could replace hundreds of individual PCR-reactions and remove the need for a clear clinical hypothesis regarding a suspected pathogen. We have established such a diagnostic platform for random amplification and subsequent microarray identification of viral pathogens in clinical samples. We show that Phi29 polymerase-amplification of a diverse set of clinical samples generates enough viral material for successful identification by the Microbial Detection Array, demonstrating the potential of the microarray technique for broad-spectrum pathogen detection. We conclude that this method detects both DNA and RNA virus, present in the same sample, as well as differentiates between different virus subtypes. We propose this assay for diagnostic analysis of viruses in clinical samples.
Highlights
A common method of choice for clinical pathogen identification is polymerase chain reaction (PCR) [1] which is a sensitive and specific method
We show that Phi29amplification generates enough material for successful identification by the Lawrence Livermore Microbial Detection Array (LLMDA) [19,20], demonstrating the potential of the microarray technique for broad-spectrum pathogen detection in clinical samples
It has been demonstrated that filtration, DNase and RNase treatment can increase the efficiency of downstream amplifications [26,27]
Summary
A common method of choice for clinical pathogen identification is polymerase chain reaction (PCR) [1] which is a sensitive and specific method. Each PCR reaction only provides identification of one specific virus, or a group of related viruses. With some clinical syndromes or symptoms, it might be difficult to determine which viruses to test for in order to establish the correct diagnosis since many viruses can present similar symptoms. For this reason various multiplexed assays with virus panels, utilizing PCR or other methods, have been established. A microarray could be viewed as an extended pathogen panel, covering all viruses and bacteria, increasing the chance of the correct identification of one or several pathogens even in situations without any prior knowledge or suspicion about the etiology
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