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Background  Mosquito-borne viruses (MBVs) and parasites (MBPs) are transmitted through hematophagous arthropods-mosquitoes to homoiothermous vertebrates. This study aims at developing a detection method to monitor the spread of mosquito-borne diseases to new areas and diagnose the infections caused by MBVs and MBPs.    Methods  In this assay, an ultra-sensitive chemiluminescence (CL) detection method was developed and used to simultaneously detect 19 common MBVs and MBPs. In vitro transcript RNA, virus-like particles (VLPs), and plasmids were established as positive or limit of detection (LOD) reference materials.    Results  MBVs and MBPs could be genotyped with high sensitivity and specificity. The cut-off values of probes were calculated. The absolute LODs of this strategy to detect serially diluted in vitro transcribed RNAs of MBVs and serially diluted plasmids of MBPs were 10 2 –10 3 copies/μl and 10 1 –10 2 copies/μl, respectively. Further, the LOD of detecting a strain of pre-quantified JEV was 10 1.8 –10 0.8 PFU/ml, fitted well in a linear regression model (coefficient of determination = 0.9678).    Conclusions  Ultra-sensitive CL imaging DNA hybridization was developed and could simultaneously detect various MBVs and MBPs. The method described here has the potential to provide considerable labor savings due to its ability to screen for 19 mosquito-borne pathogens simultaneously.    Electronic supplementary material  The online version of this article (doi:10.1186/s13071-017-1975-1) contains supplementary material, which is available to authorized users.

Ultra-sensitive chemiluminescence imaging DNA hybridization method in the detection of mosquito-borne viruses and parasites