Rapid and Non-Invasive Detection of Aedes aegypti Co-Infected with Zika and Dengue Viruses Using Near Infrared Spectroscopy

Abstract

The transmission of dengue (DENV) and Zika (ZIKV) has been continuously increasing worldwide. An efficient arbovirus surveillance system is critical to designing early-warning systems to increase preparedness of future outbreaks in endemic countries. The Near Infrared Spectroscopy (NIRS) is a promising high throughput technique to detect arbovirus infection in Ae. aegypti with remarkable advantages such as cost and time effectiveness, reagent-free, and non-invasive nature over existing molecular tools for similar purposes, enabling timely decision making through rapid detection of potential disease. Our aim was to determine whether NIRS can differentiate Ae. aegypti females infected with either ZIKV or DENV single infection, and those coinfected with ZIKV/DENV from uninfected ones. Using 200 Ae. aegypti females reared and infected in laboratory conditions, the training model differentiated mosquitoes into the four treatments with 100% accuracy. DENV-, ZIKV-, and ZIKV/DENV-coinfected mosquitoes that were used to validate the model could be correctly classified into their actual infection group with a predictive accuracy of 100%, 84%, and 80%, respectively. When compared with mosquitoes from the uninfected group, the three infected groups were predicted as belonging to the infected group with 100%, 97%, and 100% accuracy for DENV-infected, ZIKV-infected, and the co-infected group, respectively. Preliminary lab-based results are encouraging and indicate that NIRS should be tested in field settings to evaluate its potential role to monitor natural infection in field-caught mosquitoes.

Keywords: Aedes aegypti; Near Infrared Spectroscopy (NIRS); Zika; chikungunya; dengue; diagnostic; early-warning system; surveillance.

Figure 1

Average NIR spectra of Co-infected, ZIKV, DENV and uninfected mosquitoes (control) collected at 14 days post-infection by the Labspec 4i NIR spectrometer. In particular the absorbance value of uninfected mosquitoes (Blue line) was slightly higher than those of infected mosquitoes.

Figure 2

The number of viral copies in Aedes aegypti females at 14 dpi when mosquitoes were either single infected with ZIKV or DENV, or simultaneously co-infected with both ZIKV/DENV. Different letters indicate statistically significant differences (Wilcoxon rank sum test (U-test), p < 0.001). Each circle represents an individual mosquito; negative control samples (uninfected Ae. aegypti from laboratory colony) are not shown because they did not produce any amplification signal.

Figure 3

The predictive accuracy of NIRS for samples that were used for training and validation of the ANN model.

Figure 4

The predictive accuracy of NIRS for samples that were used to test the ANN model.

Figure 5

The second derivative spectra of Aedes aegypti mosquitoes showing overtones of infected and uninfected groups. The black arrows in the figure indicate the wavelength (nm) band regions that are important for differentiating the groups.