Label-free visualization of internal organs and assessment of anatomical differences among adult Anopheles, Aedes, and Culex mosquito specimens using bidirectional optical coherence tomography

Mosquitoes transmit several fatal human diseases and constitute a global threat to the fight against infectious diseases. Thus, it is crucial to identify the diseases transmitted by mosquitoes, analyze their internal organs, investigate the life cycles of the viruses and pathogens they carry, and elucidate the anatomical changes they cause inside the host without dissecting them. Here we have demonstrated a method for label-free visualization of the internal organs of adult Anopheles, Aedes, and Culex mosquitoes using swept-source optical coherence tomography (SS-OCT). To overcome the limitation of depth-dependent signal-to-noise ratio (SNR) reduction, imaging was conducted using a dynamic rotational OCT scanner to acquire images of the top and bottom surfaces of the specimens. The internal structure and organ images of all the mosquito specimens had constant resolvability and higher SNR than in those obtained via conventional OCT. Furthermore, a depth profiling algorithm was developed to obtain quantitative information about the internal organs. Several internal organs, such as the salivary glands, heart, midgut, dorsal and ventral crop, and abdominal ganglia, were precisely identified and analyzed noninvasively using OCT. The average thicknesses of the heart, midgut, dorsal and ventral crop, and abdominal ganglia of Anopheles, Aedes, and Culex mosquitoes were 72.1, 107.3, 87.3, and 63.4 μm, respectively. This study demonstrates the applicability of OCT in entomology research for high-resolution microscopic analysis. The findings of this study can guide future studies requiring nondestructive assessment of internal organs to evaluate the morphological differences among various virus-transmitting mosquito specimens.