An Efficient and Simple Method for Collecting Haemolymph of Cerambycidae (Insecta: Coleoptera) Adults

SIMPLE SUMMARY: The lack of efficient methods to extract haemolymph from adult cerambycid beetles limits physiological and biochemical research on these important pest species. To enable the collection of large amounts of pure haemolymph from Cerambycidae adults efficiently and easily, we developed an innovative extraction method, termed net centrifugation. Anoplophora chinensis, Monochamus saltuarius and Saperda populnea, three cerambycid beetle species with substantial differences in adult size, were used to evaluate the newly developed method. Compared with estimates for other commonly used methods for coleopteran haemolymph collection, this method generated a larger amount of pure haemolymph in a shorter time. The net centrifugation method is therefore highly efficient, providing a solid foundation for further studies of cerambycid beetles. ABSTRACT: Cerambycid beetles (Cerambycidae) are major forest pests, posing a serious threat to the security of forest resources worldwide. Extensive research has focused on the control of cerambycid beetles from physiological and biochemical perspectives. Despite the important roles of insect haemolymph in physiological processes, efficient collection methods for Cerambycidae are lacking. For the efficient and easy collection of large amounts of pure haemolymph from adult cerambycid beetles, a new method, named net centrifugation, was developed. Three species of cerambycid beetles with large differences in size, Anoplophora chinensis, Monochamus saltuarius and Saperda populnea, were selected for the study. Haemolymph was collected by the newly developed net centrifugation method—in which an inner nylon net is used during centrifugation under optimised conditions, and a relatively small wound is generated on the insect—as well as the traditional tearing method and double centrifugation method. Among the three methods evaluated, the net centrifugation method caused the least damage to cerambycid beetles during the whole operation. This method resulted in the most haemolymph from a single beetle, with the lowest turbidity, mostly pure haemocytes in the precipitate, the clearest haemolymph smears by microscopy and the highest quality of RNA extracted from haemocytes. The net centrifugation method has a high collection efficiency, providing important technical support for haemolymph extraction and entomological research.