We ZJU-CHINA 2015 are pursuing an effective and eco-friendly way to control termites. It is unwarranted if we only engage in idle theorizing about termites behavior and only do the genetic engineering experiments. Thus we carry out a series of termites experiments in order to achieve our goals, which are:
- Our baits mixed with bacteria can attract the termites
- Worker termites carry the baits in their mouth and return to the nest, being alive
- Worker termites feed other termites in the nest through the process called trophallaxis
- The termites that receive food die because of the toxin in the baits
To kill the termites, first of all, we have to make sure that termites will eat our baits. In any case, we cannot pry open termites' mouth and stuff in our food. Thanks to our advisor Professor Mo, who is an authority on termites control, we are provided with fantastic bait for termites. In order to confirm its attractiveness, we put a pinch of the bait in the petri dish together with a pinch of normal pine wood powder and see how the termites are attracted and distributed in the dish. If the baits are more attractive, most termites will gather around it.
Figure 1 Examination of infected D.saccharalis midgut by scanning electron microscopy.
Chosen compound toxicity verification
Before the gene manipulation and capsulation experiments begin, it is important to make sure that the toxins we choose are toxic enough to kill termites. Avermectin is naturally produced in Streptomyces avermitilis. Toxic proteins TcdA1, TcdB1, Plu0840 and Plu1537 are natural products of Photorhabdus luminescens strain TT01. Accordingly, we feed the termites with S. avermitilis and TT01 and note down the mortality rate and action speed. Besides, other insects are tested under the same condition to detect the toxicity to these insects.
Cellulase and lysozyme activity measurement
It is vital for worker termites to keep alive when they are carrying the food back to their nest. If they die on the way, the toxin cannot be brought and may spread into the environment. Thus we encapsulate the bacteria within cellulose. To ensure that the cellulose can be digested efficiently so as to release the bacteria into the nest after a period of time, we measure the cellulase activity in different parts of termite's gut. Besides, lysozyme activity is measured to ensure that bacteria can be broken in termite's gut and the toxin can be released.
Verification of the release delay effect due to the CNC capsulation
To ensure the termites will not die during the carrying stage, we design the CNC capsulation and expect a release delay effect. Thus, we feed the termites with capsulated Streptomyces avermitilis and assume that the dying speed will be lower, which illustrates that the capsulation process has the release delay effect.
Trophallaxis has been well proved exists in the Coptotermes formosanus Shiraki. However, we hope to have a concise demonstration so that everyone can have a clear understanding of the trophallaxis process. Therefore, we conduct an experiment to feed termites with stained food. In detail, a group of termites are dyed in blue and the other in red. We mix the termites from these two groups and observe the color change. Besides, soldier termites who can only get food by trophallaxis are also added in each group.
Figure 2 top and side view of the tcdA1 prepore (left) and pore (right) pentamer.
Final product effectiveness measurement
Our final product is the genetic modified bacteria capsulated with cellulose. Considering about safety, we are not allowed to test our product or device in the field. Instead, we build up a nice model to predict the result of the use of our product. To optimize our model and find the suitable parameters, supporting experiment are conducted.
1 Fujita, A., Shimizu, I. & Abe, T., Distribution of lysozyme and protease, and amino acid concentration in the guts of a wood-feeding termite, Reticulitermes speratus (Kolbe): possible digestion of symbiont bacteria transferred by trophallaxis. PHYSIOL ENTOMOL 26 116 (2001).
2 Lax, A. R. & La Osbrink, W., United States Department of Agriculture - Agriculture Research Service - research on targeted management of the Formosan subterranean termite Coptotermes formosanus Shiraki (Isoptera : Rhinotermitidae). PEST MANAG SCI 59 788 (2003).
3 Huang, Q., Wang, W., Mo, R. & Lei, C., Studies on feeding and trophallaxis in the subterranean termite Odontotermes formosanus using rubidium chloride. ENTOMOL EXP APPL 129 210 (2008).
4 Xing, L. et al., Observations of grooming and trophallaxis in a Chinese subterranean termite, Reticulitermes aculabialis Tsai et Hwang (Isoptera: Rhinotermitidae). Shengtaixue Zazhi 33 149 (2014).
5 Zhang, J., Wang, W., Li, W., Zhuang, T. & Chen, L., Analysis on the trophallaxis of Coptotermes formosanus Shiraki. Huazhong Shifan Daxue Xuebao (Ziran Kexue Ban) 37 90 (2003).
6 Ruan, G., Li, J. & Mo, J., Effects of bait for controlling Reticulitermes chinensis colonies. Journal of Zhejiang a&f University 31 768 (2014).