Researchers in China studied Epicauta chinensis blister beetles to learn how they produce cantharidin, a defensive toxin that is highly bitter and toxic to most animals. It has also been found inhibit several types of cancer cells, leading to interest in potential artificial production of the molecule. (Photo credit: Yalin Zhang, Ph.D.)

By John P. Roche

Blister beetles produce cantharidin, a 10-carbon isoprenoid molecule that is highly bitter and is toxic to most animals. Because cantharidin is so bitter, it discourages predators, and its production may have been selected for in blister beetles because of this bitterness. The molecule is found in only two insect groups: blister beetles, which are members of the family Meloidea, and oedemerid beetles, which are members of the family Oedemeridae.

Cantharidin is thought to be produced via a common and important pathway in living organisms—the isoprenoid pathway. Isoprenoids are molecules made up of five-carbon building blocks, and they include many important compounds in organisms, including heme molecules, which are part of hemoglobin molecules, and cholesterol molecules, which are a precursor to steroid hormones. HMG-CoA reductase (HMGR) is a key enzyme in the isoprenoid pathway, which is why cholesterol-lowering medications called statins target HMGR.

Previous work by Yalin Zhang and colleagues at Northwest Agriculture and Forestry University in China confirmed that HMGR is an important component of the cantharidin-synthesis pathway in blister beetles. But not much is known about where or when cantharidin is synthesized in blister beetles, or how cantharidin production is related to HMGR transcripts. In a study published in April in the open-access Journal of Insect Science, Zhang and colleagues worked on adding to our knowledge of the synthesis and distribution of cantharidin in blister beetles by measuring amounts of HMGR and cantharidin in various tissues in male and female Epicauta chinensis blister beetles. “Our project sought to study how blister beetles produce cantharidin in their bodies,” Zhang commented, “and study how we can follow the pathway to produce cantharidin outside of the beetles.” Cantharidin has been used in traditional Chinese medicine for over 2,000 years for treating cancer. Cantharidin has been observed to inhibit human leukemic cells and breast cancer cells in vitro, and derivatives of cantharidin have been found to inhibit several types of cancer in cancer cell lines, including cervical, colon, and prostate cancers.

By studying both adult and multiple instar stages of Epicauta chinensis blister beetles, researchers in China found evidence that an enzyme known as HMGR is involved in production of cantharidin, the defensive, blister-causing toxin that gives the beetles their common name. (Photo credit: Yalin Zhang, Ph.D.)

Blister beetles of the genus Epicauta produce high levels of cantharidin. In Epicauta blister beetles, the first instar larva, called triungulin larvae because they have three-claws on each foot, eat grasshopper eggs. The adults eat plants. Adults can be serious pests of crops such as alfalfa, and when hay containing blister beetles is fed to horses, the cantharidin in the beetles can be harmful or even deadly.

Male Epicauta beetles synthesize cantharidin but females produce almost none. Males transfer cantharidin to females during copulation. A lot of cantharidin is found in eggs during oviposition, added to eggs by females, which protects eggs from predation.

Zhang and his group collected Epicauta chinensis blister beetles in the wild in Shaanxi, China, and reared them in the laboratory, feeding them alfalfa or soybean leaves. They then took adult pairs just after copulation, kept the pairs separate, and reared their larvae. They then measured cantharidin levels and HMGR transcription levels in different tissues and different stages of development. They measured cantharidin levels with gas chromatography and they measured HMGR expression levels using quantitative real-time PCR. Tissues tested included the Malpighian tubules, which eliminate wastes; the fat body, which provides energy storage and metabolic regulation; and the accessory gland, which provides secretions that aid reproduction.

In their study, Zhang and colleagues confirmed that there was a high positive correlation between HMGR transcription and production of cantharidin. In males, levels of HMGR transcript were high in the fat body and Malpighian tubules. In females, HMGR transcript levels were high in the ovary and the accessory gland. Zhang concluded that the high HMGR levels in the fat body and Malpighian tubules of males correlated with cantharidin production, whereas the high HMGR levels in the ovary and the accessory gland of females correlated with the development of the ovary and accessory gland. Interestingly, cantharidin was high in first-instar larvae, lower in the second and third instars, and higher again in the fourth and fifth instars. HMGR levels were elevated in the third and fourth instars, which is consistent with the hypothesis that this rise in HMGR levels in the third and fourth instars triggered the subsequent rise in cantharidin. As a control, the investigators also examined HMGR transcript levels in mealworms, Tenebrio molitor (family Tenebrionidae), a species that does not synthesize cantharidin. They found that HMGR levels in Tenebrio were low in Malpighian tubules, the fat body, and the accessory gland, consistent with the hypothesis that HMGR transcript is involved in cantharidin production.

This study adds to our understanding of this vital biochemical pathway in insects, which could lead to further developments in our knowledge of the synthesis of isoprenoids in general and the production of blister beetle cantharidin in particular. Because of its established anti-cancer properties, an increased understanding of cantharidin synthesis could have valuable medical applications.

John P. Roche is a science writer and author with a Ph.D. in the biological sciences. He has served as a senior scientist and adjunct professor at Boston College, as an editor-in-chief of periodicals at Indiana University and Boston College, and as a science writer at Indiana University and the University of Massachusetts Medical School. He has published more than 170 articles and has written and taught extensively about science. For more information, visit http://authorjohnproche.com/.