Gall-Inducing Wasps Have Enlarged Venom Glands, Study Finds

Closeup of a small black wasp with brown legs and antennae perched on a flat green leaf, with its ovipositor on the bottom of its abdomen piercing the surface of the leaf. — A female gall wasp lays eggs in the midrib of a red oak leaf. Gall wasps of the family Cynipidae use compounds to stimulate the growth of protective growths, called galls, in which their young can feed and grow in a protective enclosure. In a new study on gall wasps by researchers at Penn State University the investigators examined the anatomy of cynipid species and found that gall-inducing wasps have especially large venom glands, implicating venom in gall induction. (Photo by Antoine Guiguet, Ph.D.)

By John P. Roche, Ph.D.

Some plant-eating insects interfere with growth pathways in plants to create abnormal growths, called galls, in which their young can feed and grow in a protective enclosure. Groups that have gall-producers include moths, aphids, midges, and the wasps—particularly in the family Cynipidae.

The mechanism of gall induction in insects remains poorly understood. To explore potential sources of substances inducing these galls, researchers at Penn State University described and compared the anatomy of secretory glands of female cynipid wasps in a study published in October in Insect Systematics and Diversity. The team investigated the anatomy of 69 species of cynipids that produce galls and 29 species of cynipids that do not. Led by postdoctoral researcher Antoine Guiguet, Ph.D., the researchers found that gall-inducing Cynipidae have especially large venom glands, implicating venom in gall induction.

The Cynipidae family of wasps evolved from an ancestral group of parasitoid wasps. “Parasitoid wasps inject substances from the venom gland to stun their insect hosts and manipulate their natural immunity,” says Heather Hines, Ph.D., associate professor of biology and entomology at Penn State and senior author on the study. In gall wasps, females inject substances not into other insects but into plants to trigger the growth of galls.

“Our study shows that the gall-inducing Cynipidae have increased the size of these [venom] glands sometimes to very large portions,” Guiguet says. “The largest venom glands among the oak gallers are found in the mealy oak gall wasp Disholcaspis cinerosa, whose venom gland when unfolded is 10 times longer than its abdomen, and in the oak rough bulletgall wasp (Disholcaspis quercusmamma), whose venom reservoir occupies a third of its abdomen volume.”

Collage image of 12 plant galls caused by Cynipid wasps, in three rows of four. The galls vary greatly in appearance: some appear on stems and some appear on leaves; some are green, some brown, some white, and others yellowish green with red spots; some are smooth, some are bumpy, and some have hairs or spikes. — Gall wasps of the family Cynipidae use compounds to stimulate the growth of protective growths, called galls, in which their young can feed and grow in a protective enclosure. The appearance and physical characteristics of these plant galls produced by cynipid gall wasps can vary greatly, as illustrated here. (Image originally published in Guiguet et al 2023, *Insect Systematics and Diversity*)

Recent publications suggest gall induction likely evolved several times independently in Cynipidae. One group of cynipids form galls in roses; another uses plants of the composite family, mint family, and poppy family; another uses Rubus bramble berries; and the most species-rich lineage induces galls in oaks. Guiguet and colleagues found that gall wasp species that induced gall formation have a relatively larger venom apparatus than gall wasp species that do not induce galls. They also found that another secretory organ, the accessory glands, can be enlarged in some gall-inducing species.

“Rose gallers appear to have basically lost the venom gland,” Hines says, “but do have expanded accessory glands, suggesting that other secretions may have evolved in some lineages.” The larger size of these secretory glands in some galling species supports the hypothesis that they may also be involved in stimulating gall production.

Although these glands are large across most gall-inducers, this study also found that the anatomy of venom glands and accessory glands, and the content of these two types of secretory glands, varied among gall-producing cynipids. “When we investigated different glands at a morphological and histological level,” Guiguet says, “we found a diversity of venom colors, venom particles, and secretory structures.” The authors conclude that this suggests that molecular effectors involved in gall formation likely have shifted over time.

Two-part image. Top image shows a closeup of a wasp that is rusty brown in color with black stripes, photographed on a white surface and background. Bottom image is a 3D microCT scan visualization of the wasp's abdomen, with two glands highlighted, while the rest is translucent. The venom gland is colored in yellowish orange and resembles a long squiggly tube folded over several times, located near the hind end of the abdomen. The venom reservoir is colored in fuschia and shaped like a U-shaped sac, appearing to fill nearly half of the abdomen, also toward the rear end. — In wasps that generate abnormal plant growths called galls, a new study finds they commonly have large venom glands relative to body size, suggesting their venom plays a key role in inducing plant gall formation. Shown here is a female oak rough bulletgall wasp (*Disholcaspis quercusmamma*) and a microCT scan image of the anatomy of its venom gland and venom reservoir, which occupies a third of its abdomen volume. (Photo and image by Antoine Guiguet, Ph.D.)

It is not yet definitely known which compounds gall-producing species use to trigger galls to form. Two species of oak gallers produce hormones also produced by plants: auxins, which regulate growth in plants, and cytokinins, which stimulate cell division in plants. These hormones are likely candidates for compounds that could affect plant growth when injected by wasps.

It is also not yet definitely known whether molecules gall wasps introduce during egg laying or molecules introduced by gall wasp larvae, or both, trigger gall formation. One study found that in Liposthenes glechomae, surgical removal of larva caused gall growth to stop. In a different study, in herb gallers, galls began forming before eggs hatched, indicating that compounds introduced by egg laying, not by larvae, stimulated the growth of galls in that species. Data collected by Guiguet and colleagues support the hypothesis that secretions introduced in egg laying, especially venoms, likely have a role in gall formation.

Research on gall induction by cynipids is ongoing in Hines’s group. “Our lab is pursuing the mechanism,” Hines says, “by comparing the transcriptomes of these glands in species across the clade with the goal of better understanding key constituents and constituent diversity across species.”

The Cynipidae evolved a wide radiation of species with an impressive diversity of ways to utilize plants as hosts. Future research will provide important discoveries about the specific mechanisms used by different gall-wasp subgroups and insights into how those subgroups arose and radiated.

“Comparative anatomy of venom glands suggests a role of maternal secretions in gall induction by cynipid wasps (Hymenoptera: Cynipidae)”

Insect Systematics and Diversity

John P. Roche, Ph.D., is an author, biologist, and science writer with a Ph.D. and postdoctoral fellowship in the biological sciences and a dedication to making rigorous science clear and accessible. He authors books and articles and writes materials for universities, scientific societies, and publishers. Professional experience includes serving as a scientist and scientific writer at Indiana University, Boston College, and the UMass Chan Medical School.