Research entomologist Jana Lee, Ph.D., of the U.S. Department of Agriculture Agricultural Research Service in Corvallis, Oregon, grinds up an insect in preparation for an assay that will provide information about the insect’s energetic level. Such tests can reveal insights into insect physiology, ecology and behavior. A new review of entomological assays by Lee in Annals of the Entomological Society of America is part of a special collection on “Tools of the Behavior and Biocontrol Trade.” (Photo credit: Gracie Galindo/USDA-ARS)

Leslie Mertz, Ph.D.

By Leslie Mertz, Ph.D.

You may see a certain type of beetle among the marigolds, or a little wasp perched on a daisy, but what is it really doing there? That is an important question for entomologists who investigate insects, including pests. Fortunately, a collection of tools is available to get the inside scoop on insects’ lives and reveal possible ways to curtail some problematic species.

The tools, called assays, are lab procedures in which an insect is crushed in a solution, to which another substance is added to induce a chemical reaction, such as a color change. The chemistry and details vary from assay to assay, but these reactions can be used to measure the amounts of different fats (lipids), sugars, and other compounds in an insect’s body. Researchers can then use this information to determine where the insect gets its energy and how it uses that energy, according to Jana Lee, Ph.D., research entomologist at the U.S. Department of Agriculture Agricultural Research Service in Corvallis, Oregon. Lee summarizes the range of assays available to entomologists in a new paper published in the Annals of the Entomological Society of America. The paper is part of a special collection released today in Annals titled “Tools of the Behavior and Biocontrol Trade.”

To help researchers discover and learn about available tools and how to use them, the Annals of the Entomological Society of America has published a special collection of review papers that focus on tools researchers are using to study arthropod behavior and biological control. Read the collection now.

A few of the many assays noted in her paper include:

The Van Handel assays. Developed in the 1980s by Emile Van Handel of the University of Florida, the two Van Handel assays remain staples in entomology labs today because they are relatively simple to do, they don’t need out-of-the-ordinary lab equipment, and they provide a good range of information, Lee says. “You basically grind up the bugs, add certain solutions to separate the fat and sugars from the glycogen (sugar that is stored in the body for later use), and, through a series of other steps, you can then read the glycogen and sugar with one reaction that turns green and the fats with another reaction that turns pink,” she explains.

The Foray assay. First described in 2012 by Vincent Foray and others from the Université de Lyon, the Foray assay yields measurements on lipid, glycogen, sugars and proteins simultaneously, Lee says. She is intrigued by researchers who are using the assay to explore how herbivorous insects adjust their behavior in the presence of predators “and, in some cases, feed more and then acquire more nutrition.”

Enzymatic assay. Developed by Craig Phillips and others at AgResearch of New Zealand in 2018, this assay provides information about trehalose, the main blood sugar in insects (human blood sugar is glucose). By tracking trehalose, scientists can learn how insects metabolize the food they eat and, therefore, how much energy the insect actually gets from its food.

Assay data have many applications, Lee says. For instance, farmers may try to fight pest insects by enticing other pest-killing insects, called parasitoids, into their fields. Common parasitoids are various species of small wasps that attack pest insects. By using the assays, researchers can identify which plant’s nectar the adult parasitoids are consuming, and consequently they can help farmers determine what wildflowers to plant alongside their crops to lure parasitoids.

Research entomologist Jana Lee, Ph.D., of the U.S. Department of Agriculture Agricultural Research Service in Corvallis, Oregon, uses various assays to study major crop pests, including brown marmorated stink bugs (Halyomorpha halys) and spotted-wing drosophila (Drosophila suzukii). For her doctoral work, she used the assays to study parasitoid wasps Diadegma insulare, which attack diamondback moths (Plutella xylostella). Infestations of diamondback moth larvae can cause considerable damage to cabbage, broccoli, and cauliflower. (Photo credit: Gracie Galindo/USDA-ARS)

In other cases, researchers use assays to study insect behavior. After observing insect activity, such as laying eggs, looking for food, or just sitting still, researchers run the assay to determine the insects’ energy levels. “This can help with predictive modeling, so it can tell you that parasitoids that have a medium level of energy, for instance, will look for places to lay eggs, whereas those that are hungry will spend their time looking for food, and those that are very full will just do nothing,” Lee says.

In her own research, Lee uses the assays to understand pest species, notably brown marmorated stink bugs (Halyomorpha halys) and fruit flies known as spotted-wing drosophila (Drosophila suzukii) that are wreaking havoc on U.S. crops. Specifically, she is deciphering which lures work best when the insects first emerge from overwintering. “They are very hungry, and [the assays] are helping us to understand how to attract them with a food source as a way to trap them,” she says. Farmers use trap counts to determine the timing and extent of pest infestation, so they can effectively fight the pests with pesticides or other management methods.

Assays can also provide insights into the general ecology of insects, including their dispersal. “It’s very helpful to see if insects stay put or how far they disperse with certain energy levels,” Lee says. “For that, researchers might fly them on a flight mill in a lab, so they get the insects to ‘pretend fly,’ and then run the assay and measure their nutrients as a way to infer things like how much flight energy comes from glycogen versus fat.” Researchers can do similar experiments in the field by marking insects, tracking their flight movements, and then running the assays, she adds.

With so many different assays that can be used for such a diversity of research projects, Lee hopes her paper will provide a good summary of the options. “Since I had already been teaching people how to do some of these assays, including the different logistical things to think about, it just seemed natural to write it all down,” she says. “For someone who is starting a project, for example, they have to think about the assay procedures, the kinds of research questions that are important to them, and whether they need very precise data from a [more complicated] assay or can get enough data from a low-tech, easy assay,” she says. “The idea with this paper is to help readers decide which option fits their needs.”

Leslie Mertz, Ph.D., teaches summer field-biology courses, writes about science, and runs an educational insect-identification website, www.knowyourinsects.org. She resides in northern Michigan.