When Que Lan passed away unexpectedly of complications from gastric cancer in 2014, the UW entomology professor left behind a promising research project. Her work suggested bacteria might offer a new and effective way to repel mosquitoes.

She had started out looking for bacterial compounds that could kill mosquitoes. Before her passing, Lan discovered that, although one particular bacterial extract was not lethal to mosquitoes, when it was put in their food, the insects refused to eat.

Lan’s colleague, entomology professor Susan Paskewitz, helped secure additional funding to keep the study going and found a scientist, Mayur Kajla, interested in carrying the work forward. In January 2019, their findings were published in the journal Science Advances. Paskewitz and Kajla, an assistant scientist in the Department of Entomology at the time of publication, describe the first mosquito-repelling compounds to be derived from microbes.

“We didn’t come at it thinking we would find a repellent,” says Paskewitz, chair of the entomology department and director of the Midwest Center of Excellence for Vector-Borne Disease. “It was a bit of serendipity.”

These compounds, purified from extracts from the bacterium Xenorhabdus budapestensis, appear to work at lower doses than repellents currently on the market, including DEET and picaridin. The study showed the compounds to be effective against Aedes aegypti, Anopheles gambiae, and Culex pipiens — mosquito species known to transmit diseases such as Zika, West Nile, malaria, and chikungunya, which afflict millions of people worldwide.

Whether these natural chemical compounds, called fabclavines, are suitable for human use remains to be determined. But the study opens up a new area of exploration in the search for insect-repelling and insect-killing compounds, Paskewitz says.

When Kajla joined the project, he designed a set of experiments to test the repellent potential of the bacterial extract and identify the compounds responsible. He modified a commercial mosquito feeding system to more closely mimic a mosquito feeding on a human. For instance, he selected a skin-like membrane to contain a special, red-dyed mosquito diet that simulates human or animal blood. He also tested a variety of cloth coverings to sit atop the membrane, which would be coated with the repellents being screened.

Kajla coated the cloth with water, DEET, or picaridin and allowed mosquitoes to feed for 30 minutes before freezing them and counting the number that were fed (engorged with red liquid) or unfed. The mosquitoes did not feed when the cloth was coated in repellent.

He then tested purified extracts from the bacteria and, with assistance from UW–Madison co-author Gregory Barrett-Wilt, director of mass spectrometry and proteomics at the UW Biotechnology Center, found that an extract dominated by two fabclavine molecules effectively deterred mosquitoes from feeding.

When compared for effectiveness against picaridin and DEET, which is found in more than 500 insect repellents registered with the U.S. Environmental Protection Agency, the bacterial extract was effective at doses eight times and three times lower than each, respectively.

“If you can use less of an active ingredient in a formulation, it may be less expensive,” says Paskewitz, who, with Kajla, has filed for a patent related to this work through the Wisconsin Alumni Research Foundation (WARF).

The scientists are not quite sure how the bacterial extract deters mosquitoes from feeding, but the reason may be simply that it tastes bad.

Paskewitz and Kajla are now collaborating with colleagues at the UW–Madison School of Medicine and Public Health to test the safety of the compounds in human cell culture. Preliminary work suggests the presence of some toxicity, but the investigation continues.

“I am itching to put it on my hand and put it in a cage of mosquitoes, but I can’t do it yet,” says Paskewitz.

DEET is the most widely used insect repellent in the United States and has repeatedly been shown to be safe and effective, yet the public continues to express concern about its use, especially in young children. This is why some scientists continue to search for alternatives, Paskewitz says. Most research so far has looked to plants. But bacteria are common sources of antibiotics and other pharmaceuticals, and the species Bacillus thuringiensis is often used in agriculture to deter insects.

Kajla continues to explore the potential of the compounds extracted from Xenorhabdus budapestensis and says they may yet prove useful for other applications.

Paskewitz adds, “These compounds might end up being more effective against a wider array of biting arthropods. DEET works against ticks, but it’s not as good as it is with mosquitoes. We will test the bacterial compounds against other kinds of biting insects and their relatives.”

This study was funded by the National Institutes of Health grant no. AI123719.