Amy Wray doesn’t expect everyone to love bats like she does, but she hopes to help people understand how essential they are to our ecosystem. Illustration by Danielle Lamberson Philipp / College of Agricultural and Life Sciences

Bats are flying throughout Amy Wray’s Twitter account. Her pinned tweet: “Live every week like it’s #batweek.”

What’s Bat Week? Wray, a doctoral candidate in wildlife ecology, is glad you asked. Observed (naturally) Oct. 24–31, Bat Week is an international celebration designed to raise awareness about the need for bat conservation. Wray just got back from the North American Society for Bat Research conference in Kalamazoo, Michigan. It’s a good time to hang (get it?) with fellow bat lovers — and also stock up on bat craft supplies.

“Especially stickers — you can’t have enough bat stickers,” Wray says.

She doesn’t expect everyone to love bats like she does, but Wray hopes to help people understand how essential they are to our ecosystem. What better time than Halloween?

Bat expert Amy Wray, a doctoral candidate in wildlife ecology, in a research lab in Russell Laboratories. Photo: Jeff Miller

Clearly you love bats. Why?

Bats are fascinating creatures  — they are the only flying mammal, they are super intelligent, and they are absolutely adorable. There are a lot of different kinds of bats too: white bats, speckly bats, cuddly bats, bats that catch fish, and even bats that hunt other bats. Sometimes we think of “bats” like they are all one type, when in reality, there are currently 1,411 different species.

When did this adoration begin?

When I was a little kid I was obsessed with the book “Is Your Mama a Llama?” and my favorite part of the story is about a bat, which had a very cute illustration. (Side note: The bat’s name is Dave.) And then, of course, “FernGully.”

“Bats can totally see,” Wray says. “Some fruit-eating bats have very big, puppy-like eyes and they can see well even in the dark. Other bats use echolocation to create a highly accurate sound-based image of the world around them, but those guys can also see.”

Settle it once and for all: “blind as a bat” — true or false?

False! Bats can totally see. Some fruit-eating bats have very big, puppy-like eyes and they can see well even in the dark. Other bats use echolocation to create a highly accurate sound-based image of the world around them, but those guys can also see.

Be honest. Do they want … to suck … my blood?

No! There are only three bat species that are true “vampires” who actually feed on blood. These species also much prefer to snack on animals like cows. It’s also not fair to think that only bats are vampires — insects are much more likely to feed on human blood, and in terms of other blood-feeding animals, there are also birds like the vampire finch (although, vampire finches don’t really bite humans).

What’s the most ridiculous thing you’ve heard about bats?

A new one that I just heard is that if you throw a marshmallow in the air at night, bats will come and catch it. So far, there have been no verified reports of such behavior.

I think the worst myth about bats is the myth that many of them carry rabies. Like most mammals, bats can carry rabies, but it’s actually pretty rare. It’s not a good idea to ever touch a bat without gloves (partly to protect the bat from our germs, too), but by simply treating them (and other wildlife) respectfully there is little risk of contracting diseases from them.

“One thing I really enjoy about bats is that they totally play hard to get,” Wray says. “Studying them has a lot of unique challenges since they are super wily, can be really small, and they’re kind of tricky to find or observe.”

Also, bats do not get stuck in people’s hair. I think this myth comes partly because sometimes if you are sitting outside where bats are foraging, they’re super curious and they may fly somewhat nearby to check out what you’re doing. They may also be foraging around insects that are trying to bite you. However, usually bats are not actually flying that close (it might seem like it because they’re flappy and hard to see). Bats are exceptionally agile, good fliers and they also have incredibly refined echolocation (like, they can “see” a single strand of a spider web or the pattern on a leaf), so they are definitely not going to fly into you.

Why do you think people are afraid?

I think fear usually stems from things that we don’t understand. Bats are kind of mysterious — flying around at night, hiding in small spaces, etc., so I think a lot of people hear myths and misinformation, but may be less likely to see bats up close. For a lot of people, an experience like having a bat stuck in their house, where the bat is going to be super afraid, is their only encounter with them. (If you do have a bat in your house, open a window and they can usually find their way out.) One thing that surprises people a lot is that up close, most bats (including all of the species in Wisconsin) are much smaller and more delicate than you might think they are.

“One thing that surprises people a lot is that up close, most bats (including all of the species in Wisconsin) are much smaller and more delicate than you might think they are,” Wray says.

Are people always giving you bat-related gifts?

The best bat-related gift I ever get is from citizen scientists who send me bat guano and/or data for my research. But in terms of bat-related stuff, people have gotten me some really good gifts like a bat-shaped cookie cutter and some cute bat dog toys. One of my lab-mates also got me some fake eyelashes with bats on them, which I love.

You’re not alone in having a fondness for bats. What do you think bat lovers have in common?

There are a few common threads that I think attract people to bats. First, for anyone curious about the natural world, there are so many different bats that do really interesting things, and yet we have so much to learn about them still. There is a real need for more bat scientists and more people interested in helping them.

Bats also get an unfair reputation as being scary, so they’re great animals for anyone who loves an underdog story. One thing I really enjoy about bats is that they totally play hard to get. Studying them has a lot of unique challenges since they are super wily, can be really small, and they’re kind of tricky to find or observe. Bats are all around us, but we don’t necessarily see them all the time, so it also feels really special to have opportunities to see them up close or even just be around them.

“Like a lot of wildlife, bats don’t really want to mess with people at all, they just want to keep on doing their own batty thing,” Wray says. “If we just leave them alone or handle them appropriately when necessary, there is no reason to be afraid of them.”

I’m sure it’s hard to choose, but do you have a favorite bat?

I have a few favorites, but one of them at the very top of my list is the proboscis bat (Rhynchonycteris naso). These bats are found in Central and South America, and they often roost on trees near the water, sitting in a neat little line, occasionally doing some wiggles. Their faces are outrageously cute in a super goofy way, where it looks like they kind of have an overbite, and it always makes me laugh to look at them.

Tell me again. Why shouldn’t I be afraid?

First step: google “baby flying fox,” then “bat eating banana,” and then “Honduran white bat.” These are the ultimate gateway bats. Like a lot of wildlife, bats don’t really want to mess with people at all, they just want to keep on doing their own batty thing. If we just leave them alone or handle them appropriately when necessary, there is no reason to be afraid of them. Bats also play really important roles in the ecosystem, and a lot of them need our help.

You’ve said a world without bats would be frightening. Why?

Bats are super important for the ecosystem. They do a lot of important things like pollinating plants, promoting forest diversity by dispersing seeds, and controlling insect pests that threaten agricultural systems. There are also so many mysteries about bats that are yet to be discovered, and I think it’s absolutely tragic to lose species before we can even learn about them.

Wray conducts a test with a sample vial of bat guano. Wray is using next-generation DNA sequencing to study the diets of insect-eating bats in Wisconsin. The findings will help her assess the extent to which bats feed on agricultural pests and how the spread of diseases may impact the utility of bats in pest control. Photo: Jeff Miller

The chemical, piperonyl butoxide or PBO for short, is widely used as a “synergist” in household and agricultural insecticides to make the toxic effects of the insecticide longer lasting and to reduce the amount of actual insecticide in a product.

Despite its widespread and growing use, as well as its ubiquity in the environment, the chemical has been little studied. Now, a team led by Robert Lipinski, a professor of comparative biosciences in the University of Wisconsin-Madison’s School of Veterinary Medicine, reports that PBO interferes with the critical signaling pathway dubbed by scientists as sonic hedgehog, resulting in stunted forebrain development and signature facial abnormalities. The study is published this week (Oct. 23, 2019) in the journal Environmental Health Perspectives.

A pair of fetal pig specimens with an abnormal proboscis structure and double-globe cyclopia, both more extreme forms of a birth defect known as holoprosenchephaly. Photo by: Jeff Miller

Robert Lipinski, professor of comparative biosciences in the University of Wisconsin-Madison’s School of Veterinary Medicine, looks at specimens. Photo by: Jeff Miller

This is a specimen of a calf’s head with cyclopia, the more extreme form of a birth defect known as holoprosenchephaly. Photo by: Jeff Miller

First synthesized in the 1940s, PBO was identified in 2012 as an inhibitor of the sonic hedgehog signaling pathway, a genetic pathway especially important in early development. The pathway is highly conserved, present in animals ranging from fruit flies to mice to humans.

Lipinski and his colleagues build on the discovery of PBO’s role in interfering with sonic hedgehog and show how that interference plays out in early development in mice. The signaling pathway is involved in multiple aspects of embryonic development, including the forebrain and face.

In the new study, Lipinski and his team exposed pregnant mice to PBO at a critical stage of development analogous to the period of human development when a condition known as holoprosencephaly arises.

Holoprosencephaly is exhibited in a range of developmental abnormalities, ranging from cleft lip and palate, to far more serious malformations. A dramatic manifestation of holoprosencephaly, known as cyclopia — where the median forebrain and face fail to develop, leaving a single central eye — was first linked in sheep to maternal consumption of a plant, a corn lily, that harbors the chemical cyclopamine. That chemical is also known to inhibit the sonic hedgehog developmental pathway.

“This is all about timing,” says Lipinski. “The critical period is very early in development.”

A previous study of household dust found PBO to be a common chemical contaminant, indicating the agent is ubiquitous in the environment. According to the Environmental Protection Agency, it is an ingredient in at least 1500 commercial products, including sprays and shampoos.

“It’s been said that birth defects like holoprosencephaly are caused by ‘a little bit of this and a little bit of that,’” says Lipinski, who is also the associate director of the UW–Madison Molecular and Environmental Toxicology Center. “We don’t know if PBO is contributing to birth defects in the human population, but our study suggests that more rigorous examination of PBO’s potential human health effects is warranted.”

In humans, live births with holoprosencephaly are very rare, occurring in roughly one in 10,000 newborns. However, it is believed to affect as many as one in 250 human embryos, most of which never make it to term.

PBO’s influence on human health has been the subject of relatively little scientific scrutiny, with only a handful of published studies. In humans, it can be found in “personal air samples,” and, in one study, was found in 75 percent of air samples provided by pregnant women. At present, labels of products containing PBO provide no cautionary information for exposure during pregnancy.

“Environmental PBO exposure comes from many different sources,” Lipinski says, “and human serum concentrations have not been measured. Part of our funded grant is to measure PBO concentrations in serum samples of women of childbearing age and to relate observed concentrations to those that are associated with developmental outcomes in mice.”

There is very likely a genetic component as well, says Lipinski, noting that at least 12 genes are known to be associated with holoprosencephaly, the most common being sonic hedgehog.

This research was supported by the National Institute of Environmental Health Sciences of the National Institutes of Health under award numbers R01ES026819 and T32ES007015.

Morgridge Institute for Research investigator Phillip Newmark

A research team led by Morgridge investigator Phillip Newmark reported in the Oct. 17, 2019 issue of PLOS Biology the successful characterization of this chemical, which could lead to new ways to fight the neglected tropical disease schistosomiasis. This disease, caused by schistosome infection, affects more than 240 million people in Africa, Asia and parts of South America.

In fact, schistosomiasis is second only to malaria in sheer numbers of people infected, and children are most often the victims. The disease is common in tropical areas where freshwater and human wastewater are not separated, allowing the schistosome to thrive.

The Newmark team focused on a phase of the schistosome life cycle that’s an intriguing target for preventing infection. Schistosomes seek out freshwater snails as hosts in order to produce millions of tiny fork-tailed creatures called cercariae, which are unleashed in the water and seek out mammals to infect. Their frenzied swimming allows them to penetrate human skin in minutes.

However, tiny aquatic creatures called rotifers also live on these snails and release a chemical compound that paralyzes cercariae on contact. Scientists have known about this factor for decades but have not further probed its biochemistry.

University of Wisconsin–Madison graduate student Jiarong Gao

In the new paper, the Newmark lab and collaborators in Jonathan Sweedler’s laboratory at the University of Illinois at Urbana-Champaign report their successful effort to purify and chemically define this molecule, calling it “schistosome paralysis factor” (SPF). Lead author and University of Wisconsin–Madison graduate student Jiarong Gao placed SPF in various concentrations in water and demonstrated that the compound immobilized the cercariae, which promptly sank to the bottom of the water and remained in that state. Further, she showed that cercariae exposed to SPF were unable to infect mice.

Newmark, a professor of integrative biology at UW–Madison and an investigator of the Howard Hughes Medical Institute, says the results could open a promising new path to controlling schistosomiasis. Currently only a single drug, praziquantel, is used to treat infection and is given to millions of school children each year. But it only kills adult schistosomes and does not stop reinfection.

“Any time you’re talking about treating that many people with just one drug and no alternative, you’re really concerned about the ability of the parasites to develop resistance,” Newmark says.  “And that’s becoming more and more of an issue as the geographic range of the parasite may be spreading and hybrids between human- and livestock-infecting schistosome species are being reported.”

The Phil Newmark Lab has isolated a natural chemical capable of paralyzing the parasitic worm schistosome, opening the door to new ways to combat a neglected tropical disease that sickens more than 240 million people. Video: Morgridge Institute for Research

Once schistosomes burrow inside host skin, the parasites migrate into the blood vessels and anchor on a vein that supplies the liver. During this process, the parasites reorganize their tissues, and upon reaching the liver, develop reproductive organs, pair with a mate, and grow into mature adults.

At this point, the adults can live for decades inside the host. They produce hundreds to thousands of eggs daily, many of which are not expelled through waste and become lodged in organs such as the liver. Children with the disease are often ravaged by anemia, malnutrition and pervasive learning disabilities. The World Health Organization estimates roughly 280,000 people die annually from schistosomiasis.

Schistosomiasis is second only to malaria in sheer numbers of people infected, affecting more than 240 million people in Africa, Asia and parts of South America. Children are most often the victims.

Newmark says one interesting element of SPF is that its structure resembles serotonin, a neurotransmitter most widely known for regulating mood. But it also has an impact on normal neuromuscular function and SPF may be interfering with that pathway.

This research would not have been possible without a key discovery in 1981 by Peg Stirewalt and Fred Lewis of the Biomedical Research Institute in Rockville, Maryland. They found that the rotifers — which are known for the flowing filaments on their heads that look like spinning wheels — produced a factor that paralyzed cercariae.

But until now, their sole impact on schistosome research was to keep them away from snails because they interfered with the infection process. Newmark turned the equation around and put the rotifers in the spotlight.

The Newmark lab first ventured into studying schistosomes about 10 years ago. The lab’s primary focus has been on the planarian, an organism that can regenerate its entire body from tiny fragments. The researchers recognized the many similarities between planarians and schistosomes, and then applied more than two decades of advances in planarian cell and molecular biology to study its parasitic relatives.

“All the work that we’ve done has been driven by our curiosity about these amazing planarians and all the different things that they can do,” Newmark says. “But we now have a new end game: We could actually wind up helping people in a tangible way, on a scale that we could not have considered before.”

The bald eagle was removed from the endangered species list in 2007 after recovering from a low of 412 nesting pairs in the contiguous United States. There are now about 1,700 nesting pairs in Wisconsin alone. U.S. Fish & Wildlife Service

Scientists at the University of Wisconsin–Madison, the U.S. Geological Survey  National Wildlife Health Center and the Wisconsin Department of Natural Resources found the virus while searching for the cause of Wisconsin River Eagle Syndrome, an enigmatic disease endemic to bald eagles near the Lower Wisconsin River. The newly identified bald eagle hepacivirus, or BeHV, may contribute to the fatal disease, which causes eagles to stumble and have seizures.

But BeHV was also found in eagles without symptoms of the syndrome, making a direct link between virus and disease difficult to confirm. The virus is related to human hepatitis C virus, which causes liver damage in people, and some birds with BeHV show similar effects. BeHV infects eagles from Washington to Florida but is most common in Wisconsin’s eagles.

Tony Goldberg

The researchers published their findings Oct. 18 in the journal Scientific Reports. Tony Goldberg, a UW–Madison professor of pathobiological sciences in the School of Veterinary Medicine, led the study. He collaborated with LeAnn White at the USGS National Wildlife Health Center and Sean Strom at the Wisconsin Department of Natural Resources.

Wisconsin River Eagle Syndrome (WRES) was first described in the 1990s. The river attracts eagles year-round because its open waters allow the birds to fish through the winter. Observers near the river spotted eagles vomiting or staggering, and all of these birds either succumbed to the disease or were euthanized.

Officials at the USGS National Wildlife Health Center conducted necropsies to determine the cause of death and found liver damage in most of the birds. Damage to brain tissue was also common. Searches for environmental toxins underlying the disease came up empty.

A microscopic image of a liver from a bald eagle that died from Wisconsin River Eagle Syndrome. The large white spaces demonstrate tissue damage characteristic of the disease. Photo: Marie Pinkerton

“Everything was pointing toward some unknown virus, but standard diagnostic techniques couldn’t find one,” says Goldberg.

Goldberg’s lab specializes in discovering new viruses. By analyzing all of the genetic material in infected tissue, his team can scan for the genetic signature of any virus within the sample. The technique does not require prior knowledge of what kind of virus might lurk within an animal.

Working with White and Strom, Goldberg’s lab analyzed liver tissue from nine birds diagnosed with WRES. The team first spotted the new virus in a bald eagle collected in 2002 in Sauk County. This family of viruses had not been found in birds before, although it has since been identified in ducks as well.

Testing 47 eagles from 19 states across the contiguous United States, the research team found that 32 percent harbored BeHV. Researchers found the virus in seven states, including several in the Midwest, but also in states as far apart as Washington and Florida. Eagles in Wisconsin were nine times more likely to carry BeHV than birds from other states, and the virus was 14 times more common in counties surrounding the Lower Wisconsin River than elsewhere.

Yet the presence of BeHV in birds collected outside of Wisconsin — none of which had been diagnosed with WRES — suggests the virus may not be responsible for the enigmatic disease, or that the story is more complex.

“Is BeHV the cause of WRES?” asks Goldberg. “Or is it more complicated than that?”

There is a long list of possible explanations. Infected birds may be more likely to die of starvation or other illnesses before they show symptoms of WRES, but eagles collected from outside Wisconsin did not show the liver damage characteristic of WRES.  It’s also possible that the prime conditions near the Wisconsin River allow birds to survive long enough for the disease to progress to its end stage.

“It is curious that the liver pathology of the eagles resembles the damage to human livers caused by hepaciviruses,” says White, branch chief for wildlife epidemiology and emerging diseases at the USGS National Wildlife Health Center. “But since the lesions are nonspecific there’s more that needs to be explored if we want to understand the virus itself or really get at what is the cause of WRES.”

The virus and the syndrome do not appear to endanger the resurgence of the bald eagle, which was removed from the endangered species list in 2007 after recovering from a low of 412 nesting pairs in the contiguous United States. There are now about 1,700 nesting pairs in Wisconsin alone.

“We don’t think this virus is having a serious impact on the bald eagle population, but the fact that WRES is an unknown condition keeps our interest,” says Strom, an environmental toxicologist with the DNR. “This study is another piece of the puzzle. Hopefully we can find more pieces and figure out what is happening.”

“This study has opened our eyes to glaring knowledge gaps about infection in a species of great national importance,” says Goldberg. “It’s a more complicated story than we thought it might be at first, but that makes it more interesting.”

This work was supported in part by the U.S. Geological Survey (award number G14AC00364).

The silver-containing MicroLyte bandage is thinner than a human hair and able to be absorbed into the wound. The material is shown here on a fabric background. Photo: Imbed Biosciences

Millions of people with severe burns or diabetic skin ulcers could benefit from an experimental enhancement to a next-generation covering that is already healing difficult wounds.

A $1.5 million, two-year Small Business Innovation Research grant announced today will test whether adding gallium metal ions to an ultra-thin material carrying antimicrobial silver can defeat the “biofilms” that shield bacteria from antibiotics.

Persistent wounds are distressingly common, says Ankit Agarwal, founder and CEO of Imbed Biosciences, the Fitchburg, Wisconsin-based University of Wisconsin–Madison spinoff that makes and sells the silver-bearing covering.

Ankit Agarwal

According to a scientific report published in August 2019, “In the United States, chronic ulcers affect more than 6 million people, with increasing numbers in the growing elderly and diabetic populations.”

In many cases, the chronic wounds remain open for months, due to difficult-to-treat infections that are often shielded by a biofilm.

Imbed’s silver-covering technology was invented in the lab of Nicholas Abbott, then a professor of chemical engineering at UW–Madison. Abbott guided Agarwal’s postdoctoral studies on the technology.

Imbed’s existing product, called MicroLyte, was cleared by the FDA for marketing in the U.S. in August 2016. MicroLyte is on the formulary of five major hospital systems, including UW Health, and is used in more than a dozen other hospitals across the country. “The formulary is a list of medications readily available for use at a hospital or health system,” Agarwal says.

MicroLyte contains a tiny dose of metallic silver particles on an absorbable polymeric multilayer film. Flexible and only 25 micrometers thick (thinner than a human hair), the covering places silver in direct contact with bacteria.

MicroLyte can eliminate the painful process of repeated removal that is needed when conventional bandages are replaced on tenacious wounds.

Silver’s antimicrobial activity rests on its ability to punch holes in the bacterial cell wall, but a biofilm can block that exposure. Chemically, gallium ions resemble a form of iron that cells use to gain chemical energy through the process of reduction.

“Bacteria inside the biofilm are looking for more iron, so they end up taking up gallium,” Agarwal says. But gallium ions cannot be reduced, and the bacteria atrophy for want of energy, helping to break up the biofilm. “So in return for accepting a worthless ‘Trojan horse,’ the bacteria are exposed to silver — the second punch — and they will die.

By placing nanometer-sized particles of both metals on the ultra-thin film, “we are using levels of both metals that are so low that they are not toxic to human cells, placed on a film that actually supports growth of skin cells,” Agarwal says.

Under the new grant, silver plus gallium will be tested on full-thickness wounds in pigs in the UW–Madison School of Veterinary Medicine.

Before: A venous stasis ulcer on the leg on a 74-year-old man did not respond to standard care over 35 days. In the U.S., chronic ulcers affect more than 6 million people.

After: Weekly application of Imbed Biosciences’ silver-bearing bandage jump-started the healing and the wound was closed in six weeks. Photos: Imbed Biosciences

“To simulate chronic wounds, we will surgically create wounds and inoculate them with pre-established biofilms of antibiotic-resistant bacteria,” says Jonathan McAnulty, chief of surgery at the vet school. “Healing in porcine skin wounds is similar to healing in humans.”

McAnulty will head the animal studies in collaboration with Charles Czuprynski, chair of pathobiological sciences. Both researchers have worked for years with the silver-based dressings.

“We see animals with skin injuries that get colonized by bad actors,” says McAnulty, “including different strains of antibiotic-resistant staphylococcus. These wounds don’t want to heal, despite our best efforts, and can go weeks or months without progress. This clinical scenario is almost identical to human cases.”

Although the silver-bearing covering is highly effective in healing infections and closing many difficult wounds, the gallium is intended to enhance that activity by defeating bacterial biofilms. “Behind a biofilm, bacteria are more protected against any agent trying to get in and kill them,” says McAnulty. “If we can prevent the biofilm from forming, or if it is present and we can get it to dissolve, it will expose the bacteria to killing agents.”

Research at the veterinary school has already shown that the combination film can “completely disperse the bacteria biofilms created by a mix of two pathogenic bacteria,” Agarwal says. “Treatment with a commercial, silver-based dressing — not MicroLyte — had no effect.”

Chronic ulcers affect more than 6 million people in the U.S. In many cases, the wounds remain open for months due to difficult-to-treat infections that are often shielded by a biofilm.

Angela Gibson, a burn surgeon and medical director of UW Health wound healing services, has used MicroLyte on patients of all ages when conventional products have failed. “The property that I find unique in MicroLyte is the total conformation to the wound bed as the thin film melts into all the crevices of a wound likely where the bacteria are hiding and preventing healing. It is also very easy to apply and may decrease irritation, such as itching, caused by other products.”

Gibson says she looks forward to a clinical trial of the product to determine if her experience holds true in a larger study of patients.

Finding metal ions to defeat bacteria is simpler than the usual approach of finding or inventing molecules, Agarwal says. “We like to keep it simple. We already had silver working, but we thought, ‘Let’s take another metal, one that’s has been used in other FDA-approved formulations, and put it into the matrix in combination with silver, and see if it gets into the biofilm and sensitizes the bacteria to silver, which will kill it.”

An aging population, rising rates of diabetes, and a growing problem of antibiotic-resistant bacteria all portend a bigger problem with persistent wounds. If gallium’s Trojan horse strategy proves able to defeat biofilms, that will be a first, Agarwal says. “Today there is no commercially available formulation that is indicated for dispersal of biofilm in a wound. Zero.”

The research is funded by a phase II Small Business Innovation Research  grant #2R44AR073710-02 from the National Institute of Arthritis and Musculoskeletal and Skin Diseases at the National Institutes of Health.

Researchers at the University of Wisconsin–Madison have addressed many of those problems by packing a gene-editing payload into a tiny, customizable, synthetic nanocapsule. They described the delivery system and its cargo today (Sept. 9, 2019) in the journal Nature Nanotechnology.

Sarah Gong

“In order to edit a gene in a cell, the editing tool needs to be delivered inside the cell safely and efficiently,” says Shaoqin “Sarah” Gong, a professor of biomedical engineering and investigator at the Wisconsin Institute for Discovery at UW–Madison. Her lab specializes in designing and building nanoscale delivery systems for targeted therapy.

“Editing the wrong tissue in the body after injecting gene therapies is of grave concern,” says Krishanu Saha, also a UW–Madison biomedical engineering professor and steering committee co-chair for a nationwide consortium on genome editing with $190 million in support from the National Institutes of Health. “If reproductive organs are inadvertently edited, then the patient would pass on the gene edits to their children and every subsequent generation.”

Most genome editing is done with viral vectors, according to Gong. Viruses have billions of years of experience invading cells and co-opting the cell’s own machinery to make new copies of the virus. In gene therapy, viruses can be altered to carry genome-editing machinery rather than their own viral genes into cells. The editing machinery can then alter the cell’s DNA to, say, correct a problem in the genetic code that causes or contributes to disease.

Krishanu Saha

“Viral vectors are attractive because they can be very efficient, but they are also associated with a number of safety concerns including undesirable immune responses,” says Gong.

New cell targets can also require laborious alterations of viral vectors, and manufacturing tailored viral vectors can be complicated.

“It is very difficult — if not impossible — to customize many viral vectors for delivery to a specific cell or tissue in the body,” Saha says.

Gong’s lab coated a gene therapy payload — namely, a version of the gene-editing tool CRISPR-Cas9 with guide RNA designed in Saha’s lab — with a thin polymer shell, resulting in a capsule about 25 nanometers in diameter. The surface of the nanocapsule can be decorated with functional groups such as peptides which give the nanoparticles the ability to target certain cell types.

The nanocapsule stays intact outside cells — in the bloodstream, for example — only to fall apart inside the target cell when triggered by a molecule called glutathione. The freed payload then moves to the nucleus to edit the cell’s DNA. The nanocapsules are expected to reduce unplanned genetic edits due to their short lifespan inside a cell’s cytoplasm.

This project is a collaboration combining UW–Madison expertise in chemistry, engineering, biology and medicine. Pediatrics and ophthalmology professor Bikash R. Pattnaik and comparative biosciences professor Masatoshi Suzuki and their teams worked to demonstrate gene editing in mouse eyes and skeletal muscles, respectively, using the nanocapsules.

Because the nanocapsules can be freeze-dried, they can be conveniently purified, stored, and transported as a powder, while providing flexibility for dosage control. The researchers, with the Wisconsin Alumni Research Foundation, have a patent pending on the nanoparticles.

“The small size, superior stability, versatility in surface modification, and high editing efficiency of the nanocapsules make them a promising platform for many types of gene therapies,” says Gong.

The team aims to further optimize the nanocapsules in ongoing research for efficient editing in the brain and the eye.

This research was supported by grants from NIH (UG3 NS111688, R01EY024995, 1R35GM119644-01 and R01NS091540) and National Science Foundation (CBET-1350178 and CBET-1645123).

As outbreaks on Pacific islands and in the Americas in recent years made Zika virus a pressing public health concern, the Zika virus’s close similarity to dengue presented the possibility that one infection may exacerbate the other.

Dawn Dudley Photo: Bryce Richter

Dengue virus infections are infamous for being bad the first time around. But following infection with one of the four variants (called serotypes) of dengue with an infection by a different serotype can amplify the already dangerous symptoms — high temperature, fatigue and pain — and make dengue fever even more life-threatening.

“When that second dengue virus occurs, antibodies kind of recognize it, but not in a way that allows them to take the virus out of the system and neutralize it like normal,” says Dawn Dudley, a scientist in the University of Wisconsin–Madison’s Department of Pathology and Laboratory Medicine and one of the authors of the new Zika study. “Instead, they have kind of a secondary effect, where by binding to the virus loosely they actually enhance the ability of the virus to get into other cells in the body and replicate more.”

Studies in tissue cultures and mice of back-to-back Zika and dengue infections suggested that the two members of the Flaviviruses — a genus that also includes West Nile virus and yellow fever virus — could interact to enhance each other. Data collected from human infections since the UW–Madison group began its work in 2017 appeared to contradict those tissue culture and mouse findings.

Meghan Breitbach

The study of 21 Wisconsin National Primate Research Center macaque monkeys, in which animals infected with one virus were challenged with another within nine to 12 months, supports the human epidemiological results.

“Whether it was a primary infection with one of the dengue serotypes followed by a Zika infection, or Zika first with a later dengue infection, we didn’t see anything unusual in those secondary infections,” says UW–Madison pathology research specialist Meghan Breitbach, also an author of the study.

Monkey weights, body temperatures, red and white blood cell counts, liver function and markers of cell damage did not stray significantly from typical infection levels.

“Because we’ve done several prior studies of Zika virus infections, we have a lot of historical data on what a typical infection looks like in these animals,” says Christina Newman, study author and UW–Madison scientist in Pathology and Laboratory Medicine. “For the animals that were experiencing a secondary Zika virus infection after primary dengue infection, their viral loads were almost indistinguishable from animals that were only ever infected with Zika.”

Christina Newman

That news, which was published today in the journal PLOS Pathogens, is a positive development. But it comes with a caveat important to Zika: none of the study’s monkeys were pregnant. Zika’s most visible and troubling results are neurological problems in babies whose mothers were infected during pregnancy, though those complications vary widely.

“The immune system is different in pregnancy,” Dudley says. “Previous dengue immunity may still be one of the reasons that some women have severe congenital Zika syndrome outcome in their infant while another woman with a known Zika infection doesn’t.”

A UW–Madison study of pregnant monkeys encountering both viruses could soon help describe whether back-to-back infections are more dangerous for the monkeys and their offspring.

The newly published study, which was supported by the National Institutes of Health, also represents a snapshot of monkeys encountering infections roughly one year apart, Newman says.

Dengue fever is enhanced by an earlier dengue infection only during certain conditions dependent on the serotypes of dengue involved, whether the immune memory produced by the initial infection was relatively strong or weak, and how much the antibodies created may have faded over months or years. The complicating factors have led to caution in development of Zika and dengue vaccines for fear of sparking more severe infections later.

“Our study suggests that that is unlikely,” Newman says. “But as we learn more about people whose infections come two or three years apart, we may see we need to combine a Zika vaccine with a good vaccine against all four serotypes of dengue virus to prevent enhancement of either virus.”

One major challenge in treating cancers and other disorders of the brain is ensuring that medicines reach their targets. A team of biomedical engineers and clinician-scientists at the University of Wisconsin–Madison and the University of Texas at Austin borrowed molecules from the immune system of the parasitic sea lamprey to deliver anti-cancer drugs directly to brain tumors.

Lampreys and humans have similar immune systems. But instead of producing antibodies to neutralize threats (that’s how vaccines help protect us against measles), they produce small defensive molecules. iStock Photo

They published their results today (May 15, 2019) in the journal Science Advances.

Unlike most currently used medicines, which target specific features on or inside individual cells in our body’s organs and tissues, the lamprey-derived molecules take aim at a different target — the extracellular matrix, a tangled mesh of proteins and sugars that supports and surrounds all cells in the brain.

The researchers believe the molecules could be adapted and combined with a wide array of other therapies, offering hope to treat numerous brain ailments beyond tumors, such as multiple sclerosis, Alzheimer’s disease or even traumatic injuries.

“This set of targeting molecules appears somewhat agnostic to the disease,” says Eric Shusta, a professor of chemical and biological engineering at UW–Madison. “We believe it could be applied as a platform technology across multiple conditions.”

The technology takes advantage of the fact that many diseases disrupt one of the body’s natural defense mechanisms: the blood-brain barrier, which lines the blood vessels of the central nervous system and protects the brain from potential threats such as circulating toxins or pathogens.

Many drugs — including the lamprey-derived molecules — cannot reach targets in the brain when they are injected into the bloodstream, because the blood-brain barrier normally prevents large molecules from leaving the blood vessels in the brain.

Chemical and biological engineering professor Eric Shusta, right, and postdoctoral researcher Ben Umlauf at work in the lab, where they helped develop molecules from parasitic lamprey for use in treating disorders in the brain. UW–Madison photo by Sam Million-Weaver

Yet, in conditions such as brain cancer, stroke, trauma and multiple sclerosis, the barrier becomes leaky in and around the disease locations. A leaky barrier offers a unique point of entry. It will allow the matrix-targeting lamprey molecules to access the brain and deliver drugs precisely on target.

“Molecules like this normally couldn’t ferry cargo into the brain, but anywhere there’s a blood-brain barrier disruption, they can deliver drugs right to the site of pathology,” says Shusta.

Knowing that brain tumors often cause the barrier to leak, the researchers linked the lamprey-derived molecules to a Food and Drug Administration-approved chemotherapy called doxorubicin. The treatment prolonged survival in mouse models of glioblastoma, the incurable brain cancer that afflicted Senators John McCain and Ted Kennedy.

The matrix-targeting strategy means a wide variety of therapies could be linked to the lamprey-derived molecules. They could also be combined with techniques that temporarily open the blood brain barrier at specific brain sites. And it’s possible that drugs delivered to the matrix could accumulate to a much higher therapeutic dose than medicines aimed at the inside of cells.

“Similar to water soaking into a sponge, the lamprey molecules will potentially accumulate much more of the drug in the abundant matrix around cells compared to specific delivery to cells,” says collaborator John Kuo, a neurosurgeon-scientist and professor of neurosurgery in the Dell Medical School at the University of Texas at Austin.

Additionally, brain cells actively pump out many chemicals — a useful trick to protect against toxic compounds, but a major headache for achieving effective therapeutic doses for medicines.

Targeting the matrix that surrounds the cells sidesteps that pumping problem.

“This could be a way to hold therapies in place that don’t otherwise accumulate well in the brain so they can be more effective,” says Ben Umlauf, a postdoctoral scholar in Shusta’s group who isolated the lamprey-derived molecules.

In the future, the researchers plan to link the matrix-targeting molecules to additional anti-cancer drugs, such as immunotherapy agents that activate a patient’s own immune system to destroy tumors.

Lampreys and humans have similar immune systems. But instead of producing antibodies to neutralize threats (that’s how vaccines help protect us against measles), they produce small crescent-shaped defensive molecules called VLRs. To obtain their drug-delivery molecules, the researchers “vaccinated” lampreys with components of the brain extracellular matrix and then hunted through many thousands of VLRs to find one that stuck specifically to the brain matrix.

Importantly, in the mouse studies, the lamprey-derived molecules circulated throughout the body without accumulating in healthy brain tissue or other organs. This targeted delivery is especially important in cancer treatments, since many therapies frequently cause debilitating adverse reactions due to indiscriminate effects on healthy cells.

In the future, the researchers plan to link the matrix-targeting molecules to additional anti-cancer drugs, such as immunotherapy agents that activate a patient’s own immune system to destroy tumors.

They also see promise in using the molecules as diagnostic tools to detect blood-brain barrier disruption by linking the matrix binders with probes for advanced imaging with PET scanners or MRI machines.

And because the molecules appear to be quite adaptable, the researchers speculate that many other medicines for the brain could become more effective if they were targeted to the matrix.

“I’m excited about trying this strategy in different disease model systems,” says Kuo. “There are several disease processes that disrupt the blood-brain barrier and we could conceive of delivering a variety of different therapies with these molecules.”

Other collaborators on the study include Paul Clark, Jason Lajoie, Julia Georgieva and Samantha Bremner at UW–Madison and Brantley Herrin at Emory University.

The work was supported by grants from the National Institutes of Health (NS091851 and NS099158) and the Defense Threat Reduction Agency (HDTRA1-15-1-0012) and Falk Medical Research Trust Catalyst Award.

Common marmosets at the Wisconsin National Primate Research Center. A new study found that marmosets whose medical histories included inflamed colons had more of a Parkinson’s disease-related protein in their intestines. Photo: Jordana Lenon / WNPRC

The intestinal linings of monkeys with inflamed bowels show chemical alterations similar to abnormal protein deposits in the brains of Parkinson’s patients, lending support to the idea that inflammation may play a key role in the development of the degenerative neurological disorder.

A study published by University of Wisconsin–Madison researchers today in the Journal of Inflammation Research found phosphorylated alpha-synuclein — a modified version of a protein common to nerve cells — in samples from common marmosets kept in a tissue bank at the Wisconsin National Primate Research Center.

“It’s not entirely clear what its function is, but the typical version of the protein alpha-synuclein occurs normally in all neurons,” says Marina Emborg, a professor of medical physics in the UW School of Medicine and Public Health and a Parkinson’s disease researcher. “A lot of neurodegenerative disorders seem to be related to the aggregation of certain proteins. When you have Parkinson’s, alpha-synuclein changes its shape and aggregates with other proteins into masses called Lewy bodies.”

Marina Emborg

The Lewy bodies aren’t necessarily a cause of Parkinson’s, but they are a hallmark of the disease, which affects more than 10 million people worldwide. Parkinson’s progressively degrades the nervous system, causing characteristic tremors and dangerous loss of muscle control.

Parkinson’s patients also suffer gastrointestinal problems, symptoms described in detail as early as 1817 by James Parkinson in his original essay on the disorder that would come to bear his name.

People who suffer from inflammatory bowel disorders are more likely to be diagnosed with Parkinson’s — one of several clues that inflammation and oxidative stress may be involved in the disease. Inflammation had been proposed as a possible trigger for the alteration of normal alpha-synuclein into the phosphorylated alpha-synuclein found in Lewy bodies.

“The colon, the gastrointestinal tract overall, has this dense network of nervous tissue, the enteric nervous system, which is sometimes called the gut brain,” says Emborg, whose work is supported by the National Institutes of Health and the Parkinson’s Foundation. “This has lots of neurons, and those neurons — like all neurons — have alpha-synuclein.”

When the Emborg research group heard from primate center pathologists that marmosets sometimes deal with inflamed bowel problems like colitis, they decided to test marmoset tissue samples for changes in alpha-synuclein. The researchers found that marmosets whose medical histories included inflamed colons had more of the phosphorylated alpha-synuclein in their intestines.

“It shows us the relationship between inflammation and Parkinson’s-like alpha-synuclein pathology,” says Emborg. “It doesn’t mean if you have inflammatory bowel disorder, you will get Parkinson’s. The development of a neurodegenerative disorder is multifactorial. But this could be a contributing factor.”

Former UW–Madison undergraduate student Henry Resnikoff and graduate student Jeanette Metzger are lead authors on the study.

This research was supported by grants from the Parkinson’s Foundation and the National Institutes of Health (P51OD011106, UL1TR000427, R24OD019803, F31HL136047).

David Vail, right, professor and oncologist at the UW School of Veterinary Medicine, delivers the very first vaccine in the Vaccination Against Canine Cancer Study to Trilly, a nine-year-old Gordon Setter, as she is held by veterinarian Kara Magee, left, and veterinary technician Rubi Hayem. Photo by Meghan Lepisto

On Thursday, May 2, the very first dog received the very first vaccine intended to protect her from cancer. And soon after the 9-year-old Gordon setter named Trilly received her shot, so, too, did Norton, a 9-year-old rat terrier mix.

“We’re testing a totally novel way of creating an anti-cancer immune response,” says David Vail, a professor and board-certified oncologist at the University of Wisconsin–Madison School of Veterinary Medicine. “The holy grail would be to prevent cancer as opposed to waiting for it to start and then treating it.”

Veterinary technician Abbey Ace holds her dog Norton, a nine-year-old rat terrier mix, after he received one of the first vaccines in the Vaccination Against Canine Cancer Study, a five-year clinical trial to evaluate a vaccine strategy for the prevention of cancer in dogs. The UW School of Veterinary Medicine is one of three participating institutions. Photo by Meghan Lepisto

If the vaccine works in dogs, Vail says, it may not only provide a new strategy for addressing a critical canine health concern, it might also work in people.

The Vaccination Against Canine Cancer Study, now underway, will evaluate a vaccine strategy for the prevention of cancer in dogs. Much like an influenza vaccine bolsters the body’s readiness to fight the flu, this preventative vaccine follows the same principle, “to have the immune system primed such that if a cancer cell develops, it will attack,” Vail says.

More than 800 patients are enrolled in the study, making it the largest clinical trial conducted to date for canine cancer, and across the history of veterinary medicine. The UW School of Veterinary Medicine is one of three participating institutions.

Cancer is the number one cause of illness and death in the aging dog population, with approximately one out of every three dogs affected and six million new cancer diagnoses made in dogs each year.

“The vaccine may not be effective, but this is probably the only approach to this type of vaccine, so we feel we have to try it. The implications of success would be quite large — for dogs and people,” says Stephen Albert Johnston, professor and director of the Center for Innovations in Medicine at Arizona State University, who developed the technology behind the vaccine.

Traditional vaccines work by introducing into the body a protein found on the surface of the virus that the vaccine is protecting against. The immune system sees the protein as a threat, establishes a memory of it, and, if there is a later infection, recognizes that protein and is primed to react.

“It’s almost like putting up a wanted poster,” Vail explains. “When that virus infects you, the immune cells recognize it because of the ‘poster.’ Then, the immune cells go out and kill it.”

Trilly, a nine-year-old Gordon Setter, received the very first vaccine in the Vaccination Against Canine Cancer Study, a five-year clinical trial to evaluate a vaccine strategy for the prevention, rather than the treatment, of cancer in dogs. With more than 800 patients enrolled, it is the largest clinical trial conducted to date across the history of veterinary medicine. Photo by Meghan Lepisto

The anti-cancer vaccine being tested targets approximately 30 abnormal proteins found on the surface of cancer cells. These proteins, a result of improperly coded RNA — so-called frame-shift mutations — are generally only found in patients with cancer (in dogs and people).

By injecting this cluster of proteins into healthy patients, along with a substance that stimulates an immune response, researchers believe the vaccine could serve as a universal defender against cancer by “turning on” the immune system.

The vaccine will target several cancers common to dogs, including lymphoma, a cancer of the lymphatic system; osteosarcoma, or bone cancer; hemangiosarcoma, a deadly cancer that originates in the blood vessels and is almost exclusive to dogs; and mastocytomas, or mast cell tumors.

The potential to preventively target several types of cancer with a single vaccine series would be a major paradigm shift in veterinary and human medicine, explains Vail, who is also a member of the UW Carbone Cancer Center.

Researchers at UW–Madison and elsewhere have developed a number of anti-cancer vaccines in recent decades for the treatment of cancer after it has been diagnosed. But those vaccines are tailored to a specific cancer type and are often produced for an individual patient, making broad deployment worldwide both time- and cost-prohibitive.

This year, nine million people around the world will die from cancer, Vail explains, and 70 percent of them are poor or living in developing countries where access to treatment may be limited. A preventative vaccine, if available to people, could be “applied globally at low expense,” he says, adding: “It’s a whole new way of looking at anti-cancer vaccines. The key is that you don’t have to personalize the vaccine to an individual, which is a very expensive proposition.”

Initial trials of the vaccine in mice suggest the new strategy could be successful, but many in the scientific community remain unsure.

“It’s so outside of the current box. Based on what we know right now about the immune system, there are several reasons why this vaccine shouldn’t work and there are several reasons why it could work,” says Vail. “I go in with healthy skepticism, but if this works, or even if this is one step in the correct direction, I will be thrilled.”

The trial is slated to run over five years. Cancer-free, healthy dogs between the ages of six to 10 will be randomized to receive either a series of the investigational vaccine or placebo vaccines. (In fact, researchers don’t know which version Trilly and Norton received)

Two sets of vaccines will be given every two weeks, for a total of four treatments, and then annually. Researchers have not observed any side effects other than those typical to any vaccine, such as moderate local pain or swelling at the site of injection, in mice or dogs to date, but the study will characterize any unanticipated adverse reactions in the larger study population.

Participating dogs will live at home and be checked two-to-three times yearly for five years after enrollment to monitor them for the development of any cancers. Their medical care will be covered by the study, which is funded by a $6 million grant from the Open Philanthropy Project. It is also supported by Calviri, Inc., a company Johnston started to commercialize the vaccine if it is effective.

“Even if a patient is randomized to the placebo group, they will enjoy five years of the best medical care available at no cost,” Vail notes. “And funds are available to diagnose and treat cancers that may develop in dogs in both the treatment and placebo group.”

About 280 dogs will be treated at UW Veterinary Care, with the remainder of participants at the other two trial sites, Colorado State University and the University of California, Davis.

The School of Veterinary Medicine is a national leader in the field of research known as comparative oncology, studying naturally occurring cancers in pets as models for human disease to advance new therapies and diagnostics that could benefit animals and people.

The array of information gleaned throughout the trial will also advance scientists’ understanding of cancer, the immune system, and how it responds, says Vail. “This is a team project involving dozens of researchers across the country and hundreds of pets and pet owners. We’re so thankful that they’re with us and that they want to strive to push the envelope as far as what we can do to prevent cancer in the future.”