Post provided by Hal Holmes

Each year Methods in Ecology and Evolution awards the Robert May Prize to the best paper in the journal by an author at the start of their career. Hal Homes has been shortlisted for their article ‘Focused ultrasound extraction (FUSE) for the rapid extraction of DNA from tissue matrices’. In this blog, Hal discusses how their paper came to be and the future applications of FUSE technology.

Applying biomedical technology to conservation challenges

My soon-to-be PI, Dr. Eli Vlaisavljevich, had just started a clinical trial using the novel ultrasound technology he’d spent years developing as a non-invasive therapy for liver cancer. I asked him if this technology could break down timber and other tough tissues in the same way it could erode tumor tissue into an acellular debris. He thought I was joking…

I had just finished the first draft of my PhD thesis in Bioengineering, which developed new microfluidic technology to enable rapid DNA testing that could be performed in the field to identify material from unknown species. This technology would be useful for those on the frontline of conversation as a screening tool to prevent illegal timber and wildlife trafficking, which is a critical driver of biodiversity loss. Through this work, I learned one of the greatest challenges for routine genetic testing in the field was not performing the actual test itself – but was simplifying and accelerating the arduous process of isolating DNA from the sample. This challenge is particularly acute for mechanically robust tissues like timber and bone.

During my PhD I had utilized technologies initially developed by biomedical engineers for use in medicine and global health to solve problems in conservation, and I considered how my background in biomedical engineering might now be applied to solve the problem of rapid DNA extraction. Dr. Eli Vlaisavljevich had been an early mentor to me and trained me in cell culture when we were undergraduate researchers. While on a catch-up call he described the advantages of his new technology for liver cancer therapy as “non-thermal” (DNA is destroyed by excessive heat) and able to “rapidly” erode tissue to an “acellular debris”. The penny dropped, I realized this technology may be the solution and began applying for opportunities to investigate the use of focused ultrasound technology for rapid DNA extraction. I was awarded a Schmidt Science Fellowship to conduct this work in Dr. Vlaisavljevich’s lab, I wasn’t joking.

Testing DNA extraction using focused ultrasound

Focused ultrasound technology is a form of histotripsy or “tissue breakdown”, which translates from the Greek histo– for tissue or structure and –tripsy for crushing. Histotripsy is accomplished by focusing pulses of ultrasonic energy at a focal location (similar to how light can be focused with a magnifying lens to start a fire), which then generates intense negative pressure that forms cavities or “void space” in the liquid medium. Cavitation can be imagined as looking like clouds of bubbles, and when these bubbles collapse they tear the tissue into microscopic particles.

My hypothesis was that the microscopic particles released during the cavitation process from a tissue sample would accelerate the release of entrapped DNA into the solution. This would speed up the extraction process via chemical digestion and eliminate the need for pulverizing robust tissue such as bone and timber, which requires cumbersome equipment or manual techniques that cannot be easily performed in the field. Ultimately, I believe this technology will enable rapid, routine genetic tests at the frontline of conservation and expand critical capacity for researchers to perform genetic sequencing at remote biodiversity hotspots in field laboratories.

Focus of ultrasound at a focal point creates an intense cavitation cloud. Credit: Hal Holmes

However, one question remained unanswered – what happens to target DNA in samples treated with this focused ultrasound technique? We knew that cavitation could shear DNA, as sonicators have been used to fragment DNA for library preparation in some sequencing procedures, but we did not know if these intense cavitation clouds would destroy the DNA contained with a tissue sample.

Using tissue samples from Atlantic salmon (Salmo salar) as a model, we were able to determine that FUSE technique could yield target DNA that was sufficient for both amplicon sequencing and shotgun sequencing. We observed the DNA acquired from FUSE treatment was of a quality and quantity of the gold standard DNA extraction process. Furthermore, we also observed that this technique could yield amplifiable DNA without any needing any biological or chemical compounds to break down cells and release DNA, such as digestion enzymes or permeabilizers. Our findings indicate that FUSE has the potential to readily integrate with microfluidics and offer more options for rapid sample-to-result testing.

Next steps and future work

The results of this initial study were quite promising which gave the green light to invest substantial development and engineering effort to begin adapting this technology for specific applied cases in conservation and genetics. Our team at Virginia Tech , together with key collaborators at University of Washington and Conservation X Labs, received support from the Gordon and Betty Moore Foundation to further this work, with a focus on adapting focused ultrasound technology for more challenging timber and plant tissues.

One of the primary goals of this collaboration is to miniaturize the large transducer arrays used for histotripsy and FUSE, which were originally designed for treating human patients, into smaller format transducer modules that are easy-to-use, affordable, and can even be transported into the field. Our work also addresses the breakdown mechanics and ultrasound connections with mechanically robust plant tissues. Ultimately, we aim to provide a rapid, easy-to-use system that can streamline and accelerate the bottlenecks of genetic testing and explore new applications for FUSE technology.

Discover more about the articles that were shortlisted for this year’s Robert May Prize here.