The Genetics Behind RNAi Technology for Viral Treatments in Honey Bees

By: Seanna Wengryn, TTP Summer Student

Honey bees are one of the most economically important pollinators and contribute approximately 6.1 billion dollars annually to the Canadian economy in pollination services (1). Alongside this, Canadian honey bees produce upwards of 75 million pounds of honey each year, adding another 253 million dollars to this contribution (1). Unfortunately, like many other agricultural commodities, the beekeeping industry is declining in its number of producers (2). This is because there has been a major increase in colony mortality throughout Canada and the world, with upwards of 45% of honey bee losses annually, averaging around 27% in the past 15 years (1). These losses have forced many hobbyist and commercial beekeepers to leave the industry, while hindering business planning and expansion for commercial producers that have endured (2). One of the biggest reasons for the decline in managed honey bees is extensive and unpredictable colony death due to the increased presence of virulent pathogens, including viruses, bacteria, and fungi (3). Viruses are a particularly challenging pathogen in honey bees, as there are currently no commercialized treatment options for producers (4). Additionally, viruses are transmitted through the varroa mite vector; an increasingly prevalent ectoparasite that has caused a significant burden to beekeepers worldwide. This highlights the importance of developing strategies to mitigate these pathogens, which is crucial for the health and welfare of honey bees, the livelihood of producers, and the Canadian economy.

The Central Dogma of Molecular Biology. The central dogma illustrates the flow of genetic information from DNA to RNA to protein. DNA is transcribed into RNA, which is then translated into a protein. It also includes reverse transcription, where RNA is reverse transcribed back into DNA, and DNA replication, where DNA is duplicated before cell division.

Ideally, the development of a viable viral treatment may reduce the need for frequent interventions, as improper chemical use, inadequate diagnostics, or mismanagement of pesticides can exacerbate bee diseases rather than alleviate them (2). Alternatively, a more sustainable approach would be to prevent the spread of disease altogether using genetic technologies. Genetics, a branch of biology that studies the inheritance of traits in organisms, offers promising avenues in this regard (5).

Before we delve into genetic technologies, we will briefly touch on the principles of genetics and gene flow, and explore how these principles can be utilized in the development of viral treatments. A gene is the basic unit of inheritance, containing genetic material that determines specific characteristics. Genes are passed from parent to offspring during reproduction. A gene is made up of DNA, which can be found in almost every cell in the body. DNA functions as a code that specialized enzymes can read. This code is transcribed into a small useful segment, known as messenger RNA, or mRNA (like DNA, but has an easier code to read), which carries the genetic information. Subsequently, other enzymes translate mRNA into a protein (Figure 1.) (5). These proteins constitute the building blocks of all living beings on Earth, including bees! The genome is the sum of all the genetic material in a cell and has many similarities and differences from one individual to the next. Genetic variation is what contributes to these differences, which primarily occurs through inheriting different versions of genes, also known as alleles (5). As a result, gene expression varies from bee to bee and can be influenced through natural inheritance or artificial manipulation. Gene expression changes over time; for instance, a viral infection can lead to an increase in the expression of genes related to immunity or disease protection (5). Scientists have been able to manipulate these biological processes to study diseases, breed animals towards more efficient and sustainable targets, and develop animals that are resistant, resilient, or tolerant to pathogen challenges (6). Along with the health and welfare benefits associated with an animal’s ability to cope with disease challenges, genetic biotechnology can optimize production and performance levels, therefore reducing potential production losses (6).

. The Basic RNAi Pathway in Honeybees. RNAi pathway starts with an enzyme cleaving double-stranded RNA (dsRNA) into small interfering RNAs (siRNAs). These siRNAs are then incorporated into a cutting complex, which targets and degrades viral messenger RNA (mRNA), leading to decreased gene expression.

The use of genetic tools for viral treatments in honey bees may be an innovative way to either reduce the spread of disease or completely remove it altogether. There are various tools already being explored, such as CRISPR-Cas9 gene editing, estimated breeding values within quantitative genetics, or more accurate bioinformatic databases. However, a promising technology in the beekeeping world is RNA Interference, or RNAi. RNAi is a natural antiviral immune mechanism already found in bees, along with other invertebrates, plants, and mammals (7). Scientists have been able to create an artificial version of this technology that allows them to precisely target viral mRNA and cut it into pieces (Figure 2.). They make use of the body’s natural production of small interfering RNA, or siRNA, that is made to match the virus’ mRNA. The siRNA then forms a ‘cutting’ complex with host enzymes and guides the group to the viral mRNA of interest. Finally, the complex cuts up the genetic material, resulting in non-functional, degraded viral mRNA that can no longer encode for a protein (7). This is crucial, as the virus will no longer have a method to replicate, which will in turn protect the honey bee from infection.

The main concern is always the safety of the animals along with the humans consuming their food products. RNAi is a natural process already present in bees, with the genetic technology primarily making use of building blocks within the animal. In the lab, double-stranded RNA (dsRNA), which is the precursor to siRNA, can be produced and easily consumed orally by honey bees in the field, making it feasible to implement at the production level (7). Although this genetic material must be artificially introduced into the bee, it breaks down quickly and will not cause permanent genetic changes to the animal.

. A honeybee infected with Deformed Wing Virus (DWV). This condition impairs the bee’s ability to fly due to its characteristic crumpled and misshapen wings. This virus strongly impacts the honeybee’s role in the hive and overall colony health. Photo taken by Shelley Hoover.

RNAi is a technology to help combat viruses as they are acquired by leveraging the body’s natural antiviral defense mechanisms (7). If implemented effectively, this technology may completely wipe out viral diseases altogether due to the newfound lack of susceptibility in honey bee hosts. In terms of honey, there should be no siRNA residues in the product, as again, the technology makes use of natural processes within the bee and degrades rapidly post-application. RNAi has already been shown to be highly effective in the treatment of Sacbrood Virus (SBV), Deformed Wing Virus (DWV), and Acute Paralysis Virus (APV) (Figure 3.) (3,8,9). However, more research is needed to further explore feasibility at a commercial level, viral resistance to siRNA, and the risk of off-target effects (7).

RNAi and genetic biotechnology have the power to combat any virus a bee may encounter, revolutionizing modern beekeeping throughout Canada and the world. The use of these technologies may help reduce the extensive amount of colony losses and significant disease-related challenges that beekeepers continue to face. With the agriculture sector constantly evolving towards more efficient and sustainable goals, the use of genetic tools within the beekeeping industry may be crucial for developing long-term disease management solutions. It may also help continue to grow the economy and provide further revenue for all producers, reducing the loss of small hobbyist farmers and continuing to support growing commercial beekeepers. Further exploration of genetic biotechnology within the beekeeping industry may lead to insights that enhance animal, economic, and producer outcomes.

 References

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