Lauren Fuge
Bee-reaking news: Belgian scientists have developed a simple method to cryopreserve the semen of drones, the male equivalent of honey bees (Apis mellifera).
As part of the FreezeBEE project at the University of Liège, the researchers then thawed the semen and successfully artificially inseminated queens. It resulted in a female brood with an equivalent viability to bees inseminated with fresh semen.
Unlike existing preservation methods, this didn’t require antibiotics for success, opening up new opportunities in apiculture and bee conservation.
Let’s get down to bees-ness
“Bees play a crucial role in the ecosystem and agriculture,” explains lead author of the research Sophie Egyptien, from the University of Liège.
About 35% of global crop production relies on bees. But they face a host of threats, including habitat loss, climate change, pesticides, fertilisers and disease, such as the parasitic mite Varroa destructor that targets the European honey bee.
“The conservation of their genetic material, such as drone semen, is a key strategy to preserve biodiversity and support breeding programmes,” Egyptien says.
Winging it
It may surprise you to know that researchers have experimented with storing bee semen for decades.
The early 20th century saw the development of artificial insemination techniques for honey bees, and since then it’s become integral to breeding and research programs around the world. Finding ways of storing drone semen was the necessary next step.
(Are you wondering how, exactly, researchers collect this genetic material from a bee? According to entomologist Brad Hopkins from the United States Department of Agriculture, “You pretty much just squeeze them, and the stuff pops out.” Unfortunately, he adds, drones die after successful mating.)
In 1960, US entomologists reported in Science that they had successfully inseminated honey bee eggs with sperm that had been stored at above-freezing temperatures for 68 days. By the 1980s, several protocols for freezing drone semen had been developed, though they were often complex and required the use of antibiotics to control microbial contamination. Their outcomes were also not always reproducible.
One of the last papers published on cryopreservation of honey bee semen in the 1980s indicated that the method was not yielding good enough results to pursue it further, and so researchers returned to the traditional method of storing semen – at room temperature, where it remains viable for about a fortnight.
In the 2000s, researchers again turned their attention again to cryogenic freezing. The method had proven successful in preserving the semen of other animals like cows, though drone semen is different to mammal semen so the method had to be refined.
The cryopreserve buzz
Up until now, all cryopreservation methods for drone semen used antimicrobial agents.
“We wanted to see if an antibiotic-free recipe could be a viable option,” says Stefan Deleuze, from the University of Liège. “We also wanted to test a single-step procedure as we ideally would like to be able to offer the technique to beekeepers and not for research purposes only.”
After collecting the sample of semen and analysing it to check viability, Deleuze and colleagues diluted some in a medium containing cryoprotectants. When a cell is frozen and thawed, the ice crystals that form can damage cell structures – but cryoprotectants can help protect them by reducing ice formation.
The team then slowly froze the dilution. Meanwhile, they used the rest of the still-fresh semen to immediately inseminate 13 queens to create a control group. To achieve this, the queens were anesthetised with CO2 for 5 minutes and inseminated with a specialised syringe.
The next day, the frozen semen was thawed and its viability was again checked. It was then used to inseminate 16 queens.
“Despite a 37% loss of sperm viability during freezing, 5 out of the 8 queens inseminated with this frozen semen produced female brood,” says Deleuze. “These results are comparable to those obtained with fresh semen in our study.”
Breeding across time and space
This is the first antibiotic-free method for semen preservation to prove effective in female brood production.
This may greatly benefit apiculture, as beekeepers can freeze the sperm of bee lineages that are locally adapted, or even resistant to disease.
The new protocol also means that live bees do not need to be shipped around to inseminate other colonies, reducing the spread of pathogens. It also allows breeding programs to access greater genetic diversity.
But the field is not yet at the stage of developing bee ‘sperm banks’, which have become common for breeding other animals such as horses and cattle.
“Freezing drone semen is still very much limited to research,” Deleuze says. “A cryobank has recently been started in Germany, but to the best of our knowledge this one mostly aims at banking specific lineages of bees, which for example are tolerant to some diseases.”
The study is published in the journal Insects.