While we continued our activities during the Corona years, all beekeepers, volunteers and Arista employees enjoyed the regained freedom to – literally – work closely together on the diverse sets of tasks in our breeding program. The good selection results are “contagious”, which is reflected in the number of beekeepers that would like to join the program. For this reason, we need to expand our fundraising efforts as those beekeepers are most effective and efficient when they can be supported by a project leader from Arista.
In this update, you will find an overview of the 2022 results. 2022 was the 9th year of Arista, 2023 will already be our 10th anniversary! A good moment not only to reflect on what has been achieved but also to start planning what our direction could be for the next decade.
Progress in the breeding program
In 2022 we welcomed Marian Meyer in Beers (the Netherlands) as a new Project Leader. He received his Bachelor of Horticultural Sciences in Berlin (Germany) where he already worked on VSH behavior. Marian has been a passionate beekeeper for 6 years and went through a steep learning curve last year to acquire the necessary skills in the VSH breeding program. He is now supporting the Dutch beekeepers in the NL-EU SIB funded VSH breeding program. He also monitors and supports the breeding done with our Arista (donor/sponsor) hives.
The good weather early in the season, all over Europe, allowed a quick start in breeding and production of vital queens and drones for the many insemination sessions. Throughout Europe, Arista supported about a dozen groups with the insemination of around 1000 queens. A thousand other queens were inseminated by beekeepers from the participating groups. Three years ago, Arista started with inseminations courses for the groups to ensure more and more queens can be inseminated by the beekeepers themselves.
Before the season started, we also prepared and shared a high-quality semen diluent, essential for instrumental insemination. A total of 6 liters was produced at the facilities of InHolland University of Applied Sciences in Amsterdam, providing us with a sterile environment to do so. The recipe has been used successfully by Dr. Guillaume Misslin for more than 20 years and allows the conservation of semen, under the right conditions, for at least 6 weeks.
Single Drone Insemination results
This year 130 breeders participated in the Single Drone part of the breeding program, divided into 31 groups in 7 countries. We assessed a total of 1065 test colonies. This was the first time we passed the bar of 1000 colonies, a great milestone for all participating beekeepers.
Again, the results were very good: 34% of these colonies (367 in total) are considered high VSH, meaning such colonies can fight the mite on their own and do not need any treatment against Varroa. Out of these 367 high VSH colonies, 43% (149 in total) are fully resistant which implies that they remove all reproducing Varroa mites. So, even after adding 100-150 mites to these colonies, not a single reproducing Varroa mite is found in the brood (after opening 300-600 cells…). These colonies are classified as 100% VSH.
Multi Drone Insemination and Land Mating Stations results
Every year selected high VSH SDI test colonies are sent to Italy. The local beekeepers produce daughters and let those mate in a dedicated mating station populated with high VSH drone colonies. The first year the colonies grow and are assessed on mite infestation. The colonies with less than 3% mite infestation are wintered without treatment and assessed the second year for all important traits like swarm tendency, gentleness, honey production and, of course, Varroa infestation.
The best performing colonies, usually one or two per original SDI colony, are kept to produce drones the third year. So, in May 2022, 3 project leaders and 5 volunteers again went to Northern Italy to collect semen from the best performing Varroa resistant colonies.
This extremely valuable semen was then distributed back to the participating groups that originally supplied the SDI colonies. The collected semen allowed the insemination of 682 queens. The colonies developed from those queens will once again be assessed by the beekeepers for all the important traits, including Varroa infestation. This Italian selection step ensures that we not only select for VSH (in relatively small Single Drone colonies), but also select for all the important traits in large production colonies under standard field conditions. Of the queens which were inseminated in Belgium, and sampled later on during the season, 85% had very low mite levels and did not need any treatment.
The transnational Belgian-Luxembourgish, Varroa resistant (land) mating station of Sélange hosted a total of 412 virgin queens during 5 periods of two-weeks each. The results of the matings varied from one period to another, e.g. the first period turned out to be least favorable for mating with better results in the consecutive periods. Overall mating success was a little below 60%.
All those queens will be assessed in production in 2023, like the Italy project. Those colonies with the best overall bee-qualities (honey-harvest, low swarming tendency, gentle) and very low mite counts will be used in 2024 as drone colonies. The first results of the queens which were mated in 2021, look very good in terms of production quality and Varroa resistance. More data have to be collected for proper quantification.
Year after year, more and more naturally mated queens are evaluated in Sélange. The queens tested are a mix of older (longer selection, higher VSH) and younger lines (with new bloodlines, for more biodiversity, but generally lower VSH). Of 251 colonies sampled in 2022, 62% were Varroa resistant (<3% infestation). As with the other land-mating stations, there is still a “dilution” of our high VSH drones with non-VSH drones from non-VSH colonies not controlled by Arista beekeepers.
This is not a problem for the breeding program as we only use the drones from the best colonies in the case of land mating stations; with drone eggs being un-fertilized, they only represent the mother. When more and more VSH ends up in the non-Arista colonies close to the mating station, the higher the resistance level of the queens mated on the land mated station will become. It is of course the end goal of the participating beekeepers to not have to treat any of their colonies against Varroa.
In Luxembourg, located around 3 km as the bee flies from Sélange, is the (land) mating station of Fingig. This mating station is provided with drone colonies from the Luxembourg VSH breeding program. The mating station hosted 409 mating nucs which allowed 347 queens (85%) to be successfully mated. Here, the mite levels of the colonies from the VSH breeding program are also monitored through mite washes. More than half of these colonies show high levels of resistance against Varroa.
As in the previous year, we kept the land mating station at Bronlaak in operation to produce queens for production colonies. Mainly beekeepers from neighboring bee-associations participate, by first grafting larvae from high VSH colonies at Arista in Beers and then bringing the mating nucs to the mating station around 2 weeks later. Several of these beekeepers do queen breeding and preparation of mating nucs for the first time. From the queens mated in 2021 we have a large range in the measured mite infestation levels. Several queens score very nicely with scores below 3% and often even 0% but at least one third of the queens has also much higher mite counts (5-15% in September). For the breeding program the best queens can be used, but for the participating beekeepers it is not nice if a significant part of the queens is still not protected against the Varroa. During the last two years we have unfortunately identified several, previously unknown to us, beekeepers (with non-VSH bees) very close to the station
This made us decide to move the station to Linden, a small town very close to the Arista headquarters and partly surrounded by water. In cooperation with a local beekeeper and the bee-association Bernheze we will manage this mating station. In addition to the better geographical location, we will also increase the number of drone colonies on the station to further improve the VSH mating results.
The Flevoland breeding group and mating station has decided to fully concentrate on the VSH breeding program, supported by Arista. So, 2022 was used to make several combinations (MDI inseminations), to test colonies (for mite infestations) and to produce the VSH drone colonies for their mating station. This implies that the mating station will change its role to become more of a tool in the overall VSH breeding program, from as it was used more for distribution of material in the past.
Education for the breeding programs
Both during the season and winter we educate beekeepers on certain topics for a successful breeding program, using various different channels. In Belgium we performed webinars concerning breeding and selection for different strains of bees (Black bee, Carnica and Buckfast) and on the Arista VSH breeding program.
Furthermore, we provided courses both on location and online on topics such as selection and preparation of drone colonies, selection and management of pedigree data in our in-house developed Queenbase software and insemination as well. The level of support we can give with our full-time project leaders in a country, depends on the amount of financial support (either private or governmental) available in that particular country.
Research & Development
Genetic marker for VSH
This project started in 2018 as a collaboration between Arista, Van Hall Larenstein, Inholland and Bejo Seeds under a “RAAK-pro” grant. Each year, Arista collects samples of bees from dozens of colonies, during the many VSH counting sessions. From all these samples 75 very high VSH and 75 very low VSH colonies were selected for further analysis, so those with the biggest differences in VSH levels. From each of these 150 colonies, the full genome was sequenced and mapped (each sample being 20 workers). The resulting genetic code from each of the colonies in the two pools were assessed, comparing the high VSH colonies with low VSH colonies, to find differences that would hopefully be responsible for the VSH behavior in the high VSH colonies (or the lack of it in the low VSH colonies).
We are very excited to report that we indeed found several significant differences between the genomes of the two groups. This is very good news! However, this is not yet proof that the genes or regions identified are responsible for the VSH behavior itself. They could be, but it could also just be areas on the genome that are more or less prevalent in high VSH or low VSH bees. In case of the latter, they could still be of use as a marker, but great caution should be taken, as the genome of the honeybee has a high rate of recombination. For that reason a certain sequence might only be related to high VSH for a limited amount of time or limited part of the population.
A lot of progress has been made, because we have not only identified these regions, but we also have a large database with the full genome of a large number of colonies. Now, with the identified areas and the full genome in place we can semi-automatically search for small pieces of genetic code that are unique for the identified regions on the genome. These small pieces of genetic code are the basis for creation of probes: markers that can be used in a (quick) laboratory test. Both the high-VSH-linked code and the low-VSH-linked-code can be used to produce small pieces of DNA that “glue” to either the high-VSH or low-VSH regions. By attaching a fluorescent compound to the small pieces of DNA, which has a different color for high and low VSH, a marker test is created.
We are currently developing candidate markers which we will first test on existing samples, with known VSH levels, to determine the predictive value of the markers. We will then select the most promising markers, to create a panel that we can use to test a much larger set of samples, to be collected in the months and years to come.
The genetic analysis and markers are currently performed and created based on mainly one strain of bees (Buckfast). If the markers work, we will have to show that this is the case for other strains of bees as well. Depending on how long ago the genetic trait has been established, more or less adaptations would have to be made to make the tests work on other strains of bees as well.
So, a very large step has been taken (identification of areas on the genome). Now the development of probes/markers has to show, whether this can indeed be translated into a successful test for the selection of resistant honeybees. Such a test would be of extremely high value, as we could both screen far more colonies for our breeding program (having to do only a relative simple laboratory test) and at the same time collect very detailed information of the level of resistance per queen including possible missing genes in case of partial resistance. In these cases, “matching” partners could be identified with the same test to increase the level of resistance in the next generation.
Arista, Inholland and Bejo Seeds will work together on these genetic markers, supported by the approved NL-EU-SIB-government grant. Together, we will also look for further funding and partners to develop this very promising tool on the widest possible basis (different strains of bees and geographically).
In summary, we have not find the holy grail yet but we did find pieces of the map leading to its discovery. We have become even more motivated to keep looking for it. We trust on Aristaeus to help us on this exciting journey, but can use any additional support in this treasure hunt…
VSH rapid test
We have started to work on a rapid test that could be used by a beekeeper to determine the level of resistance/VSH of a colony without doing the elaborate brood count or having to wait for a long time to see if the mite levels remains low. Basically, such a test kit could be another holy grail, if it would reliably predict the VSH level.
One of the approaches for such a test is the use of odors, compounds that could be linked to the VSH behavior. Application of such compounds to the brood would ideally trigger the brood removal behavior – specifically in VSH colonies and not in non-VSH colonies.
The UBO test is one of the first developed potential tests, developed by Kaira Wagener. UBO stands for Unhealthy Brood Odor. In 2022 Arista had its own project in Netherlands, but also supported a project in Belgium with Buckfast Flanders and University of Leuven as main contractors. A large number of colonies (>60) with different levels of VSH (as measured in our brood assay) were studied for their generic hygienic behavior using the two standard tests; the Freeze-killed brood test and the Pin-killed brood test, in addition to that the UBO test (a spray with selected compounds) and the standard VSH brood count were performed (infestation of brood with Varroa and count of remaining reproducing and non-reproducing mites). In line with a lot of research done in the last few decades, the Freeze-killed and Pin-killed brood tests showed measurable, but low correlations to both the VSH brood test as well as the UBO test. The UBO test itself showed only very low or no correlation to the VSH brood test.
For the hygienic tests it is known that the environmental circumstances (honey flow etc.) can impact the results of the tests. So, the UBO test could possibly give higher scores in spring than in summer (when we did the tests, as summer is also the period when we do our VSH brood tests). However, Arista is not interested in just hygienic behavior, which has shown to be only weakly correlated to Varroa resistance. We also do not believe we should rely on mechanisms that would only be prevalent in short periods of the year or under very specific circumstances. That is why Arista has decided to not continue the work with the UBO compounds.
Whilst we were working on these UBO compounds, a paper by Fanny Mondet on a different set of compounds was published: the “VPS” compounds (Varroa Parasitization Specific compounds). Fanny Mondet has been able to show these compounds are only present in the cell when young mites are present. This is very interesting as this is also the base of the VSH behavior: the removal of brood that has young mites present…. Supported by the Goeie Grutten grant and advice from Fanny Mondet we have now started to work on this approach. We hope we can do the first experiments this year to see whether these compounds could indeed be used for the development of a VSH rapid test.
The 22 Arista hives in the Netherlands are playing an increasingly important role in our activities. While the financial contribution is already 1/3 of our budget, the colonies are also a bridge to new beekeepers and local associations throughout the country. As these colonies are often taken care of by local beekeepers, they often become ambassadors of our program in their region.
In addition to the Arista hives in the Netherlands, we have been able to get the support for our first Arista hive in Belgium! The Arista hives also are a good source for our breeding program. This spring we brought several queens back to the Arista headquarters in Beers as they headed the best performing colonies. In addition to the very low mite levels, they performed well on honey harvest, low swarm tendency and gentleness.
Now back at Arista these queens are part of the “top-league” and used by the participating beekeepers in the breeding program. Thank you, Arista hive donors and sponsors!
Overall the Arista hives are doing very well on the mite levels. Combining the data of the last 2 years, on average the mite levels stay around or below 1 % from the start until the end of the season (obviously without any treatment). These very low mite levels also are reflected in how the colonies get through the winter. This spring we wintered out 41 high VSH inseminated queens in total; the Arista hives plus reserve queens on Mini-plus hives. Only one colony did not make it to spring! This is a very good result compared to the overall, national percentages of winter losses of colonies in the Netherlands and Belgium, which are in the range of 30 to 40% (with most beekeepers doing several treatments per year). And the differences are not just in the number of colonies surviving the winter but also in the strength of those colonies . We also see very low winter losses counted as frames of bees in most of the high VSH colonies. Basically winter-in = winter-out. For non-VSH colonies most beekeepers consider it “normal” if a colony reduces in strength by half between before and after the winter. We now experience the overwintering of colonies as it was before the Varroa started its destructive work in the 1980’s…
The next decade
At the end of this year, Arista will be active for 10 years. What did we learn and what are the plans?
There are several lessons we learned during this journey.
The first one is that “it works”. Running a breeding program, based on the inventions of the USDA combined with modern bee breeding selecting methods, results in fully Varroa resistant honeybees with good beekeeping traits. It is a lot of work, but the results are very good and make all the investments in blood, sweat and tears well worth it! Now we need to set ourselves up for doing this for a broad selection of the honeybees (strains, bloodlines) and broad geographical representation.
The second lesson is the need for cooperation. Those beekeepers that have organized themselves in breeding groups were most successful. Being in a breeding group ensures that important pieces of information are available and shared. Lessons are learned collectively, successes are celebrated together. Whereas beekeepers are sometimes known for working alone, basically the common enemy has brought us together. The model where a group of passionate beekeepers is supported by Arista project leaders for support and coordination, is working very well.
The third lesson is that funding those Arista project leaders works best if national bee-associations become actively engaged in supporting the acquisition of national, EU and private subsidies. The region of Wallonia has been the most successful so far. But bee-associations in the Netherlands have also actively supported the first NL-EU subsidy for Arista.
More progress could and should be made in other countries, like the large bee-country Germany. This spring in Germany a very encouraging initiative was started during a large conference (https://varroaresistenzprojekt.eu), organized by all the major bee-associations.
Representatives of neighboring countries were also invited, as well as the bee-institutes. The conference is currently being followed up by regular workshop sessions to prepare written proposals that can be submitted to the German authorities and to form a plan to breed a Varroa resistant honeybee in a large cooperative setting. During the conference the ambitious target was set to convert the German honeybees towards Varroa resistance by 2033! Arista takes part in the working groups and is very motivated to support this ambition.
Looking more to the west we have to conclude that, despite the fact that the VSH was discovered in the USA (USDA, Baton Rouge), only a few initiatives have been started with limited implementation so far. Given the experiences of the last 10 years in Europe, as well as the progress we are making towards a genetic marker, Arista has reached out to the USDA to re-start a technical transfer of the breeding techniques to the US bee-industry. It is our objective to start a combined set of initiatives, using the expertise of both the USDA and Arista. This program will address the development of tools as well as the creation of a pilot project producing fully Varroa resistant production queens.
And then looking more eastwards we have recently been approached by parties in New Zealand and Australia to see what can be done to start a Varroa resistance breeding program. It would be very rewarding if the knowledge built up so far could also be applied on this side of the planet!
So, the next decade basically can be described as “geo-expansion”. While developing new tools like the genetic marker, we are now ready to involve more bee-associations, beekeepers and bee-dependent companies in a more global setting.
The big steps forward in the VSH breeding have only been possible by the hundreds of volunteers who, are active in the program. A very large, diverse set of tasks is being executed by a large number of volunteers.
Arista teamwork: volunteers work on filling and labeling honey jars and painting of hives
Firstly, the beekeepers that participate and produce quality queens and drones for the SDI and MDI sessions, spending their precious time throughout the year on this demanding task alongside their other beekeeping, professional and private activities. One test colony requires the equivalent of more than a full working day. Weeks of free-time are invested each year, to produce a new generation of potential breeder queens. Secondly, the selection would not have been possible without the support of the numerous volunteers, beekeepers and non-beekeepers that asses the colonies by carefully examining the infested frames, but also support the program in 1000 other ways like preparing the counting room, catering for the counting and insemination days, the overall logistics, preparing frames and hives, extracting honey, doing mite-washes, processing data etc. …
A big thank you also goes to the various governments that support us, and to our sponsors, private donors and funders that enable us to support the beekeepers with the professional support of our project leaders. In addition, these funds make it possible to do the research to develop new tools for the selection program.
Thank you all for your contributions that make it possible to keep us passionately pushing for healthy bees in more and more regions and countries.
The Arista team