1. Which strain?

We are often asked why breeding instructors in French-speaking Switzerland work with only one strain (Carnica): why not another, and why not several? In this article, we will briefly answer these questions and thereby clarify an area of uncertainty for many beekeepers.

Before the advent of modern beekeeping (movable-frame hives, around the 1800s), bees lived quietly within their region of origin and had been adapting to their local environment for millions of years. This natural selection led to specific ecotypes depending on how a species adapted to its environment. These subspecies are what we refer to in beekeeping as bee strains.

The best known in our area are:

• Apis mellifera mellifera (the black bee, also called Nigra) ;
• Apis mellifera linguistica (Ligustica, also called the Italian bee) ;
• Apis mellifera carnica (Carnica, also called the grey bee).

Each of these strains has developed particular characteristics. For example, Ligustica stores little for winter and overwinters with a large population. The black bee and Carnica, by contrast, overwinter with substantial stores and a small population, which facilitates overwintering in cold regions.

Modern beekeeping developed breeding methods and enabled humans to proceed as in livestock breeding or agriculture: crossing subspecies to create new lines (e.g. Africanised bee). However, these crosses have only rarely brought positive outcomes because, by diluting the genetically anchored traits of the different strains, numerous defects appeared: aggressiveness, low resistance to disease, poor yields. Moreover, due to the bees’ reproductive system (the queen is mated by roughly fifteen drones), these undesirable traits spread rapidly in nature. To such an extent that, in the 1950s, the Société romande d’apiculture (SAR) approached the Federal Council to establish a working group whose objective was to combat this hybridisation problem. This working group highlighted that the only solution was to return to a pure strain. The difficulty was that the local bee had already been hybridised almost entirely; they therefore decided not to continue with that strain (see genomic past of the Swiss bee). After testing five subspecies in different apiaries, the working group decided to use Carnica.

For more than seventy years now, the SAR breeding commission, in partnership with the groups of breeding instructors in the French-speaking cantons, has worked tirelessly to maintain the purity of this strain, improve it (in particular with regard to disease resistance), and make it available to every beekeeper. We will discuss in more detail the work carried out by breeding instructors in upcoming articles.

In conclusion, the choice to maintain a pure strain and to work with a single strain is therefore not arbitrary. It followed a drift in beekeeping to which we had to respond, and we have been working ever since to preserve the quality of these bees. As for the question of why not use several strains of bees in one region, it is now clear that this would create a serious hybridisation problem and would return us to the same catastrophic situation as at the beginning of the twentieth century.

2. Bee genetics

This month, we will focus on bee genetics. Without going into too much detail, we will attempt to summarise its basic principles and their implications for selection. Interested readers will find more information online.

As you know, a bee colony consists of a queen, workers, and a few males, called drones. Their respective developmental pathways are very different.

The queen can choose whether or not to fertilise the egg she lays. A fertilised egg produces a female; an unfertilised egg produces a male. After three days, the egg hatches. It is then nutrition that determines whether a fertilised egg becomes a worker or a queen.

A larva destined to become a queen is fed exclusively with royal jelly until the cell is sealed (day 9 after laying). On day 16, the young queen emerges. After a few days, she will leave the hive to be mated by 10 to 25 drones from other colonies, thereby ensuring the genetic diversity of her offspring.

By contrast, a worker larva is fed for three days with brood food, then with honey and pollen. During the three days when it is fed brood food, the female larva can still become a queen if the bees decide to feed it royal jelly (for example, if they have lost their queen in the meantime and therefore need to raise a new one). This is the phenomenon used in breeding by giving very young larvae to queenless bees.

Drone larvae are fed in the same way as worker larvae. However, as indicated earlier, the male develops from an unfertilised egg, which is a specificity in bees. Queens and workers have 32 chromosomes, 16 from the mother and 16 from the father. They are diploid like most animals and plants. Drones, on the other hand, have only 16 chromosomes, coming solely from the mother. They are haploid: each chromosome is present in a single copy inherited only from the mother. Consequently, the spermatozoa of a given male are all strictly identical; they are clones—unlike the queen’s eggs, which are all different, each egg being a unique combination of the various genes of the queen and the father.

 

In a hive, we therefore have a queen that has been mated by several males (blue, yellow, red and green in the illustration below). Sons inherit only the genetic material of their mother. Daughters are half-sisters or super-sisters. Half-sisters do not have the same father. Super-sisters have the same father and, since the male is haploid, they have received from him exactly the same genetic material.

 

Thus, only the mother’s genetics are transmitted to males, and males retransmit them in full when they mate with a young queen. Our selection programme is based on this specificity to disseminate the best possible genetics in the mating stations. In each A mating station, the males are all grandsons of a queen awarded within the selection programme (testing apiaries). This makes it possible to have a single genetic pool within a given mating station, to control inbreeding, and to manage crosses between lines while respecting genetic diversity. We will return to these points in more detail in upcoming editions.

3. Purity of the Carnica strain

As already explained in the previous chapters, the primary objective of SAR breeding is to conserve the purity of the Carnica strain selected by Liebefeld. How do breeding instructors measure this purity (or rather the absence of hybridisation)? Is this work carried out over decades effective? What are the prospects for simplifying the work of breeding instructors? This is the agenda for today’s article.

From the start of SAR breeding, the only method used by scientists to determine the strain (subspecies) of a bee relied on morphological measurements. The literature mentions several criteria: cubital index, discoidal shift, width of the tomentum, tongue length, abdomen colouration, etc.

 

In Europe, the most widely used criterion is the cubital index. It is easy to measure and makes it possible to discriminate hybridisation with Ligustica (Italian bee) and Mellifera (black bee). The cubital index is the ratio between the lengths of segments “a” and “b” of the forewing:

 

Average cubital index values in Switzerland:

• Carnica: 2.3 – 3.2
• Ligustica: 2.0 – 2.7
• Mellifera: 1.4 – 2.1

To be sure there is no crossing with the other two strains, breeding instructors therefore set a minimum cubital index value of 2.8 for our Carnica selection. We quickly introduced an additional criterion because almost all our queens met these criteria and we wanted to increase the precision of our selection. This is how the tongue-length criterion was introduced.

Every queen that a breeding instructor wishes to use for breeding must reach this value. Thanks to this strict threshold, we have been able to maintain the purity of our bees for so many years. Each year, the breeding instructor must take a series of bees from the colony they want analysed and send this sample to the laboratory. The laboratory measures the wing and the tongue of about fifteen bees and provides an average value for the cubital index and tongue length. If all these criteria reach the extremely high level required, the breeding instructor can breed from this queen and continue the selection process. This high selection level has made it possible to exclude any hybridisation for more than seventy years.

For about ten years now, we have also used genetic analyses to verify whether our approach is correct and whether there is indeed no hybridisation within our lines, and also to check the security of the mating stations. This tool, although more complex, will probably allow us in the near future to reduce the number of morphological measurements and thus simplify the work of breeding instructors.

As you can see, ensuring the purity of our stock is a long-term task that must be repeated year after year; otherwise, the traits naturally present in our Carnica bees would be diluted and lost in hybrid bees.

Back up :

To calculate the cubital index, measurements are taken on 100 different right forewings of workers, of the two vein portions forming an obtuse angle at the posterior base of the third cubital cell, and the ratio segment a/segment b is calculated.

The discoidal shift index is measured on the veins of a worker’s forewing by drawing a straight line connecting the two ends of the radial cell. Then a perpendicular line is drawn through the intersection of the veins of the radial cell and the 3rd cubital cell.

 

If this straight line passes through point A or to the left (proximally) of it, this is referred to as a negative discoidal shift and therefore A.m. mellifera. Conversely, if this straight line passes to the right (distally) of point A, this is referred to as a positive discoidal shift and therefore A.m. carnica.

 

Measurement of tomentum width (hairy area) is taken on the 4th tergite (dorsal segment of the abdomen) and corresponds to the width of the hair on this segment.

4. Selection criteria

How enjoyable it is to work with gentle and productive bees. Isn’t this the goal of every beekeeper? However, obtaining and, above all, maintaining these characteristics in a bee population does not happen automatically. This is why any queen-breeding programme must be based on robust assessments. In this February section, we will present the selection criteria retained by the SAR breeding commission (CE-SAR) for the Carnica strain.

The CE-SAR selection criteria initially numbered five: gentleness, frame holding, swarming resistance, disease resistance, and yield. They were recently complemented by three new criteria: colony strength at the start of overwintering, overwintering, and spring development.

Each of these criteria is assessed using a measurement scale common to all breeders.

• For gentleness, there must be no flying up, no attacking, and no stinging to obtain the maximum score. This assessment is complemented by frame holding, i.e. the absence of clustering and movement on the frame. The evaluation of these two criteria must be carried out over at least four visits, then averaged.
• Swarming resistance is measured every 5 to 9 days during the swarming period. The maximum score can be given if there are no signs of swarming: no empty or laid queen cells.
• To assess disease resistance, hygienic behaviour is evaluated using the pin test: 50 sealed brood cells (pupal stage with pink to light brown eyes) are pierced with a needle; after a delay of 8 to 12 hours, the percentage of cleaned cells is counted.
• Yield is the easiest criterion to measure, by the difference in weight between a full and an empty honey super.

Three new criteria were introduced with the aim of selecting colonies that tend to become rapidly productive in spring. To this end, the colony must be strong at the start of overwintering, with, in September, good brood and many emerging bees. After overwintering, at cherry blossom, the colony must occupy at least five seams of bees and be clean (no dysentery, bottom well cleaned by the bees). Spring development is then assessed during the following three visits. It is optimal if brood is distributed regularly by age and if the colony quickly draws foundation, increases its population exponentially, and is ready when honey supers are added.

And what about Varroa resistance? Can it be used as a selection criterion? How? To date, unfortunately, there is still no clear answer to this question. Within the SAR selection framework, we strive to identify colonies with the lowest Varroa loads and to take this into account. However, there is no scientific proof regarding the heritability of such resistance or of related behaviours. Moreover, a low Varroa load is not necessarily due to colony behaviour: it may also be the result of environmental conditions, notably the absence of heavily infested hives nearby.

Varroa resistance is therefore the most complex criterion to develop. Let us hope that heritable resistance behaviours can be identified soon. In any case, selection must continue according to the various other criteria presented above so that we can continue to benefit from gentle and productive bees.

5. How to control the origin of male genes

In any breeding programme, in order to cross and transmit very specific traits, it is necessary to control where the carriers of male and female genes come from. In most livestock species, this is fairly simple because one male mates with one female. Crosses can be controlled easily. In bees, however, the situation is completely different because of polyandry. The female is mated by a multitude of males (around fifteen). How can the origin of male genes in bees be controlled in order to determine crosses precisely?

There are two methods to address this difficulty:

1. Artificial insemination

This procedure makes it possible, using a microscope and drone semen, to inseminate the queen. This technique provides 100% certainty regarding mating, but it is difficult, remains limited to a few people, and yields a success rate of around 80% for experts. Only a small number of queens can be inseminated in this way, which does not allow large-scale dissemination of the genetics selected by breeders.

2. Mating stations

Another solution is to isolate, in a remote location, the queens to be mated and a series of drone lines containing only brothers from the same family. The specific topography of the Alps allows such groups of bees to be isolated in remote valleys and thus protected from the arrival of undesired drones. These stations are a wonderful working tool for breeders because they allow a large number of queens to be naturally mated (propagation of genetics) with a technical level that is easily accessible to any beekeeper. These stations are protected by cantonal agricultural law; it is therefore prohibited to place hives within a perimeter of around 6 km around the station in order to avoid genetic pollution.

 

Valais has three stations managed by SAR breeding instructors: Bonatchiesse (reserved for breeding instructors), located at the foot of the Mauvoisin dam Les Toules, located at the foot of the Les Toules dam Moiry, located at the foot of the Moiry dam (see photo)

 

Breeding instructors in Valais work tirelessly to ensure that all beekeepers can benefit each year from the genetics of the lines of the SAR breeding commission (CE-SAR). A line is present for two consecutive years in the same station. (Detailed information at https://favr.ch/elevage/stations-de-fecondation/)

If you too would like to benefit from these stations, we invite you to contact the breeding instructor of your section or to participate in the breeding course in your region. This will enable you to breed from selected brood and to obtain all the information needed to set up your mating nucs in the station according to good practice.

 

6. Testing apiaries

In the previous chapter, we presented the SAR mating stations and their role in selection. Thanks to the six A stations of the SAR, the purity of the Carnica strain has been maintained for more than 70 years. We will now explain how the best genetics from our selection are made available within the stations.

It is through the testing apiaries that the best-performing queens are identified. Each year, between 10 and 15 tester beekeepers, spread across French-speaking Switzerland, receive in July 12 queens to test. They do not know which lines these queens come from. Testing is therefore carried out as impartially as possible.

The testers will introduce these queens into standardised packages of bees or standardised nucleus colonies. These colonies are placed in the same apiary and managed identically in order to avoid bias. The testers will evaluate them over the following 12 months, based on the selection criteria presented in our February article: gentleness, frame holding, swarming resistance, disease resistance, yield, etc.

All assessments are recorded in the testing logbook and then transmitted to the SAR breeding commission. Based on this, a ranking of the tested lines is established. Three to four lines are awarded each year. The best queen of each awarded line is given to the managers of the drone colonies of the mating stations that need to renew their line. The same line remains in station for a maximum of two years. 

The managers of the station drone colonies will then rear a large number of daughter queens from the awarded queen received. The objective is to have at least 40 colonies originating from the awarded queen in order to be able, over the next two years, to place the 20 best colonies in the station. In spring, a drone frame is introduced so that there will be as many mature drones as possible in these colonies when the station opens.

 

Illustrative table: example from the 2022 testing apiary

 

There is thus in each mating station a specific genetic pool derived from an awarded queen from the testing apiaries (see also our December article on bee genetics). The Beebreed database then makes it possible to calculate the inbreeding rate of this genetic pool with that of the brood made available by breeding instructors for your queen rearing. Beebreed also allows estimation of the chances of improving a given trait (gentleness, production, etc.) depending on the station chosen.

This year, the new lines made available in the Valais stations are SLO97 at Bonatchiesse and CC95 at Les Toules. At Moiry, we will find SM89 for the second year. Do not hesitate to drive a few extra kilometres to reach another SAR A station: Hongrin (above Aigle VD), Petit-Mont (above Charmay FR), or Vermeilley (above Nyon VD). Breeding instructors are available to advise you on your choice.

 

7. Breeding for one’s own use

May has arrived, and with it the swarming season is in full swing! Bees naturally raise new queens to split the colony into two, or even several swarms, thus ensuring the survival of the population. Swarming is one of the three situations in which a hive will raise queens. In the case of supersedure or the sudden death of a queen, the colony will also raise queens.

The breeder will use these natural predispositions and add a few techniques to simplify and encourage breeding for personal use. Humans become the trigger for the bees’ breeding work and will steer it in order to be able to obtain, in the end, several queens that can be used to replace ageing queens, form nucleus colonies, or replace an underperforming queen.

In this article, we will explain a simple method for obtaining several queens with minimal work and with a basic technical level. Next month, we will provide more complete information on a more advanced breeding method that makes it possible to obtain more queens and to take them to a mating station.        

First, since the objective is to obtain F1 (queens descending from a pure F0 queen and mated at the apiary—see article from November 2023), we need a good colony that is strong, healthy, and headed by an F0 queen. We must also prepare an empty 6-frame nucleus box.

We will make the mother hive queenless by taking 2 frames of stores, 2 frames of sealed brood, and 1 frame of pollen, while making sure to take the F0 queen. These frames are placed in the empty nucleus box, which is moved further away within the apiary. Depending on weather and nectar flow, a light stimulating syrup can be given to the mother colony. It is important to check that there is fresh brood (eggs or very young larvae) in the mother colony so that it can properly raise new queens.

2–3 hours later, this colony will naturally begin raising numerous queens. The next day, one can observe a series of initiated queen-cell cups filled with royal jelly.

Six days later, the cells will be sealed and can be transferred into nucleus colonies or into mating nucs that have been populated in advance. To cut out the queen cells so that they can be transferred, you can use a small knife whose blade you will regularly heat using a blowtorch. You can leave a generous margin of wax around the cell. A toothpick can be used to fix the cell on one of the frames of the nucleus. A small feed of syrup for the colonies that will continue to tend the cells for another six days is recommended. Queens should emerge about twelve days after the start of the operation. On day 13 or 14, you can check that the queen cells have been opened. In case of a problem (for example, if a queen did not emerge), either you have another queen cell or queen available and can introduce it, or you can unite this nucleus with another. If they have emerged, all that remains is to wait patiently for about two weeks for the queen to mate and begin laying.

There are many variations of this method on the internet and in the literature: cutting the brood frame so that it is easier to cut out cells, splitting the colony to make the method even easier but then you cannot make more than 1–2 nucleus colonies… It is up to you to try and find the method that suits you.

 

8. Breeding with a “starter”

Like May, June is favourable for queen rearing. It is also the month when mating stations open. In our May article, we presented a simple breeding method consisting of dividing a colony and then cutting out queen cells to transfer them into nucleus colonies or mating nucs. Today we will present another method that allows more queens to be reared. We will also remind you of the rules that must be followed when taking nucs to a mating station and the rationale for them.

Breeding begins by making part of the hive queenless: a “starter” is created. This starter must contain sufficient nurse bees, honey, pollen, water, and above all young larvae from a good F0 queen. In principle, the starter is created on a Friday afternoon or at the latest on a Saturday morning, for example Saturday morning 1 June 2024, the date of our breeding course. This makes it possible to take nucs to the mating station two weeks later (stations being open on Saturdays).

A simple way to create a starter is to remove from a very strong hive the queen together with 3–4 frames of young brood (eggs and small larvae). The queen and the frames of young brood, with the bees, are placed in a 6-frame nucleus box that can be left in the apiary or taken to another apiary. The rest of the hive constitutes the starter. Another way to create a starter is to insert an airtight divider into the original hive or to use a double hive: on one side, the queen with the young brood, and on the other side, the rest of the colony (starter).

In both cases, it is necessary to ensure that there is as little young brood as possible in the starter. To do this, a vertical queen excluder can be introduced one week in advance to prevent young brood from being present in the part that will become the future starter. There are many other methods of creating a starter that you can find online or in volume 3 of “Apiculture une fascination”.

The more nurse bees there are in the starter, the more queen cells there will be. Nurse bees are generally found on frames of young brood. You can reinforce your starter by taking a frame of young brood from another hive; make sure the queen is not on this frame; brush the bees into the starter; then return the brood frame to its original hive.

One to two hours later, young larvae must be introduced into the starter. To do this, you will visit a breeding instructor with a grafting frame and new plastic queen cups. He can graft young larvae from his best F0 queen. You can also do this yourself if you have trained beforehand in grafting (picking). The breeding instructor will also provide you with the signed declaration form for taking nucs to the station, certifying the purity of the brood made available and thereby allowing you access to an A Carnica SAR mating station.

 

The grafting frame must then be wrapped in a damp cloth to prevent the larvae from drying out during transport. The grafting frame is then introduced into your starter. If you took your starter with you (or part of it in a swarm box, for example), you can introduce the grafting frame directly and bring everything back to your apiary. Feed your starter well if natural intake is low, especially at the beginning of the rearing phase, and even beforehand.

24 hours later (Saturday afternoon or Sunday morning 2 June 2024 in the example of our breeding course), you can open your starter to check the number of initiated queen cells. If necessary, you can return to the breeding instructor for another grafting and introduce additional cups into your starter.

5–6 days later (Wednesday 5 or Thursday 6 June), the queen cells will be sealed. You will then be able to place protective rollers around the queen cells to prevent the first queen to emerge from destroying the others. Take the opportunity to destroy any “wild” queen cells drawn on other frames. It is important to place the protective rollers at this time and not later, because the pupae will then enter a metamorphosis phase during which they become very sensitive to shocks and cold. The starter must therefore not be opened during the following days (days 7 to 9, i.e. from 7 to 9 June).

Meanwhile, or even before if you are well prepared, you must prepare the mating nucs. The mini-frames must be fitted with a strip of foundation of about one third of the frame height. It is fixed by melting a little wax. Each mating nuc must contain three mini-frames. The feeder is then filled with candy. This candy must not contain honey to avoid disease transmission in the mating station (honey may contain foulbrood spores). Candy without honey can be purchased commercially or made at home (10 kg icing sugar, 3.8 kg invert syrup).

 

10 days after creating the starter (Monday 10 June), it is time to populate the mating nucs. Bees are taken from various hives and/or from the breeder colony. They are sprayed with a little oxalic acid to avoid bringing Varroa into the mating stations. They are then gently brushed into a large funnel placed above a 6-frame nucleus box topped by a super, with a queen excluder between the two. The number of frames to brush depends on the number of mating nucs to populate.

One can count about one side of a frame per mating nuc. The bees are then gently pushed with a board and a little smoke towards the bottom of the super; they pass through the queen excluder and settle in the 6-frame nucleus box. This filters out drones: they cannot pass through the queen excluder. They are released only once the mating nucs have been populated. This avoids having drones end up in the mating nucs, which must be strictly free of drones to avoid genetic pollution at the mating station.

 

To populate the mating nucs, the equivalent of one yoghurt cup of bees per nuc is collected. The mating nuc is turned upside down, the bottom is opened, the bees are poured in, and everything is quickly closed without crushing bees. The mating nuc is then turned back upright. These operations are easier to carry out with two people. The mating nucs are then placed in a cool location (cellar). During the night, the bees will feel queenless. The next day, they will be ready to accept a new queen.

On day 11 (Tuesday 11 June), queen cells are taken from the starter and introduced into the mating nucs. Queens will emerge on day 12 or 13. It is then important to keep the nucs closed so that colony cohesion can form around the new queen during days 13 and 14. On day 15 (Saturday 15 June), the nucs can be taken to a mating station and nature is allowed to do its work. The nucs are collected two weeks later. As a reminder, the Les Toules station opens on 8 June and the Moiry station on 15 June. Do not forget to contact the station manager before going, and bring the “Déclaration de montée en station 2024” form.

 

The breeding calendar is an essential tool to plan rearing properly and intervene at the right time; otherwise, the rearing will fail. It is available in electronic form (xlsx). You can enter the desired date for taking nucs to the station in cell D1. Here is the calendar for our breeding course on 1 June.

 

You will find more details and information on this rearing method in the document “ Becoming a breeder beekeeper ” by Julien Balet, which you can download free of charge.

We wish you much pleasure and every success with your queen rearing.

 

QR code breeding calendar

 

 

QR code Becoming a breeder beekeeper

 

9. Managing mating nucs after returning from the station and introducing queens

At the station, by looking through the plexiglass of the nuc, one can already see whether the combs are well built, sometimes even with construction in the feeder. The queen is probably present and laying. However, if the mini-frames are poorly built and there are few bees, there is probably no queen. She will have been lost during the mating flight or eaten by a bird.

After returning from the station, the nucs are placed at home or in the apiary, if possible not too exposed to the sun. The least populated nucs should be inspected first. If there is no laying, the mini-frames are poorly built, and there are drone cells, there is no queen. The nuc can then be emptied by brushing the bees outside. They will seek refuge in other hives or nucs.

The other nucs can then be inspected carefully. Take care not to frighten young queens: they may fly off and be lost. One must therefore proceed gently, lifting only the first mini-frame. If there is laying, all is well. Do not inspect further; close the nuc.

Subsequently, remember to check that the nucs have enough food. From mid-July, external intake decreases and the nucs need to be refed with candy regularly: check at least every two weeks.

Before taking a queen from a nuc to introduce her into a hive, it is preferable to wait until the first daughters of that queen have emerged, or at least until there is sealed brood in the nuc. The queen will be more readily accepted than if she has only just started laying.

Be cautious when collecting the queen from the nuc. Avoid frightening her, otherwise she may fly off and be lost. Queen-collection tubes using suction are very practical, especially when the queen is not on a mini-frame and is hiding at the bottom of the nuc. Once the queen has been collected, mark her and place her in a cage. If you do not introduce her the same day, add three attendant bees to the cage.

There are several methods for introducing queens. First, you must be sure that the colony is truly queenless. It may contain two queens if supersedure has occurred and the old queen is coexisting with her daughter. In that case, if you remove only the old queen, the daughter remains and the station queen you want to introduce will not be accepted. A good way to avoid such mishaps is to wait 7 to 9 days after removing the old queen before introducing the station queen. You can then check that there are queen cells, which proves the hive is queenless and is expecting a new queen. The station queen can then be introduced in a cage sealed with candy. You may destroy the queen cells, but this is not essential.

1 to 2 days after introducing the cage, you must check that the queen has been able to exit. Sometimes the candy is too dry and the bees do not eat it. In that case, remove most of the candy and replace the cage. Check the next day whether the queen has been able to exit. It is also possible to release the queen gently between two frames.

Then wait at least one week before checking acceptance of the queen. During this control visit, it is also necessary to proceed gently so as not to frighten the queen. Do not use smoke, or only very little. It is not necessary to see the queen. As soon as you observe fresh eggs, all is well. You can calmly close the hive.