Use Of Carbon Dioxide

Carbon dioxide not only serves as an anesthetic, but also stimulates queens to begin laying. Proper treatment with this gas causes virgin queens, as well as instrumentally inseminated queens to begin laying practically as soon as naturally mated queens, whereas without such treatment only about 20 percent start earlier than 30 days after emergence.

At least two anesthetizations are necessary and a third may be given as a margin of safety without apparent harm. They may or may not be accompanied by insemination. An anesthetic treatment without insemination is given by placing the caged queen for 10 minutes in a convenient container through which carbon dioxide is flowing (fig. 5). This gas is heavier than air, and as it fills the container from the bottom, the air is forced out through an escape tube at the top, insuring a high concentration. (Any wide-mouth container with a loosely fitted lid will suffice for carbon dioxide treatments. ) Ovi-position starts 2 to 6 days after the second treatment and a day or two later than would be the case after natural mating. Although treatments given as early as the second or third day are effective, they do not cause laying to start earlier than the normal age. Since exposure during insemination may be too short or the anesthetization incomplete, it is recommended that two treatments be given after a single insemination and one treatment after two inseminations. These exposures to carbon dioxide appear to have no ill effect, but since anesthesia has been shown to change the behavior of worker bees, overexposure should be avoided. The minimum effective dose in time of concentration has not been determined.

Insemination Procedure

The best way to learn the insemination technique is to study under an experienced operator. A microscope arrangement permitting the student and the instructor to observe the same image is ideal.

Since an instructor may not be available, the procedure is here described in sufficient detail for the reader to learn the technique on his own. The description applies specifically to the apparatus that has just been described. The reader can adapt it to any other equipment he might be using.

The student's task will be made easier by the following suggestions:

1. Work with queens reared from several different breeder queens. Daughters of some breeders will be easier to inseminate that those of others. Try queens of different races if available.

2. Use queens reared under the most favorable conditions because large queens are easier to inseminate than small ones.

3. Use virgin queens that have had the free run of the nucleus and that are at least 7 days old. Such queens have less feces in their rectums than queens maintained any other way. A distended rectum pushes the reproductive tract ventrally making it more difficult to hold back the sting and to find the entrance to the median oviduct with the syringe tip.

4. Select a humid day if possible. In a dry atmosphere the exposed tissue of the queen will dry quickly making it more difficult to insert the syringe tip. Also, the semen may dry at the end of the syringe tip clogging the passage before it can be inserted or before semen can be taken from another drone. It may be necessary to moisten these parts occasionally. An experienced operator will work so fast that he will have little difficulty, but a beginner may find these inconveniences very disturbing.

5. Practice the semen collection and injection procedures separately using water or saline solution instead of semen for the injection procedure.

Preparing The Anesthetic

The reduction valve of the carbon dioxide cylinder should be adjusted to a delivery pressure of about 5 pounds per square inch and the flow of gas to the queen holder stopper adjusted to a very small stream. It should be just enough to keep the queen quiet. The flow can best be judged by dipping the stopper in water.

Preparing The Syringe

The syringe is prepared by filling the syringe tip with a physiological salt solution to serve as a liquid plunger. A 0.9 percent sodium chloride solution is satisfactory. An antibiotic may be added to help prevent infection. A plastic squeeze bottle of the type used for dispensing liquids in laboratories is an aid in forcing the liquid into the tip. The socket of the adapter with diaphragm in place is then filled with the saline solution and the tip screwed into it firmly against the diaphragm. Be careful to exclude air because it tends to nullify the purpose of the liquid plunger. The extension is then screwed into the metal syringe body. By advancing the plunger, sufficient liquid is forced out to permit taking in the desired amount of semen when the plunger is retracted.

The syringe is placed in the syringe holder and swung around until the tip is away from the operator so that it will not interfere with the mounting of the queen.