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The Effect of Formic Acid in the Sealed Brood Cells

By Bernd Adelt and Karl Heinz Kimmich

Translated from the Allgemeine Deutsche Imkerzeitung, 1986.

By A.E.McArthur MIL

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During the treatments of honeybee colonies using formic acid, which were carried out in the years 1983 and 1984 at the Institute for Apicultural Research in Stuttgart-Hohenheim, a delayed-action effect was observed again and again. The mite fall occurring in the second week after the insertion of the Illertisser formic acid pad increased noticeably. Since the formic acid evaporates within a 24 hour period, it was thought that the acid vapour killed not only the mites on the adult bees during the treatment, but also reached the mites in the sealed brood cells where it either killed or weakened them. (Liebig et al 1984). The following arguments are submitted in explanation (Liebig, 1985)

Brood Chamber Trials

During the summer of 1985 the actual effect of the Formic Acid was investigated more closely.

Pieces of comb with circa 200 sealed brood cells were placed in a 12x15x22cm glass container.

Formic acid was applied at different strengths (fig.1) by soaking a beer mat which was then placed in the glass container, which was then hermetically sealed. The ambient temperature of the brood box was maintained at 35°C for all tests. After each test was finished the glass container seal was replaced with a bee tight wire mesh and half of the brood comb removed.

*The effect of the formic acid on the mite mortality in the sealed brood relative to the concentration
*The trial was conducted using a brood box with a glass container and lasted 24 hours.*

The death rate of the mature (dark) mites in the cells of the removed comb was established by opening the cells and withdrawing the brood, brood and cells were examined for mites. Mites which failed to respond to a movement stimulus were deemed to be dead. The other half of the comb which was left in the brood box served to demonstrate the emergence rate of the brood. After all the brood had emerged the mite death rate was investigated here as well. Here a note was made of the mites still alive on the emerged adult bee and the mites which dropped when the bee emerged. Only the mature (dark) mites were counted.

The effectiveness of the formic acid in the sealed brood stage was measured over time and concentration in a series of trials. All tests were conducted using a ‘control’. The quantities of formic acid used were calculated in ppm (fig.1)

The series of tests for concentration were conducted over a 24 hour period,  the natural mite fall in the ‘controls’ was found to be between  0 – 10% (fig.1) At an acid concentration of 43 ppm (corresponding to 0.25ml 60% formic acid per glass container) a death rate of 100% in the worker brood was recorded. Even at double this concentration this result could not be achieved in the drone brood. Using 0.5ml, 60% formic acid a mite death rate of only 95% could be achieved. This difference in the mortality rate in drone and worker brood was confirmed statistically using a sample analysis, thus it must be assumed that when treating drone brood against Varroa a higher dosage of formic acid is required.

The time ‘series’ of tests were carried out using 0.25ml and 0.5ml 60% formic acid.  In the ‘controls’ the natural mite fall in the brood under investigation varied between 0 and 6%. It was found that with a dosage residence time of one hour that there was no significant difference in the mite mortality rate, relative to concentration level. A significant effect was noted only after a period of three hours. Opposed to this finding, the mite mortality rate using 0.25ml and 0.5ml formic acid at 60% after a period of 6 hours was 100% in both cases (compare Fig 2).

*The effect of the formic acid on the mite mortality in the sealed brood relative to the treatment period length*
*The trial was conducted using a brood box with a glass container, using 0.25 ml formic acid* (=43 ppm)

The mite mortality rate was also checked after the emergence of the brood.  In this case also differences between the acid concentration tests and the ‘time’ tests were found. Strikingly, after the emergence of the bees an increased mortality rate was noted relative to the tests on the brood samples, which had been previously investigated. We assume that a few mites die at the time of treatment or sampling of the brood and up to the emergence of the bees as a result of the treatment.

 

The Mite Mortality Rate is Higher at the Edges of the Brood Nest.

The rate of emergence of the treated brood as well as the mite mortality rate was checked. An evaluation of the recorded emergence rates was not carried out, since in many cases among the untreated control colonies total collapse occurred, therefore a meaningful statement is not possible. This was made up for in the field trials carried out in 1986. (See figs 2 and 3).

*Mite mortality at the centre of the brood nest after treatment in August using an Illertisser plate ( placed on the frame tops)*

The results achieved from the brood chamber tests were rechecked in 1985, by removing brood from colonies, which had been treated with formic acid. In each colony, 24 hours after treatment, samples were removed from the centre of the brood nest and also from the upper region of the brood nest closest to the inserted formic acid plate and evaluated. In every case the mite mortality indicated a higher death rate in the samples taken from the upper region of the brood nest than at the centre of the brood nest (fig 3). This means that where colonies are treated using formic acid the mites in the sealed cells are also killed, but not uniformly in the whole brood nest.

As a result of this finding practical trials were carried out in 1986 using brood combs without adult bees.

A box measuring 40 x 49 x 29 cm (inside dimensions!) was constructed from 4 cm thick ‘Styrodur’ sheets, which was capable of holding 10 ‘Zander hive’ size frames. The treatment was carried out using 1.5mm thick porous plastic plates of different sizes and using different amounts of 60% formic acid. These plates were placed flat over the full complement of frames.

By weighing the plates both before and after the treatment it was established how much formic acid was evaporated. Moreover the temperature and the concentration of the formic acid (in ppm) in the box were recorded. Using test equipment supplied by the firm Dräger.   The mite death rate was established by checking the brood cells immediately after treatment and after the emergence of the treated bees( see fig 3). Moreover the emergence rate and the longevity of the treated bees was compared with that of the untreated bees. To effect this directly before and after treatment a piece of brood comb with circa 200 sealed brood cells was out and placed in the brood box. Fifty emerged bees were maintained in ‘Liebefeld’ boxes in the brood chamber. A trial in a colony using marked treated winter bees and marked untreated winter bees is still underway. In some trials the open brood in the treated frames was marked and a few days after emergence the empty or sealed brood cells checked by counting

The decisive factor for effectiveness of the formic acid is the amount of acid evaporated and this factor is dependant :

With the use of a 40 x 11 cm plate soaked with 30 ml 60% formic acid, it is possible after a period of 90 minutes to achieve a mortality rate of between 80 – 95% of mites in the worker brood cells.  (measured at the emergence of the bees). During the 90 minute period, 7 – 8g of acid is evaporated, where the plate is so placed that the acid is able to evaporate freely both upwards and downwards. To achieve this the plate is supported on slips of wood and not directly on the frame tops and sized to cover all of the spaces between the frames. An air gap of 5cm should be left between the underside of the box cover and the top side of the plate. If the plate is placed directly on the frames and covered from above, considerably less acid is evaporated and the effect is very low.

The treatment using the 30ml 60% amount of acid and a treatment time of 90 minutes has virtually no effect on the hatching rate of the treated bees compared with control colonies: only bees which emerge during the treatment period are visibly damaged or killed.

Bees: No Difference in Longevity

There is no difference in longevity between treated and untreated bees. Open brood develops as a rule quite normally, compared with untreated colonies, treated open brood showed only a negligible increase in the number of larvae removed.

When the plate is soaked using 40 ml 60% acid a higher evaporation rate can be achieved, resulting in a 100% mite mortality rate. The hatching rate of the brood also increased by up to 30%.

The older brood, which is between one and three days from emergence is particularly prone to damage with this treatment level.  The open brood also showed a higher removal rate at the higher evaporation level.

While a 7 – 8g acid evaporation quantity is sufficient to achieve a good treatment success in worker brood, in the drone brood a 9 – 10g acid quantity has to be evaporated during the 90 minute treatment period to achieve a suitable mite kill in the sealed brood. A success rate of 90% was achieved using 50ml 60% acid with  a 40 x 15 cm plate.

It is recommended when using the above treatments that the procedure is carried out quite soon after the bulk of the brood on the combs to be treated is sealed, to reduce the damage, which the mites might cause the brood, to a minimum and also to avoid bees in the cells in the critical stage of hatching being exposed to the acid.

When the effect of the treatment is being checked directly after the procedure, it should noted that mites which still show faint signs of life, as a rule do not recover and die. (see above).

* Varroa Treatment Box in Crossection. The box measures 40cm wide x 49 cm long x 29cm deep including the lid and contains 10 Zander design frames *
A - Lid
B - Circa 5 cm free space
C - Absorbent plate (Illertisser!)
D - Wooden separation blocks
E - Support for frames
F - Zander frames

 

The plate should be soaked with acid at least a day before the treatment, so that the acid can become evenly distributed. The box should be constructed such that sufficient space is available between the cover and the frames (5cm) to allow the formic acid to evaporate without hindrance. (fig 4).

The ambient during treatment should be at least 20°C or higher. The internal temperature inside the box will be around 25 – 33°C at this condition.

In conclusion, from the experience gathered during the last 2 years, we ought to describe how we explain the effect of the formic acid

During the treatment of colonies and also the isolated brood frames in the fume box we measured the atmospheric acid concentration. As a rule this concentration invariably lay under 180ppm. In the treatment of brood combs, from a purely theoretical aspect, the atmospheric acid concentration resulting from the acid quantity evaporating from the plate should have reached a level of 30,000 ppm: instead of this the level measured was only a fraction of this.  What was the reason for this result? The question is able to be easily answered using a pH-paper.

All the surfaces of the treatment volume show an acid reaction. The acid evaporates from the plate surface, the air becomes totally saturated with the acid vapour,  the saturation of the air results in a condensation of the acid on the comb surfaces and the box internals. The acid diffuses from the comb surface through the air permeable cell capping into the cell interior. This process – the spreading and filling of the acid vapour in the box and the diffusion through the cell capping – requires time, especially if the effect is also to be experienced by the comb or comb components, which are quite distant from the acid plate. Due to this ‘delayed action’ phenomenon within the box the effective time of 90 minutes cannot be reduced. In the bee colony, therefore the acid plate should be in as close proximity to the brood nest as possible. In the coming years methods will be researched, which will make use of the information about this characteristic of formic acid to improve the application technique.