3.3.4.3. Pin-killed brood assay

The pin test method is recommended in Europe as a standard in field selection programmes, because it shows a significant correlation with the removal of varroa infested brood, can be standardized and is easily handled by beekeepers. A statistical tool has been established to include pin test data in the estimation of breeding values for varroa tolerance (see 4.1). For the pin-killed brood assay protocol, see Fig. 16 while following the numbered protocol below. Additionally, Fig. 17 shows images of the protocol being applied in the field.

  1. A rhomboid frame of a 10×10 cell wide template (Fig. 16, number 2) is placed on a brood comb containing young pupae (Fig. 16, number 1)
  2. The upper left and lower right cells are marked with a colour felt-tip pen (Fig. 16, number 3)
  3. 50 capped brood cells are pierced (Fig. 16, number 4) row by row from left to right with a fine insect pin (entomological pin size No 2).
  4. Cell 51 is marked to identify the treated brood area (Fig. 16, number 3).
  5. The comb is marked on the top bar and placed back to the brood nest in its former position.
  6. After 7-15 hours (uniform for all colonies within the comparison) the removal progress is checked. All cells that are still sealed or contain the remains of brood are counted and subtracted from 50. The percentage of completely cleaned cells is noted in the protocol.
  7. The highest discriminatory power of the test is reached when all of the test colonies remove an average of 50 % of the pupae within the time interval. Therefore, the time interval between piercing the cells and checking should be adapted to the average removal response of the test population. If the average removal rate is much lower than 50 %, the time interval should be prolonged to yield higher differences between colonies with high and low hygienic behaviour.  If the average removal is much higher than 50 %, a shorter time interval should be realized in further test repetitions.
  8. The test should be repeated 2-3 times during the main brood season.


Fig. 16. Pin-killed test for hygienic behaviour. The numbers correspond to text references in Section 3.3.4.3.

Figure 16

Fig. 17. Pin test: a. Piercing 50 cells containing young pupae; b. Control of brood removal after about 8 hours, many cells are opened but not removed; c. Nearly all cells are completely cleaned. Photos: R Büchler.

Figure 17