&   SEARCH
David A. Cushman logo
Testing For Apistan Resistant Varroa Mites
On Honey Bees

Strains of varroa mites that are resistant to the pyrethroid compounds fluvalinate and flumethrin, which are the active ingredients in the treatment strips Apistan and Bayvarol that are licensed in UK have developed... Starting Devon in August 2001 and becoming more widespread as of 2006.

Strains of varroa mites that are resistant to treatments used against them are a challenge to beekeepers. Since 2003 treatment Apiguard has been licensed and although it is not as strongly effective as Apistan and Bayvarol, it has good action agaist mites that are resistant to the pyrethroids and providing other measures are taken, like oxalic acid dribbling authorised for use in the UK to control varroa. Further cases of pyrethroid resistant varroa have been detected in apiaries in England since 2001.

Beltsville Apistan/Coumaphos double test

This test can be used to determine mite resistance when a beehive doesn't appear to respond to chemical mite control measures. The test uses Apistan® and CheckMite+TM strips and can be conducted simultaneously on the same group of colonies by taking two samples from each hive.

Materials required to carry out the test:

500ml jar with lid (wide-mouth pint canning jar)
light metal mesh cover for the jar (8mesh to the inch hardware cloth)
index card or similar and stapler
3/8" X 1" piece of a new Apistan®and CheckMite+TM strip
1/4cup or 2oz. measure to scoop up bees
25% alcohol or windshield washing fluid
straining cloth and 6 clothes pins
plastic or rubber gloves
plastic bucket
sheet of white paper
Step 1

Staple a 3/8" X 1" section of an Apistan® and CheckMite+TM strip to the center of an index card. Make sure to handle the Apistan® and CheckMite+TM with gloves. Place the card in the jar with a section of the Apistan® or CheckMite+TM strip facing inwards. Replace the solid, round metal section of the canning jar lid with a piece of wire mesh. The holes in the mesh should be large enough to easily let varroa through.

Shake bees from one or two brood combs into an up-turned hive lid, bucket or box. Scoop up 1/4 cup or 2oz. of bees (~150 bees) and place them into each jar, being careful not to damage the bees. Screw the lids onto the jars to stop the bees from escaping.

Step 3
Place the jars in an incubator or a warm 30°C room in the dark for 6 hours. Make sure the lids are not covered so air gets to the bees.

Step 4
After 6 hours, hold the jar about 10cm above the piece of white paper and turn it so the mesh lid is facing downwards. Hit the jar with the palm of your hand three times. Count the number of mites that fall on the paper.

Step 5
Knock the bees to the bottom of the jar. Remove the index card with the attached strip and fill the jar half-way with the alcohol or washer fluid. This should be done outside using gloves. Remove the mesh lid and replace with the original solid lid for the jar. Shake the jar vigorously for 5 min.

Step 6
Remove the solid lid and replace it with the mesh lid. Pour the fluid into the straining cloth pinned to the bucket. Refill the jar with fluid, swirl the bees around and pour through the strainer again.

Step 7
Count the number of mites recovered on the cloth. If the total number of mites recovered in both samplings (Apistan® CheckMite+TM) is less than 5, the results should be discarded.

Step 8
To calculate the percentage of mites killed by Apistan® and CheckMite+TM, divide the number of mites that initially fell on the white paper before the bees were killed, by the total number of mites (total mites = white paper mite count + mite count from bee washing).

%kill by Apistan® or CheckMite+TM = initial kill, divided by total mites x 100

Resistance test with bees
If more than 50% of the mites were killed by the Apistan® or CheckMite+TM after 6 hours, the mites should be susceptible and adequate mite control can be expected. If less than 50% of the mites are killed after 6 hours by Apistan?or CheckMite+TM, the mites may be resistant to Apistan® or CheckMite+TM.

Critical Factors for the Success of the Resistance Test
Prescreen hives using the ether roll technique (250-300 bees) and test only those hives yielding 5 or more mites. This test gives meaningful results only when performed on hives with adequate mite levels. Do not expect levels of resistance to be the same among hives. Select 12 hives per apiary. More hives are better. This test is not designed to identify individual hives showing resistance. Use apiary averages to assess the results.
Perform the test exactly as described. Jar size, size of Apistan® or CheckMite+TM pieces and temperature are important.
Ensure that bees are mobile in the jars so they contact the strips. Cool temperatures may cause the bees to cluster away from the strips. If using darkened incubator, it may be helpful to open the incubator periodically to admit light and fresh air to encourage bee movement.
It is best not to reuse strip pieces or index cards. Wash jars between tests.

Note: Do not expose jars with Apistan® or CheckMiteTM to sunlight for any length of time. It is best to keep the jars in their storage boxes before and after filling until they are incubated. Sample bees from brood frames. For accruacy and to avoid bee injury, use a measuring scoop. Do not scrape bees direclty into jars. Disclaimer: This assay is intended to screen for resistant mites and is not intended to indicate the exact level of resistance.

The bioassay involves collecting two groups of bees with mites and ushering them into glass canning jars capped with fine-mesh lids. Other materials include index cards that hold strips treated with either coumaphos or fluvalinate. By using two samples of bees, the beekeeper can test for resistance to both compounds simultaneously.

Bees are exposed to the test strips for 6 hours. Then, the jars are inverted and gently tapped to cause dead mites to fall out onto a light-colored surface, where they can be counted. The beekeeper counts any surviving mites by washing them off of the bees in the jar.

Next, the percentage of mites killed by the treatments is calculated. "Mite kills below 50 percent indicate approaching resistance," Pettis says. "Kills below 25 percent generally indicate full resistance."

Pettis verified the bioassay's reliability at commercial apiaries in Maine and Florida and on hives at ARS's Beltsville bee lab.

Mite resistance prompted by continuous use of coumaphos and fluvalinate, Pettis notes, "is forcing beekeepers to adopt integrated pest-management approaches, using selective stocks of honey bees resistant to the mite, other chemicals like formic acid and thymol, and rotation?lternating between coumaphos and fluvalinate."

In some states, Pettis reports, bee inspectors have used the bioassay results to apply for emergency-use exemptions from the U.S. Environmental Protection Agency, which requires documented mite resistance before allowing use of alternative control products.

State inspectors are also using the bioassay to conduct mite surveillance. Says Pettis: "Inspectors want to have a handle on where mite resistance is spreading within a state to give beekeepers a heads-up."?y Jan Suszkiw, Agricultural Research Service Information Staff.

This research is part of Crop Production and Protection, an ARS National Program (#305) described on the World Wide Web at www.nps.ars.usda.gov.

Jeffery S. Pettis and Mark F. Feldlaufer are with the USDA-ARS Bee Research Laboratory, 10300 Baltimore Ave., Bldg. 476, Beltsville, MD 20705-2350; phone (301) 504-8205, fax (301) 504-8736

CSL Apistan Resistance Test

Apistan Resistance Test
This sheet describes a simple test (a variation of the USDA Beltsville method) to check if varroa mites are resistant to Apistan. NE. Apistan resistant mites will almost invariably be cross-resistant to Eayvarol.
1 Cut a 9mm x 25mm piece from an Apistan strip and staple it to the centre of a piece of thin card about 75mm x 125mm in size.
2. Place the card in a 500ml jar, or lib. honey jar, with the strip facing inwards.
3 Prepare a 2-3mm mesh cover to close the jar. Plastic green house shading mesh is ideal. Cut a piece larger than the opening so that it can be folded back over the open end of the jar and secured using a strong elastic band.
4 Take precautions not to sample the queen. Shake adult bees from l or 2 brood combs into a container such as an upturned roof or washing up bowl. Gently scoop up bees with the jar until it is halt.
5 Place a sugar cube in the jar and seal off using the mesh. Store in the dark at room temperature with the mesh uppermost.
6 After 24 hours hit the upturned jar with the palm of your hand over white paper. Repeat two more times to dislodge any mites Count the mites knocked out.
7. Immerse the bees in the jar into a solution of water and washing up liquid (strong washing up strength).
8 Wash the dead bees to remove any remaining mites. Place the bees in a coarse kitchen sieve that will hold bees but let varroa mites through. Secure a honey, straining cloth or jelly bag under the sieve to retain mites. Place under a fast running tap or tap with a shower fitting to wash any remaining mites off the bees. Count the number of mites washed off Dispose of the dead bees and mites in a suitable way, such as composting.
9. If the total number of mites is less than 5 discard the results. ~
10. Calculate the efficacy as a percentage. Multiply the number of mites knocked - down by 100 and divide by the total number ofmites ie. the number knocked down plus those washed off If the answer is less than 50% it indicates that a resistance problem is likely.
11. Please complete the Voluntary Varroa Resistance form VVR T( I) and return it to the National Bee Unit. A copy of this form is on the reverse, or is obtainable from the CSL website www.csl.gov.uk, the NBU or the below named.
12. Thank you for your help and co-operation. If you have any queries please contact the below named, or the NBU on 01904462510.