Video Tracking Protocol to Screen Deterrent Chemistries for Honey Bees

Summary

The loss of honey bee colonies presents a challenge to crop pollination services. Current pollinator protection practices warrant an alternative approach to minimize the contact of honey bees to harmful pesticides using repellent chemistries. Here, we provide detailed methods for a visual tracking protocol to screen deterrents for bees.

Abstract

The European honey bee, Apis mellifera L., is an economically and agriculturally important pollinator that generates billions of dollars annually. Honey bee colony numbers have been declining in the United States and many European countries since 1947. A number of factors play a role in this decline, including the unintentional exposure of honey bees to pesticides. The development of new methods and regulations are warranted to reduce pesticide exposures to these pollinators. One approach is the use of repellent chemistries that deter honey bees from a recently pesticide-treated crop. Here, we describe a protocol to discern the deterrence of honey bees exposed to select repellent chemistries. Honey bee foragers are collected and starved overnight in an incubator 15 h prior to testing. Individual honey bees are placed into Petri dishes that have either a sugar-agarose cube (control treatment) or sugar-agarose-compound cube (repellent treatment) placed into the middle of the dish. The Petri dish serves as the arena that is placed under a camera in a light box to record the honey bee locomotor activities using video tracking software. A total of 8 control and 8 repellent treatments were analyzed for a 10 min period with each treatment was duplicated with new honey bees. Here, we demonstrate that honey bees are deterred from the sugar-agarose cubes with a compound treatment whereas honey bees are attracted to the sugar-agarose cubes without an added compound.

Introduction

The European honey bee, Apis melliferaL., is an economic and agriculturally important insect that provides pollination services that are valued at more than $200 billion globally¹. In the United States and Europe, honey bee colony numbers have been declining. The United States has lost ca. 60% of managed honey bee colonies from 1947-2008 whereas Europe has lost ca. 27% from 1961-2007^2,3. There are a number of factors that might be responsible for the increased number of colony losses, including but not limited to, parasite infestations, pathogen infections, be....

Protocol

1. Prepare Sugar-agarose Cubes

1. Weigh out 8 g of sugar and place into a 50 mL Erlenmeyer flask.
2. Fill the Erlenmeyer flask with 20 mL of de-ionized water. Dissolve the sugar by swirling the flask.
3. Weigh 170 mg of agarose and add it to the sugar solution.
4. Heat the sugar-agarose solution in a microwave on high for 25 s. Dissolve the agarose into the sugar solution.
5. Allow the flask and sugar-agarose solution to cool.
   NOTE: The flask should be cool to the touch, but do not allow the solution to solidify.
   1. To prepare a sugar-agarose cube for the control treatment, pour the semi-cooled sugar-....

Results

A visual tracking protocol was developed to record the amount of time the honey bees spent in a target zone with either sugar-agarose (control treatment) or sugar-agarose-compound cube (deterrent treatment). The recorded time was analyzed using a statistical software program and the mean time spent ± standard error in the target zone is reported as a bar graph. DEET, the gold standard for insect repellent/deterrent testing, was used in this protocol as a positive control. The honey bees p.......

Discussion

This visual tracking protocol provides a simple approach to screen chemical deterrents for honey bees in a relatively quick and easy manner. There are no recommended protocols for the laboratory testing of chemical deterrents for honey bees. Previous semi- and full-field studies have examined honey bee repellents^14,15; however the described protocols are time consuming, labor intensive, and require additional facility resources outside of a general laboratory. Th.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
50 mL Erlenmeyer flask	Kimax	26500-50	used for making the sugar/agarose cubes
Sugar	Kroger		any similar product will sufffice
Deionized water			acquired in house
Agarose	Apex	20-102	used for making the sugar/agarose cubes
Mold for agarose cubes (Weigh Boat)			any mold that will provide the researcher with a 1.5 cm x 1.5 cm x 0.3 cm sugar/agarose cube will suffice
EthoVision XT	Noldus		visual tracking software
633 nm LEDs	Cyron	HTP904E	These lights were placed into a constructed light box to illuminate the arenas from below.  The box was a simple wooden structure with a frosted plastic/plexi glass cover that allowed the light to disperse upwards without any glare.
Laptop or PC	Dell	Inspiron One 2305	necessary for video tracking software. Any pc device capable of runnin tbe visual tracking software will suffice
Bee Keeping protective clothing	Dadant & Sons Inc	V0126	any protective hood and jacket will suffice
Hive tool	Dadant & Sons Inc	M00757	used to open honey bee hive
Container for honey bees			any container suitable for housing and storing honey bees will suffice
Featherweight forceps narrow tip	Bioquip	4748	used to select individual honey bees
9 cm (diameter) Petri dish	Fisher Scientific	S01778	arena used to contain individual honey bees during video tracking
Recording Device (Camera)	Basler	acA-1300-60gm	any device that can record the subject clearly and transfer the file to a computer will suffice
GraphPad Prism	Graphpad		any statistical software package will suffice