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Miscellaneous standard methods for Apis mellifera research

Summary:

A variety of methods are used in honey bee research and differ depending on the level at which the research is conducted. On an individual level, the handling of individual honey bees, including the queen, larvae and pupae are required. There are different methods for the immobilising, killing and storing as well as determining individual weight of bees. The precise timing of developmental stages is also an important aspect of sampling individuals for experiments. In order to investigate and manipulate functional processes in honey bees, e.g. memory formation and retrieval and gene expression, microinjection is often used. A method that is used by both researchers and beekeepers is the marking of queens that serves not only to help to locate her during her life, but also enables the dating of queens. Creating multiple queen colonies allows the beekeeper to maintain spare queens, increase brood production or ask questions related to reproduction. On colony level, very useful techniques are the measurement of intra hive mortality using dead bee traps, weighing of full hives, collecting pollen and nectar, and digital monitoring of brood development via location recognition. At the population level, estimation of population density is essential to evaluate the health status and using beelines help to locate wild colonies. These methods, described in this paper, are especially valuable when investigating the effects of pesticide applications, environmental pollution and diseases on colony survival.

    1. 2.1. Standard methods for immobilising, killing and storing adult Apis mellifera in the laboratory
    2. 2.1.1. Introduction
    3. 2.1.2. Immobilising adults
    4. 2.1.2.1. Physical immobilisation
    5. 2.1.2.2. Chemical and physical immobilisation
    6. 2.1.2.2.1. Carbon dioxide
    7. 2.1.2.2.2. Chilling
    8. 2.1.2.2.3. Anaesthesia considerations
    9. 2.1.3. Killing adults
    10. 2.1.3.1. Thermal killing
    11. 2.1.3.1.1. Cold
    12. 2.1.3.1.2. Heat
    13. 2.1.3.2. Mechanical killing
    14. 2.1.3.3. Chemical killing
    15. 2.1.4. Storing dead adults
    16. 2.2. Determination of individual bee weight
    17. 2.2.1. Balance required for weighing individual bees or larvae or body parts
    18. 2.2.2. Weighing of larvae
    19. 2.2.3. Weighing of adult honey bees
    20. 2.2.4. Weighing body parts
    21. 2.2.5. Determining dry weight
    22. 2.3. Microinjection
    23. 2.3.1. Introduction
    24. 2.3.2. Microinjection using a Hamilton syringe
    25. 2.3.3. Microinjection of small volumes using the Nanoject device and other micro injectors
    26. 2.3.4. Perspectives
    27. 2.4. Marking honey bee queens
    28. 2.4.1. Colour-marking queens
    29. 2.4.1.1. Marking type
    30. 2.4.1.2. Procedure for paint marking
    31. 2.4.1.3. Procedure for marking with Opalith discs
    32. 2.4.1.4. Colour marking code
    33. 2.4.2. Clipping queens’ wings
    34. 2.5. Obtaining brood and adults of known age
    35. 2.5.1. Obtaining brood of known age
    36. 2.5.1.1. Procedure to obtain worker or drone brood of known age
    37. 2.5.1.2. Procedure to obtain queen brood of known age
    38. 2.5.2. Obtaining pupae of known age
    39. 2.5.3. Recognising the instar of larvae
    40. 2.5.4. Recognising the age of larvae
    41. 2.5.5. Recognising the age of pupae
    42. 2.5.6. Obtaining workers of known age
    43. 2.5.7. Conclusion
    1. 3.1. Using a haemocytometer to estimate the concentration of cells, spores or sperms
    2. 3.1.1. Total or microscopic count
    1. 4.1. Weighing full hives
    2. 4.1.1. Introduction
    3. 4.1.2. The Capaz hive scale
    4. 4.1.3. The honey meter
    5. 4.1.4. The use of the data from an electronic scale
    6. 4.2. Using beelines to locate wild honey bee colonies
    7. 4.2.1. Introduction
    8. 4.2.2. Suggested materials
    9. 4.2.3. Establishing a beeline
    10. 4.2.3.1. Setting up a feeding station
    11. 4.2.4. Following the beeline
    12. 4.2.4.1. Observing beelines
    13. 4.2.4.2. Tracking the beeline
    14. 4.2.4.3. Using a mobile feeding station
    15. 4.2.5. Locating the honey bee nest
    16. 4.2.6. Alternative methods
    17. 4.2.6.1. Following bees from water sources
    18. 4.2.6.2. Beelining with a bee box
    19. 4.2.6.3. Triangulating with feeding stations
    20. 4.2.6.4. Calculating the distance between a honey bee nest and feeding station by timing a forager’s round trip
    21. 4.3. Honey bee colony density estimations
    22. 4.3.1. Determining a colony density index using feeding stations
    23. 4.3.1.1. Material used
    24. 4.3.1.2. Procedure
    25. 4.3.1.3. Index data
    26. 4.3.1.4. Statistical analyses
    27. 4.3.2. Determination of honey bee colony density using genetic markers
    28. 4.3.3. Sampling
    29. 4.3.3.1. Drone sampling
    30. 4.3.3.2. Worker sampling
    31. 4.3.3.3. Genotyping
    32. 4.3.3.4. Genetic diversity measures and reconstruction of queen genotypes
    33. 4.3.3.5. Non-detection and non-sampling errors
    34. 4.3.3.6. Density estimation
    35. 4.3.4. Future research needs and perspectives
    36. 4.4. Estimating the number of dead honey bees expelled from a honey bee colony with a trap
    37. 4.4.1. Aim of using dead bee traps
    38. 4.4.2. Limitations of using dead bee traps
    39. 4.4.3. Types of dead bee traps
    40. 4.4.4. Dead bee traps requirements as gathered from the literature
    41. 4.4.5. Recommended dead bee traps to use
    42. 4.4.6. Building a dead bee trap
    43. 4.4.7. Protocol for calibrating dead bees in traps
    44. 4.4.8. Protocol for using a dead bee trap
    45. 4.4.9. Dead bee trap trade-offs
    46. 4.5. Creating multiple queen colonies
    47. 4.5.1. Mandible clipping procedure
    48. 4.5.2. Preparation of colonies destined to host the multiple queens
    49. 4.5.3. Steps for maintenance of an artificially established multiple-queen social organisation
    50. 4.6. Digital monitoring of brood development via location recognition
    51. 4.6.1. Introduction
    52. 4.6.2. Procedure for data acquisition
    53. 4.6.2.1. Software requirements
    54. 4.6.2.2. Before starting the project
    55. 4.6.2.3. Image acquisition
    56. 4.6.3. Image analysis
    57. 4.6.3.1. Analysis of the first image (BFD 00)
    58. 4.6.3.2. For all consecutive images (BFD + 05, 10, 16, 22)
    59. 4.6.4. Finalisation of the analysis
    60. 4.6.5. Conclusion
    61. 4.7. Collecting pollen and nectar from bees and flowers
    62. 4.7.1. Introduction
    63. 4.7.2. Methods for pollen collection
    64. 4.7.3. Nectar collection
    65. 4.7.3.1. Collecting nectar from honey bees
    66. 4.7.3.2. Nectar collection from flowers
    67. 4.7.4. Precautions when sampling pollen and nectar for residue analyses
    68. 4.7.4.1. Collection of fresh pollen from flowers
    69. 4.7.4.1.1. Using paper bags to collect fresh pollen
    70. 4.7.4.1.2. Manual collection of fresh pollen
    71. 4.7.4.1.3. Using a paint brush for collection of fresh pollen
    72. 4.7.4.1.4. Collection of fresh pollen from smaller flowers such as canola
    73. 4.7.4.2. Collection of bee collected pollen using pollen traps
    74. 4.7.4.3. Ensuring quality of bee collected pollen