Detection and surveillance of Tropilaelaps
Tropilaelaps mites are indigenous brood parasites of giant Asian honey bees that have spread to the Western honey bee, Apis mellifera, following their introduction to Asia. Infestations of A. mellifera by Tropilaelaps mercedesae have been reported in areas far beyond the range of their native hosts, leading to severe diseases in A. mellifera colonies. Additionally, the rise in global trade and new transmission pathways make these mites a significant emerging global threat to honey bees. In England and Wales, Tropilaelaps spp. are classified as a statutory notifiable pest. The National Bee Unit (NBU) and Fera Science ltd oversee a national scheme for monitoring and diagnosing Tropilaelaps spp. in honey bee colonies, adhering to standard operating procedures (SOPs). This initiative aimed to evaluate the sensitivity and effectiveness of current national SOPs compared to new detection methods for Tropilaelaps spp. mites in honey bee colonies.
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Distribution of giant Asian honey bees (Apis dorsata) and the current confirmed distribution of Tropilaelaps mercedesae (correct Jan 2025).
Methods
Sixty honey bee (A. mellifera) colonies in Chiang Mai province, Thailand were monitored for the level of T. mercedesae using existing SOP methods, which included brood uncapping, floor debris sampling, comb bumping and the use of sticky floor inserts. Additional novel methods, traditionally used for Varroa monitoring were also trialled which included alcohol wash, CO2 sampling and using icing sugar roll of 300 adult honey bees.
Brood uncapping -
The SOP method uncapped 200 sealed drone brood cells containing pink eyed pupae using an uncapping fork. However, this technique damaged the pupae making identification of Tropilaelaps spp extremely difficult as the mites became trapped by exudate from the damaged pupae. As a result the method was adapted and a 10 x 10 rhombus template of 100 brood cells was placed over a section of sealed brood. Larvae or pupa were removed from each cell using forceps and carefully examined them for the presence of Tropilaelaps.
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Dead brood on sticky floor insert 24 hours after 'bump' method surveillance had been carried out on colony.
Comb bump method -
Adult bees were shaken from every brood frame of a colony. frames were then held horizontally over a white tray and the wooden edge of the brood frame was firmly 'bumped' (hit) on the edge of the white tray. This allowed for phoretic mites to be dislodged into the tray which could then be examined for the presence of mites.
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It should be noted that when sticky floor inserts were removed from colonies 24 hours after the bump method had been used significant amounts of dead brood were observed on the inserts and on further examination this brood had been removed by the bees from the 'bumped' frames.
Sticky floor inserts -
Sticky floors, made up of a sticky board (oil or commercially available sticky paper insert) protected by a shallow wooden frame and topped with a plastic mesh covering were deployed in colonies. Acaricide strips were not utilised as the ambient temperatures were outside the manufacturers guidance, therefore, only natural mite drop after 24 hours was recorded.
Floor debris sampling -
Hive floor debris was collected and sent for laboratory analysis. Endecott sieves were stacked in decreasing size (1.7mm, 1mm, 0.7mm, 0.35mm) and the hive debris were washed through the sieves, resulting in size separation of the debris. Although we initially placed the sieve and contents in ethanol, we quickly discovered that Tropilaelaps didn’t float as expected. Switching to demineralised water revealed that while Tropilaelaps did float in demineralised water, identifying them among the other debris proved to be impractical.
Alcohol wash, CO2, and icing sugar roll -
A Varroa EasyCheck was used to sample ~300 bees and assess mite infestation levels. The adult bees from two frames were shaken into a bowl. Three separate ½ cup scoops of bees were sampled and added to 3 separate Varroa EasyChecks. One contained 300ml of alcohol and another contained 25g of icing sugar respectively. CO2 was gradually added to the third until the bees were anaesthetised. The bees in all three EasyChecks were then agitated or rolled gently to dislodge mites. The alcohol was assessed for Tropilaelaps and Varroa, while the icing sugar and CO2 EasyChecks were shaken onto a white tray and assessed for the mites.
Alcohol wash, CO2, and an icing sugar roll of ~300 bees using a Varroa EasyCheck.
Results
Out of the 60 colonies examined, the average size was 5.3 frames of bees and 3.8 frames of brood T. mercedesae was found in 80% of the colonies and in all of the apiaries (Fig 1). A total of 53 colonies were evaluated for the effectiveness of alcohol wash, brood bump, CO2, uncapping, and sugar roll. Mite drop was monitored using oiled floors in 24 colonies and sticky traps in 29 colonies. Ethanol flotation was not applied to floor debris samples since preliminary tests showed that Tropilaelaps did not consistently float in ethanol, making this method ineffective.
Fig 1. The number of T. mercedesae mites observed per colony and by apiary.
When comparing detection methods across all 53 colonies, it was clear that both uncapping brood and a sugar roll of bees were more likely to detect Tropilaelaps mites than an alcohol wash of adult bees in ethanol. Bumping the brood and using CO2 to knock down the mites on adult bees were not better than alcohol wash (Fig 2A). When restricting to those colonies where mite drop was monitored using sticky traps, the sticky traps were significantly more likely to detect the presence of Tropilaelaps mites when compared to an alcohol wash of adult bees (Fig 2B).
Fig 2. The odds ratio of T. mercedesae detection using different methods when compared to alcohol washing mites from ~300 adult bees.
The study emphasizes the necessity of conducting 'real-life' field trials to evaluate the effectiveness of detection methods for exotic pests. Our findings have modified the Tropilaelaps surveillance policy in England and Wales, resulting in more reliable detection methods for these harmful invasive mites.
While earlier research established effective detection techniques, such as the bump method and brood examination, it is crucial to consider the practicality and willingness of beekeepers to engage with such methods. Additionally, one should not assume that surveillance techniques developed in experimental settings will be adequate for real-world scenarios, especially when they originate from regions with significantly different beekeeping practices and climates.
This project was generously funded by Bee Disease Insurance. You can find out more about the work on the Bee Disease Insurance YouTube channel.
Dr Bajaree Chuttong of Chiang Mai University kindly hosted the team and supported our work.
This project was carried out in collaboration with the Animal Plant Health Agency's National Bee Unit and Fera Science limited and with the support of DEFRA.
