As we know, honey bees face many pests and diseases that impact colonies, including trypanosomatids. Trypanosomatids are single-celled, flagellated eukaryotic parasites that are found in many different hosts ¹. In honey bees, the most common trypanosomatid is Lotmaria passim. This species colonizes the hindgut of honey bees and can influence physiology, behaviour and survival. 

What is Lotmaria passim?   

Lotmaria passim was first formally described in 2015 after being distinguished from Crithidia mellificae using molecular and morphological evidence ². The parasite exists as flagellated promastigotes, and as a non-mobile round spheroid that adheres to the gut lining in honey bees ². Morphologically, L. passim cells have a lanceolate or tear‑drop shape with a single flagellum ². L. passim colonizes the ileum and rectum of the honey bee hindgut, as both motile and stationary forms ². The stationary forms are spheroids that form single-layered sheets along the gut wall ². 

Arismendi et al. (2022) examined L. passim prevalence across eggs, larvae, pupae, newly emerged bees, nurses and foragers. L. passim was detected in 21–100% of developmental stages except eggs, where it was absent ³. Pupae, nurse bees and foragers showed high prevalence (>92%) of L. passim ³. Young larvae, old larvae and newly emerged bees showed lower prevalence (<71%) compared to foragers ³. Parasite loads ranged from 1.91 × 10¹ to 7.39 × 10⁵ cell equivalents per bee, with foragers carrying the highest loads ³. These findings indicate that L. passim is present in nearly all developmental stages except eggs ³.

Figure 1: Images of isolated cells of Lotmaria passim, wet slides (A), May Grunwald-Giemsa-stained slides (B) and scanning electron microscope (C) (photo:  Rudelli et al. 2023)

The parasite colonizes the ileum and rectum, where its cells can be shed into the gut lumen and passes to other bees through normal social behaviours ². Transmission occurs when bees ingest infected material during trophallaxis, grooming, or while cleaning contaminated brood cells ³. Honey bees frequently exchange food and contact each other during colony tasks, which are behaviours that create efficient routes for parasite movement between individuals ¹. Forager bees can also pick up fecal material on flowers, allowing the parasite to spread outside of the hive. Pollen brought back to the hive has been shown to contain L. passim, suggesting that contaminated floral resources serve as an environmental reservoir for transmission ³. 

Infected bees show reduced survival and increased sucrose responsiveness, indicating energetic stress and altered feeding behaviour ⁴. At the colony level, L. passim infection lowers vitellogenin expression and contributes to precocious foraging, accelerating physiological aging and reducing worker longevity ⁴.

It is important for beekeepers to understand the different diseases honey bees face, including emerging parasites whose full impacts are still being uncovered. Lotmaria passim is a relatively new trypanosomatid described in 2015, and although its long‑term effects on colonies are not yet fully known, it has already been detected across Canada, the United States, South America and Europe ^2,3,4.

 Written by Kaitlyn Newton, ATTTA Seasonal Apiculturist

Connecting with ATTTA Specialists

If you’d like to connect with ATTTA specialists or learn more about our program, you can:

visit our website at https://www.perennia.ca/portfolio-items/honey-bees/

Email attta@perennia.ca

References:

1. Wallace, F., 1966. The Trypanosomatid Parasites of Insects and Arachnids. Experimental Parasitology, 18, p. 124-193. 

2. Schwarz, R.S., Bauchan, G.R., Murphy, C.A., Ravoet, J., de Graaf, D.C. and Evans, J.D., 2015. Characterization of Two Species of Trypanosomatidae from the Honey Bee Apis mellifera: Crithidia mellificae Langridge and McGhee, and Lotmaria passim n. gen., n. sp. Journal of Eukaryotic Microbiology, 62(5), pp.567-583.

3. Arismendi, N., Castro, M.P., Vargas, M., Zapata, C. and Riveros, G., 2022. The trypanosome Lotmaria passim prevails in honey bees of different ages and stages of development. Journal of Apicultural Research, 61(1), pp.63-69.

4. MacInnis, C.I., 2024. How honey bees (Apis mellifera L.) respond to infection with Nosema ceranae and Lotmaria passim.

5. Rudelli, C., Isani, G., Andreani, G., Tedesco, P. and Galuppi, R., 2023. Detection of Lotmaria passim in honeybees from Emilia Romagna (Italy) based on a culture method. Journal of Invertebrate Pathology, 201, p.108007.

Honey bees face a wide range of threats from visible pests to microscopic pathogens, all of which can impact their health and survival. While some diseases like Varroa destructor or Nosema spp. are well known among beekeepers, others remain less recognizable but are still important to understand. One disease is caused by the protozoan parasite Malpighamoeba mellificae, which infests the bee’s excretory system.

Understanding Malpighamoeba mellificae 

This amoeba disease of honey bees was first discovered over a century ago in Germany ¹. Malphighamoeba mellificae is a protozoan parasite that infects an essential organ in honey bees known as the Malpighian tubules which is part of the digestive system. It is transmitted through the fecal-oral route ². Usually, infection occurs when the feces of infected bees get on the comb, which then gets ingested ³.  

The life cycle of Malphighamoeba mellificae begins when honey bees ingest the cysts. Once inside the bee, the cysts transform into trophozoites, which are the active feeding stage of the amoeba ¹. These trophozoites invade and damage the Malpighian tubules. Eventually, they form new cysts that pass through the honey bee’s hind gut and are excreted.

The Malpighian tubules, as part of the excretory system in the bee’s abdomen, play a vital role in osmoregulation.  Osmoregulation is a cellular process which maintains fluid and salt balance within the bee’s body ¹. The tubules also remove nitrogenous waste from the hemolymph by passing it into the hindgut for excretion ¹. Additionally, the Malpighian tubules help filter out foreign substances that could cause harm to the bees, like insecticides, fungicides or antibiotics ⁴. When these tubules are damaged, their inability to function properly can significantly compromise the honey bee’s overall health. 

Figure 1: Malpighamoeba mellificae disease a) showing amoebic cysts in fecal samples from western honey bee. b) Cysts observed in Malpighian tubules under a light microscope. c) Normal appearance of Malpighian tubules in a non-infected honey bee and d) Discolored Malpighian tubules of honey bees who were infected with Malpighamoeba mellificae (image: Iredale et al, 2023 ²). 

When infected by Malpighamoeba mellificae, the Malpighian tubules become swollen and filled with cysts, which impairs their function and weakens the bee ⁴. This enlargement may result from thinning of the epithelial layer, caused by the accumulation of cysts or by trophozoites feeding on the tissue ⁴. At the colony level, this disease may lead to population decline, reduced lifespan, and signs of dysentery ³. It is often found alongside Nosema spp. infections ¹.

Diagnosing Malpighamoeba mellificae requires a compound light microscope and follows a similar method to screening for Nosema. About ten bees should be collected from the front of a hive and placed into a sealable bag with 10 mL of distilled water ³. The bees are then crushed to release internal contents, and a drop of the resulting liquid is placed on a microscope slide for examination of the 8-10 μm in diameter cysts at 400x magnification ³. However, this technique can also have its limitations. The amoeba can remain in its trophozoite stage for extended periods and only form cysts when its environmental conditions are poor ¹. While cysts are easy to identify due to their consistent shape, the trophozoites are much harder to recognize, especially in mild infections ¹.

Figure 2: Distribution of M. mellificae showering where disease is present (yellow), suspected presence (teal) and no presence/unreported (purple) (image: Gurland et al, 2024 ³). 

This disease has been reported in various regions where European honey bees are kept, including Canada ³. The disease may be seasonal, with higher prevalence in winter and early spring ³. Despite its long history, Malpighamoeba mellificae remains understudied, and no specific treatments are available. Malphigamoeba mellificae has not been proven to be a problem for beekeepers in our region. But we do need to understand the presence of the pathogen and the implications for honey bee health in the Maritimes.

 Written by Kaitlyn Newton, ATTTA Seasonal Apiculturist 

Connecting with ATTTA Specialists

If you’d like to connect with ATTTA specialists or learn more about our program, you can:

visit our website at https://www.perennia.ca/portfolio-items/honey-bees/

Email attta@perennia.ca

References:

1. Schäfer, M.O., Horenk, J. and Wylezich, C., 2022. Molecular detection of Malpighamoeba mellificae in honey bees. Veterinary Sciences, 9(3), p.148.
2. Iredale, M.E., Viadanna, P.H., Subramaniam, K., Tardif, E., Bonning, B.C. and Ellis, J.D., 2023. Report of amoebic disease in a colony of Western honey bees (Apis mellifera). Veterinary Pathology, 60(5), pp.709-713. 
3. Gurland, H.N., Fulton, J.C., Iredale, M.E. and Vu, A.T., 2024. Monitoring for Amoebic Disease (Malpighamoeba mellificae) in Honey Bee Colonies: ENY2112/IN1431, 10/2024. EDIS, 2024(5).
4. Rossi, M., Ott, S.R. and Niven, J.E., 2020. Malpighamoeba infection compromises fluid secretion and P-glycoprotein detoxification in Malpighian tubules. Scientific Reports, 10(1), p.15953.

American Foulbrood (AFB) is a highly contagious bacterial brood disease in honey bees. The disease is caused by the bacterium Paenibacillus larvae which produce spores that are viable in the environment for decades. Protecting honey bees from AFB is extremely important and recently a new product has been developed that demonstrates providing some degree of immunity to the disease. The Canadian Food Inspection Agency (CFIA) has conditionally licensed the product, under veterinary supervision, and it is expected to be distributed on a limited basis to commercial beekeepers in Canada starting this Spring. To learn more about AFB, and the potential benefits this new product could provide, read this week’s blog.

Product for American Foulbrood Protection Approved for Canada's Honey Bees

American foulbrood (AFB) is caused by the Paenibacillus larvae bacterium which produce spores that are viable in the environment for decades. Worker bees can transport the spores and spread the disease, but it is the larval stage that shows clinical symptoms of infection. Infected brood usually die at the pre-pupal or pupal stage, severely weakening the colony and eventually killing it. Because of its virulent nature and detrimental effects on honey bee colonies, AFB is classified as a notifiable disease worldwide.

Antibiotics, such as Oxytetracycline, are largely ineffective at eradicating the disease as they are only effective against the vegetative state of the bacteria, which is when bacteria can grow and reproduce. Once a hive shows the clinical symptoms of the disease, the only effective way to eradicate it and prevent the spread of the disease is by burning the equipment, and the colony.

Therefore, it is extremely important to protect honey bees from AFB, and there is now a new product that could help. This product is being called a vaccine in the media and throughout published research. In an original study done by Dickel et al. (2022) the group of researchers demonstrated that the oral administration of an inactivated AFB bacterin to the queen bee is safe and induces some level of protection in the next generation of larvae against AFB infection in a hybrid lab and field study. The suggested delivery of this new product is that the inactive bacteria is mixed into powered sugar and glucose syrup for the worker bees to eat, and their secretions are fed to the queen. Then the next generation of developing bees, produced from the queen, have increased immunity to AFB.

The research trails, funded by Dalan Animal Health, showed between 30% to 50% decrease in AFB infection among colonies that received the product compared to colonies that received the placebo product in the hybrid lab and field study. The researchers fed the queens the product for 8 days in queen cages in the lab, and those queens were released into nucleus hives. Then, 18 days post-queen placement, frames of brood were collected and fed food with a known concentration of AFB spores and assessed for mortality over time in the lab.

It is known that this is an antibody-free immunological response since insects lack antibodies. It is proposed that information about the disease agent is transferred to the next generation with the help of the egg yolk protein Vitellogenin carrying immune elicitors, such as pieces of bacteria. Another proposed mechanism of action is a transfer of mRNA and proteins as well as epigenetic factors. Further research is needed to determine this product’s mechanism of action.

©Dalan Animal Health

This product could be beneficial in protecting honey bees from AFB and decrease the economic impact of the disease. That being said, the efficacy of the product is still largely unproven both from a multitude of research trials, and through beekeeping practices. In Fall 2023, the Canadian Food Inspection Agency (CFIA) conditionally licensed the product, under veterinary supervision, and it is expected to be distributed on a limited basis to commercial beekeepers in Canada starting this Spring. The United States Department of Agriculture (USDA) conditionally licensed the product by Dalan Animal Health back in January 2023. Some key differences exist between having a product licensed by CFIA versus USDA, including the level of efficacy a product must demonstrate. Since there is differences in the standard a product must meet, it is typical that CFIA will license a product after USDA. Both CFIA and USDA only license products that have been proven to be safe for livestock and humans, when following manufacturer instructions.

Although this product could provide multiple benefits to the Canadian honey bee industry, it is important to recognize that beekeepers must still remain vigilant in detecting potential AFB infections. Beekeepers should examine brood for discolored larvae; dead larvae; brood that appears greasy, sunken, or punctured; and poor brood pattern. Additionally, beekeepers should perform the rope test regularly, and especially on any abnormal looking larvae. Finally, if a beekeepers does have reason to suspect an AFB infection this must be reported to the provincial apiculturist immediately.

References

Dickel, F., Bos, N.M.P., Hughes, H., Martín-Hernández, R., Higes, M., Kleiser, A. and Freitak, D., 2022. The oral vaccination with Paenibacillus larvae bacterin can decrease susceptibility to American Foulbrood infection in honey bees—A safety and efficacy study. Frontiers in Veterinary Science, 9.

Connecting with ATTTA Specialists

If you’d like to connect with ATTTA specialists or learn more about our program, you can:

visit our website at https://www.perennia.ca/portfolio-items/honey-bees/

Email abyers@perennia.ca

An IPM Series: The Importance of Monitoring for Pests and Diseases

Beekeepers in Atlantic Canada are all too familiar with managing a variety of honey bee pests and diseases. Fortunately, they have a wide variety of tools available to ensure the health of their colonies. Using a variety of strategies to manage pests, including chemical, biological, cultural, and physical controls, as well as limiting the frequency of chemical treatments, is part of integrated pest management (IPM). Integrated pest management is the best management practice for beekeepers, as it aids in preventing an overreliance, and potential reduced efficacy, of chemical treatments. A key component to any beekeeper’s IPM plan is monitoring the population levels of pests and diseases. This week’s blog will discuss Varroa mite monitoring options, the advantages and disadvantages of each, and the overall importance of assessing pest population levels before and after any treatment.

An IPM Series: The Importance of Monitoring for Pests and Diseases

In Atlantic Canada, the pest most monitored for is Varroa destructor. Varroa are parasitic mites of honey bees that cause severe damage to colonies if left untreated. They cause damage through direct feeding on bees, and by the transmission of a variety of viruses. That is why it is important to monitor Varroa mites and treat against the pest when population levels reach the economic threshold. There are a variety of ways to sample colonies, both to determine if mites are present at levels that are damaging to the colony, and to assess the effectiveness of control measures.

Washing samples of adult bees is a common way to determine Varroa mite infestation levels. Both alcohol washes and ether rolls require a sample of 300 bees (1/2 cup) from the brood nest. These nurse bees are the most likely to have adult Varroa mites parasitizing them. However, the brood nest is also the most likely location of the queen, so it is important to either isolate her or make sure she is not on the frame being sampled. For directions on how to perform an alcohol wash or ether roll refer to ATTTA’s “Summer Disease and Pest Monitoring in Honey Bees” fact sheet. For an alcohol wash in May, the economic threshold is 2 mites per 100 bees, and in August it is 3 mites per 100 bees. For an ether roll in May, the economic threshold is 1 mite per 100 bees, and in August it is 2 mites per 100 bees. One advantage to washes is that there is a predetermined economic threshold to indicate if treatment is required. Additionally, washes only require one trip to the bee yard and results can be obtained instantly. A disadvantage of washes is that 300 or more bees are killed during the sampling. Also, when doing washes, only a small sample of the colony population is being assessed.

Alcohol wash (ATTTA©2021).

Sticky boards are an effective and easy way to assess mite levels in a colony. The sticky board is placed in a screen bottom board and after 24 hours it is removed, and the number of mites is counted. In May the economic threshold is 9 mites per 24 hours, and in August it is 12 mites per 24 hours. This method of monitoring also doubles as a physical (non-chemical) treatment to reduce Varroa mite numbers. A main advantage of sticky boards is the entire hive is the sample. Additionally, sticky boards do not kill bees or endanger the queen. A disadvantage of sticky boards is that they require two trips to the bee yard and potentially additional equipment.

There are also a couple of other less common monitoring options. The sugar shake method dislodges mites from bees. Like the washes, 1/2 a cup of bees is collected in a jar, and then the bees are coated in sugar which dislodges the mites. Once mites have been shaken out of the jar, the sugar-coated bees are returned to the hive. Another less common monitoring option is the CO₂ method. In the CO₂ method, bees and mites are anesthetized by exposure to carbon dioxide gas. The sample of bees is then gently shaken causing the mites to fall from the bees. The bees are then returned to the hive. The supposed advantage of both methods is that the bees are returned to the colony unharmed. This assumption of bees surviving the procedure is unproven. A disadvantage of both methods is that although it will detect the presence of mites, there is no standard economic threshold in Atlantic Canada for these methods.

It is essential to monitor for pests and disease not only before treating colonies, but also following treatment to know if the method of treatment was successful. For more information about the importance and challenges of IPM, be sure to read the next two blogs in this series.

Connecting with ATTTA Specialists

If you’d like to connect with ATTTA specialists or learn more about our program, you can:

visit our website at https://www.perennia.ca/portfolio-items/honey-bees/

Email abyers@perennia.ca