Testing Pollen of Single and Stacked Insect-Resistant Bt-Maize on In vitro Reared Honey Bee Larvae

Abstract American foulbrood (AFB) is a highly virulent disease afflicting honey bees (Apis mellifera). The causative organism, Paenibacillus larvae, attacks honey bee brood and renders entire hives dysfunctional during active disease states, but more commonly resides in hives asymptomatically as inactive spores that elude even vigilant beekeepers. The mechanism of this pathogenic transition is not fully understood, and no cure exists for AFB. Here, we evaluated how hive supplementation with probiotic lactobacilli (delivered through a nutrient patty; BioPatty) affected colony resistance towards a naturally occurring AFB outbreak. Results demonstrated a significantly lower pathogen load and proteolytic activity of honey bee larvae from BioPatty-treated hives. Interestingly, a distinctive shift in the microbiota composition of adult nurse bees occurred irrespective of treatment group during the monitoring period, but only vehicle-supplemented nurse bees exhibited higher P. larvae loads. In vitro experiments utilizing laboratory-reared honey bee larvae showed Lactobacillus plantarum Lp39, Lactobacillus rhamnosus GR-1, and Lactobacillus kunkeei BR-1 (contained in the BioPatty) could reduce pathogen load, upregulate expression of key immune genes, and improve survival during P. larvae infection. These findings suggest the usage of a lactobacilli-containing hive supplement, which is practical and affordable for beekeepers, may be effective for reducing enzootic pathogen-related hive losses.

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Anti-Virulence Strategy against the Honey Bee Pathogenic Bacterium Paenibacillus larvae via Small Molecule Inhibitors of the Bacterial Toxin Plx2A

Toxins ◽

10.3390/toxins13090607 ◽

2021 ◽

Vol 13 (9) ◽

pp. 607

Author(s):

Julia Ebeling ◽

Franziska Pieper ◽

Josefine Göbel ◽

Henriette Knispel ◽

Michael McCarthy ◽

...

Keyword(s):

Small Molecules ◽

Honey Bee ◽

Small Molecule ◽

Bacterial Infections ◽

Small Molecule Inhibitors ◽

Paenibacillus Larvae ◽

Bacterial Toxin ◽

American Foulbrood ◽

Honey Bee Larvae

American Foulbrood, caused by Paenibacillus larvae, is the most devastating bacterial honey bee brood disease. Finding a treatment against American Foulbrood would be a huge breakthrough in the battle against the disease. Recently, small molecule inhibitors against virulence factors have been suggested as candidates for the development of anti-virulence strategies against bacterial infections. We therefore screened an in-house library of synthetic small molecules and a library of flavonoid natural products, identifying the synthetic compound M3 and two natural, plant-derived small molecules, Acacetin and Baicalein, as putative inhibitors of the recently identified P. larvae toxin Plx2A. All three inhibitors were potent in in vitro enzyme activity assays and two compounds were shown to protect insect cells against Plx2A intoxication. However, when tested in exposure bioassays with honey bee larvae, no effect on mortality could be observed for the synthetic or the plant-derived inhibitors, thus suggesting that the pathogenesis strategies of P. larvae are likely to be too complex to be disarmed in an anti-virulence strategy aimed at a single virulence factor. Our study also underscores the importance of not only testing substances in in vitro or cell culture assays, but also testing the compounds in P. larvae-infected honey bee larvae.

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Flumethrin at honey-relevant levels induces physiological stresses to honey bee larvae (Apis mellifera L.) in vitro

Ecotoxicology and Environmental Safety ◽

10.1016/j.ecoenv.2019.110101 ◽

2020 ◽

Vol 190 ◽

pp. 110101 ◽

Cited By ~ 3

Author(s):

Suzhen Qi ◽

Lizhen Zhu ◽

Donghui Wang ◽

Chen Wang ◽

Xiaofeng Chen ◽

...

Keyword(s):

Apis Mellifera ◽

Honey Bee ◽

Honey Bee Larvae

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Combined Toxicity of Insecticides and Fungicides Applied to California Almond Orchards to Honey Bee Larvae and Adults

Insects ◽

10.3390/insects10010020 ◽

2019 ◽

Vol 10 (1) ◽

pp. 20 ◽

Cited By ~ 28

Author(s):

Andrea Wade ◽

Chia-Hua Lin ◽

Colin Kurkul ◽

Erzsébet Ravasz Regan ◽

Reed M. Johnson

Keyword(s):

Larval Development ◽

Honey Bee ◽

Honey Bees ◽

Pesticide Exposure ◽

Larval Mortality ◽

Combined Toxicity ◽

Pollination Services ◽

Almond Orchards ◽

Honey Bee Larvae

Beekeepers providing pollination services for California almond orchards have reported observing dead or malformed brood during and immediately after almond bloom—effects that they attribute to pesticide exposure. The objective of this study was to test commonly used insecticides and fungicides during almond bloom on honey bee larval development in a laboratory bioassay. In vitro rearing of worker honey bee larvae was performed to test the effect of three insecticides (chlorantraniliprole, diflubenzuron, and methoxyfenozide) and three fungicides (propiconazole, iprodione, and a mixture of boscalid-pyraclostrobin), applied alone or in insecticide-fungicide combinations, on larval development. Young worker larvae were fed diets contaminated with active ingredients at concentration ratios simulating a tank-mix at the maximum label rate. Overall, larvae receiving insecticide and insecticide-fungicide combinations were less likely to survive to adulthood when compared to the control or fungicide-only treatments. The insecticide chlorantraniliprole increased larval mortality when combined with the fungicides propiconazole or iprodione, but not alone; the chlorantraniliprole-propiconazole combination was also found to be highly toxic to adult workers treated topically. Diflubenzuron generally increased larval mortality, but no synergistic effect was observed when combined with fungicides. Neither methoxyfenozide nor any methoxyfenozide-fungicide combination increased mortality. Exposure to insecticides applied during almond bloom has the potential to harm honey bees and this effect may, in certain instances, be more damaging when insecticides are applied in combination with fungicides.

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Horizontal Honey-Bee Larvae Rearing Plates Can Increase the Deformation Rate of Newly Emerged Adult Honey Bees

Insects ◽

10.3390/insects12070603 ◽

2021 ◽

Vol 12 (7) ◽

pp. 603

Author(s):

Juyeong Kim ◽

Kyongmi Chon ◽

Bo-Seon Kim ◽

Jin-A Oh ◽

Chang-Young Yoon ◽

...

Keyword(s):

Apis Mellifera ◽

Honey Bee ◽

Honey Bees ◽

Deformation Rate ◽

Sublethal Effects ◽

Horizontal Position ◽

Control Groups ◽

Honey Bee Larvae ◽

Ideal Method

Rearing honey bee larvae in vitro is an ideal method to study honey bee larval diseases or the toxicity of pesticides on honey bee larvae under standardized conditions. However, recent studies reported that a horizontal position may cause the deformation of emerged bees. Accordingly, the purpose of this study was to evaluate the emergence and deformation rates of honey bee (Apis mellifera ligustica) larvae reared in horizontal and vertical positions. The study was conducted under the same laboratory conditions with three experimental groups, non-capped or capped horizontal plates and capped vertical plates. However, our results demonstrated that the exhibited adult deformation rates of the horizontal plates were significantly higher (27.8% and 26.1%) than those of the vertical plates (11.9%). In particular, the most common symptoms were deformed wings and an abnormal abdomen in the horizontal plates. Additionally, adults reared on horizontal plates were substantially smaller (10.88 and 10.82 mm) than those on vertical plates (11.55 mm). Considering these conclusions, we suggest that a vertical rearing method is more suitable when considering the deformation rates of the control groups to verify the sublethal effects of pesticides on honey bees.

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Inhibitory effect of indole analogs against Paenibacillus larvae, the causal agent of American foulbrood disease

Journal of Insect Science ◽

10.1093/jisesa/iex080 ◽

2017 ◽

Vol 17 (5) ◽

Cited By ~ 5

Author(s):

Israel Alvarado ◽

Joseph W Margotta ◽

Mai M Aoki ◽

Fernando Flores ◽

Fresia Agudelo ◽

...

Keyword(s):

Honey Bee ◽

Spore Germination ◽

Inhibitory Effect ◽

Antagonistic Effect ◽

Paenibacillus Larvae ◽

American Foulbrood ◽

Bacterial Proliferation ◽

American Foulbrood Disease ◽

Honey Bee Larvae

Abstract Paenibacillus larvae, a Gram-positive bacterium, causes American foulbrood (AFB) in honey bee larvae (Apis mellifera Linnaeus [Hymenoptera: Apidae]). P. larvae spores exit dormancy in the gut of bee larvae, the germinated cells proliferate, and ultimately bacteremia kills the host. Hence, spore germination is a required step for establishing AFB disease. We previously found that P. larvae spores germinate in response to l-tyrosine plus uric acid in vitro. Additionally, we determined that indole and phenol blocked spore germination. In this work, we evaluated the antagonistic effect of 35 indole and phenol analogs and identified strong inhibitors of P. larvae spore germination in vitro. We further tested the most promising candidate, 5-chloroindole, and found that it significantly reduced bacterial proliferation. Finally, feeding artificial worker jelly containing anti-germination compounds to AFB-exposed larvae significantly decreased AFB infection in laboratory-reared honey bee larvae. Together, these results suggest that inhibitors of P. larvae spore germination could provide another method to control AFB.

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