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Bioactivity studies of porphyrinoids against microsporidia isolated from honeybees
Katarzyna Buczek1, Mariusz Trytek2, Kamil Deryło3
1Department of Industrial and Environmental Microbiology, Institute of Biological Sciences, Faculty of Biology and Biotechnology, Maria Curie-Skłodowska University, Akademicka 19, 20-033, Lublin, Poland.
Abstract:
Microsporidian infections are dangerous to honeybees due to the absence of an efficient treatment for nosemosis. In the present work, the abilities of several porphyrins to directly inactivate microsporidia derived from Nosema-infected honeybees were studied in vitro. Amide derivatives of protoporphyrin IX (PPIX) conjugated with one and two amino acid moieties were synthesized, and their activities were compared with those of two cationic porphyrins, TMePyP and TTMePP. The most active porphyrins, PP[Lys-Asp]2, PP[Lys-TFA]2, PP[Asp(ONa)2]2 and PP[Lys-Lys]2 at concentrations as low as 10-50 µM exerted significant effects on microsporidia, reducing the number of spores by 67-80% compared to the control. Live-cell imaging of the spores treated with porphyrins showed that only 1.6% and 3.0% of spores remained alive after 24 h-incubation with 50 µM PP[Asp(ONa)2]2 and PP[Lys-Asp]2, respectively. The length of the amino acid side chains and their identity in the PPIX molecules affected the bioactivity of the porphyrin. Importantly, the irradiation of the porphyrins did not enhance their potency in destroying Nosema spores. We showed that the porphyrins accumulated inside the living spores but not inside dead spores, thus the destruction of the microsporidia by non-metallated porphyrins is not dependent on photosensitization, but is associated with their active transport into the spore cell. When administered to honeybees in vivo, PPIX[Lys-TFA]2 and PPIX[Lys-Lys]2 reduced spore loads by 69-76% in infected individuals. They both had no toxic effect on honeybees, in contrast to zinc-coordinated porphyrin.
Insights
New porphyrins effectively treat honeybee nosemosis by directly inactivating microsporidia. These compounds, when administered in vivo, significantly reduce spore loads without harming bees, offering a promising solution for this dangerous honeybee infection.
Area of Science:
- Apiculture
- Veterinary Entomology
- Biochemistry
Background:
- Honeybee populations face significant threats from microsporidian infections like nosemosis.
- Current treatments for nosemosis are limited, necessitating the development of novel therapeutic strategies.
Purpose of the Study:
- To investigate the in vitro efficacy of synthesized porphyrins against microsporidia responsible for honeybee nosemosis.
- To compare the activity of novel amide derivatives of protoporphyrin IX (PPIX) with existing cationic porphyrins.
- To evaluate the in vivo therapeutic potential and safety of selected porphyrins in infected honeybees.
Main Methods:
- Synthesis of amide derivatives of protoporphyrin IX (PPIX) conjugated with amino acids.
- In vitro inactivation assays of microsporidia using various porphyrins at different concentrations.
- Live-cell imaging to assess spore viability after porphyrin treatment.
- In vivo administration of porphyrins to infected honeybees to determine spore load reduction and toxicity.
Main Results:
- Several porphyrins, including PP[Lys-Asp]2 and PP[Lys-Lys]2, significantly reduced microsporidia spore counts by 67-80% in vitro at 10-50 µM concentrations.
- Live-cell imaging confirmed high efficacy, with only 1.6-3.0% of spores remaining viable after 24h incubation with specific porphyrins.
- In vivo studies showed that PPIX[Lys-TFA]2 and PPIX[Lys-Lys]2 reduced honeybee spore loads by 69-76% without observable toxic effects.
- Porphyrin activity was linked to active transport into living spores, not photosensitization.
Conclusions:
- Synthesized porphyrins, particularly PPIX derivatives, demonstrate potent direct inactivation of microsporidia, offering a novel treatment for honeybee nosemosis.
- The efficacy of porphyrins is influenced by amino acid side chain modifications and is independent of photosensitization.
- Selected porphyrins show excellent in vivo therapeutic potential and safety for honeybees, addressing a critical need in apiculture.
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