Bal arılarının kutikulasından izole edilen Bifidobacterium sp. ERSapi20 suşunun Varroa destructor Oudemans, 1904 (Acari: Varroidae)’a karşı antiparazitik potansiyelinin değerlendirilmesi

Öz

Varroa destructor Oudemans, 1904 (Acari: Varroidae), bal arısı kolonilerinin kaybına ciddi katkıda bulunduğu bilinmektedir. Aktinobakteri türleri ise geniş bir dağılıma sahiptir ve bakteriler ve parazitler gibi çeşitli patojenlere karşı biyolojik olarak aktif bileşikler üretirler. Bu temele dayanarak, bu çalışma Apis mellifera L., 1758 (Hymenoptera: Apidae) ile ilişkili aktinobakteri simbiyontları tarafından üretilen biyolojik olarak aktif bileşikleri araştırmayı ve bunların V. destructor'a karşı akarisit potansiyelini değerlendirmeyi amaçlamaktadır. Kültür bazlı bir yöntem kullanılarak işçi arılardan Bifidobacterium sp. ERSapi20 suşu Antalya, Türkiye lokasyonundan 2022 yılında izole edilmiş ve tanımlanmıştır. Antiparazitik aktiviteyi belirlemek için, suşun ekstraselüler ve intraselüler ekstraktı, böcek Ringer çözeltisinin değişen konsantrasyonlarında (%0, %25, %50, %75 ve %100) püskürtme yöntemi kullanılarak V. destructor'a uygulanmıştır. Ekstraselüler çözeltinin 16 saat içinde %90 oranında akar ölümüne yol açtığı, intraselüler ekstraktın ise 18 saat içinde %100 oranında akar ölümüne neden olduğu belirlenmiştir. Bu, aktinobakteriyel suşların intraselüler ekstraktı kullanılarak V. destructor'a karşı akarisit aktivitesinin rapor edildiği ilk çalışmadır. Bu araştırma, bal arılarının varroa akarlarına karşı etkili bileşikler üreten aktinobakterilerle ilişki kurduğu hipotezini desteklemekte ve V. destructor'u kolonilerde kontrol etmek için doğal ürünlerin kullanılmasına dair bilgiler sunmaktadır.

Anahtar Kelimeler

• Aktinobakteri
• antiparazitik aktivite
• Apis mellifera
• sekonder metabolit
• Varroa destructor

Assessing the antiparasitic potential of Bifidobacterium sp. ERSapi20 isolated from the cuticle of honeybees against Varroa destructor Oudemans, 1904 (Acari: Varroidae)

Abstract

The mite Varroa destructor Oudemans, 1904 (Acari: Varroidae) is a major contributor to honeybee colony losses worldwide. Actinobacteria are known for their wide distribution and production of biologically active compounds effective against various pathogens, including bacteria and parasites. This study aims to investigate the bioactive compounds produced by actinobacterial symbionts associated with Apis mellifera L., 1758 (Hymenoptera: Apidae) and evaluate their acaricidal potential against V. destructor. In 2022, a strain of Bifidobacterium sp. ERSapi20 was isolated from worker bees in Antalya, Türkiye, using a culture-based method. The antiparasitic activity of the extracellular solution and intracellular extract of the strain was assessed through direct spraying on V. destructor at varying concentrations of insect Ringer solution (0%, 25%, 50%, 75%, and 100%). The extracellular solution achieved 90% mite mortality within 16 hours, while the intracellular extract resulted in 100% mite mortality within 18 hours. This is the first report demonstrating acaricidal activity against V. destructor using intracellular extracts of actinobacterial strains. These findings support the hypothesis that honeybee-associated actinobacteria produce compounds effective against varroa mites, highlighting the potential of natural products for controlling V. destructor in honeybee colonies.

Keywords

• Actinobacteria
• antiparasitic activity
• Apis mellifera
• secondary metabolite
• Varroa destructor

References

1. Aizenberg-Gershtein, Y., I. Izhaki & M. Halpern, 2013. Do honeybees shape the bacterial community composition in floral nectar? PloS One, 8 (7): e67556.
2. Alberoni, D., F. Gaggìa, L. Baffoni & D. Di Gioia, 2016. Beneficial microorganisms for honeybees: problems and progresses. Applied Microbiology and Biotechnology, 100 (22): 9469-9482.
3. Anderson, K. E., T. H. Sheehan, B. M. Mott, P. Maes, L. Snyder & M. R. Schwan, 2013. Microbial ecology of the hive and pollination landscape: bacterial associates from floral nectar, the alimentary tract and stored food of honeybees (Apis mellifera). PloS One, 8 (12): e83125.
4. Asama, T., T. H. Arima, T. Gomi, T. Keishi, H. Tani & Y. Kimura, 2015. Lactobacillus kunkeei YB 38 from honeybee products enhances IgA production in healthy adults. Journal of Applied Microbiology, 119 (3): 818-826.
5. Barka, E. A., P. Vatsa, L. Sanchez, N. Gaveau-Vaillant, C. Jacquard & H. P. Klenk, 2016. Taxonomy, physiology, and natural products of Actinobacteria. Microbiology and Molecular Biology Reviews, 80 (1): 1-43.
6. Běhal, V., 2000. Bioactive products from Streptomyces. Advances in Applied Microbiology, 47: 113-156.
7. Beveridge, T. J., 2001. Use of the gram stain in microbiology. Biotechnic & Histochemistry, 76 (3): 111-118.
8. Chandler, D., G. Davidson, J. K. Pell, B. V. Ball, K. Shaw & K. D. Sunderland, 2000. Fungal biocontrol of Acari. Biocontrol Science and Technology, 10 (4): 357-384.

9. Chandler, D., K. D. Sunderland, B. V. Ball & G. Davidson, 2001. Prospective biological control agents of Varroa destructor n. sp., an important pest of the European honeybee, Apis mellifera. Biocontrol Science and Technology, 11 (4): 429-448.
10. Chaudhary, H. S., B. Soni, A. R. Shrivastava & S. Shrivastava, 2013. Diversity and versatility of actinomycetes and its role in antibiotic production. Journal of Applied Pharmaceutical Science, 3 (8): 83-94.
11. Crotti, E., L. Sansonno, E. M. Prosdocimi, V. Vacchini, C. Hamdi & A. Cherif, 2013. Microbial symbionts of honeybees: a promising tool to improve honeybee health. New Biotechnology, 30 (6): 716-722.
12. Daffe, M., P. J. Brennan & M. McNeil, 1990. Predominant structural features of the cell wall arabinogalactan of Mycobacterium tuberculosis as revealed through characterization of oligoglycosyl alditol fragments by gas chromatography/mass spectrometry and by 1H and 13C NMR analyses. Journal of Biological Chemistry, 265 (12): 6734-6743.
13. De Grandi-Hoffman, G., V. Corby-Harris, E. W. De Jong, M. Chambers & G. Hidalgo, 2017. Honeybee gut microbial communities are robust to the fungicide Pristine® consumed in pollen. Apidologie, 48 (3): 340-352.
14. De Guzman, L. I., T. E. Rinderer & L. D. Beaman, 1993. Survival of Varroa jacobsoni Oud. (Acari: Varroidae) away from its living host Apis mellifera L. Experimental & Applied Acarology, 17 (4): 283-290.
15. De Piano, F. G., M. D. Maggi, F. R. Meroi Arcerito, M. C. Audicio, M. J. Eguaras & S. R. Ruffinengo, 2020. Effects of bacterial cell-free supernatant on nutritional parameters of Apis mellifera and their toxicity against Varroa destructor. Research Inst Pomology Floriculture; Journal of Apicultural Science, 64 (1): 55-66.
16. Duquesne, S., D. Destoumieux-Garzón, J. Peduzzi & S. Rebuffat, 2007. Microcins, gene-encoded antibacterial peptides from enterobacteria. Natural Product Reports, 24 (4): 708-734.
17. Engelberg-Kulka, H. S. Amitai, I. Kolodkin-Gal & R. Hazan, 2006. Bacterial programmed cell death and multicellular behavior in bacteria. PLoS Genetics, 2 (10): e135.
18. Evans, J. D., 2003. Diverse origins of tetracycline resistance in the honeybee bacterial pathogen Paenibacillus larvae. Journal of Invertebrate Pathology 83 (1): 46-50.
19. Ganeshprasad, D. N., J. K. Lone, K. Jani, Y. S. Shouche, K. A. Khan, S. Sayed, M. Shukry, S.A. Dar, M. Mushtaq & A.H. Sneharani, 2022. Gut Bacterial Flora of Open Nested Honeybee, Apis florea. Frontiers in Ecology and Evolution, 10 (3): 837381.
20. Gliński, Z. F. & J. Jarosz, 1990. Serratia marcescens, artificially contaminating brood and worker honeybees, contaminates the Varroa jacobsoni mite. Journal of Apicultural Research, 29 (2): 107-111.
21. Goulson, D., E. Nicholls, C. Botías & E. L. Rotheray, 2015. Bee declines driven by combined stress from parasites, pesticides, and lack of flowers. Science, 347 (6229): 1255957.
22. Hamiduzzaman, M. M., A. Sinia, E. Guzman-Novoa & P. H. Goodwin, 2012. Entomopathogenic fungi as potential biocontrol agents of the ecto-parasitic mite, Varroa destructor, and their effect on the immune response of honeybees (Apis mellifera L.). Journal of Invertebrate Pathology, 111 (3): 237-243.
23. Higes, M., R. Martín-Hernández, C. S. Hernández-Rodríguez & J. González-Cabrera, 2020. Assessing the resistance to acaricides in Varroa destructor from several Spanish locations. Parasitology Research, 119 (11): 3595-3601.
24. Hong, Y., H. S. Yang, J. Li, S. K. Han, H. C. Chang & H. Y. Kim, 2014. Identification of lactic acid bacteria in salted Chinese cabbage by SDS‐PAGE and PCR‐DGGE. Journal of the Science of Food and Agriculture, 94 (2): 296-300.
25. Hussain, R., S. Farooq, M. Kalsoom & H. U. Rehman, 2018. Prevalence of Varroa destructor on honeybees hives in district Karak, Khyber Pakhtunkhwa, Pakistan. Journal of Entomology and Zoology Studies, 6 (1): 169-171.
26. Kanbe, K., Y. Mimura, T. Tamamura, S. Yatagai, Y. Sato & A. Takahashi, 1992. AB3217-A, a novel anti-mite substance produced by a strain of Streptomyces platensis. The Journal of Antibiotics, 45 (4): 458-464.
27. Khasabuli, O. Y. & A. N. Kibera, 2014. Isolation, characterization and primary screening of soil actinomycetes from Kenyatta University arboretum grounds for antibacterial activities. Journal of Applied Biosciences, 74: 6072-6079.
28. Kwong, W. K. & N. A. Moran, 2016. Gut microbial communities of social bees. Nature Reviews Microbiology, 14 (6): 374-384.
29. Lodesani, M. & C. Costa, 2005. Limits of chemotherapy in beekeeping: development of resistance and the problem of residues. Bee World, 86 (4): 102-109.
30. Maggi, M. D., S. R. Ruffinengo, P. Negri & M. J. Eguaras, 2010. Resistance phenomena to amitraz from populations of the ectoparasitic mite Varroa destructor of Argentina. Parasitology Research, 107 (5): 1189-1192.
31. Manivasagan, P., J. Venkatesan, K. Sivakumar & S. K. Kim, 2014. Pharmaceutically active secondary metabolites of marine actinobacteria. Microbiological Research, 169 (4): 262-278.
32. Martin, S. J., 2004. Acaricide (pyrethroid) resistance in Varroa destructor. Bee World, 85 (4): 67-69.
33. Martinson, V. G., B. N. Danforth, R. L. Minckley, O. Rueppell, S. Tingek & N. A. Moran, 2011. A simple and distinctive microbiota associated with honeybees and bumble bees. Molecular Ecology, 20 (3): 619-628.
34. Meixner, M. D., 2010. A historical review of managed honeybee populations in Europe and the United States and the factors that may affect them. Journal of Invertebrate Pathology, 103 (Supplement): S80-S95.
35. Monod, J., 1949. The growth of bacterial cultures. Annual review of microbiology 3 (1): 371-394.
36. Muñoz-Colmenero, M., I. Baroja-Careaga, M. Kovačić, J. Filipi, Z. Puškadija & N. Kezić, 2020. Differences in honeybee bacterial diversity and composition in agricultural and pristine environments-a field study. Apidologie 51 (6): 1018-1037.
37. Peng, C. Y., X. Zhou & H. K. Kaya, 2002. Virulence and site of infection of the fungus, Hirsutella thompsonii, to the honeybee ectoparasitic mite, Varroa destructor. Journal of Invertebrate Pathology, 81 (3): 185-195.
38. Pettis, J. S., 2004. A scientific note on Varroa destructor resistance to coumaphos in the United States. Apidologie, 35 (1): 91-92.
39. Reyes-Quintana, M., L. G. Espinosa-Montaño, D. Prieto-Merlos, G. Koleoglu, T. Petukhova, A. Correa-Benítez & E. Guzman-Novoa, 2019. Impact of Varroa destructor and deformed wing virus on emergence, cellular immunity, wing integrity and survivorship of Africanized honeybees in Mexico. Journal of Invertebrate Pathology, 164 (2019): 43-48.
40. Rosenkranz, P., P. Aumeier & B. Ziegelmann, 2010. Biology and control of Varroa destructor. Journal of Invertebrate Pathology, 103 (1): 96-119.
41. Roshan, K., A. Tarafdar, K. Saurav, S. Ali, S. A. Lone & S. Pattnaik, 2013. Isolation and screening of bioactive compound from actinomycetes isolated from salt pan of Marakanam district of the state Tamil Nadu, India. Elixir International Journal, 61: 16826-16831.
42. Roy, D., 2001. Media for the isolation and enumeration of bifidobacteria in dairy products. International Journal of Food Microbiology, 69 (3): 167-182.
43. Saccà, M. L. & M. Lodesani, 2020. Isolation of bacterial microbiota associated to honeybees and evaluation of potential biocontrol agents of Varroa destructor. Beneficial Microbes, 11 (7): 641-654.
44. Saitou, N. & M. Nei, 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution, 4 (4): 406-425.
45. Shaw, K. E., G. Davidson, S. J. Clark, B. V. Ball, J. K. Pell, D. Chandler & K. D. Sunderland, 2002. Laboratory bioassays to assess the pathogenicity of mitosporic fungi to Varroa destructor (Acari: Mesostigmata), an ectoparasitic mite of the honeybee, Apis mellifera. Biological Control, 24 (3): 266-276.
46. Shen, M., L. Cui, N. Ostiguy & D. Cox-Foster, 2005. Intricate transmission routes and interactions between picorna-like viruses (Kashmir bee virus and sacbrood virus) with the honeybee host and the parasitic varroa mite. Journal of General Virology, 86 (8): 2281-2289
47. Shirling, E. T. & D. Gottlieb, 1966. Methods for characterization of Streptomyces species. International Journal of Systematic Bacteriology, 16 (3): 313-340.
48. Simon-Delso, N., G. San Martin, E. Bruneau, L. A. Minsart, C. Mouret & L. Hautier, 2014. Honeybee colony disorder in crop areas: the role of pesticides and viruses. PloS One, 9 (7): e103073.
49. Stecher, G., K. Tamura & S. Kumar, 2020. Molecular evolutionary genetics analysis (MEGA) for macOS. Molecular Biology and Evolution, 37 (4): 1237-1239.
50. Tamura, K., G. Stecher & S. Kumar, 2021. MEGA11: molecular evolutionary genetics analysis version 11. Molecular Biology and Evolution, 38 (7): 3022-3027.
51. Tejerina, M. R., M. R. Benítez-Ahrendts & M.C. Audisio, 2020. Lactobacillus salivarius A3iob Reduces the Incidence of Varroa destructor and Nosema Spp. in Commercial Apiaries Located in the Northwest of Argentina. Probiotics & Antimicro. Prot., 12: 1360-1369.
52. Tsagou, V., A. Lianou, D. Lazarakis, N. Emmanouel & G. Aggelis, 2004. Newly isolated bacterial strains belonging to Bacillaceae (Bacillus sp.) and Micrococcaceae accelerate death of the honeybee mite, Varroa destructor (V. jacobsoni), in laboratory assays. Biotechnology Letters, 26 (6): 529-532.
53. Tutun, H., N. Koç & A. Kart, 2018. Plant essential oils used against some bee diseases. Turkish Journal of Agriculture-Food Science and Technology, 6 (1): 34-45.
54. ul Hassan, S. S. & A. L. Shaikh, 2017. Marine actinobacteria as a drug treasure house. Biomedicine & Pharmacotherapy, 87 (2017): 46-57.
55. Vilarem, C., V. Piou, F. Vogelweith & A. Vétillard, 2021. Varroa destructor from the Laboratory to the Field: Control, Biocontrol and IPM Perspectives - A Review. Insects, 12 (9): 800 (1-26).
56. Weerahandi, S. & C.-R. Yu, 2020. Exact distributions of statistics for making inferences on mixed models under the default covariance structure. Journal of Statistical Distributions and Applications, 7: 4 (1-14).
57. Zemene, M., B. Bogale, S. Derso, S. Belete, S. Melaku & H. Hailu, 2015. A review on varroa mites of honeybees. Academic Journal of Entomology, 8 (3): 150-159.
58. Zimmermann, W., 1990. Degradation of lignin by bacteria. Journal of Biotechnology, 13 (2-3): 119-130.
59. Zwietering, M. H., I. Jongenburger, F. M. Rombouts & K. J. A. E. M. Van't Riet, 1990. Modeling of the bacterial growth curve. Applied and Environmental Microbiology, 56 (6): 1875-1881.