Bacillus megaterium'un Fusarium Türlerine Karşı Antifungal Etkinliğinin Araştırılması

Yıl 2023, Cilt: 33 Sayı: 2, 183 - 191, 30.06.2023

Esra Nur Keleş Keyfoğlu , Ayşe Feyza Tufan Dülger , Emre Yörük

https://doi.org/10.29133/yyutbd.1237451

Öz

Birçok Fusarium türü, dünya çapında küçük taneli tahıllarda ciddi patojenler olarak ortaya çıkmaktadır. Fungisitlerin kullanımı Fusarium hastalıklarıyla mücadelede kısa vadeli bir stratejidir. Biyokontrol ajanlarının kullanımı, ekonomik olmasının yanı sıra çevreye verilen kimyasal girdiyi azaltması nedeniyle cazip bir alternatif stratejidir. Bacillus türleri, biyokontrol ajanları olarak dikkat çekmektedir. Bu çalışmada, Bacillus megaterium CTBmeg1 ve HMA5 suşlarının Fusarium culmorum UK99 ve F. graminearum PH-1 izolatları üzerindeki antagonistik aktiviteleri in vitro ve moleküler düzeyde incelenmiştir. İkili kültür testinin 7. gününde, B. megaterium suşlarının her ikisi de, %72,7 ile %77,7 arasındaki inhibisyon oranıyla çok yüksek antifungal aktivite ile Fusarium izolatlarının misel büyümesini önemli ölçüde azaltmıştır. Benzer şekilde, her iki suş da uçucu organik bileşik (VOC) analizinde %52,1 ile %62,4 arasında yüksek antifungal aktiviteye neden olmuştur. İturin, surfaktin ve fengycin genleri, bakterinin antagonist aktiviteye sahip metabolitler üretme yeteneğini belirlemek için PCR ile analiz edildi. Bu genlerin bakteri suşlarında bulunduğu belirlendi. Moleküler düzeyde, test edilen tüm gruplarda, trikotesen üretimi ile ilişkili tri5 geninin transkript seviyeleri azalırken, bir antioksidan gen olan cat ve apoptoz ile ilişkili bir gen olan mst20'nin transkript seviyeleri artmıştır. Bu çalışmadan elde edilen bulgular, B. megaterium CTBmeg1 ve HMA5 suşlarının dünya çapındaki fitopatojenler F. culmorum ve F. graminearum'a karşı oldukça etkili biyokontrol ajanları olarak kabul edilebileceğini göstermektedir.

Anahtar Kelimeler

Bacillus megaterium, biocontrol, Fusarium culmorum, Fusarium graminearum, gene expression

Investigation of the Antifungal Activity of Bacillus megaterium Against Fusarium Species

Öz

Several Fusarium species are emerging as serious pathogens on small grain cereals worldwide. The use of fungicides is a short-term strategy in the fight against Fusarium diseases. The use of biocontrol agents is an attractive alternative strategy by reducing the chemical input to the environment as well as being economical. Bacillus species have received attention as biocontrol agents. In this study, the antagonistic activities of Bacillus megaterium CTBmeg1 and HMA5 strains on Fusarium culmorum UK99 and F. graminearum PH-1 isolates were investigated in vitro and at molecular level. On the 7th day of the dual culture assay, both of B. megaterium strains significantly reduced the mycelial growth of Fusarium isolates, with very high antifungal activity with the inhibition rate between 72.7% and 77.7%, respectively. Similarly, both strains caused high antifungal activity in the volatile organic compound (VOC) analysis between 52.1% and 62.4%, respectively. At the molecular level, in all tested groups, transcript levels of the tri5 gene, which is associated with trichothecene production, decreased, while the transcript levels of cat, an antioxidant gene, and mst20, a gene related to apoptosis, increased. Findings from this study showed that B. megaterium CTBmeg1 and HMA5 strains could be accepted as highly effective biocontrol agents against worldwide phytopathogens F. culmorum and F. graminearum.

Anahtar Kelimeler

Bacillus megaterium, biocontrol, Fusarium culmorum, Fusarium graminearum, gene expression

Kaynakça

• Akcay, K., & Kaya, Y. (2019). Isolation, characterization and molecular identification of a halotolerant Bacillus megaterium CTBmeg1 able to grow on halogenated compounds. Biotechnology & Biotechnological Equipment, 33(1), 945–953. https://doi.org/10.1080/13102818.2019.1631717
• Aksoy H. M., Saruhan I., Kaya, Y., & Ozturk, M. (2018). Morphological changes caused by Bacillus megaterium on adult emergence of fall webworm’s pupa, Hyphantria cunea (Drury)(Lepidoptera: Erebidae). Journal of Agricultural Sciences, 24(4), 539–546. https://doi.org/10.15832/ankutbd.349474
• Bernardo, A., Bai, G., Guo, P., Xiao, K., Guenzi, A. C., & Ayoubi, P. (2007). Fusarium graminearum-induced changes in gene expression between Fusarium head blight-resistant and susceptible wheat cultivars. Functional & Integrative Genomics, 7, 69-77. https://doi.org/10.1007/s10142-006-0028-1
• Bivi, M. R., Farhana, M. S. N., Khairulmazmi, A., & Idris, A. (2010). Control of Ganoderma boninense: A causal agent of basal stem rot disease in oil palm with endophyte bacteria in vitro. International Journal of Agriculture and Biology, 12(6), 833–839.
• Bolivar-Anillo, H. J., González-Rodríguez, V. E., Cantoral, J. M., García-Sánchez, D., Collado, I. G., & Garrido, C. (2021). Endophytic bacteria Bacillus subtilis, isolated from Zea mays, as potential biocontrol agent against Botrytis cinerea. Biology, 10(6), 492. https://doi.org/10.3390/biology10060492
• Cantoro, R., Palazzini, J. M., Yerkovich, N., Miralles, D. J., & Chulze, S. N. (2021). Bacillus velezensis RC 218 as a biocontrol agent against Fusarium graminearum: effect on penetration, growth and TRI5 expression in wheat spikes. BioControl, 66(2), 259–270. https://doi.org/10.1007/s10526-020-10062-7
• El-Gremi, S. M., Draz, I.S., & Youssef, W. A. E. (2017). Biological control of pathogens associated with kernel black point disease of wheat. Crop Protection, 91, 13–19. https://doi.org/10.1016/j.cropro.2016.08.034
• Gao, Z., Zhang, B., Liu, H., Han, J., & Zhang, Y. (2017). Identification of endophytic Bacillus velezensis ZSY-1 strain and antifungal activity of its volatile compounds against Alternaria solani and Botrytis cinerea. Biological Control, 105, 27–39. https://doi.org/10.1016/j.biocontrol.2016.11.007
• Gazdağlı, A., Sefer, Ö., Yörük, E., Varol, G. İ., Teker, T., & Albayrak, G. (2018). Investigation of camphor effects on Fusarium graminearum and F. culmorum at different molecular levels. Pathogens, 7(4), 90. https://doi.org/10.3390/pathogens7040090
• Grosu, A. I., Sicuia, O. A., Dobre, A., Voaideş, C., & Cornea, C. P. (2015). Evaluation of some Bacillus spp. strains for the biocontrol of Fusarium graminearum and F. culmorum in wheat. Agriculture and Agricultural Science Procedia, 6, 559–566. https://doi.org/10.1016/j.aaspro.2015.08.085
• Jones, R. K. (2000). Assessments of Fusarium head blight of wheat and barley in response to fungicide treatment. Plant Disease, 84(9), 1021-1030. https://doi.org/10.1094/PDIS.2000.84.9.1021
• Khan, N., Maymon, M., & Hirsch, A. (2017). Combating Fusarium infection using Bacillus-based antimicrobials. Microorganisms, 5(4), 75. https://doi.org/10.3390/microorganisms5040075
• Legrand, F., Picot, A., Cobo-Díaz, J. F., Chen, W., & Le Floch, G. (2017). Challenges facing the biological control strategies for the management of Fusarium head blight of cereals caused by F. graminearum. Biological Control. 113, 26–38. https://doi.org/10.1016/j.biocontrol.2017.06.011
• Li, X., Wang, X., Shi, X., Wang, B., Li, M., Wang, Q., & Zhang, S. (2020). Antifungal effect of volatile organic compounds from Bacillus velezensis CT32 against Verticillium dahliae and Fusarium oxysporum. Processes, 8(12), 1674. https://doi.org/10.3390/pr8121674
• Livak, K.J., & Schmittgen, T.D. (2001). Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods, 25(4), 402–408. https://doi.org/10.1006/meth.2001.1262
• Matny, O. N. (2015). Fusarium head blight and crown rot on wheat & barley: losses and health risks. Advances in Plants & Agriculture Research, 2(39), 10-15406. Ol
• Moya-Elizondo, E. A., & Jacobsen, B.J. (2016). Integrated management of Fusarium crown rot of wheat using fungicide seed treatment, cultivar resistance, and induction of systemic acquired resistance (SAR). Biological Control, 92: 153–163. https://doi.org/10.1016/j.biocontrol.2015.10.006
• Nalam, V.J., Sarowar, S., & Shah, J. (2016). Establishment of a Fusarium graminearum infection model in Arabidopsis thaliana leaves and floral tissues. Bio-protocol 6(14), e1877. https://doi.org/10.21769/BioProtoc.1877
• Nayak, S. K., Nayak, S., & Mishra, B. B. (2017). Antimycotic role of soil Bacillus sp. against rice pathogens: a biocontrol prospective. Microbial Biotechnology: Volume 1. Applications in Agriculture and Environment, 29-60. https://doi.org/10.1007/978-981-10-6847-8_2
• Özsoy, E., Kesercan, B., & Yörük, E. (2020). Antifungal activity of specific plant essential oils against Fusarium graminearum. Journal of Plant Science and Phytopathology, 4, 060-062. https://doi.org/10.29328/journal.jpsp.1001052
• Pan, D., Mionetto, A., Tiscornia, S., & Bettucci, L. (2015). Endophytic bacteria from wheat grain as biocontrol agents of Fusarium graminearum and deoxynivalenol production in wheat. Mycotoxin Research, 31(3), 137–143. https://doi.org/10.1007/s12550-015-0224-8
• Pasquali M., & Migheli, Q. (2014). Genetic approaches to chemotype determination in type B-trichothecene producing Fusaria. International Journal of Food Microbiology, 189, 164–182. https://doi.org/10.1016/j.ijfoodmicro.2014.08.011
• Raza, W., Wang, J., Wu, Y., Ling, N., Wei, Z., Huang, Q., & Shen, Q. (2016). Effects of volatile organic compounds produced by Bacillus amyloliquefaciens on the growth and virulence traits of tomato bacterial wilt pathogen Ralstonia solanacearum. Applied Microbiology and Biotechnology, 100, 7639–7650. https://doi.org/10.1007/s00253-016-7584-7
• Tahir, H. A. S., Gu, Q., Wu, H., Raza, W., Hanif, A., Wu, L., Colman, M. V., & Gao, X. (2017). Plant growth promotion by volatile organic compounds produced by Bacillus subtilis SYST2. Frontiers in Microbiology, 8, 171. https://doi.org/10.3389/fmicb.2017.00171
• Teker, T., Sefer, Ö., Gazdağlı, A., Yörük, E., Varol, G. İ., & Albayrak, G. (2021). α-Thujone exhibits an antifungal activity against F. graminearum by inducing oxidative stress, apoptosis, epigenetics alterations and reduced toxin synthesis. European Journal of Plant Pathology, 160(3), 611–622. https://doi.org/10.1007/s10658-021-02269-w
• Toh, S. C., Samuel, L., & Awang, A. S. A. H. (2016). Screening for antifungal-producing bacteria from Piper nigrum plant against Phytophthora capsici. International Food Research Journal, 23(6), 2616-2622.
• Tufan, F., Uçarlı, C., Tunalı, B., & Gürel, F. (2017). Analysis of early events in barley (Hordeum vulgare L.) roots in response to Fusarium culmorum infection. European Journal of Plant Pathology, 148, 343–355. https://doi.org/10.1007/s10658-016-1087-3
• Uluhan, E., Keleş, E. N., & Tufan, F. (2019). Analysis of WRKY transcription factors in barley cultivars infected with Fusarium culmorum. International Journal of Life Sciences and Biotechnology, 2, 165–174. https://doi.org/10.38001/ijlsb.588730
• Wu, Y., Yuan, J., Raza, W., Shen, Q., & Huang, Q. (2014). Biocontrol traits and antagonistic potential of Bacillus amyloliquefaciens strain NJZJSB3 against Sclerotinia sclerotiorum, a causal agent of canola stem rot. Journal of Microbiology and Biotechnology, 24(10), 1327–1336.
• Wu, Y., Zhou, J., Li, C., & Ma, Y. (2019). Antifungal and plant growth promotion activity of volatile organic compounds produced by Bacillus amyloliquefaciens. MicrobiologyOpen, 8(8), e813. https://doi.org/10.1002/mbo3.813
• Yörük, E. (2018). Tetraconazole leads to alterations in Fusarium graminearum at different molecular levels. Applied Ecology and Environmental Research, 6, 6155–6167. http://dx.doi.org/10.15666/aeer/1605_61556167
• Zalila-Kolsi, I., Mahmoud, A. B., Ali, H., Sellami, S., Nasfi, Z., Tounsi, S., & Jamoussi, K. (2016) Antagonist effects of Bacillus spp. strains against Fusarium graminearum for protection of durum wheat (Triticum turgidum L. subsp. durum). Microbiology Research, 192, 148–158. https://doi.org/10.1016/j.micres.2016.06.012
• Zhao, Y., Selvaraj, J. N., Xing, F. et al. (2014) Antagonistic action of Bacillus subtilis strain SG6 on Fusarium graminearum. PLoS ONE, 9(3), e92486. https://doi.org/10.1371/journal.pone.0092486
• Zhou, Y., Chen, J., Zhu, X., Wang, Y., Liu, X., Fan, H,, Duan, Y., & Chen, L. (2020). Efficacy of Bacillus megaterium strain Sneb207 against soybean cyst nematode (Heterodera glycines) in soybean. Pest Management Science, 77(1), 568–576. https://doi.org/10.1002/ps.6057