Biological control of citrus canker by endophytic bacteria

Authors

DOI:

https://doi.org/10.14719/pst.5633

Keywords:

Bis(2-ethylhexyl) phthalate, Biocontrol, Bacillus amyloliquefaciens, Bacillus subtilis, metabolic pathways, siderophore

Abstract

Citrus is an important fruit crop in India, playing a significant role in the agricultural economy and experiencing high demand due to its rich nutritional content. Despite its economic importance, citrus canker, caused by the Gram-negative bacterium Xanthomonas citri subsp. citri, poses a significant threat to citrus production globally, including in India. This study focuses on using endophytic bacteria, specifically Bacillus amyloliquefaciens ESK-8 and Bacillus subtilis EPM-3, for the biological management of this pathogen. A roving survey in major citrus cultivating areas of Tamil Nadu revealed varying disease prevalence rates (21% - 65%). The citrus canker pathogen and endophytic Bacillus isolates were isolated from the surveyed locations and identified based on biochemical and molecular analyses using 16S rDNA sequences. In vitro assays through agar well diffusion method using culture filtrates of various Bacillus isolates revealed significant inhibition rates, with Bacillus amyloliquefaciens ESK-8 and Bacillus subtilis EPM-3 exhibiting inhibition diameters of 14.1 mm and 11.6 mm, respectively, compared to the control. GC-MS analysis of Bacillus amyloliquefaciens ESK-8 and Bacillus subtilis EPM-3 unraveled important antibacterial compounds such as bis(2-ethylhexyl) phthalate, n-hexadecanoic acid, D-erythro-pentose, 2-deoxy, hexadecanoic acid, octadecanoic acid, and tridecane. Furthermore, the metabolic pathways related to these compounds include glycerolipid metabolism, glutamate metabolism, and tryptophan metabolism, all of which play significant roles in plant growth and antagonism-promoting activities. Additionally, pot culture studies confirmed the antagonistic potential of Bacillus amyloliquefaciens ESK-8 and Bacillus subtilis EPM-3 against the citrus canker pathogen. This research highlights the potential of endophytic Bacillus isolates for the sustainable management of citrus canker disease.

Downloads

Download data is not yet available.

References

Sanofer AA. Role of citrus fruits in health. Journal of Pharmaceutical Sciences and Research. 2014 Feb 1;6(2):121.

Anonymous. Horticulture Statistics at a Glance New Delhi: NHM, Government of India. 2015; p. 237.

Das AK. Citrus canker-A review. Journal of Applied Horticulture. 2003 Jan;5(1):52-60. https://doi.org/10.37855/jah.2003.v05i01.15

Ali Y, Inusa I, Sanghvi G, Mandaliya VB, Bishoyi AK. The current status of phage therapy and its advancement towards establishing standard antimicrobials for combating multi drug-resistant bacterial pathogens. Microbial Pathogenesis. 2023 Aug 1;181:106199. https://doi.org/10.1016/j.micpath.2023.106199

Gouda S, Kerry RG, Das G, Paramithiotis S, Shin HS, Patra JK. Revitalization of plant growth promoting rhizobacteria for sustainable development in agriculture. Microbiological Research. 2018 Jan 1;206:131-40. https://doi.org/10.1016/j.micres.2017.08.016

Kumar P, Pahal V, Gupta A, Vadhan R, Chandra H, Dubey RC. Effect of silver nanoparticles and Bacillus cereus LPR2 on the growth of Zea mays. Scientific Reports. 2020 Nov 23;10(1):20409. https://doi.org/10.1038/s41598-020-77460-w

Hosni K, Zahed N, Chrif R, Abid I, Medfei W, Kallel M, et al. Composition of peel essential oils from four selected Tunisian Citrus species: Evidence for the genotypic influence. Food Chemistry. 2010 Dec 15;123(4):1098-104. http://dx.doi.org/10.1016/j.foodchem.2010.05.068

Wu L, Wu HJ, Qiao J, Gao X, Borriss R. Novel routes for improving biocontrol activity of Bacillus based bioinoculants. Frontiers in Microbiology. 2015 Dec 10;6:1395. https://doi.org/10.3389/fmicb.2015.01395

An SQ, Potnis N, Dow M, Vorhölter FJ, He YQ, Becker A, et al. Mechanistic insights into host adaptation, virulence and epidemiology of the phytopathogen Xanthomonas. FEMS Microbiology Reviews. 2020 Jan;44(1):1-32. https://doi.org/10.1093/femsre/fuz024

Gottwald TR, Graham JH, Schubert TS. Citrus canker: the pathogen and its impact. Plant Health Progress. 2002;3(1):15. https://doi.org/10.1094/PHP-2002-0812-01-RV

McKinney HH. A new system of grading plant diseases. J Agric Res.1923;26(2):195-218.

Graham JH, Gottwald TR, Cubero J, Achor DS. Xanthomonas axonopodis pv. citri: factors affecting successful eradication of citrus canker. Molecular Plant Pathology. 2004 Jan;5(1):1-5. https://doi.org/10.1046/j.1364-3703.2004.00197.x

Francis MI, Peña A, Graham JH. Detached leaf inoculation of germplasm for rapid screening of resistance to citrus canker and citrus bacterial spot. European Journal of Plant Pathology. 2010 Aug;127:571-78. http://dx.doi.org/10.1007/s10658-010-9620-2

Graham JH, Bassanezi RB, Dawson WO, Dantzler R. Management of huanglongbing of citrus: Lessons from São Paulo and Florida. Annual Review of Phytopathology. 2024 Mar 1;62. https://dx.doi.org/10.2139/ssrn.4745163

Daungfu O, Youpensuk S, Lumyong S. Endophytic bacteria isolated from citrus plants for biological control of citrus canker in lime plants. Tropical Life Sciences Research. 2019 Jan;30(1):73. https://doi.org/10.21315/tlsr2019.30.1.5

Trivedi P, Spann T, Wang N. Isolation and characterization of beneficial bacteria associated with citrus roots in Florida. Microbial Ecology. 2011 Aug;62:324-36. https://doi.org/10.1007/s00248-011-9822-y

Ogolla FO, Neema DB. Cultural, morphological and biochemical identification of Xanthomonas spp. the causative agent of bacteria leaf spot in tomatoes in Wanguru, Mwea, Kirinyaga County, Kenya. International Journal of Research and Innovation in Applied Science (IJRIAS). 2019;4.

Naqvi SF, Inam-ul-Haq M, Khan MA, Tahir MI, Zahid Ali ZA, Rehman HM. Morphological and biochemical characterization of Xanthomonas campestris (Pammel) Dawson pv. sesami and it's management by bacterial antagonists. Pakistan Journal of Agricultural Sciences. 2013;50:229-35.

Huatang W, Xinnian Z, Peipei X, Liu Y. Rapid DNA extraction methods for direct-PCR detection Citrus Huanglongbing. Plant Diseases and Pests. 2015 Jun 1;6(3).

Chowdappa A, Kousalya S, Kamalakannan A, Gopalakrishnan C, Venkatesan K. Efficacy of plant oils against Xanthomonas axonopodis pv. punicae. Advances in Research. 2018 Oct 27;17(1):1-5. http://dx.doi.org/10.9734/AIR/2018/45031

Isokar S. Molecular and biochemical characterization of Xanthomonas citri subsp. citri. 2019. http://dx.doi.org/10.13140/RG.2.2.19197.74729

He Y, Jia R, Qi J, Chen S, Lei T, Xu L, et al. Functional analysis of citrus AP2 transcription factors identified CsAP2-09 involved in citrus canker disease response and tolerance. Gene. 2019 Jul 30;707:178-88. https://doi.org/10.1016/j.gene.2019.04.021

Tamura K, Stecher G, Kumar S. MEGA11: molecular evolutionary genetics analysis version 11. Molecular Biology and Evolution. 2021 Jul 1;38(7):3022-27. https://doi.org/10.1093/molbev/msab120

Ali MR, Hasan MF, Lia RS, Akter A, Sumi MS, Hossain MF, et al. Isolation and characterization of a canker disease causing pathogen from Citrus aurantifolia and evaluation of its biological control measure. Journal of Entomology and Zoology Studies. 2017;5(6):1526-32.

Ambrico A, Trupo M. Efficacy of cell free supernatant from Bacillus subtilis ET-1, an Iturin A producer strain, on biocontrol of green and gray mold. Postharvest Biology and Technology. 2017 Dec 1;134:5-10. https://doi.org/10.1016/j.postharvbio.2017.08.001

Buragohain T, Dey P, Osborne WJ. In vitro studies on the inhibition of microbial pathogens by PPDHMP synthesized by Bacillus sp.; an endophyte of Citrus limon (Kaji nemu). Food Bioscience. 2023 Oct 1;55:103003. http://dx.doi.org/10.1016/j.fbio.2023.103003

Balogun OO, Ugoh SC, Oladosu PO. Antimicrobial activity and GC-MS based analysis of the extract of Bacillus subtilis subsp. subtilis 168 isolated from a river bank. Innovations in Microbiology and Biotechnology. 2022 Aug 25;7:126-45. https://doi.org/10.9734/mrji/2022/v32i230374

Manju P, Subramanian S. Effect of Bacillus spp. on Gerbera plant growth and control of Meloidogyne incognita. Journal of Applied and Natural Science. 2017 Sep 1;9(3):1644-50. http://dx.doi.org/10.31018/jans.v9i3.1415

Wang X, Liang L, Shao H, Ye X, Yang X, Chen X, et al. Isolation of the novel strain Bacillus amyloliquefaciens F9 and identification of lipopeptide extract components responsible for activity against Xanthomonas citri subsp. citri. Plants. 2022 Feb 7;11(3):457. https://doi.org/10.3390/plants11030457

Wheeler BE. An introduction to plant diseases. John Willey and Sons Ltd. London, UK. 1969;301.

Wang N. The citrus huanglongbing crisis and potential solutions. Molecular Plant. 2019 May 6;12(5):607-09. https://doi.org/10.1016/j.molp.2019.03.008

Bora P, Sharma P, Saikia A, Ahmed SS. Canker-induced shifts in microbial diversity for antagonist mediated disease management: A mini-review. Research and reviews. Journal of Agricultural Science and Technology. 2021;10(2):20-27.

Roper MC, Greve LC, Warren JG, Labavitch JM, Kirkpatrick BC. Xylella fastidiosa requires polygalacturonase for colonization and pathogenicity in Vitis vinifera grapevines. Molecular Plant-microbe Interactions. 2007 Apr;20(4):411-19. https://doi.org/10.1094/mpmi-20-4-0411

Graham JH, Gerberich KM, Davis CL. Injection–infiltration of attached grapefruit with Xanthomonas citri subsp. citri to evaluate seasonal population dynamics in citrus canker lesions. Journal of Phytopathology. 2016 Aug;164(7-8):528-33. https://doi.org/10.1111/jph.12478

Mansfield J, Genin S, Magori S, Citovsky V, Sriariyanum M, Ronald P, et al. Top 10 plant pathogenic bacteria in molecular plant pathology. Molecular Plant Pathology. 2012 Aug;13(6):614-29. https://doi.org/10.1111/j.1364-3703.2012.00804.x

Fonseca NP, Felestrino ÉB, Caneschi WL, Sanchez AB, Cordeiro IF, Lemes CG, et al. Detection and identification of Xanthomonas pathotypes associated with citrus diseases using comparative genomics and multiplex PCR. PeerJ. 2019 Oct 1;7:e7676. https://doi.org/10.7717/peerj.7676

González-González MG, Gómez-Sanchis J, Blasco J, Soria-Olivas E, Chueca P. CitrusYield: A dashboard for mapping yield and fruit quality of citrus in precision agriculture. Agronomy. 2020 Jan 15;10(1):128. https://doi.org/10.3390/agronomy10010128

Li Y, Liu S, Zhao C, Zhang Z, Nie D, Tang W, Li Y. The chemical composition and antibacterial and antioxidant activities of five citrus essential oils. Molecules. 2022 Oct 19;27(20):7044. https://doi.org/10.3390/molecules27207044

Kloepper JW, Ryu CM, Zhang S. Induced systemic resistance and promotion of plant growth by Bacillus spp. Phytopathology. 2004 Nov;94(11):1259-66. https://doi.org/10.1094/phyto.2004.94.11.1259

Al-Saleh MA. Evaluation of Saudi fluorescent Pseudomonas isolates as a biocontrol agent against citrus canker disease caused by Xanthomonas citri subsp. citri. Egyptian Academic Journal of Biological Sciences, G Microbiology. 2014 Dec 1;6(2):1-7. https://doi.org/10.21608/eajbsg.2014.16493

Singh A, Thakur A, Sharma S, Gill PP, Kalia A. Bio-inoculants enhance growth, nutrient uptake and buddability of citrus plants under protected nursery conditions. Communications in Soil Science and Plant Analysis. 2018 Nov 13;49(20):2571-86. http://dx.doi.org/10.1080/00103624.2018.1526946

Zhang M, Yang C, Powell CA, Avery PB, Wang J, Huang Y, Duan Y. Field evaluation of integrated management for mitigating citrus huanglongbing in Florida. Frontiers in Plant Science. 2019 Jan 31;9:1890. https://doi.org/10.3389/fpls.2018.01890

Zhou Y, Ma J, Xie J, Deng L, Yao S, Zeng K. Transcriptomic and biochemical analysis of highlighted induction of phenylpropanoid pathway metabolism of citrus fruit in response to salicylic acid, Pichia membranaefaciens and oligochitosan. Postharvest Biology and Technology. 2018 Aug 1;142:81-92. http://dx.doi.org/10.1016/j.postharvbio.2018.01.021

Zheng K, Zheng H, Yu Y, Su J, Chen L, Zheng M, et al. Simultaneous determination of four pesticides residues in rice by modified QuEChERS coupled with GC-MS/MS. Journal of Food Composition and Analysis. 2024 Jun 12;106396. http://dx.doi.org/10.1016/j.jfca.2024.106396

Ajilogba CF, Babalola OO. GC–MS analysis of volatile organic compounds from Bambara groundnut rhizobacteria and their antibacterial properties. World Journal of Microbiology and Biotechnology. 2019 Jun;35:1-9. https://doi.org/10.1016/j.jfca.2024.106396

Giassi V, Kiritani C, Kupper KC. Bacteria as growth-promoting agents for citrus rootstocks. Microbiological Research. 2016 Sep 1;190:46-54. https://doi.org/10.1016/j.micres.2015.12.006

Chen K, Tian Z, He H, Long CA, Jiang F. Bacillus species as potential biocontrol agents against citrus diseases. Biological Control. 2020 Dec 1;151:104419. https://doi.org/10.1016/j.biocontrol.2020.104419

Qian J, Zhang T, Tang S, Zhou L, Li K, Fu X, Yu S. Biocontrol of citrus canker with endophyte Bacillus amyloliquefaciens QC-Y. Plant Protection Science. 2020 Dec 3;57(1):1-3. https://doi.org/10.17221/62/2020-PPS

Ke X, Wu Z, Liu Y, Liang Y, Du M, Li Y. Isolation, antimicrobial effect and metabolite analysis of Bacillus amyloliquefaciens ZJLMBA1908 against citrus canker caused by Xanthomonas citri subsp. citri. Microorganisms. 2023 Dec 6;11(12):2928. https://doi.org/10.3390/microorganisms11122928

Ma X, Wang X, Cheng J, Nie X, Yu X, Zhao Y, Wang W. Microencapsulation of Bacillus subtilis B99-2 and its biocontrol efficiency against Rhizoctonia solani in tomato. Biological Control. 2015 Nov 1;90:34-41. http://dx.doi.org/10.1016/j.biocontrol.2015.05.013

Shaheed KA, AlGaraawi NI, Alsultany AK, Abbas ZH, Khshayyish IK, Al Khazali MT. Analysis of bioactive phytochemical compound of (Cyperus iria L.) By using gas chromatography–mass spectrometry. In: IOP Conference Series: Earth and Environmental Science. IOP Publishing; 2019 Nov 1;388(1):012064. https://iopscience.iop.org/article/10.1088/1755-1315/388/1/012064

Georgiadis MP. Products from furans. 1. Synthesis and anticoccidial and antimicrobial activity of 5-amino-5, 6-dihydro-6-methoxy-2-methyl-2-(4'-biphenylyl)-2H-pyran-3 (4H)-ones and related compounds. Journal of Medicinal Chemistry. 1976 Feb;19(2):346-49. https://doi.org/10.1021/jm00224a033

Okore GJ, Oguzie EE, Ogukwe CE, Akalezi CO. GC-MS analysis of phytochemicals from the extract of Hibiscus sabdariffa grown in Northern Nigeria. Journal of Chemical Society of Nigeria. 2021 Apr 11;46(2). https://doi.org/10.46602/jcsn.v46i2.613

Sánchez-Hernández E, Cáceres-González C, Peña-Delgado V, García-Valdecasas Medina JI, Casao A, Valdez-Ayala S, et al. Proximate analysis and GC-MS phytochemical profiling of aqueous extracts of Doryopteris raddiana, a plant used by the Mbya-Guaraní as a contraceptive. Natural Product Research. 2024 May 7;1-8. https://doi.org/10.1080/14786419.2024.2352869

Abu-Gharbia M, M Mohamed R, A Awad M. Metabolic profiling of endophytic Bacillus subtilis US2 isolated from Rosmarinus officinalis leaves with potential antimicrobial activity. Journal of Pharmaceutical and Applied Chemistry. 2020 Jan 1;6(1):26-37. https://jpac.journals.ekb.eg/article_202184.html

Adebiyi JA, Njobeh PB, Adebo OA, Kayitesi E. Metabolite profile of Bambara groundnut (Vigna subterranea) and dawadawa (an African fermented condiment) investigation using gas chromatography high resolution time-of-flight mass spectrometry (GC-HRTOF-MS). Heliyon. 2021 Apr 1;7(4). https://doi.org/10.1016/j.heliyon.2021.e06666

Khan FA, Abdeltawab AA, Al-Deyab SS, Ali J, Ullah R, Qureshi MN, et al. Comparative evaluation of physiochemical and GC-MS analysis of sour oranges and sweet oranges peels oil. Life Science Journal. 2013;10(10s):205-09.

Ragupathi V, Stephen A, Arivoli D, Kumaresan S. Antibacterial activity, in vitro antioxidant potential and GC-MS characterization of methanolic extract of Gymnopilus junonius, a wild mushroom from Southern Western Ghats, India. Eur J Biomed. 2018 Mar;5:650-57.

Sampathkumar Y, Elumali S, Halith AM. GCMS determination of anticancer, anti-inflammatory and anti-bacterial compounds from salt tolerance microalgae (Lyngbya sp. Nostoc sp. and Phormidium sp.) Isolated from Marakkanam Salt Pan, Tamil Nadu, India. Tamil Nadu, India. 2020;11:14. http://dx.doi.org/10.1007/s42770-023-01111-1

Murniasih T, Masteria Yunovilsa P, Untari F. Antibacterial activity and GC–MS based metabolite profiles of Indonesian marine Bacillus. Indones J Pharm. 2022 Jul 1;33:475-83. https://doi.org/10.22146/ijp.3504

Awan ZA, Shoaib A, Schenk PM, Ahmad A, Alansi S, Paray BA. Antifungal potential of volatiles produced by Bacillus subtilis BS-01 against Alternaria solani in Solanum lycopersicum. Frontiers in Plant Science. 2023 Jan 26;13:1089562. https://doi.org/10.3389/fpls.2022.1089562

Wong CH, Chen ST, Hennen WJ, Bibbs JA, Wang YF, Liu JL, et al. Enzymes in organic synthesis: use of subtilis in and a highly stable mutant derived from multiple site-specific mutations. Journal of the American Chemical Society. 1990 Jan;112(3):945-53. https://doi.org/10.1021/ja00159a006

Koilybayeva M, Shynykul Z, Ustenova G, Waleron K, Jo?ca J, Mustafina K, et al. Gas chromatography–mass spectrometry profiling of volatile metabolites produced by some Bacillus spp. and evaluation of their antibacterial and antibiotic activities. Molecules. 2023 Nov 12;28(22):7556. https://doi.org/10.3390/molecules28227556

Published

19-12-2024

How to Cite

1.
Shanmugapriya V, Mareeswari P, Revathy N, Yesuraja I, Manonmani K, Anandhan M, Mini M, Ayyandurai M. Biological control of citrus canker by endophytic bacteria. Plant Sci. Today [Internet]. 2024 Dec. 19 [cited 2024 Dec. 22];11(sp4). Available from: https://horizonepublishing.com/journals/index.php/PST/article/view/5633