Pathological phenotypes of Xanthomonas citri pv. citri causal agent of citrus bacterial canker and in vitro efficiency assessment of copper compounds in its inhibition

Document Type : Research Article

Authors

1 Department of Plant protection, College of Agriculture, Shiraz University, Iran

2 Dept. of Plant Protection Shiraz University

Abstract

As a consequence of the abundant use of copper bactericides, some species of plant pathogenic bacteria, including Xanthomonas citri pv. citri (Xcc), have developed resistance to copper. This study evaluates pathological phenotypes and in vitro efficiency of prevalent copper compounds in inhibiting Xcc. Sampling was conducted across citrus orchards during 2021-2022. One hundred and seventeen citrus orchards were surveyed and 132 strains identified as Xanthomonas citri pv. citri based on phenotypic and genotypic characteristics. Different pathogenic phenotypes were observed by inoculating the representative strains on different cultivars of lime, orange, citrumelo and grapefruit, but tangerine cultivars did not show disease symptoms. All the strains showed an intermediate level of sensitivity and grew in the medium containing 100 and 150 mg/l of copper sulfate. In addition higher concentrations (250, 300, 350 and 400 mg/liter) showed inhibition rate more than 90%. Copper carboxylate at the concentration of 0.75 g/liter prevented the growth of more than 85% of the strains. Also, Copper oxychloride showed the best inhibition rate in concentrations higher than 3 g/liter. 1.5 ml/liter of Bordeaux mixture controlled more than 80% of the strains. None of copper resistance gene clusters copLAB and copABCD were detected in tested Xanthomonas citri strains. The proper characterization of pathological phenotypes and sensitivity to copper leads more confident decisions in citrus canker disease management.

Keywords


Adriko J., Mbega E. R., Mortensen C. N., Wulff E. G., and Lund O. S. 2014. Improved PCR for identification of members of the genus Xanthomonas. European Journal of Plant Pathology 138: 293–306. 10.1007/s10658-013-0329-x
Alizadeh A. and Rahimian H. 1990.  Citrus canker in Kerman Province. Iranian Journal of Plant Pathology 26:1-4.
 Areas M. S., Gonçalves R. M., Soman J. M., Souza Filho R. C., Gioria R., Silva Junior T. A. F. and  Maringoni A. A. 2018. Resistance of Xanthomonas euvesicatoria strains from Brazilian pepper to copper and zinc sulfates. Annals of the Brazilian Academy of Sciences 90: 2375-2380. https://doi.org/10.1046/j.1439-0434.2002.00746.x
Behlau F., Canteros B. I., Minsavage G. V., Jones J. B. and Graham J. H. 2011. Molecular characterization of copper resistance genes from Xanthomonas citri subsp. citri and Xanthomonas alfafae subsp. citrumelonis. Applied and Environmental Microbiology 77:4089-4096. 10.1128/AEM.03043-10
Behlau F., Canteros B.I., Minsavage G.V., Jones J.B. and Graham J.H. 2011. Molecular characterization of copper resistance genes from Xanthomonas citri subsp. citri and Xanthomonas alfalfae subsp. citrumelonis. Applied and Environmental Microbiology 77:4089–4096. 10.1128/AEM.03043-10
Behlau F., Gochez A.M., Lugo A.J., Elibox W., Minsavage G.V., Potnis N., White F.F., Ebrahim M., Jones J.B. and Rumsubhag A. 2017a. Characterization of a unique copper resistance gene cluster in Xanthomonas campestris pv. campestris isolated in Trinidad, West Indies. European Journal of Plant Pathology 147: 671–681. https://doi.org/10.1007/s10658-016-1035-2
Behlau F., Graham J.H. and Jones J.B. 2012. Copper resistance genes from different xanthomonads and citrus epiphytic bacteria confer resistance to Xanthomonas citri subsp. citri. European Journal of Plant Pathology 133: 949–963. https://doi.org/10.1007/s10658-012-9966-8
Behlau F., Hong J.C., Jones J.B. and Graham J.H. 2013. Evidence for acquisition of copper resistance genes from different sources in citrus-associated xanthomonads. Phytopathol­ogy 103: 409–418. http://dx.doi.org/10.1094/ PHYTO-06-12-0134-R
Behlau F., Scandelai L.H.M., Silva Junior G.J. and Lanza F.E. 2017b. Soluble and insoluble copper formulations and metallic copper rate for control of citrus canker on sweet orange trees. Crop Protection 94:185–191. https://doi.org/10.1016/j.cropro.2017.01.003
Chillappagari S., Seubert A., Trip H., Kuipers O.P., Marahiel M.A. and Miethke M. 2010. Copper stress affects iron homeostasis by destabilizing iron sulfur cluster formation in Bacillus subtilis. Journal of Bacteriology 192: 2512–2524. https://doi.org/10.1128/jb.00058-10
Coletta-Filho H. D., Takita M. A., de Souza A. A., Neto J. R., Deste fano S. A. L., Hurtung J. S., and Machado, M. A. 2006. Primers based on rpf gene region provide improved detection of Xanthomonas axonopodis pv. citri in naturally and artificially infected citrus plants. Applied Microbiology 100: 279–285. 10.1111/j.1365-2672.2005.02787.x
Escalon A., Javegny S., Vernière C., Noel L.D., Vital K., Poussier S., Hajri A., Boureau T.,  Pruvost O., Arlat M., Gagnevin L. 2013. Variations in type III effector repertoires, pathological phenotypes and host range of Xanthomonas citri pv. citri pathotypes. Molecular Plant Pathology 14: 483–496. https://doi.org/10.1111/mpp.12019
Fan X., Saleem T. and Zou H. 2022. Copper resistance mechanisms in plant pathogenic bacteria. Phytopathologia Mediterranea 61: 129-138. https://doi.org/10.36253/phyto-13282
Figueiredo J., Minsavage G., Graham J., White F. and Jones J. 2011. Mutational analysis of type III effector genes from Xanthomonas citri subsp. citri. European Journal of Plant Pathology. 130: 339–347. 10.1007/s10658-011-9757-7
Graham J. H., Dewdney M. M., and Yonce H. D. 2011. Comparison of copper formulations for control of citrus canker on ‘Hamlin’ orange. Proceedings of the Florida State Horticultural Society 124: 79-84.
Graham J.H., Brooks C. and Yonce H.D. 2016. Importance of early season copper sprays for protection of Hamlin orange fruit against citrus canker infection and premature fruit drop. Proceedings of the Florida State Horticultural Society 129: 74–78.
Heydarpanah S., Rezaei R., Taghavi S. M. and charehgani H. 2019. Efficacy of different copper compounds in the control of Xanthomonas citri subsp. citri pathotypes A and A*. Journal of phytopathology 168: 73-80. 10.1111/jph.12869
Hoshino N., Kimura T., Yamaji A. and Ando T. 1999. Damage to the cytoplasmic membrane of Escherichia coli by catechin-copper (II) complexes. Free Radical Biology and Medicine 27: 1245–1250. 10.1016/s0891-5849(99)00157-4
Izadiyan M. and Taghavi S. M. 2023. Diversity of copper resistant Xanthomonas citri subsp. citri strains, the causal agent of Asiatic citrus canker, in Iran. European Journal of Plant Pathology 168:593-606. https://doi.org/10.1007/s10658-023-02786-w
Khodakaramian G. and Swings J. 2002. AFLP fingerprinting of the strains of Xanthomonas axonopodis inducing citrus canker disease in southern Iran. Journal of Phytopathology 150: 227–231. https://doi.org/10.1046/j.1439-0434.2002.00746.x
Lai Y.R., Lin C.H., Chang C.P., Ni H.F., Tsai W.S. and Huang C.J. 2021. Distribution of copper resistance gene variants of Xanthomonas citri subsp. citri and Xanthomonas euvesicatoria pv. perforans. Plant Protection Science 57: 206–216. 10.17221/160/2020-PPS
Lamichhane J.R., Osdaghi E., Behlau F., Köhl J., Jones J.B. and Aubertot J. N. 2018. Thirteen decades of antimicrobial copper compounds applied in agriculture. A review. Agronomy for Sustainable Development 38: 28-46. https://doi.org/10.1007/s13593-018-0503-9
Lee Y. A., Sung A., N., Liu T. F., and Lee Y. S. 2009. Combination of chromatic differential medium and estA-specific PCR for isolation and detection of phytopathogenic Xanthomonas spp. Applied and Environmental Microbiology 75: 6831–6838. 10.1128/AEM.01653-09
Licciardello G., Caruso P., Bella P., Boyer C., Smith M.W., Pruvost O., Robene I., Cubero J. and Catara V. 2022. Pathotyping Citrus Ornamental Relatives with Xanthomonas citri pv. citri and X. citri pv. aurantifolii Refines Our Understanding of Their Susceptibility to These Pathogens. Microorganisms 10: 986-1000. https://doi.org/10.3390/microorganisms10050986
Marco G. M. and Stall R. E. 1983. Control of bacterial spot of pepper initiated by strains of Xanthomonas campestris pv. vesicatoria that differ in sensitivity to copper. Plant Disease 67: 779-781. 10.1094/PD-67-779
Marin T.G.S., Galvanin A. L., Lanza F. E. and Behlau F. 2019. Description of copper tolerant Xanthomonas citri subsp. citri and genotypic comparison with sensitive and resistant strains. Plant Pathology 68: 1088–1098. https://doi.org/10.1111/ppa.13026
Mostofizadeh-Ghalamfarsa R. and Rahimian H. 1996. Incidence of the Asiatic form of citrus canker in Iran. Iranian Journal of Plant Pathology 32: 189 (short report).
 Moura Duin I., Aparecida Forasteiro T., Hitomi Sugahara Rodrigues V., Isabel Balbi-Peña M. and Pereira Leite Júnior R. 2022. In vitro sensitivity of Xanthomonas vasicola pv. vasculorum and the control of bacterial leaf streak of corn with copper oxychloride alone and in mixtures with mancozeb and fluxapyroxad. European Journal of plant pathology 164: 263-268. https://doi.org/10.1007/s10658-022-02556-0
Pernezny K., Raid R.N., Stall R.E., Hodge N.C., Collins J. 1995. An outbreak of bacterial spot of lettuce in Florida caused by Xanthomonas campestris pv. vitians. Plant Diseases 79: 359-360. 10.1094/PD-79-0359
Pohronezny K., Stall R.E., Canteros B.I., Kegley M., Datnoff L.E. and Subramanya R. 1992. Sudden shift in the prevalent race of Xanthomonas campestris pv. vesicatoria in pepper fields in southern Florida. Plant Disease 76: 118–120. 10.1094/PD-76-0118
Pruvost O., Goodarzi T., Boyer K., Soltaninejad H., Escalon A., Alavi S. M., Javegny S., Boyer C., Cottyn B., Gagnevin L. and Verniere C. 2015. Genetic structure analysis of strains causing citrus canker in Iran reveals the presence of two different lineages of Xanthomonas citri pv. citri pathotype A*. Plant pathology 64: 776–784: 10.1111/ppa.12324.
Rehman M. A., Ali Sh., Khan M. N., Ahmad S. and Ali M. A. 2020. Comparative efficacy of different antibiotics, fungicides and botanical extract for the control of citrus canker of kinnow mandarin. Pakistan journal of phytopathology 32: 113-119. 10.33866/phytopathol.030.02.0573
Richard D., Ravigné V., Rieux A., Facon B., Boyer C., Boyer K., Grygiel P., Javengy S., Terville M., Canteros B.I., Robene I., Verniere C., Chabirand A., Pruvost O. and Lefeuvre P. 2017. Adaptation of genetically monomorphic bacteria: evolution of copper resistance through multiple horizontal gene transfers of complex and versatile mobile genetic elements. Molecular Ecology 26: 2131–2149. 10.1111/mec.14007
Schaad N.W., Jones J.B. and Chun W. 2001. Laboratory Guide for Identification of Plant Pathogenic Bacteria, Third edition. APS, St. Paul, MN, USA, 379 pp.
Stall R. E., Loschke D. C., and Jones J. B. 1986. Linkage of copper resistance and avirulence loci on a self-transmissible plasmid in Xanthomonas campestris pv. vesicatoria. Phytopathology 76: 240-243. 10.1094/Phyto-76-240
Terumi Itako A., Batista Tolentino Júnior J., Fernandes da Silva Júnior T. A., Marcelo Soman J. and Carlos Maringoni A. 2015. Chemical products induce resistance to Xanthomonas perforans in tomato. Brazilian Journal of Microbiology 46: 701-706. http://dx.doi.org/10.1590/S1517-838246320140177
Vernière C., Devaux M., Pruvost O., Couteau A. and Luisett J. 1991. Studies on the biochemical and physiological variation among strains of Xanthomonas campestris pv. citri the causal agent of citrus bacterial canker disease. Fruits 46: 160-170.
Yan Q. and Wang N. 2012. High-throughput screening and analysis of genes of Xanthomonas citri subsp. citri involved in citrus canker symptom development. Molecular Plant–Microbe Interactions. 25: 69–84. 10.1094/MPMI-05-11-0121
Yousefi Kopaei F., Taghavi S. M., Shiotani H. and Tesuyumu Shinji. 2014. A PCR-based assay for differentiating A and A* Type strains of Xanthomonas citri subsp. citri, the cusal agent of Asiatic citrus canker. Journal of General Plant Pathology 80: 85–89. 10.1007/s10327-013-0494-y