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Comparison of Resistance Acquisition and Mechanisms in Erwinia amylovora against Agrochemicals Used for Fire Blight Control

  • Hyeonheui Ham (Crop Protection Division, National Institute of Agricultural Sciences, Rural Development Administration) ;
  • Ga-Ram Oh (Crop Protection Division, National Institute of Agricultural Sciences, Rural Development Administration) ;
  • Yong Hwan Lee (Crop Protection Division, National Institute of Agricultural Sciences, Rural Development Administration) ;
  • Yong Hoon Lee (Division of Biotechnology, Jeonbuk National University)
  • Received : 2024.07.17
  • Accepted : 2024.09.02
  • Published : 2024.10.01

Abstract

Agrochemicals containing antibiotics are authorized to manage fire blight that has been occurring in Korea since 2015. The minimum inhibitory concentration (MIC) of each antibiotic against Erwinia amylovora, the causal pathogen of fire blight, has increased over the years due to the pathogen's frequent exposure to antibiotics, indicating the necessity to prepare for the emergence of antibiotic resistance. In this study, E. amylovora was exposed to stepwise increasing concentrations of eight different agrochemicals, each containing single or mixed antibiotics, and gene mutation and changes in MIC were assessed. Streptomycin and oxolinic acid induced an amino acid substitution in RpsL and GyrA, respectively, resulting in a rapid increase in MIC. Oxytetracycline initially induced amino acid substitutions or frameshifts in AcrR, followed by substitutions of 30S small ribosomal protein subunit S10 or AcrB, further increasing MIC. E. amylovora acquired resistance in the order of oxolinic acid, streptomycin, and oxytetracycline at varying exposure frequencies. Resistance acquisition was slower against agrochemicals containing mixed antibiotics than those with single antibiotics. However, gene mutations conferring antibiotic resistance emerged sequentially to both antibiotics in the mixed formulations. Results suggested that frequent application of mixed antibiotics could lead to the emergence of multidrug-resistant E. amylovora isolates. This study provided essential insights into preventing the emergence of antibiotic-resistant E. amylovora and understanding the underlying mechanisms of resistance acquisition.

Keywords

Acknowledgement

This study was carried out with the support of Cooperative Research Programs (Project no. RS-2020-RD009337) from the Rural Development Administration, Republic of Korea.

References

  1. Anes, J., McCusker, M. P., Fanning, S. and Martins, M. 2015. The ins and outs of RND efflux pumps in Escherichia coli. Front. Microbiol. 6:587.
  2. Beabout, K., Hammerstrom, T. G., Perez, A. M., Magalhaes, B. F., Prater, A. G., Clements, T. P., Arias, C. A., Saxer, G. and Shamoo, Y. 2015. The ribosomal S10 protein is a general target for decreased tigecycline susceptibility. Antimicrob. Agents Chemother. 59:5561-5566.
  3. Brodersen, D. E., Clemons, W. M. Jr., Carter, A. P., Morgan-Warren, R. J., Wimberly, B. T. and Ramakrishnan, V. 2000. The structural basis for the action of the antibiotics tetracycline, pactamycin, and hygromycin B on the 30S ribosomal subunit. Cell 103:1143-1154.
  4. Chiou, C.-S. and Jones, A. L. 1995. Molecular analysis of high-level streptomycin resistance in Erwinia amylovora. Phytopathology 85:324-328.
  5. Deng, W., Li, C. and Xie, J. 2013. The underling mechanism of bacterial TetR/AcrR family transcriptional repressors. Cell. Signal. 25:1608-1613.
  6. Entenza, J. M., Giddey, M., Vouillamoz, J. and Moreillon, P. 2010. In vitro prevention of the emergence of daptomycin resistance in Staphylococcus aureus and enterococci following combination with amoxicillin/clavulanic acid or ampicillin. Int. J. Antimicrob. Agents 35:451-456.
  7. Escursell, M. M., Roschi, A., Smits, T. H. M. and Rezzonico, F. 2021. Characterization and direct molecular discrimination of rpsL mutations leading to high streptomycin resistance in Erwinia amylovora. J. Plant Pathol. 103:99-108.
  8. Forster, H., McGhee, G. C., Sundin, G. W. and Adaskaveg, J. E. 2015. Characterization of streptomycin resistance in isolates of Erwinia amylovora in California. Phytopathology 105:1302-1310.
  9. Grossman, T. H. 2016. Tetracycline antibiotics and resistance. Cold Spring Harb. Perspect. Med. 6:a025387.
  10. Gu, R., Li, M., Su, C. C., Long, F., Routh, M. D., Yang, F., McDermott, G. and Yu, E. W. 2008. Conformational change of the AcrR regulator reveals a possible mechanism of induction. Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 64:584-588.
  11. Ham, H., Oh, G.-R., Lee, B. W., Lee, Y. H. and Lee, Y. H. 2023. Assessment of oxytetracycline and oxolinic acid resistance of Erwinia amylovora isolated from 2019 to 2022 in Korea. Korean J. Pestic. Sci. 27:283-292 (in Korean).
  12. Ham, H., Oh, G.-R., Lee, B. W., Lee, Y. H. and Lee, Y. H. 2024. Changes of sensitivity to streptomycin in Erwinia amylovora isolated from 2019 to 2023 in Korea. Res. Plant Dis. 30:199-205 (in Korean).
  13. Ham, H., Oh, G.-R., Park, D. S. and Lee, Y. H. 2022. Survey of oxolinic acid-resistant Erwinia amylovora in Korean apple and pear orchards, and the fitness impact of constructed mutants. Plant Pathol. J. 38:482-489.
  14. Herbert, A., Hancock, C. N., Cox, B., Schnabel, G., Moreno, D., Cavelho, R., Jones, J., Paret, M., Geng, X. and Wang, H. 2022. Oxytetracycline and streptomycin resistance genes in Xanthomonas arboricola pv. pruni, the causal agent of bacterial spot in peach. Front. Microbiol. 13:821808.
  15. Hirata, T., Saito, A., Nishino, K., Tamura, N. and Yamaguchi, A. 2004. Effects of efflux transporter genes on susceptibility of Escherichia coli to tigecycline (GAR-936). Antimicrob. Agents Chemother. 48:2179-2184.
  16. Izghirean, N., Waidacher, C., Kittinger, C., Chyba, M., Koraimann, G., Pertschy, B. and Zarfel, G. 2021. Effects of ribosomal protein S10 flexible loop mutations on tetracycline and tigecycline susceptibility of Escherichia coli. Front. Microbiol. 12:663835.
  17. Kleitman, F., Shtienberg, D., Blachinsky, D., Oppenheim, D., Zilberstaine, M., Dror, O. and Manulis, S. 2005. Erwinia amylovora populations resistant to oxolinic acid in Israel: prevalence, persistence and fitness. Plant Pathol. 54:108-115.
  18. Krajewska, J., Tyski, S. and Laudy, A. E. 2023. Mutant prevention concentration, frequency of spontaneous mutant selection, and mutant selection window-a new approach to the in vitro determination of the antimicrobial potency of compounds. Antimicrob. Agents Chemother. 67:e0137322.
  19. Lee, M. S., Lee, I., Kim, S. K., Oh, C.-S. and Park, D. H. 2018. In vitro screening of antibacterial agents for suppression of fire blight disease in Korea. Res. Plant Dis. 24:41-51 (in Korean).
  20. Maeda, Y., Kiba, A., Ohnishi, K. and Hikichi, Y. 2004. Implications of amino acid substitutions in GyrA at position 83 in terms of oxolinic acid resistance in field isolates of Burkholderia glumae, a causal agent of bacterial seedling rot and grain rot of rice. Appl. Environ. Microbiol. 70:5613-5620.
  21. Maeda, Y., Kiba, A., Ohnishi, K. and Hikichi, Y. 2007. Amino acid substitutions in gyrA of Burkholderia glumae are implicated in not only oxolinic acid resistance but also fitness on rice plants. Appl. Environ. Microbiol. 73:1114-1119.
  22. Manulis, S., Kleitman, F., Dror, O. and Shabi, E. 2000. Isolation of strains of Erwinia amylovora resistant to oxolinic acid. IOBC WPRS Bull. 23:89-92.
  23. Manulis, S., Kleitman, F., Shtienberg, D., Shwartz, H., Oppenheim, D., Zilberstaine, M. and Shabi, E. 2003. Changes in the sensitivity of Erwinia amylovora populations to streptomycin and oxolinic acid in Israel. Plant Dis. 87:650-654.
  24. Martin, R. G. and Rosner, J. L. 2001. The AraC transcriptional activators. Curr. Opin. Microbiol. 4:132-137.
  25. McGhee, G. C. and Sundin, G. W. 2011. Evaluation of kasugamycin for fire blight management, effect on nontarget bacteria, and assessment of kasugamycin resistance potential in Erwinia amylovora. Phytopathology 101:192-204.
  26. McManus, P. S. and Jones, A. L. 1994. Epidemiology and genetic analysis of streptomycin-resistant Erwinia amylovora from Michigan and evaluation of oxytetracycline for control. Phytopathology 84: 627-633.
  27. McManus, P. S., Stockwell, V. O., Sundin, G. W. and Jones, A. L. 2002. Antibiotic use in plant agriculture. Annu. Rev. Phytopathol. 40:443-465.
  28. Miller, T. D. and Schroth, M. N. 1972. Monitoring the epiphytic populations of Erwinia amylovora on pear with a selective medium. Phytopathology 62:1175-1182.
  29. Mortimer, P. G. and Piddock, L. J. 1993. The accumulation of five antibacterial agents in porin-deficient mutants of Escherichia coli. J. Antimicrob. Chemother. 32:195-213.
  30. Mouton, J. W., Muller, A. E., Canton, R., Giske, C. G., Kahlmeter, G. and Turnidge, J. 2017. MIC-based dose adjustment: facts and fables. J. Antimicrob. Chemother. 73:564-568.
  31. Schnabel, E. L. and Jones, A. L. 1999. Distribution of tetracycline resistance genes and transposons among phylloplane bacteria in Michigan apple orchards. Appl. Environ. Microbiol. 65:4898-4907.
  32. Stockwell, V. O. and Duffy, B. 2012. Use of antibiotics in plant agriculture. Rev. Sci. Tech. 31:199-210.
  33. Sundin, G. W., Peng, J., Brown, L. E., Zeng, Q., Forster, H. and Adaskaveg, J. E. 2023. A novel IncX plasmid mediates high-level oxytetracycline and streptomycin resistance in Erwinia amylovora from commercial pear orchards in California. Phytopathology 113:2165-2173.
  34. Sundin, G. W. and Wang, N. 2018. Antibiotic resistance in plant-pathogenic bacteria. Annu. Rev. Phytopathol. 56:161-180.
  35. Wang, G., Inaoka, T., Okamoto, S. and Ochi, K. 2009. A novel insertion mutation in Streptomyces coelicolor ribosomal S12 protein results in paromomycin resistance and antibiotic overproduction. Antimicrob. Agents Chemother. 53:1019-1026.
  36. Wiegand, I., Hilpert, K. and Hancock, R. E. W. 2008. Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances. Nat. Protoc. 3:163-175.
  37. Yoshida, H., Bogaki, M., Nakamura, M. and Nakamura, S. 1990. Quinolone resistance-determining region in the DNA gyrase gyrA gene of Escherichia coli. Antimicrob. Agents Chemother. 34:1271-1272.