Detection and characterization of endophytic bacteria causing knot in young olive trees


  • Maryam KHEZRI Assistant Professor of Plant Pathogenic Prokaryotes, Department of Plant Protection, Faculty of Agriculture, Urmia University, Urmia, Iran
  • Abolghasem GHASEMI



Pseudomonas savastanoi pv. savastanoi, Phenotypic identification, 16S rRNA, rpoB, cultivar resistance


Olive knot is an important disease in most countries where olives are commercially grown. In the spring of 2015, some galls were observed on the trunk and branches of 4-year-old olive trees in the north of Iran. The bacteria were isolated from galls and all isolates were gram-negative, aerobic, and capable of producing florescent pigment. Other phenotypic characteristics of the isolates were assessed. Pathogenicity tests were carried out on olive branches incubated with different isolates. Primary symptoms were observed after two weeks. Sequences of 16S rRNA and RNA polymerase beta subunit genes of pathogenic isolates were completely similar to Pseudomonas savastanoi pv. savastanoi (Smith 1908) Young et al. 1978 in GenBank. Based on the results from phenotypic analyses, pathogenicity tests and phylogenetic data, the isolates were identified as P. savastanoi pv. savastanoi. The host range of our isolates was specific to olive trees. None of the inoculated oleander (Nerium oleander L.), winter jasmine (Jasminum nudiflorum Lindl.), Japanese privet (Ligustrum japonicum Thunb.) and ash (Fraxinus excelsior L.) developed disease symptoms. No difference in disease resistance was observed between six studied olive cultivars. There was no olive tree or orchard around the studied orchard as far as more than one kilometer. As the disease agent listed in Iran’s foreign quarantine pests and diseases list, appropriate quarantine and phytosanitary measures were undertaken to eradicate the disease.

Author Biographies

  • Maryam KHEZRI, Assistant Professor of Plant Pathogenic Prokaryotes, Department of Plant Protection, Faculty of Agriculture, Urmia University, Urmia, Iran
    Department of Plant Protection, Faculty of Agriculture, Urmia University, Urmia, Iran
  • Abolghasem GHASEMI
    Department of Plant Pathology Research, Iranian Research Institute of Plant Protection (IRIPP), Tehran, Iran
    Director General, Bureau of Plant Quarantine, Plant Protection Organization (PPO), Tehran, Iran.


Addy, H. S., & Wahyuni, W. S. (2016). Nucleic acid and protein profile of bacteriophages that infect Pseudomonas syringae pv. glycinea, bacterial blight on soybean. Agriculture and Agricultural Science Procedia, 9, 475-481.

Agrios, G. N. (2005). Plant Pathology. 5th ed., New York, NY: Academic Press.

Benie, C. K. D., Dadie, A., N’Golo, D. C., Guessennd, N., Aka, S., Dje, K. M., & Dosso, M. (2016). Comparative evaluation of molecular detection performance of Pseudomonas aeruginosa based on phylogenetic markers 16S RNAr, recA, rpoB and ITS1. Clinical Microbiology, 5(6), e269.

Caballo-Ponce, E., Murillo, J., Martinez-Gil, M., Moreno-Perez, A., Pintado, A., & Ramos, C. (2017). Knots untie: molecular determinants involved in knot formation induced by pseudomonas savastanoi in woody hosts. Frontiers in Plant Science, 8, e1089.

Campos, A., da Costa, G., Coelho, A. V., & Fevereiro, P. (2009). Identification of bacterial protein markers and enolase as a plant response protein in the infection of Olea europaea subsp. europaea by Pseudomonas savastanoi pv. savastanoi. European Journal of Plant Pathology, 125, e603.

Case, R. J., Boucher, Y., Dahllof, I., Holmstrom, C., Doolittle, W. F., & Kjelleberg, S. (2007). Use of 16S rRNA and rpoB genes as molecular markers for microbial ecology studies. Applied and Environmental Microbiology, 73(1), 278-288.

EPPO. (2006). Pathogen-tested olive trees and rootstocks. EPPO Bulletin, 36, 77-83.

Ghasemi, A., Salehi, S., Shahriari, D., & Baniameri, V. (2006). Occurrence of oleander knot disease (Nerium oleander) in Tehran. Iran Journal of Plant Pathology, 42, 703-704.

Hall, T. A. (1999). BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/ 98/NT. Nucleic Acid Symposium Series, 41, 95-98.

Hassani, D., Buonaurio, R., & Tombesi, A. (2003). Response of some olive cultivars, hybrid and open pollinated seedling to Pseudomonas savastanoi pv. savastanoi. In N. S. Iacobellis, A. Collmer, S. Hutcheson, J. Mansfield, C. E. Morris, J. Murillo, N. W. Schaad, …(Eds.), Pseudomonas syringae and Related Pathogens (pp, 489-494). The Netherlands, Kluwer Academic Publishers.

Iacobellis, N. S. (2001). Olive knot. In O. C. Maloy & T. D. Murray (Eds.), Encyclopedia of Plant Pathology (pp, 714-715). New York, NY: John Wiley & Sons, Inc.

Kieffer, M., Neve, J., & Kepinski, S. (2010). Defining auxin response contexts in plant development. Current Opinions in Plant Biology, 13, 12-20.

Kimura, M. (1980). A simple method for estimating evolutionary rate of base substitutions through comparative studies of nucleotide sequences. Journal of Molecular Evolution, 16, 111-120.

Krid, S., Rhouma, A., Quesada, J. M., Penyalver, R., & Gargouri, A, (2009). Delineation of Pseudomonas savastanoi pv. savastanoi strains isolated in Tunisia by random-amplified polymorphic DNA analysis. Journal of Applied Microbiology, 106, 886-894.

Lelliott, R. A., Billing, E., & Hayward, A. C. (1966). A determinative scheme for the fluorescent plant pathogenic pseudomonads. Journal of Applied Bacteriology, 29, 470-489.

Llop, P., Caruso, P., Cubero, J., Morente, C., & Lopez, M. M. (1999). A simple extraction procedure for efficient routine detection of pathogenic bacteria in plant material by polymerase chain reaction. Journal of Microbiology Methods, 37, 23-31.

Marcelo, A., Fernandes, M., Fatima Potes, M., & Serrano, J. F. (1999). Reactions of some cultivars of Olea europaea L. to experimental inoculation with Pseudomonas syringae pv. savastanoi. Acta Horticulturae, 474, 581-584.

Marchi, G., Mori, B., Pollacci, P., Mencuccini, M., & Surico, G. (2009). Systemic spread of Pseudomonas savastanoi pv. savastanoi in olive explants. Plant Pathology, 58, 152-158.

Marchi, G., Viti, C., Giovannetti, L., & Surico, G. (2005). Spread of levan-positive populations of Pseudomonas savastanoi pv. savastanoi, the causal agent of olive knot, in central Italy. European Journal of Plant Pathology, 112, 101-112.

Marques, D. S. A., & Samson, R. (2016). Population dynamics of Pseudomonas savastanoi pv. phaseolicola in bean, throughout the epiphytic and pathogenic phases. Pesquisa Agropecuaria Brasileira, 51(5), 623-630.

Mehri, I., Turki, Y., Daly, I., Ben Rjab, A., Hassen, A., & Gtar, M. (2013). Molecular identification and assessment of genetic diversity of fluorescent pseudomonads based on different polymerase chain reaction (PCR) methods. African Journal of Microbiology Research, 7(19), 2103-2113.

Moretti, C., Vinatzer, B. A., Onofri, A., Valentini, F., & Buonaurio, R. (2017). Genetic and phenotypic diversity of Mediterranean populations of the olive knot pathogen, Pseudomonas savastanoi pv. savastanoi. Plant Pathology, 66, 595-605.

O’Brien, J. A., & Benková, E. (2013). Cytokinin cross-talking during biotic and abiotic stress responses. Front in Plant Science, 4, e451.

Patten, C. L., Blakney, A. J., & Coulson, T. J. (2013). Activity, distribution and function of indole-3-acetic acid biosynthetic pathways in bacteria. Critical Reviews in Microbiology, 39, 395-415.

Penyalver, R., Garcia, A., Ferrer, A., Bertolini, E., & Lopez, M. M. (2000). Detection of Pseudomonas savastanoi pv. savastanoi in olive plants by enrichment and PCR. Applied and Environmental Microbiology, 66(6), 2673-2677.

Penyalver, R., Garcia, A., Ferrer, A., Bertolini, E., Quesada, J. M., Salcedo, C. I., … Lopez , M. M. (2006). Factors affecting Pseudomonas savastanoi pv. savastanoi plant inoculations and their use for evaluation of olive cultivar susceptibility. Phytopathology, 96, 313-319.

Quesada, J. M., Garcia, A., Bertolini, E., Lopez, M. M., & Penyalver, R. (2007). Recovery of Pseudomonas savastanoi pv. savastanoi from symptomless shoots of naturally infected olive trees. International Microbiology, 10, 77-84.

Quesada, J. M., Penyalver, R., Pérez-Panadés, J., Salcedo, C. I., Carbonell, E. A., & López, M. M. (2010). Comparison of chemical treatments for reducing epiphytic Pseudomonas savastanoi pv. savastanoi populations and for improving subsequent control of olive knot disease. Crop Protection, 29, 1413-1420.

Rademaker, J. L. W., & de Bruijn, F. J. (1997). Characterization and classification of microbes by Rep-PCR genomic fingerprinting and computer assisted pattern analysis. In: G. Caetano-Anollés, & P. M. Gresshoff (Eds.), DNA Markers: Protocols, Applications and Overviews. New York, NY: John Wiley & Sons, Inc.

Rajwar, A., & Sahgal, M. (2016). Phylogenetic relationships of fluorescent pseudomonads deduced from the sequence analysis of 16S rRNA, Pseudomonas-specific and rpoD genes. 3 Biotech. 6, e80.

Ramos, C., Matas, I. M., Bardaji, L., Aragon, I. M., & Murillo, J. (2012). Pseudomonas savastanoi pv. savastanoi: some like it knot. Molecular Plant Pathology, 13, 998-1009.

Renesto, P., Lorvellec-Guillon, K., Drancourt, M., & Raoult, D. (2000). rpoB gene analysis as a novel strategy for identification of spirochetes from the genera Borrelia, Treponema, and Leptospira. Journal of Clinical Microbiology, 38(6), 2200-2203.

Rodriguez-Moreno, L., Jimenez, A. J., & Ramos, C. (2009). Endopathogenic lifestyle of Pseudomonas savastanoi pv. savastanoi in olive knots. Microbial Biotechnology, 2, 476-88.

Saitou, N., & Nei, M. (1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees. Journal of Molecular Evolution, 4, 406-425.

Schaad, N. W., Jones, J. B., & Chun, W. (2001). Laboratory Guide for Identification of Plant Pathogenic Bacteria. 3th ed. Minnesota. MN: APS Press.

Sisto, A., Cipriani, M. G., & Morea, M. (2004). Knot formation caused by Pseudomonas savastanoi subsp. savastanoi on olive plants is hrp-dependent. Phytopathology, 94, 484-489.

Spaepen, S., Vanderleyden, J., & Remans, R. (2007). Indole-3-acetic acid in microbial and microorganism-plant signaling. FEMS Microbiology Reviews, 31, 425-448.

Taghavi, M., & Hasani, S. (2012). Occurrence of Pseudomonas savastanoi the causal agent of winter jasmine gall in Iran. Iran Agricultural Research, 31(1), 39-48.

Tamura, K., Dudley, J., Nei, M., & Kumar, S. (2007). MEGA4: molecular evolutionary genetics analysis (MEGA) software version 4.0. Molecular Biology and Evolution, 24, 1596-1599.

Tayeb, L. A., Ageron, E., Grimont, F., & Grimont, P. A. D. (2005). Molecular phylogeny of the genus Pseudomonas based on rpoB sequences and application for the identification of isolates. Research in Microbiology, 156, 763-773.

Tegli, S., Gori, A., Cerboneschi, M., & Grazia Cipriani, M. (2011). Type three secretion systems in Pseudomonas savastanoi pathovars: does timing matter? Genes, 2, 957-979.

Thompson, J. D., Higgins, D. G., & Gibson, T. J. (1994). CLUSTALW: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position specific gap penalties and weight matrix choice. Nucleic Acids Research, 22, 4673-4680.

Weisburg, W. G., Barns, S. M., Pelletier, D. A., & Lane, D. J. (1991). 16S ribosomal DNA amplification for phylogenetic study. Journal of Bacteriology, 173, 697-703.



1. 04. 2019



Agronomy section

How to Cite

KHEZRI, M., GHASEMI, A., & AHANGARAN, A. (2019). Detection and characterization of endophytic bacteria causing knot in young olive trees. Acta Agriculturae Slovenica, 113(1), 109–119.

Similar Articles

1-10 of 261

You may also start an advanced similarity search for this article.