European Journal of Plant Pathology

, Volume 127, Issue 4, pp 571–578 | Cite as

Detached leaf inoculation of germplasm for rapid screening of resistance to citrus canker and citrus bacterial spot



A detached leaf protocol for rapid screening of germplasm for resistance to citrus canker (Xanthomonas citri subsp. citri, Xcc) and citrus bacterial spot (Xanthomonas alfalfae subsp. citrumelonis, Xac) was developed to evaluate limited quantities of leaf material. Bacterial inocula of Xcc or Xac at 104, 105, or 108 cfu ml−1 were injection-infiltrated into the abaxial surface of disinfested, immature leaves of susceptible and resistant genotypes. Inoculated detached leaves were placed on the surface of 0.5% water agar plates and incubated at 28°C under a 12 h photoperiod. Likewise, inocula were infiltrated into attached leaves of greenhouse plants. At high inoculum concentrations of Xcc or Xac (108 cfu ml−1), resistant cultivars of kumquat developed a hypersensitive-like reaction within 3 days post inoculation (dpi). At 105 cfu ml−1, populations 14 dpi were <104 per inoculation site. In canker-susceptible Citrus spp. (‘Duncan’ grapefruit and ‘Rough’ lemon), water-soaked areas occurred by 3 dpi and typical canker lesions developed by 7 to14 dpi. Concentration of Xcc recovered from inoculation sites was approximately 105 cfu ml−1 by 14 dpi. In citrus bacterial spot-susceptible citrus (‘Swingle’ citrumelo and grapefruit), symptoms developed within 7 dpi. Populations of Xac after inoculation at 105 cfu ml−1 were comparable to Xcc in susceptible hosts 14 dpi (>105). The detached leaf assay is useful for the characterization and differentiation of lesion phenotype for each Xanthomonas pathogen permitting rapid screening of germplasm resistance based on the quantification of number of lesions and bacterial concentration.


Citrus germplasm screening Bacterial disease resistance Xanthomonas alfalfae subsp. citrumelonis Xanthomonas citri subsp. citri 


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Copyright information

© KNPV 2010

Authors and Affiliations

  • Marta I. Francis
    • 1
  • Alma Peña
    • 1
  • James H. Graham
    • 1
  1. 1.IFAS, Department of Plant Pathology, Citrus Research and Education CenterUniversity of FloridaLake AlfredUSA

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