Environmental and Experimental Botany 71 (2011) 41–49 Contents lists available at ScienceDirect Environmental and Experimental Botany journal homepage: www.elsevier.com/locate/envexpbot Antioxidant metabolism of grapefruit infected with Xanthomonas axonopodis pv. citri Naveen Kumar, Robert C. Ebel ∗ , Pamela D. Roberts Southwest Florida Research and Education Center, Institute of Food and Agriculture Sciences, University of Florida, Immokalee, FL 34142, USA article info Article history: Received 5 May 2010 Received in revised form 19 October 2010 Accepted 23 October 2010 Keywords: Citrus paradisi Ascorbate peroxidase Catalase Peroxidase Reactive oxygen species abstract Grapefruit is one of the most susceptible citrus genotypes to Asiatic Citrus Canker, caused by Xanthomonas axonopodis pv. citri (Xac), that can cause severe losses in citrus yield and quality. Although much is known about citrus response to Xac, little is known of the role of antioxidant metabolism. Grapefruit leaves were artificially injected with a strain of Xac obtained from a commercial grove in Florida and components of oxidative metabolism were measured. Symptoms observed included water soaking (2 dai; days after inoculation), raised and ruptured epidermis (6–8 dai), formation of necrotic lesions (16 dai), and leaf abscission (21 dai). The Xac population increased to a maximum (≈10 9 CFU/cm 2 ) 8 dai and then declined to ≈10 7 CFU/cm 2 by 20 dai. Lipid peroxidation was higher in infected leaves than uninoculated controls from 4 to 21 dai indicating greater oxidative stress. H 2 O 2 concentration demonstrated a biphasic pattern with peak concentrations at 4 and 13 dai and minimum concentrations that were lower than the controls at 10 and 20 dai. The H 2 O 2 concentration somewhat corresponded with superoxide dismutase (SOD) activity, which generates H 2 O 2 via dismutase of superoxide ions. Total SOD activity in Xac-infected leaves increased to a maximum at 4 dai, the day of highest H 2 O 2 concentration, and then declined and remained at or below controls. Mn-SOD and Fe-SOD activities both increased to maximum activities at 4 dai. Mn- SOD had four isoforms in Xac-infected leaves but only three in the controls. Fe-SOD had three isoforms in both infected and control plants. Suppression of H 2 O 2 in Xac-infected leaves also corresponded to higher activities of the H 2 O 2 catabolising enzymes catalase (CAT), ascorbate peroxidase (APOD), and peroxidase (POD). Two additional CAT isoforms were detected in infected leaves and not the controls. Three POD isoforms were detected in both control and infected leaves. Previous research has shown that Xac is sensitive to intraplant H 2 O 2 concentration, however, the pattern of Xac in this study did not correspond to H 2 O 2 concentration, which initially increased due to enhanced SOD activity, but was later suppressed apparently with the aid of peroxidases. In conclusion, Xac infection altered H 2 O 2 metabolism in grapefruit leaves by changes in the activities and isoforms of SODs, CATs, PODs and APOD. Published by Elsevier B.V. 1. Introduction Grapefruit (Citrus paradise Macfad.) is one of the important crops worldwide (Talon and Gmitter, 2008) and is one of the most impor- tant fresh fruit citrus in Florida (Francis et al., 2009). Xanthomonas axonopodis pv. citri (Xac) causes one of the most devastating dis- eases of citrus, with grapefruit being one of the most susceptible genotypes (Francis et al., 2009). Xac occurs in 30 countries including the U.S. (Del Campo et al., 2009), and outbreaks in Florida con- tributed to a decline in acreage by 2009 to 61% of the acreage in 1994 (Anon., 2009). Xac dispersal is enhanced by wind and rain and enters through wounds and stomates (Brunings and Gabriel, 2003). Canker devel- opment is governed by a sequence of systematic and coordinated ∗ Corresponding author. Tel.: +1 239 658 3400; fax: +1 239 658 3469. E-mail addresses: rcebel@ufl.edu, bob ebel@yahoo.com (R.C. Ebel). events (Brunings and Gabriel, 2003). Following artificial inocu- lation: (I) 1 day after inoculation (dai) bacterial cells occupied intercellular spaces (Al-Mousawi et al., 1982). (II) 2 dai the type III secretion system (T3S) attaches to mesophyll cells by hrp (hyper- sensitivity response and pathogenicity) pili or by type IV pili as observed during Xanthomonas pv. malvacearum–Gossypium hirsu- tum interaction (Brunings and Gabriel, 2003 and within). After the T3S is formed, effector proteins pthA (pathogenicity), Avr (avir- ulence), and Pop (pseudomonas outer protein) in turn promote hypertrophy (cell elongation) and hyperplasia (cell division) in host tissue (Brunings and Gabriel, 2003 and within). (III) 3 dai, meso- phyll cells divided and enlarge reaching a maximum at 7 dai, which coincided the highest bacterial population 10 9 CFU/cm 2 (Khalaf et al., 2007). (IV) 4 dai the infected region developed water soak- ing. (V) 8 dai the epidermis was raised and ruptured (Brunings and Gabriel, 2003). (VI) 10–14 dai and beyond tissue necrosis devel- oped (Duan et al., 1999). (VII) 21 dai leaves abscised (Khalaf et al., 2007). 0098-8472/$ – see front matter. Published by Elsevier B.V. doi:10.1016/j.envexpbot.2010.10.019