Electrolyzed sodium bicarbonate inhibits Penicillium digitatum and induces defence responses against green mould in citrus fruit Frida Fallanaj a , Antonio Ippolito a , Angela Ligorio a , Francesca Garganese a , Ciro Zavanella b , Simona M. Sanzani a, * a Dipartimento di Scienze del Suolo, della Pianta e degli Alimenti, Università degli Studi Aldo Moro, Via Amendola 165/A, 70126 Bari, Italy b Aqanat Limited, Clarence House, Thirsk Bank, Coxwold, York, UK A R T I C L E I N F O Article history: Received 18 September 2015 Received in revised form 1 December 2015 Accepted 9 December 2015 Available online xxx Keywords: Electrolysis Sodium bicarbonate Citrus Penicillium digitatum Induction of resistance A B S T R A C T In previous investigations, electrolyzed sodium bicarbonate (eNaHCO 3 ) proved to be an effective mean for controlling postharvest rots of citrus fruit. However, little is known about its mode of action. In the present study, the direct effect of eNaHCO 3 on Penicillium digitatum and the ability to induce citrus defence mechanisms were investigated. Results indicated that eNaHCO 3 reduced conidia germination and germ tube elongation of P. digitatum by 80% within 15 min of treatment, as compared to untreated control; electrolyzed water (ew) alone or non-electrolyzed NaHCO 3 showed a less marked activity . After 45 min, eNaHCO 3 almost completely suppressed spore germination. Moreover, eNaHCO 3 triggered the accumulation of reactive oxygen species (ROS), causing an oxidative stress in P. digitatum conidia, followed by a collapse of mitochondrial membrane potential and a decrease in intracellular ATP. Results of specific assays on citrus fruit showed that eNaHCO 3 was able to control P. digitatum infections when applied in wounds nearby those inoculated with the pathogen, supporting the induction of host resistance as a possible mode of action. This hypothesis was confirmed by the up-regulation of defence- related genes coding chitinase, peroxidase, and phenylalanine ammonia-lyase (PAL) at 6–12 h post- treatment (hpt). As further confirmation, eNaHCO 3 proved to increase the activity of related enzymes and of b-1,3-glucanase that, being observed at 12 hpt, suggested an early host response against pathogen by limiting tissue colonization. Concluding, both the direct inhibition of P. digitatum and the induction of fruit resistance should be considered important aspects of the multiple mode of action of eNaHCO 3 in controlling postharvest citrus rots. ã 2015 Elsevier B.V. All rights reserved. 1. Introduction Citrus is one of the most important crops in many countries, widely cultivated in the Mediterranean area. During postharvest phase fruit are susceptible to several diseases, among which green mould, caused by Penicillium digitatum (Pers.) Sacc., is one of the most serious, causing up to 90% of product losses (Macarisin et al., 2007). P. digitatum is a wound parasite, infecting fruit in the orchard, the packinghouse, and during storage, distribution and marketing, by disseminating its spores in the air (Palou et al., 2008). The control of postharvest diseases is vital for maintaining citrus quality, especially in a global market in which transport from producer to consumer may take several weeks. Although synthetic fungicides, such as imazalil, thiabendazole, pyrimethanil, fludioxonil, are mostly used to minimize postharvest decay (Kanetis et al., 2007), their extensive application has led to the proliferation of resistant strains, which compromised their effectiveness. Moreover, several other issues are associated with their use, such as risks for human and environmental health, and costs for registration/re-registration of active ingredients (Sanzani et al., 2012). Therefore, the demand for alternative control strategies is increasing. Among various non-chemical approaches, the application of physical means (Droby et al., 1993; Palou, 2009), Generally Regarded as Safe (GRAS) compounds (Youssef et al., 2014; Lachhab et al., 2014; Sanzani et al., 2014), or antagonistic microorganisms (Droby et al., 2009; Lima et al., 2015) appeared to be effective to some extent. Among them, electrolyzed water is widely used in human medicine as topical disinfectant (Tanaka et al., 1996), and for sanitation in aquaculture (Whangchai et al., 2003). Moreover, it has been also tested in food industrial processes (Kim et al., 2000), and against postharvest decay of fresh fruits and vegetables (Al-Haq et al., 2002; Cui et al., 2009; * Corresponding author. E-mail address: simonamarianna.sanzani@uniba.it (S.M. Sanzani). http://dx.doi.org/10.1016/j.postharvbio.2015.12.009 0925-5214/ ã 2015 Elsevier B.V. All rights reserved. Postharvest Biology and Technology 115 (2016) 18–29 Contents lists available at ScienceDirect Postharvest Biology and Technology journal home page: www.elsevier.com/locat e/postharvbio