Annals of Microbiology, 57 (3) 337-339 (2007) First report on the use of moderately halophilic bacteria against stem canker of greenhouse tomatoes caused by Botrytis cinerea Najla SADFI-ZOUAOUI 1 *, Badiâa ESSGHAIER 1 , Ibtissem HANNACHI 1 , Mohamed Rabeh HAJLAOUI 2 , Abdellatif BOUDABOUS 1 1 Laboratoire Microorganismes et Biomolécules Actives, Faculté des Sciences de Tunis, Campus Universitaire, 2092 Tunisie; 2 Laboratoire de Protection des Végétaux, Institut National de la Recherche Agronomique de Tunisie (INRAT), 2049 Ariana, Tunisie Received 25 January 2007 / Accepted 28 May 2007 Abstract - Experiments were conducted to study the effect of moderately halophilic bacteria isolated from different Tunisian Sebkhas (hypersaline soils), on stem canker caused by Botrytis cinerea on tomato plants grown under greenhouse conditions. Treatments per- formed with moderately halophilic isolates of Bacillus subtilis J9 and Halomonas sp. K2-5 significantly reduced stem lesion expansion by B. cinerea on tomato plants under greenhouse conditions. The use of such bacteria may constitute an important alternative to syn- thetic fungicides, which failed to suppress the development of the fungal pathogen. Key words: biocontrol, halophilic bacteria, Botrytis cinerea, tomato. INTRODUCTION Botrytis cinerea Pers.: Fr. is the causal agent of stem canker of tomato (Menzies and Jarvis, 1994; Elad, 2004), a severe disease particularly prominent in Tunisia, espe- cially under greenhouse protected conditions (Hmouni et al., 1996). The fungal pathogen infects stems, flowers and fruits by direct penetration or through wounds caused by cultivation practices (Peng et al., 1996; Elad, 2004). Pruning wounds infected by germinated spores of B. cinerea often develop stem cankers, which may eventually girdle the stem, resulting in plant death (Nicot and Baille, 1996). Recent laboratory studies showed that B. cinerea was highly resistant towards dicarboximide and benzimida- zoles, the main anti-Botrytis fungicides used in Tunisia. Furthermore, several investigations conducted all over the world, indicated that chemical control against grey mould of tomatoes was not effective due to the development of resistant strains (Locke and Fletcher, 1988; Yourman and Jeffers, 1999; Rosslenbroich and Stuebler, 2000). Alternative methods for suppressing B. cinerea are there- fore needed for commercial growers. Biological control using natural antagonistic microorganisms has been exten- sively studied, and some fungi and bacteria have been demonstrated to be effective against grey mold (Nicot et al., 1993; Sutton and Peng, 1993; Elad et al., 1994; Dick and Elad, 1999; Utkhede and Mathur, 2002; Lee et al., 2006). Biological control of stem canker of greenhouse tomatoes has been obtained using sprays of Rhodosporium diobovatum (strain S33) (Utkhede et al., 2001) and Trichoderma harzianum (Elad et al., 1995). Commercial biological products such as Prestop ® (Glicoladium catenu- latum J1446), RootShield ® , Trichodex ® (T. harzianum) and Quadra136 (Bacillus subtilis) prevented stem canker under near-commercial greenhouse conditions (Utkhede and Mathur, 2002, 2006). In recent studies, we described the isolation and identi- fication of moderately halophilic bacteria, which were shown to be highly effective agents against grey mold infection on fresh-market tomato fruits and strawberry fruits under storage (Essghaier et al., 2005; Sadfi-Zouaoui et al., 2007). Aim of this study was to evaluate the antagonistic effects of these selected isolates to suppress grey mold on stem tomato plants grown under greenhouse conditions. MATERIAL AND METHODS Tomato plants (hybrid F1 Maria) used in this study were grown in 25-cm diameter pots in a plastic house and Botrytis cinerea isolate BCt from tomato canker was used for inoculation. Eight weeks after transplanting, two lesions were made per plant by pruning petioles (about 0.5 cm long) and placing a 6-mm square agar plug from actively growing B. cinerea on PDA medium onto the resulting wound. The wounds were then covered with humidified cot- ton and sealed with parafilm to maintain high humidity and allow fungal infection. Five moderately halophilic isolates, Bacillus subtilis (M1-20, J9), Bacillus licheniformis (J24), Planococcus rifietoensis (M2-26) and Halomonas sp. (K2-5) * Corresponding author. Phone: 216-70-850553; Fax: 216-71-885480; E-mail: sadfi.najla@planet.tn