ORIGINAL ARTICLE Glycine betaine improves Listeria monocytogenes tolerance to desiccation on parsley leaves independent of the osmolyte transporters BetL, Gbu and OpuC N. Dreux 1 , C. Albagnac 1 , R.D. Sleator 3 , C. Hill 3 , F. Carlin 1 , C.E. Morris 2 and C. Nguyen-the 1 1 INRA, UMR 408, Se ´ curite ´ et Qualite ´ des Produits d’Origine Ve ´ ge ´ tale, Avignon, F-84914, France, Univ Avignon, Avignon, F-84029, France 2 INRA, Pathologie Ve ´ ge ´ tale, Domaine Saint Maurice, Montfavet, F-84140, France 3 Department of Microbiology and Alimentary Pharmabiotic Centre, University College Cork, Ireland Introduction The increase in the number of food-borne bacterial out- breaks linked to fresh fruits and vegetables over the past decade has raised concerns over the role of plants in pathogen transmission (Brandl 2006). During their resi- dence time on plants, bacterial pathogens undergo harsh physicochemical conditions that fluctuate widely and rap- idly over short periods of time (Hirano and Upper 2000). For example, aerial plant surfaces, which are poor in nutrients (Lindow and Brandl 2003), are subjected to changes in temperature, relative humidity (RH) and radi- ation that occur on time scales of seconds to hours (Hirano and Upper 2000). The survival of bacterial cells on aerial plant surfaces is therefore linked to their ability to tolerate stress, in particular, desiccation caused by low RH. A characteristic feature of the food-borne pathogen Listeria monocytogenes is its ability to survive and even proliferate under a variety of hostile environmental condi- tions, in particular, elevated osmolarity (upto 10% NaCl) and reduced temperatures (as low as )0Æ1°C) (McClure et al. 1989). This growth and survival under stressful conditions is attributed mainly to the accumulation of compatible solutes, particularly glycine betaine (GB), carnitine and proline (Sleator and Hill 1999). Studies with L. monocytogenes have shown that GB is primarily transported via a secondary transporter BetL and an ATP-binding cassette (ABC) transporter GbuABC. The Keywords compatible solutes, desiccation, fresh produce, glycine betaine, Listeria monocytogenes, relative humidity. Correspondence Nicolas Dreux, INRA, UMR 408, site Agroparc, 84914 Avignon cedex 9, France. E-mail: ndreux@avignon.inra.fr 2007 ⁄ 0629: received 20 April 2007, revised 9 August 2007 and accepted 14 September 2007 doi:10.1111/j.1365-2672.2007.03623.x Abstract Aims: To investigate the effect of glycine betaine (GB) on the survival of Liste- ria monocytogenes on leaf surfaces under low relative humidity (RH). Methods and Results: The addition of GB (‡25 mmol l )1 ) improved the sur- vival of L. monocytogenes under low RH on parsley leaves, thus suggesting that GB can improve the tolerance of L. monocytogenes to desiccation. Ten times less GB was needed to improve L. monocytogenes survival under low RH on nonbiological surfaces compared with parsley leaves, suggesting that, on the leaf surface, L. monocytogenes may have to compete for the available GB with autochthonous bacteria and ⁄ or the plant itself. Wild type and mutants carrying deletions in the three GB uptake systems, BetL, Gbu and OpuC, behaved simi- larly with and without added GB on parsley leaves (P >0Æ05). In addition, pre- accumulation of GB, triggered by osmotic stress prior to inoculation, failed to improve survival under low RH compared with osmotic stress without GB accumulation. Conclusions: Exogenous GB had a protective effect on L. monocytogenes cells from desiccation during survival on parsley leaves. This effect was independent of intracellular GB accumulation by the known uptake systems. Significance and Impact of the Study: Presence of GB could improve the survival of L. monocytogenes to desiccation on leaf surfaces and nonbiological surfaces. Journal of Applied Microbiology ISSN 1364-5072 ª 2007 The Authors Journal compilation ª 2007 The Society for Applied Microbiology, Journal of Applied Microbiology 104 (2008) 1221–1227 1221