Role of Soil, Crop Debris, and a Plant Pathogen in Salmonella enterica Contamination of Tomato Plants Jeri D. Barak*, Anita S. Liang Produce Safety and Microbiology Research Unit, Western Regional Research Center (WRRC), Agricultural Research Service (ARS), United States Department of Agriculture (USDA), Albany, California, United States of America Abstract Background: In the U.S., tomatoes have become the most implicated vehicle for produce-associated Salmonellosis with 12 outbreaks since 1998. Although unconfirmed, trace backs suggest pre-harvest contamination with Salmonella enterica. Routes of tomato crop contamination by S. enterica in the absence of direct artificial inoculation have not been investigated. Methodology/Principal Findings: This work examined the role of contaminated soil, the potential for crop debris to act as inoculum from one crop to the next, and any interaction between the seedbourne plant pathogen Xanthomonas campestris pv. vesicatoria and S. enterica on tomato plants. Our results show S. enterica can survive for up to six weeks in fallow soil with the ability to contaminate tomato plants. We found S. enterica can contaminate a subsequent crop via crop debris; however a fallow period between crop incorporation and subsequent seeding can affect contamination patterns. Throughout these studies, populations of S. enterica declined over time and there was no bacterial growth in either the phyllosphere or rhizoplane. The presence of X. campestris pv. vesicatoria on co-colonized tomato plants had no effect on the incidence of S. enterica tomato phyllosphere contamination. However, growth of S. enterica in the tomato phyllosphere occurred on co-colonized plants in the absence of plant disease. Conclusions/Significance: S. enterica contaminated soil can lead to contamination of the tomato phyllosphere. A six week lag period between soil contamination and tomato seeding did not deter subsequent crop contamination. In the absence of plant disease, presence of the bacterial plant pathogen, X. campestris pv. vesicatoria was beneficial to S. enterica allowing multiplication of the human pathogen population. Any event leading to soil contamination with S. enterica could pose a public health risk with subsequent tomato production, especially in areas prone to bacterial spot disease. Citation: Barak JD, Liang AS (2008) Role of Soil, Crop Debris, and a Plant Pathogen in Salmonella enterica Contamination of Tomato Plants. PLoS ONE 3(2): e1657. doi:10.1371/journal.pone.0001657 Editor: Ching-Hong Yang, University of Wisconsin-Milwaukee, United States of America Received November 29, 2007; Accepted January 29, 2008; Published February 27, 2008 This is an open-access article distributed under the terms of the Creative Commons Public Domain declaration which stipulates that, once placed in the public domain, this work may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. Funding: This work was supported by the U.S. Department of Agriculture (Agricultural Research Service CRIS project number 5325-42000-044-00D). Competing Interests: The authors have declared that no competing interests exist. *E-mail: jbarak@pw.usda.gov Introduction In recent years, tomatoes have been one of the most common vehicles of produce-associated Salmonellosis. In the United States, there have been 12 outbreaks caused by Salmonella enterica related to tomato consumption since 1998 [1,2]. The tomato contamination appears to originate from the fields where the tomatoes were grown and/or the packing sheds [3]. However, the route of contamination remains indefinable, although probable suspects exist: water, soil, animal waste, and insects [4]. It has been shown in a hydroponic system that S. enterica can contaminate entire tomato plants following direct root inoculations [5]. Furthermore, tomato fruits can be contaminated with S. enterica following direct flower inoculations [6]. These routes of inoculation probably do not reflect the natural contamination route of field grown tomatoes. However, the colonization or contamination of tomato plants in the absence of direct artificial inoculation has not been investigated thoroughly. S. enterica has been shown to contaminate carrots, radish, lettuce and parsley in field studies, following treatments with contami- nated manure compost or irrigation water; however, these crops were also directly contaminated [7,8]. We undertook this study to determine the ability of S. enterica to colonize or contaminate tomato plants via indirect contamination. We chose to limit our study to the contamination of tomato plants and survival of S. enterica in the phyllosphere and rhizoplane of plants preceding flower set. Other studies have addressed the contamination of tomato fruit following contamination of flowers and fruit in contact with contaminated soil [6,9]; however, examination of plant contamination by S. enterica through indirect inoculation paths prior to this developmental stage was nonexistent. We chose two paths for indirect contamination, soil contamination from an irrigation event with contaminated water or contaminated crop debris from a previous crop. Furthermore, we investigated the role of a plant pathogen in the survival and population size of S. enterica in both the phyllosphere and rhizoplane. Materials and Methods Bacterial Strains, Plasmid, and Growth Media S. enterica serovar Baildon strain 05x-02123 [2] and serovar Enteritidis strain 99A-23 (California Health Department, July PLoS ONE | www.plosone.org 1 February 2008 | Volume 3 | Issue 2 | e1657