Soil Biology & Biochemistry 39 (2007) 2150–2159 Pyruvate dehydrogenase activity is important for colonization of seeds and roots by Enterobacter cloacae Daniel P. Roberts a,Ã , Laurie F. McKenna a , Scott M. Lohrke a,1 , Stephen Rehner b , Jorge T. de Souza c a Sustainable Agricultural Systems Laboratory, USDA—Agricultural Research Service, Henry A. Wallace Beltsville Agricultural Research Center, Building 001, Rm. 140, 10300 Baltimore Avenue, Beltsville, MD 20705-2350, USA b Insect Biocontrol Laboratory, USDA—Agricultural Research Service, Henry A. Wallace Beltsville Agricultural Research Center, Beltsville, MD 20705-2350, USA c Universidade Federal do Recoˆncavo da Bahia, CCAAB, 44380-000 Cruz das Almas, BA, Brazil Received 23 December 2006; received in revised form 2 March 2007; accepted 23 March 2007 Available online 24 April 2007 Abstract Enterobacter cloacae is a plant-beneficial bacterium that shows promise for suppression of damping-off of cucumber and other crops caused by Pythium ultimum. We have been using a mutational approach to determine the E. cloacae genes important in bacterial–plant and bacterial–pathogen interactions in the spermosphere and rhizosphere. E. cloacae M43 is a transposon mutant of E. cloacae 501R3 that was significantly impaired in colonization of seeds and roots of diverse crop plants. Strain M43 did not increase in population on cucumber, sunflower, and wheat seeds and was significantly reduced in growth on pea seeds relative to strain 501R3. Populations of M43 were also dramatically lower than those of strain 501R3 in cucumber, pea, sunflower, and wheat rhizosphere in 42 d experiments. Molecular characterization of M43 demonstrated that there was a single transposon insertion in the genome of this strain and that this insertion was in a region of the E. cloacae genome with a high degree of DNA sequence identity with aceF. aceF encodes the dihydrolipoamide acetyltransferase subunit of the pyruvate dehydrogenase complex (PDHC). Cell lysates from strain 501R3 grown on minimal medium plus 50 mM glycerol and 2 mM acetate contained 0.01170.0036 U pyruvate dehydrogenase activity while cell lysates from M43 grown under identical conditions contained no detectable pyruvate dehydrogenase activity. Additionally, the nutritional use profile of M43 under aerobic and anaerobic conditions was as expected for an ace mutant. Experiments reported here strongly suggest a role for aceF and the PDHC in colonization of seeds and roots of diverse crop plants by E. cloacae. r 2007 Elsevier Ltd. All rights reserved. Keywords: Biological control; Colonization; Rhizosphere; Spermosphere 1. Introduction Beneficial bacteria applied as treatments of seeds and other plant parts have been shown to suppress plant diseases caused by seed, root, and foliar pathogens (Larkin et al., 1998; Weller, 1988; Meyer and Roberts, 2002). Colonization of the spermosphere and rhizosphere is thought to be important for effective suppression of many of these pathogens (Weller, 1988; Bull et al., 1991; Chin-A- Woeng et al., 2000; Lugtenberg et al., 2001). A number of traits, including growth by beneficial bacteria, have been shown to be important for colonization (Mazzola et al., 1992; Rainey, 1999; Lugtenberg et al., 2001; de Weert et al., 2002; Lohrke et al., 2002; Martı´nez-Granero et al., 2005). Seeds and roots support growth and other activities by microbes through the release of a complex mixture of carbohydrates, amino acids, organic acids, and other nutrients (Kraffczyk et al., 1984; Lynch and Whipps, 1990). However, there is a significant gap in our under- standing of the roles played by genes and catabolic pathways functioning in beneficial bacteria, and the nutrients released by plant seeds and roots during colonization (Roberts et al., 1999; Lohrke et al., 2002). ARTICLE IN PRESS www.elsevier.com/locate/soilbio 0038-0717/$ - see front matter r 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.soilbio.2007.03.027 Ã Corresponding author. Tel.: +1 301 504 5680; fax: +1 301 504 8370. E-mail address: dan.roberts@ars.usda.gov (D.P. Roberts). 1 Current address: Geo-Centers, Inc., Naval Health Research Center, Environmental Health Effects Laboratory, Wright-Patterson Air Force Base, Ohio, USA.