Aim: To examine the biocontrol activity of broad-range antagonists Serratia plymuthica IC1270, Pseudomonas fluorescens Q8r1-96 and P. fluorescens B-4117 against tumourigenic strains of Agrobacterium tumefaciens and A. vitis. Methods and Results: Under greenhouse conditions, the antagonists, applied via root soak prior to injecting Agrobacterium strains into the wounded stems, significantly suppressed tumour development on tomato seedlings. A derivative of P. fluorescens Q8r1-96 tagged with a gfp reporter, as well as P. fluorescens B-4117 and S. plymuthica IC1270 marked with rifampicin resistance, stably persisted in tomato tissues for at least 1 month. Mutants of P. fluorescens Q8r1-96 and S. plymuthica IC1270 deficient in 2,4-diacetylphloroglucinol or pyrrolnitrin production, respectively, also proficiently suppressed the tumour development, indicating that these antibiotics are not responsible for the observed biocontrol effect on crown gall disease. The volatile organic compounds (VOCs) produced by the tested P. fluorescens and S. plymuthica strains inhibited the growth of A. tumefaciens and A. vitis strains in vitro. Solid-phase microextraction-gas chromatography-mass spectrometry analysis revealed dimethyl disulfide (DMDS) as the major headspace volatile produced by S. plymuthica IC1270; it strongly suppressed Agrobacterium growth in vitro and was emitted by tomato plants treated with S. plymuthica IC1270. 1-Undecene was the main volatile emitted by the examined P. fluorescens strains, with other volatiles, including DMDS, being detected in only relatively low quantities. Conclusions: S. plymuthica IC1270, P. fluorescens B-4117 and P. fluorescens Q8r1-96 can be used as novel biocontrol agents of pathogenic Agrobacterium. VOCs, and specifically DMDS, might be involved in the suppression of oncogenicity in tomato plants. However, the role of specific volatiles in the biocontrol activity remains to be elucidated. Significance and Impact of the Study: The advantage of applying these antagonists lies in their multiple activities against a number of plant pathogens, including Agrobacterium.

Three Enterobacter agglomerans strains which produce and excrete proteins with chitinolytic activity were found while screening soil-borne bacteria antagonistic to fungal plant pathogens. The chitinolytic activity was induced when the strains were grown in the presence of colloidal chitin as the sole carbon source. It was quantitated by using assays with chromogenic p-nitrophenyl analogs of disaccharide, trisaccharide, and tetrasaccharide derivatives of N-acetylglucosamine. A set of three fluorescent substrates with a 4-methylumbelliferyl group linked by ␤-1,4 linkage to N-acetylglucosamine mono-or oligosaccharides were used to identify the chitinolytic activities of proteins which had been renatured following their separation by electrophoresis. This study provides the most complete evidence for the presence of a complex of chitinolytic enzymes in Enterobacter strains. Four enzymes were detected: two N-acetyl-␤-D-glucosaminidases of 89 and 67 kDa, an endochitinase with an apparent molecular mass of 59 kDa, and a chitobiosidase of 50 kDa. The biocontrol ability of the chitinolytic strains was demonstrated under greenhouse conditions. The bacteria decreased the incidence of disease caused by Rhizoctonia solani in cotton by 64 to 86%. Two Tn5 mutants of one of the isolates, which were deficient in chitinolytic activity, were unable to protect plants against the disease.

1-aminocyclopropane-1-carboxylate (ACC) deaminase activity was evaluated in the biocontrol and plant growth-promoting fungus Trichoderma asperellum T203. Fungal cultures grown with ACC as the sole nitrogen source showed high enzymatic activity. The enzyme encoding gene (Tas-acdS) was isolated, and an average 3.5-fold induction of the gene by 3 mM ACC was detected by real-time PCR. Escherichia coli bacteria carrying the intron-free cDNA of Tas-acdS cloned into the vector pAlter-EX1 under the control of the tac promoter revealed specific ACC deaminase (ACCD) activity and the ability to promote canola (Brassica napus) root elongation in pouch assays. RNAi silencing of the ACCD gene in T. asperellum showed decreased ability of the mutants to promote root elongation of canola seedlings. These data suggest a role for ACCD in the plant root growth-promotion effect by T. asperellum.

In previous research, volatile organic compounds (VOCs) emitted by various bacteria into the chemosphere were suggested to play a significant role in the antagonistic interactions between microorganisms occupying the same ecological niche and between bacteria and target eukaryotes. Moreover, a number of volatiles released by bacteria were reported to suppress quorum-sensing cell-to-cell communication in bacteria, and to stimulate plant growth. Here, volatiles produced by Pseudomonas and Serratia strains isolated mainly from the soil or rhizosphere exhibited bacteriostatic action on phytopathogenic Agrobacterium tumefaciens and fungi and demonstrated a killing effect on cyanobacteria, flies (Drosophila melanogaster), and nematodes (Caenorhabditis elegans). VOCs emitted by the rhizospheric Pseudomonas chlororaphis strain 449 and by Serratia proteamaculans strain 94 isolated from spoiled meat were identified using gas chromatography-mass spectrometry analysis, and the effects of the main headspace compounds—ketones (2-nonanone, 2-heptanone, 2-undecanone) and dimethyl disulfide—were inhibitory toward the tested microorganisms, nematodes, and flies. The data confirmed the role of bacterial volatiles as important compounds involved in interactions between organisms under natural ecological conditions.

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