Endophytes play an important role in protection of host plants from infection by phytopathogens. Endophytic bacteria were isolated from five different parts (root, stem, petiole, leaf and seed) of Panax notoginseng and evaluated for antagonistic activity against Fusarium oxysporum, Ralstonia sp. and Meloidogyne hapla, three major pathogens associated with root-rot disease complex of P. notoginseng. From 1000 endophytic bacterial strains evaluated in vitro, 104 strains exhibited antagonistic properties against at least one of these three pathogens. Phylogenetic analyses of their 16S rRNA gene sequences showed that these 104 antagonistic bacteria belong to four clusters: Firmicutes, Proteobacteria, Actinobacteria and Bacteroidetes/Chlorobi. Members of the Firmicutes, in particular the Bacillus spp., were predominant in all analyzed tissues. The root was the main reservoir for antagonistic bacteria. Of the 104 antagonists, 51 strains showed antagonistic activities to one pathogen only, while 43 and 10 displayed the activities towards two and all three pathogens, respectively. The most dominant species in all tissues were Bacillus amyloliquefaciens subsp. plantarum and Bacillus methylotrophicus, which were represented by eight strains with broad antagonistic spectrum to the all three test pathogens of root-rot disease complex of P. notoginseng.

The chemotactic response of bacteria to root exudates plays an important role in the colonization of bacteria in the rhizosphere. In this study, 420 strains of antifungal bacteria against Fusarium oxysporum f. sp. cubense (Foc) were screened for chemotaxis based on a cheA molecular diagnostic method. A total of 124 strains with antifungal efficiencies of 27.26-67.14 % generated a characteristic band of cheA. The chemotaxis of 97 bacterial strains producing a cheA band was confirmed using the drop assay and swarm plate assay using catechol, p-hydroxybenzoic acid, salicylic acid, and asparagine as the attractants. A phylogenetic analysis based on restriction fragment length polymorphisms (RFLPs) and 16S rDNA sequences indicated that the 124 chemotactic antagonists of Foc were affiliated with 18 species of Paenibacillaceae, Bacillaceae, Streptomycineae, Enterobacteriaceae, and Pseudomonadaceae. The chemical composition of banana root exudates were analyzed by GC-MS, and 62 compounds, including alkanes, alkenes, naphthalenes, benzenes, and alcohols, were evaluated. Five representative antagonists of Foc showed 1.76- to 7.75-fold higher chemotactic responses than the control to seven compounds in banana root exudates, as determination by capillary assays.

Nineteen DAB species were demonstrated to be antagonists against the M. incognita-P. nicotianae complex. Because S. flavofungini SNA26, P. putida SNB53 and S. marcescens subsp. sakuensis SNB54 significantly suppressed the infection of M. incognita and P. nicotianae in tobacco, these species have potential for development as biocontrol agents against the diseases and complex caused by these two pathogens.

Accumulated evidence suggests that root exudates have a major role in mediating plant-microbe interactions in the rhizosphere. Here, we characterized tobacco root exudates (TREs) by GC-MS and nicotine, scopoletin, and octadecane were identified as three main components of TREs. Qualitative and quantitative chemotaxis assays revealed that Pseudomonas aeruginosa NXHG29 with antagonistic activity displayed positive chemotactic responses towards TREs and their three main components (nicotine, scopoletin, octadecane) and its enhanced chemotaxis were induced by these substances in a concentration-dependent manner. Furthermore, following GC-MS and chemotaxis analysis, nicotine was selected as the target for evaluation of the effect on NXHG29 regarding antagonism, growth, root colonization and biocontrol efficiency. Results of in vitro studies showed that nicotine as a sole carbon source could enhance growth of NXHG29 and significantly increased the antagonism of NXHG29. We also demonstrated that nicotine exerted enhancing effects on the colonization ability of NXHG29 on tobacco roots by combining CLSM observations with investigation of population level dynamics by selective dilution plating method. Results from greenhouse experiments suggested nicotine exhibited stimulatory effects on the biocontrol efficiency of NXHG29 against bacterial wilt and black shank on tobacco. The stimulatory effect of nicotine was affected by the concentration and timing of nicotine application and further supported by the results of population level of NXHG29 on tobacco roots. This is the first report on the enhancement effect of nicotine from TREs on an antagonistic bacterium for its root colonization, control of soil-borne pathogens, regarding the chemotaxis and in vitro antagonism and growth.

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