| Cotton verticillium wilt caused by Verticillum dahliae is one of soil- borne diseases which is difficult to be controlled in the world. Microbial pesticides is one of the effective measures to control crop soil-borne diseases. Bacillus subtillis strain NCD-2 can effectively suppress cotton verticillium wilt in the field, and the rhizosphere colonization is the prerequisite for effective suppression against verticillium wilt. For bacteria, the colonization ability is related to its biofilm formation ability. Plant roots exudates includes rich organic material, not only provide nutrients for growth of microorganism, but also serve as signal molecules to regulate the function of rhizosphere microorganisms.Bacillus subtilis strain NCD-2 is an excellent biocontrol agent for cotton soil–borne diseases. Previous studies showed that the biocontrol efficiency of strain NCD-2 was associated with the root colonization and biofiom formation capabilities. To clarify the function of cotton root exudates on the biofilm formation in strain NCD-2, the root colonization capabilities of strain NCD-2 was firstly tested on cotton cultivars Jimian 11, Zhongmiansuo 41, Zhongzhimian 2, Lumian 29, and Pima 90. Results indicated that strain NCD-2 had different population densities in the rhizospheres of tested cultivars. The highest population density of 7.63×105 CFU/g root was achieved on cultivar Pima 90 35 d after seeding, however, the lowest population density of 6.51×104 CFU/g root was achieved on cultivar Zhongzhimian 2 35 d after seeding.Cotton root exudates were obtained using recycling hydroponic system, and the amino acides composition were analyzed by high performance liquid chromatograph(HPLC). Results showed that the kinds of amino acids are accordant, but the concentration of amino acids is different in the tested five cotton varieties. Arginine, tyrosine, valine, lysine, isoleucine and phenylalanine are the highest levels in Jimian 11 root exudates. Threonine, proline and cysteine are the highest levels in Pima 90 root exudates. While alanine and leucine are the highest levels in Lumian 29 root exudates. Methionine is the highest levels in Zhongzhimian 2 root exudates.The root exudates were collected from the five cotton cultivars and tested for the promoted abilities on the biofilm formation of strain NCD-2, respectively. Results showed that the root exudates from tested varieties could promote the biofilm formation of strain NCD-2, the root exudates from Pima90 had the strongest effect on the biofilm formation, but the root exudates from Zhongtzhimian 2 only had a slight promote effect on the biofilm formation. Six carbohydrates, 13 amino acids and 3 organic acids that existed in the cotton root exudates were selected to evaluate the promote effect on the biofilm formation of strain NCD-2. Results revealed that the glucosan, proline and malic acid had the strongest effect on the biofilm formation compared to other carbohydrate and amino acids, respectively.Chemotaxis of strain NCD-2 on different cotton root exudates were tested, and results showed that the strongest chemotaxis was achieved by root exudate from Jimian 11, and the chemotactic distance was 2.10 cm. The weakest chemotaxis of strain NCD-2 was observed on Zhongmiansuo 41, and the chemotaxis distance was only 0.04 cm. The chemotaxis of strain NCD-2 were also tested for malic acid, citric acid, succinic acid, and results indicated that the strongest chemotaxis of strain NCD-2 was achieved by malic acid, and the chemotaxis distance could reach 3.67 cm. Chemotaxis of strain NCD-2 were also tested on 15 amino acids, results showed that the chemotaxis of strain NCD-2 on arginine, alanine, lysine were obvious and the chemotaxis distance was 3.20, 3.00, 3.00 cm, respectively. In addition, the negative chemotaxis of strain NCD-2 were observed on glycine and tryptophan. The chemotaxis of strain NCD-2 were also tested on glucose, sucrose, lactose, glucan, maltose, mannose and fructose, and results indicted that the chemotaxis of strain NCD-2 were relatively weak on the test sugars.Swimming motility and twitching motility of strain NCD-2 were also tested on different compounds such as amino acids, sugars and organic acids. Results showed that proline, malic acid and glucose could promote the swimming motility of strain NCD-2 obviously, and the colony diameter was 8.00, 8.00, 7.63 cm, respectively. Valine, fructose and malic acid could promote the twitching motility of strain NCD-2 obviously, and the colony diameter was 8.00, 5.53, 5.50 cm, respectively. We also found that the glutamic acid can inhibit the growth of strain NCD-2.We also tested the controlling efficacy for verticillium wilt by strain NCD-2 on five cotton varieties. Results revealed that the best controlling efficacy of strain NCD-2 was achieved on Jimian 11 variety and the controlling efficacy reached 66.68%. The controlling efficacy of strain NCD-2 on cotton verticillium wilt of Pima 90 is poor. We also have found that the controlling efficacy of strain NCD-2 on cotton verticillium wilt of Zhongmiansuo 41 and Zhongzhimian 2 is relatively stable.To conclusion, the colonization ability of strain NCD-2 is better on Jimian 11 and Pima 90 than the others. The root exudates of Jimian 11 could promote biofilm formation of strain NCD-2, and the best controlling efficacy of strain NCD-2 on cotton verticillium wilt was achieved on Jimian 11. Proline, arginine and malic acid have significantly effect on biofilm formation, chemotaxis, swimming motility and twitching motility of strain NCD-2, and sugars have no obvious effect. We also found that the proline is the highest levels in Pima 90 and arginine is the highest levels in Jimian 11. The results provide a foundation for the application of strain NCD-2 on different cotton varieties, and provide a scientific basis for studying the bio-control bacteria to prevent soil borne disease mechanism. |
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