| Mulberry root rot caused by Lasiodiplodia theobromae was a new disease in China. In order to control the disease effectively, identification and genetic diversity of the pathogen, effects of light and dark on the growth of the pathogen, the occurrence regularity and control methods were studied and the main results were as follows:1. Twelve fungi were isolated from 870 specimens of diseased root collected from the diseased regions of Guangxi. Isolate rate of strain XY14 was 90% and the highest among the twelve fungi. The strain XY14 was identified as Lasiodiplodia theobromae (Pat.) Griffon & Maubl. based on the pathogenicity test, morphologic characteristics and the analysis of rDNA-ITS and EF1-α sequences. Mulberry root rot caused by Lasiodiplodia theobromae was firstly found in China.2. Light and dark had no significant effects on the growth of mycelia and the germination of conidia of L. theobromae based on growth rates of mycelia and germination rates of conidia under light and dark culture conditions. Continuous light was beneficial to the development of the conidia and pycnidia of L. theobromae. After two days of illumination, the bud of the fruit body could be found on the PDA plate and the fruit body gradually increased with the extension of the culture time.3. Genetic diversity of 29 L. theobromae strains from different hosts and geographic locations was studied by RAPD and ISSR molecular marked methods. Among those 29 strains,23 strains were the pathogen of mulberry Lasiodiplodia root rot respectively collected from Heng County, Yizhou County, Luzhai County and Xiangzhou County, and the other 6 strains were the pathogen of leaf spot of Agave sisalana Perr. ex Engelm collected from Wuming County in Guangxi. The results of two molecular marked methods showed that the average percentage of polymorphic loci of all tested strains was more than 93%. The genetic similarity coefficient of RAPD ranged from 0.61 to 0.92, and the genetic similarity coefficient of ISSR ranged from 0.69 to 0.99. Cluster phenograms of unweighted pair-group method with arithmetic average (UPGMA) for 29 strains were made by NTSYSpc (Version 2.10e) with RAPD and ISSR data and 29 strains could be divided into seven RAPD groups when the genetic similarity coefficient reached 0.76 and divided into five ISSR groups when the genetic similarity coefficient reached 0.746 by the UPGMA cluster dendrograms. Both the RAPD groups and the ISSR groups showed a certain degree of relationship with the geographical locations where those strains were collected. The results of two molecular marked methods indicated that the genetic similarity was high and the genetic distance was short among those Lasiodiplodia theobromae populations.4. The results of disease investigation indicated that the occurrence rate and severity of mulberry Lasiodiplodia root rot were concerned with the ages of mulberry trees, damages caused by meloidogyne and the varieties of mulberry trees. Whether young trees or adult trees could occur root rot, but the older the trees, the more serious the disease was. Almost all the diseased root had the cecidium (knot) of meloidogyne, but otherwise it was not. The incidence of mulberry Lasiodiplodia root rot of four to six-year trees of 11 varieties were investigated in Heng County, Yizhou County, Xiangzhou County and Luzhai County in Guangxi, and results showed that the incidence rate of the variety Sha 2><109 was the highest and the variety Kangqing 283 was the lowest.5. The antagonisms of Trichoderma atroviride ST-1, T. harzianum GZ-5, Bacillus subtilis Bll, B. amyloliquefaciens YZ14-3 and Pythium oligandrum against L. theobromae were studied. Results showed that five kinds of biocontrol microbes could inhibit the growth of mycelia of L. theobromae. The inhibitory rates of P. oligandrum, Bacillus subtilis Bll and B. amyloliquefaciens YZ14-3 were 88.9%,75.0% and 73.3%, respectively. The antagonistic coefficient grades of ST-1 and GZ-5 against L. theobromae were Ⅲ was found that the mycelia of P. oligandrum, ST-1 and GZ-5 tightly wrapped around the mycelia of L. theobromae and made it malformed and swelled; the mycelia contents of L. theobromae were bubble-like, malformed, easy to break after being treated with Bacillus subtilis Bll and B. amyloliquefaciens YZ14-3. The germination rates of conidia of L. theobromae in the filtrate of P. oligandrum, Trichoderma atroviride ST-1 and T. harzianum GZ-5 were 6%,20.4% and 12.2%, respectively. The conidia of L. theobromae did not germinate in the filtrate of B11 and YZ14-3, and the conidia of L. theobromae disintegrated due to the cell wall of the conidia being degraded.6. Effects of fresh juice, composted juice, boiled juice and sterile juice of leaf of 9 sisal species on the growth of mycelia and germination of conidia of L. theobromae were studied. Results showed that all the leaf juice inhibited the mycelial growth, the inhibitory rates were different, ranged from 63.4% to 100% related to sisal varieties and leaf juice treatments, the inhibition effect of 76416 sisal and Agave americana was the best with the inhibitory rates of four treatments (fresh juice, composted juice, boiled juice and sterile juice) were 100%, and followed by Agave amaniensis with the inhibitory rate 97.6%. The inhibition would drop down after fresh leaf juices of other 6 varieties were composted, boiled and sterilized.The treated mycelia of L. theobromae were malformed, swelled, broken and protoplasm leaked when observed under microscope. The inhibition of conidial germination of Lasiodiplodia theobromae by leaf juice of 9 sisal species was tested, it showed that, except A. amaniensis, H.11648 and A. angustifolia, the leaf juice of other six sisal species inhibited the conidia germinate absolutely. The average germination rates of A. amaniensis, H.11648 and A. angustifolia was only 72.4%,16.6% and 13%, respectively.7. The toxicity of 50% carbendazim WP,70% thiophanate methyl, Fenaminosulf WP, 10% Difenoconazole and 80% Dithane M-45 on L. theobromae was tested and showed that, the EC50 of five fungicides were 0.079μg/mL,0.789μg/mL,0.156μg/mL,0.165μg/mL and 42.463μg/mL, respectively. The germination rates of conidia in the EC50 of above five fungicides were 17%,37.8%,6.8%,86% and 28%.8. Field test of using T. harzianum GZ-5, P. oligandrum, B. amyloliquefaciens YZ14-3, fresh leaf juice of H.11648, Shigao (10% Difenoconazole),50% Carbendazim WP,70% Thiophanate methyl, Fenaminosulf WP and 80% Dithane M-45 to control mulberry Lasiodiplodia root rot was conducted. Result showed that, the control efficiency of Fenaminosulf WP was the highest (83.02%), followed by 50% Carbendazim WP with the control efficiency of 80.9% and the control efficiency of T. harzianum and fresh leaf juice of H.11648 was 75.62% and 74.02% respectively, significantly higher than the control efficiency of 70% Thiophanate methyl,80% Dithane M-45 and Shigao (10% Difenoconazole).Therefore, Fenaminosulf WP (600×),50% Carbendazim WP (200×), T. harzianum GZ-5 and leaf juice of H.11648 could be watered in the mulberry orchards to control the mulberry Lasiodiplodia root rot. |
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