solani was detected at all three stages of chrysanthemum development, but neverless at a lower intensity. Evidence of the presence of beneficial fungi in the rhizosphere was provided by the amplification of product from Chaetomium globosum (Band 12�C6) (Figure 2). The DGGE profiles indicated that the complexity and abundance of soil fungi was greater in the rhizosphere samples than in the bulk soil (Figure 2). A comparison between the two profiles suggested a level of similarity of 59% based on the UPGMA algorithm, and the recovery of the same fragment from duplicate samples showed that the DNA isolation, PCR, and electrophoretic procedures had all been reliable (Figure 3). Overall, Ascomycete species were the most abundant (68% of all identified species, Table 2), followed by Basidiomycetes (21%).
Figure 1The abundance of fungi in the rhizosphere and bulk soil, as indicated by a real time PCR estimation of the copy number of an 18S rDNA fragment. Standard error bars calculated from three replicates. Significant differences based on Student’s t-test indicated …Figure 2DGGE profiles of 18S rDNA fragments present in DNA extracted from bulk soil (lanes 1�C6) and rhizosphere (lanes 7�C12) sampled at various developmental stages of growing chrysanthemum plants. Lanes 1, 2, 7, and 8: seedling stage, lanes 3, …Figure 3Phylogeny of the soil microflora, derived from 18S rDNA DGGE profiles. No. 1�Cno. 6: bulk soil samples and no. 7�Cno. 12: rhizosphere samples. No. 1, no. 2, no. 7, and no. 8: soil from plants at the seedling stage, no. 3, no. 4, no. 9, and …
Table 2Most closely related sequences to those derived from selected 18S rDNA amplicons separated by DGGE.3.2. Analysis of Fungal DiversityOur results demonstrated that the fungi diversity in the rhizosphere soil was different from that in the bulk soil (Table 3). In the rhizosphere soil sampled from plants at the vegetative stage, S (35), H�� (3.48), and 1/D (29.53) were greater than in the bulk soil sampled from plants at the same stage (S = 20, H�� = 2.85, 1/D = 14.75). The value of S in the bulk soil was also lower than that in the rhizosphere soil at the seedling stage, but during the reproductive stage, S was higher in the bulk soil. However, the E parameter remained relatively constant throughout, lying in the range of 0.95�C0.99.
Table 3Diversity indices associated with the fungal flora present in the rhizosphere and bulk soil samples of continuously monocropped chrysanthemum.3.3. AV-951 The Isolation and Bioassay of Pathogens Isolated from Diseased ChrysanthemumAfter five days of in vitro culture, 15 fungal strains were isolated from various diseased plant tissues. On the basis of their ITS sequences, it was possible to identify that 11 of these 15 isolates shared 97% similarity with F. solani and the other four shared 98% similarity with F. oxysporum. One of the putative F.