Degenerated virulence and irregular development of Fusarium oxysporum f. sp. niveum induced by successive subculture

Successive cultivation of fungus has been reported to cause the sectorization, which leads to degeneration of developmental phenotype, and virulence. Fusarium oxysporum f. sp. niveum (Fon), the causal agent of watermelon Fusarium wilt, demonstrated that successive cultivation formed the degenerated sectors. In the present research, we demonstrated that subculture with aged mycelium increased the incidence of degenerations. To further investigate the differences between the Fon wild type (sporodochial type, ST) and variants (MT: mycelium type and PT: pionnotal type), developmental phenotypes and pathogenicity to watermelon were examined. Results have shown that degeneration variants (PT2, PT3, PT11 and MT6) were different from ST with mycelium growth, conidia production and chlamydospore formation. Virulence of degenerated variants on susceptible watermelon Grand Baby (GB) cultivar was determined after inoculated with Fon variants and Fon ST. In root dipping methods, all Fon variants showed slightly increased disease severity than ST. Conversely, all Fon variants showed a significant decrease in disease progression compared with ST through infested soil inoculation. The contrary results of two inoculation methods suggest that the changes of successive cultural degeneration may lead to the loss of pathogen virulence-related factors of the early stage of Fon infection process. Therefore, Cell wall-degrading enzymes (CWDEs; cellulase, pectinase, and xylanase) activities of different variants were analysed. All Fon degenerated variants demonstrated significantly decreased of CWDEs activities compared with ST. Additionally, transcripts level of 9 virulence-related genes (fmk1, fgb1, pacC, xlnR, pl1, rho1, gas1, wc1 and fow1) were assessed in normal state. The degenerated variants demonstrated a significantly low level of tested virulence-related genes transcripts except for fmk1, xlnR and fow1. In summary, the degeneration of Fon is triggered with successive subculture through aged mycelium. The degeneration showed significant impacts on virulence to watermelon, which caused by the reduction of CWDEs activities and declining expression of a set of virulence-related genes.