A Study On Biodiversity Of Endophytic Fungi Associated With Watermelon In Beijing And The Vicinity

Endophytic fungi, persistently living inside and coevolving with plants, play important roles in various ecological systems. Some of them increase host fitness through promoting growth and conferring resistance in plants to pathogens and various abiotic stresses. In order to understand the biodiversity of endophytic fungi in watermelon plants grown in an area around Beijing,60watermelon plants were collected from Beijing and Langfang in Hebei at different growth stages, including different cultivars growing in greenhouse and filed, and10other melon plants collected from Langfang. The endophytic fungi were isolated on PDA plates and identified by morphological observation and rDNA-ITS sequencing and BLAST comparison. In the results,754isolates were totally obtained,559of them were identified as39species in19genera and the others were dropped into24genera based on morphology and ITS sequence data. The145isolates the cucumber, and the68isolates the muskmelon. Isolates belonging to Cladosporium, Fusarium and Alternaria were the most frequently obtained, and Panaeolus and Lichtheimia are firstly reported here as endophytic fungi.Most of the watermelon endophytic fungi were rare taxon, which may not be occasional colonists or contaminants, but serve as true endophytic fungi, needing to be profoundly analyzed in detail.Almost all endophytic fungi similarly distributed in other plants, which were inconsistent with the challenged view that"4-5obligate endophytic fungi averagely live inside each host plants"that proposed by Petrini.The variation of endophytic fungi was analysed in watermelon plants from Daxing in Beijing. These results demonstrated that the taxon number of endophytic fungi increased with growth of watermelon plant. Fusarium species were frequent and the dominant endophytic fungi in watermelon seedlings and the relative frequency of Fusarium solani reached39.2%. Monosporascus cannonballus was the dominant endophyte in watermelon plants at vegetative stage, and the Alternaria species, the reproductive stage with account of19.4%in the total isolates. Some endophytic fungi were obtained only from samples at a specific growth stage. Most of the endophytic fungi showed organ specificity or preference in certain degree, fungal isolates belonging to some taxa existed only in certain organs. Cladosporium isolates were frequently present in stems and leaves, the Ceratorhiza and Fusarium the roots, and the Alternaria the leaves. The composition of endophyte assemblages was greatly influenced by geographical factor, the most frequently isolated species was Cladosporium oxysporum from Chaoyang, and the Fusarium solani the Daxing. Significant differences also existed in the taxa of the endophytic fungi among different melon plants, presenting a host preference. Regarding to the cultivative conditions, the diversity of endophytic fungi in the watermelon plants in the field was higher than that in greenhouse, suggesting that there is a relationship between endophyic fungi and cultivation method.The results of confront culture indicated that some endophytic fungi exhibited the activity against plant pathogenic fungi. Among the967isolates of endophytic fungi,106,106,158and215isolates were active against Sclerotinia sclerotiorum ACCC36081, Rhizoctonia solani ACCC36124, Fusarium oxysporum f. sp. cucumerinum ACCC37438, and Colletotrichum orbiculare ACCC30016, with antagonism index at20%or above, accounting for11.0%,11.0%,16.3%and22.2%of the total isolates respectively. There were19,26,49and92isolates, against Sclerotinia sclerotiorum ACCC36081, Rhizoctonia solani ACCC36124, Fusarium oxysporum f. sp. cucumerinum ACCC37438, and Colletotrichum orbiculare ACCC30016, with antagonism index at50%or above, accounting for2.0%,2.7%,5.1%and9.5%of the total isolates respectively. These results indicated that the common diseases of cucurbit crops were affected by endophytic fungi to some extent. Antagonism types were primarily recognized as the deadlock caused by mycelia contact (A) and the deadlock at a distance caused by bioactive materials produced by mycelia (B).Germinant seeds of cucumber (cultivar. Jinyan4) were pretreated with the spore suspension of endophytic Aspergillus terreus JFLu-3-1, Aspergillus ochraceus CSu-9and Penicillium oxalicum WSu-9respectively, and subsequently challenged by Fusarium oxysporum f. sp. cucumerinum ACCC37438, the pathogen of cucumber Fusarium Wilt, symptoms development of cucumber Fusarium Wilt were demonstrated to be suppressed significantly. The magnitude of the reduction in disease severity of cucumber Fusarium Wilt by sterile fermentation broth was greater than corresponding spore suspension, indicating that the effect was medicated by substance of filtrate. In30days after inoculation with Fusarium oxysporum f. sp. cucumerinum ACCC37438, most of the tested plants without inoculation with endophytic fungi in advance wilted and the disease index reached as high as86.3%. The plants treated with sterile fermentation broth of Aspergillus terreus JFLu-3-1in advance displayed mild symptoms and38.9%of the disease index, suggesting its potential use in biocontrol in future.The results of this study preliminarily revealed the overview of biodiversity of endophytic fungi and variation of endophytic mycobiota in watermelon plants and gained the initial understanding about the biological function of endophytic fungi and relationship between them and host plants, which provides some reference for further researches on and utilization of the endophytic fungi from watermelon.