Endophytic fungi associated with Monarda citriodora, an aromatic and medicinal plant and their biocontrol potential.
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ABSTRACT: The Food and Agriculture Organization has estimated that every year considerable losses of the food crops occur due to plant diseases. Although fungicides are extensively used for management of plant diseases, they are expensive and hazardous to the environment and human health. Alternatively, biological control is the safe way to overcome the effects of plant diseases and to sustain agriculture. Since Monarda citriodora Cerv. ex Lag. (Lamiaceae/Labiatae) is known for its antifungal properties, it was chosen for the study.The isolation of endophytic fungi from M. citriodora and assessing their biocontrol potential.The isolated endophytes were characterized using ITS-5.8?S rDNA sequencing. Their biocontrol potential was assessed using different antagonistic assays against major plant pathogens.Twenty-eight endophytes representing 11 genera were isolated, of which, around 82% endophytes showed biocontrol potential against plant pathogens. MC-2?L (Fusarium oxysporum), MC-14?F (F. oxysporum), MC-22?F (F. oxysporum) and MC-25?F (F. redolens) displayed significant antagonistic activity against all the tested pathogens. Interestingly, MC-10?L (Muscodor yucatanensis) completely inhibited the growth of Sclerotinia sp., Colletotrichum capsici, Aspergillus flavus and A. fumigatus in dual culture assay, whereas MC-8?L (A. oryzae) and MC-9?L (Penicillium commune) completely inhibited the growth of the Sclerotinia sp. in fumigation assay.Endophytes MC-2?L, MC-14?F, MC-22?F and MC-25?F could effectively be used to control broad range of phytopathogens, while MC-10?L, MC-8?L and MC-9?L could be used to control specific pathogens. Secondly, endophytes showing varying degrees of antagonism in different assays represented the chemo-diversity not only as promising biocontrol agents but also as a resource of defensive and bioactive metabolites.
<h4>Context</h4>The Food and Agriculture Organization has estimated that every year considerable losses of the food crops occur due to plant diseases. Although fungicides are extensively used for management of plant diseases, they are expensive and hazardous to the environment and human health. Alternatively, biological control is the safe way to overcome the effects of plant diseases and to sustain agriculture. Since Monarda citriodora Cerv. ex Lag. (Lamiaceae/Labiatae) is known for its antifun ...[more]
Project description:BACKGROUND:Present study focuses on diversity and distribution analysis of endophytic fungi associated with different tissues of the Monarda citriodora Cerv. ex Lag. (Lamiaceae/Labiatae). Anticancer and antimicrobial potential of isolated endophytes have also been investigated. RESULTS:A total of twenty eight fungal endophytes belonging to 11 different genera were isolated from this plant. All the endophytic fungi belonged to the Ascomycota phylum. The leaves were immensely rich in fungal species, while roots showed the highest tissue specific fungal dominance. Out of 28 fungal species, 72% endophytic extracts were found cytotoxic against one or more human cancer cell lines. The most prominent anticancer activity (IC50 value <10 ?g/mL) was shown by MC-14 L (Fusarium oxysporum), MC-14 F (F. oxysporum), MC-18 L (Aspergillus fumigatus), MC-24 L (Cladosporium tenuissimum), MC-25 L (Fusarium sp.), MC-26 F (F. oxysporum) extracts. 75% of the extracts showed antimicrobial activities in agar disc-diffusion assay and 27% in the tube dilution method (MIC <100 ?g/mL) respectively against the tested pathogens. Extracts of MC-14 L (F. oxysporum) and MC-18 L (A. fumigatus) displayed broad spectrum antimicrobial activity. CONCLUSIONS:These results indicated that M. citriodora harbors a rich fungal endophytic community with anticancer and antimicrobial activities. The isolated endophyte MC-24 L (C. tenuissimum) has the potential to be a source of novel cytotoxic/antimicrobial compounds. This is the first report of diversity of fungal endophytes isolated from M. citriodora.
Project description:Healthy cotton samples were collected and 93 endophytic fungal strains were isolated: 23 strains from the roots and 70 strains from the stems. Morphological characterization and ITS sequence analysis were used for the identification of these isolates. The results showed that the 93 strains including 20 species were highly diverse in terms of their taxonomy. Simpson's and Shannon's diversity indices were 0.915 and 3.848, respectively. Fusarium and Alternaria were the two dominant genera, constituting 19.4% of the total strains. Then, 72 spore-producing strains were tested for the suppression of cotton Verticillium wilt (CVW) caused by Verticillium dahliae in a greenhouse. Five strains exhibited effective suppression of CVW with average efficacy values higher than 50%. One of the effective strains, namely, Fusarium proliferatum 10R-7, was selected for the investigation of the role of fusaric acid, a secondary metabolite of strain 10R-7, in the suppression of V. dahliae and CVW. The results showed that F. proliferatum 10R-7 could produce fusaric acid, and this metabolite exhibited 100% inhibition of mycelial growth of V. dahliae at concentrations higher than 20 μg/ml. However, fusaric acid at 2.5 to 80 μg/ml was not effective in the suppression of CVW, compared with the control treatment with V. dahliae alone. F. proliferatum 10R-7 was labeled with green fluorescent protein (GFP), and the GFP-tagged strain was found to be able to colonize inside the taproots of cotton, suggesting that F. proliferatum 10R-7 is a true endophyte of cotton and endophytic colonization may play a role in the suppression of infection of cotton by V. dahliae.
Project description:Grapevine Trunk Diseases (GTDs) are a major challenge to the grape industry worldwide. GTDs are responsible for considerable loss of quality, production, and vineyard longevity. Seventy-five percent of Chilean vineyards are estimated to be affected by GTDs. GTDs are complex diseases caused by several fungi species, including members of the Botryosphaeriaceae family and Phaeomoniella chlamydospora, considered some of the most important causal agents for these diseases in Chile. In this study, we isolated 169 endophytic and 209 rhizospheric fungi from grapevines grown under organic and conventional farming in Chile. Multiple isolates of Chaetomium sp., Cladosporium sp., Clonostachys rosea, Epicoccum nigrum, Purpureocillium lilacinum, and Trichoderma sp. were evaluated for their potential of biocontrol activity against Diplodia seriata, Neofusicoccum parvum, and Pa. chlamydospora. Tests of antagonism were carried out using two dual-culture-plate methods with multiple media types, including agar containing grapevine wood extract to simulate in planta nutrient conditions. Significant pathogen growth inhibition was observed by all isolates tested. Clonostachys rosea showed 98.2% inhibition of all pathogens in the presence of grapevine wood extract. We observed 100% pathogen growth inhibition when autoclaved lignified grapevine shoots were pre-inoculated with either C. rosea strains or Trichoderma sp. Overall, these results show that C. rosea strains isolated from grapevines are promising biocontrol agents against GTDs.
Project description:Foliar diseases caused by Stemphylium and Colletotrichum species are among the major biotic factors limiting Welsh onion production in Taiwan. Owing to concerns about the environment and the development of pathogen resistance to existing fungicides, biological control using endophytes is emerging as an eco-friendly alternative to chemical control. The aim of the present study was to isolate endophytes from healthy Welsh onion leaves and investigate their antagonistic potential against the major phytopathogenic fungi associated with Welsh onion plants in Taiwan. A total of 109 bacterial and 31 fungal strains were isolated from healthy Welsh onion leaves and assigned to 16 bacterial and nine fungal genera using morphological and molecular characterization based on DNA sequence data obtained from nuclear internal transcribed spacer (nrITS) (fungi) and 16S rRNA (bacteria). Evaluation of these endophytic isolates for biocontrol activity against leaf blight pathogens Colletotrichum spaethianum strain SX15-2 and Stemphylium vesicarium strain SX20-2 by dual culture assay and greenhouse experiments resulted in the identification of two bacterial isolates (GFB08 and LFB28) and two fungal isolates (GFF06 and GFF08) as promising antagonists to leaf blight pathogens. Among the four selected isolates, Bacillus strain GFB08 exhibited the highest disease control in the greenhouse study. Therefore, Bacillus strain GFB08 was further evaluated to understand the mechanism underlying its biocontrol efficacy. A phylogenetic analysis based on six genes identified Bacillus strain GFB08 as B. velezensis. The presence of antimicrobial peptide genes (baer, bamC, bmyB, dfnA, fenD, ituC, mlna, and srfAA) and the secretion of several cell wall degrading enzymes (CWDEs), including cellulase and protease, confirmed the antifungal nature of B. velezensis strain GFB08. Leaf blight disease suppression by preventive and curative assays indicated that B. velezensis strain GFB08 has preventive efficacy on C. spaethianum strain SX15-2 and both preventive and curative efficacy on S. vesicarium strain SX20-2. Overall, the current study revealed that healthy Welsh onion leaves harbour diverse bacterial and fungal endophytes, among which the endophytic bacterial strain, B. velezensis strain GFB08, could potentially be used as a biocontrol agent to manage the leaf blight diseases of Welsh onion in Taiwan.
Project description:Endophytic fungi were first isolated from the fresh root of the Chinese medicinal plant Vernonia anthelmintica collected from the Hotan Prefecture within the Xinjiang Autonomous region of the People's Republic of China. This plant has been used in Uyghur traditional medicine to treat vitiligo, a skin condition characterized by patches of the skin losing their pigment. In total, fifteen fungal strains were isolated. Among these, four endophytic fungi were identified by their DNA sequences and registered to GenBank with accession numbers. The isolates were identified as Schizophyllum commune XJA1, Talaromyces sp. XJA4, Aspergillus sp. XJA6, Aspergillus terreus XJA8. Ethyl acetate extracts of all fungal strains were used to quantify melanin content and to identify in vitro biological activity assays including antimicrobial, antioxidant, cytotoxic, antidiabetic and tyrosinase activity on B16 cells. Among the extracts of all four identified strains, the ethyl acetate extract of the Aspergillus sp. XJA6 was chosen for further characterization because it presented the highest biological activity against these tests. In addition, twenty four volatile compounds from the petroleum ether fraction were characterized by GC-MS.
Project description:In this study, we reported thirty-nine endophytic fungi identified as Colletotrichum spp. associated with Brazilian pepper tree or aroeira (Schinus terebinthifolius Raddi. Anacardiaceae) in Paraná state, Brazil. These endophytes were identified by morphological and molecular methods, using PCR taxon-specific with CaInt/ITS4, CgInt/ITS4, and Col1/ITS4 primers, which amplify specific bands in C. acutatum, C. gloeosporioides lato sensu, and Colletotrichum boninensis, respectively, and by DNA sequence analysis of the nrDNA internal transcribed spacer region (ITS1, 5.8S, ITS2). We also assayed the presence of dsRNA particles in Colletotrichum spp. isolates. Combining both morphological characters and molecular data, we identified the species C. gloeosporioides, C. boninense, and C. simmondsii. However, we found a high genetic variability intraspecific in C. gloeosporioides which suggests the existence of several other species. Bands of double-stranded RNA (dsRNA) were detected in three of thirty-nine isolates. Identity of these bands was confirmed by RNAse, DNAse, and S1 nuclease treatments for the isolates LGMF633, LGMF726, and LGMF729. This is the first study reporting these particles of dsRNA in C. gloeosporioides.
