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An endophytic fungus isolated from finger millet (Eleusine coracana) produces anti-fungal natural products.

Mousa WK, Schwan A, Davidson J, Strange P, Liu H, Zhou T, Auzanneau FI, Raizada MN - Front Microbiol (2015)

Bottom Line: We conclude that the ancient, disease-tolerant crop, finger millet, is a novel source of endophytic anti-fungal natural products.This paper suggests the value of the crops grown by subsistence farmers as sources of endophytes and their natural products.Application of these natural chemicals to solve real world problems will require further validation.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Agriculture, University of Guelph Guelph, ON, Canada ; Department of Pharmacognosy, Mansoura University Mansoura, Egypt.

ABSTRACT
Finger millet is an ancient African cereal crop, domesticated 7000 years ago in Ethiopia, reaching India at 3000 BC. Finger millet is reported to be resistant to various fungal pathogens including Fusarium sp. We hypothesized that finger millet may host beneficial endophytes (plant-colonizing microbes) that contribute to the antifungal activity. Here we report the first isolation of endophyte(s) from finger millet. Five distinct fungal species were isolated from roots and predicted taxonomically based on 18S rDNA sequencing. Extracts from three putative endophytes inhibited growth of F. graminearum and three other pathogenic Fusarium species. The most potent anti-Fusarium strain (WF4, predicted to be a Phoma sp.) was confirmed to behave as an endophyte using pathogenicity and confocal microscopy experiments. Bioassay-guided fractionation of the WF4 extract identified four anti-fungal compounds, viridicatol, tenuazonic acid, alternariol, and alternariol monomethyl ether. All the purified compounds caused dramatic breakage of F. graminearum hyphae in vitro. These compounds have not previously been reported to have anti-Fusarium activity. None of the compounds, except for tenuazonic acid, have previously been reported to be produced by Phoma. We conclude that the ancient, disease-tolerant crop, finger millet, is a novel source of endophytic anti-fungal natural products. This paper suggests the value of the crops grown by subsistence farmers as sources of endophytes and their natural products. Application of these natural chemicals to solve real world problems will require further validation.

No MeSH data available.


Related in: MedlinePlus

HPLC chromatograms of extracts derived from endophyte WF4. Shown are chromatograms when the endophyte was cultured on (A) rice medium and (B) millet medium. The arrows show the retention time of peaks exhibiting antifungal activity.
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Figure 4: HPLC chromatograms of extracts derived from endophyte WF4. Shown are chromatograms when the endophyte was cultured on (A) rice medium and (B) millet medium. The arrows show the retention time of peaks exhibiting antifungal activity.

Mentions: Since the extract from endophyte WF4 had the most potent and the widest anti-fungal target spectrum, it was subjected to bio-assay guided purification using F. graminearum to isolate the active anti-fungal compound(s). The fermentation and extraction procedures were scaled up (see Materials and Methods) to permit purification and structural elucidation using two different solid medium (rice and millet). Flow charts that illustrate the purification procedures are included (Figures S1, S2). LC/MS analysis was conducted on the rice-derived extract (Figure 4A, Supplemental Table S3) and millet-derived extract (Figure 4B, Supplemental Table S4), and bioactive compounds eluted from the column were used as reference to track the retention time and relative abundance of these compounds relative to the total extract ingredients. The retention times (min) of each of the purified compounds were as follows: Compound 1: 14.63; Compound 2: 15.9; Compound 3: 15.98; Compound 4: 18.94 (Figure 4). Mass values (M+H+) of the purified antifungal compounds (Figures 5A,D,G,J) are shown, alongside the corresponding zoomed-in peaks from extracts of WF4 grown on either millet medium (Figures 5B,E,H,K) or rice medium (Figures 5C,F,I,L). The molecular weights of each of the purified compounds were as follows: Compound 1: 253; Compound 2: 197; Compound 3: 258; Compound 4: 272 (Figure 5). Compound 1 could be extracted from WF4 grown on both millet and rice (Figures 5B,C), whereas Compound 2 was observed on rice culture only (Figures 5E,F). Compounds 3 and 4 could be observed from millet culture only (Figures 5H,I,K,L). The purified compounds were then subjected to further spectroscopic structure elucidation. Detailed spectroscopic data for the purified compounds are included (1D-NMR, Supplemental Table S5; data for 2D-NMR are not shown). The IR, mass and 1D-NMR data were as follows:


An endophytic fungus isolated from finger millet (Eleusine coracana) produces anti-fungal natural products.

Mousa WK, Schwan A, Davidson J, Strange P, Liu H, Zhou T, Auzanneau FI, Raizada MN - Front Microbiol (2015)

HPLC chromatograms of extracts derived from endophyte WF4. Shown are chromatograms when the endophyte was cultured on (A) rice medium and (B) millet medium. The arrows show the retention time of peaks exhibiting antifungal activity.
© Copyright Policy
Related In: Results  -  Collection

License
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getmorefigures.php?uid=PMC4612689&req=5

Figure 4: HPLC chromatograms of extracts derived from endophyte WF4. Shown are chromatograms when the endophyte was cultured on (A) rice medium and (B) millet medium. The arrows show the retention time of peaks exhibiting antifungal activity.
Mentions: Since the extract from endophyte WF4 had the most potent and the widest anti-fungal target spectrum, it was subjected to bio-assay guided purification using F. graminearum to isolate the active anti-fungal compound(s). The fermentation and extraction procedures were scaled up (see Materials and Methods) to permit purification and structural elucidation using two different solid medium (rice and millet). Flow charts that illustrate the purification procedures are included (Figures S1, S2). LC/MS analysis was conducted on the rice-derived extract (Figure 4A, Supplemental Table S3) and millet-derived extract (Figure 4B, Supplemental Table S4), and bioactive compounds eluted from the column were used as reference to track the retention time and relative abundance of these compounds relative to the total extract ingredients. The retention times (min) of each of the purified compounds were as follows: Compound 1: 14.63; Compound 2: 15.9; Compound 3: 15.98; Compound 4: 18.94 (Figure 4). Mass values (M+H+) of the purified antifungal compounds (Figures 5A,D,G,J) are shown, alongside the corresponding zoomed-in peaks from extracts of WF4 grown on either millet medium (Figures 5B,E,H,K) or rice medium (Figures 5C,F,I,L). The molecular weights of each of the purified compounds were as follows: Compound 1: 253; Compound 2: 197; Compound 3: 258; Compound 4: 272 (Figure 5). Compound 1 could be extracted from WF4 grown on both millet and rice (Figures 5B,C), whereas Compound 2 was observed on rice culture only (Figures 5E,F). Compounds 3 and 4 could be observed from millet culture only (Figures 5H,I,K,L). The purified compounds were then subjected to further spectroscopic structure elucidation. Detailed spectroscopic data for the purified compounds are included (1D-NMR, Supplemental Table S5; data for 2D-NMR are not shown). The IR, mass and 1D-NMR data were as follows:

Bottom Line: We conclude that the ancient, disease-tolerant crop, finger millet, is a novel source of endophytic anti-fungal natural products.This paper suggests the value of the crops grown by subsistence farmers as sources of endophytes and their natural products.Application of these natural chemicals to solve real world problems will require further validation.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Agriculture, University of Guelph Guelph, ON, Canada ; Department of Pharmacognosy, Mansoura University Mansoura, Egypt.

ABSTRACT
Finger millet is an ancient African cereal crop, domesticated 7000 years ago in Ethiopia, reaching India at 3000 BC. Finger millet is reported to be resistant to various fungal pathogens including Fusarium sp. We hypothesized that finger millet may host beneficial endophytes (plant-colonizing microbes) that contribute to the antifungal activity. Here we report the first isolation of endophyte(s) from finger millet. Five distinct fungal species were isolated from roots and predicted taxonomically based on 18S rDNA sequencing. Extracts from three putative endophytes inhibited growth of F. graminearum and three other pathogenic Fusarium species. The most potent anti-Fusarium strain (WF4, predicted to be a Phoma sp.) was confirmed to behave as an endophyte using pathogenicity and confocal microscopy experiments. Bioassay-guided fractionation of the WF4 extract identified four anti-fungal compounds, viridicatol, tenuazonic acid, alternariol, and alternariol monomethyl ether. All the purified compounds caused dramatic breakage of F. graminearum hyphae in vitro. These compounds have not previously been reported to have anti-Fusarium activity. None of the compounds, except for tenuazonic acid, have previously been reported to be produced by Phoma. We conclude that the ancient, disease-tolerant crop, finger millet, is a novel source of endophytic anti-fungal natural products. This paper suggests the value of the crops grown by subsistence farmers as sources of endophytes and their natural products. Application of these natural chemicals to solve real world problems will require further validation.

No MeSH data available.


Related in: MedlinePlus