Enhancing the Discovery of Bioactive Secondary Metabolites From Fungal Endophytes Using Chemical Elicitation and Variation of Fermentation Media

Gakuubi, Martin Muthee; Ching, Kuan Chieh; Munusamy, Madhaiyan; Wibowo, Mario; Liang, Zhao‐Xun; Yoganathan, K.; Ng, Siew Bee

doi:10.3389/fmicb.2022.898976

Cited by 20 publications

(29 citation statements)

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“…MS data were analyzed and processed using Agilent MassHunter Qualitative Analysis Version 10.0. The presence of palmarumycins was confirmed by comparison with our in-house standard compounds library ( Gakuubi et al, 2022 ). All solvents for extraction and LC–MS were of chromatographic grade.…”

Section: Methodsmentioning

confidence: 90%

“…Lophiotrema sp. F6932 produces spirobisnaphthalene palmarumycin C 8 among other analogs such as palmarumycin CP 30 ( Gakuubi et al, 2022 ). Significantly, the biosynthesis of palmarumycins is proposed to involve phenolic oxidative dimerization of DHN, the same precursor that is involved in melanin biosynthesis in many filamentous fungi ( Bode and Zeeck, 2000 ; Mafezoli et al, 2018 ).…”

Section: Discussionmentioning

confidence: 99%

“…Our previous study on discovery of bioactive secondary metabolites from fungal endophytes resulted in the isolation of palmarumycin compounds from Lophiotrema sp. F6932 ( Gakuubi et al, 2022 ). Among the isolated compounds, palmarumycin C 8 exhibited moderate antibacterial activity against Staphylococcus aureus presenting IC 90 value of 18 μg/mL ( Gakuubi et al, 2022 ).…”

Section: Introductionmentioning

confidence: 99%

“…F6932 ( Gakuubi et al, 2022 ). Among the isolated compounds, palmarumycin C 8 exhibited moderate antibacterial activity against Staphylococcus aureus presenting IC 90 value of 18 μg/mL ( Gakuubi et al, 2022 ). In the current work, we describe the development of a CRISPR/Cas9 RNP-mediated genome editing system that has enabled us to characterize a type 1 iterative PKS (PAL) that is responsible for palmarumycin and melanin biosynthesis in fungus.…”

Section: Introductionmentioning

confidence: 99%

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CRISPR/Cas9 RNP-assisted validation of palmarumycin biosynthetic gene cluster in Lophiotrema sp. F6932

Gakuubi

Ching²,

Munusamy³

et al. 2022

Front. Microbiol.

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Lophiotrema is a genus of ascomycetous fungi within the family Lophiotremataceae. Members of this genus have been isolated as endophytes from a wide range of host plants and also from plant debris within terrestrial and marine habitats, where they are thought to function as saprobes. Lophiotrema sp. F6932 was isolated from white mangrove (Avicennia officinalis) in Pulau Ubin Island, Singapore. Crude extracts from the fungus exhibited strong antibacterial activity, and bioassay-guided isolation and structure elucidation of bioactive constituents led to the isolation of palmarumycin C8 and a new analog palmarumycin CP30. Whole-genome sequencing analysis resulted in the identification of a putative type 1 iterative PKS (iPKS) predicated to be involved in the biosynthesis of palmarumycins. To verify the involvement of palmarumycin (PAL) gene cluster in the biosynthesis of these compounds, we employed ribonucleoprotein (RNP)-mediated CRISPR-Cas9 to induce targeted deletion of the ketosynthase (KS) domain in PAL. Double-strand breaks (DSBs) upstream and downstream of the KS domain was followed by homology-directed repair (HDR) with a hygromycin resistance cassette flanked by a 50 bp of homology on both sides of the DSBs. The resultant deletion mutants displayed completely different phenotypes compared to the wild-type strain, as they had different colony morphology and were no longer able to produce palmarumycins or melanin. This study, therefore, confirms the involvement of PAL in the biosynthesis of palmarumycins, and paves the way for implementing a similar approach in the characterization of other gene clusters of interest in this largely understudied fungal strain.

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Section: Methodsmentioning

confidence: 90%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

CRISPR/Cas9 RNP-assisted validation of palmarumycin biosynthetic gene cluster in Lophiotrema sp. F6932

Gakuubi

Ching²,

Munusamy³

et al. 2022

Front. Microbiol.

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“…The study of active secondary metabolites of plant symbiotic fungi is essential for the conservation of plant diversity and the environment. However, as the limitations of endophytic fungal secondary metabolites developed from traditional plant parts, increasing numbers of researchers are turning their attention to the development of promising endophytic microorganisms from speci c environments or less studied plant parts [10].…”

Section: Introductionmentioning

confidence: 99%

Diversity of fungal communities and changes in antibacterial activity of artificially culturable fungal secondary metabolites from diseased and healthy fruits of Cinnamomum burmannii

Wang

Cai

Yang

et al. 2022

Preprint

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Background Studies on the function of endophytic fungal communities in healthy plant tissues are not new, while little is known about the functional changes of fungal communities in pathogenic plant tissues. Healthy fruits of Cinnamomum burmannii are an important medicinal resource, infection by phytopathogenic fungi causes changes in volatile components, with the corresponding, the functional characteristics and potential value of fungal communities in diseased fruits have not been reported. Consequently, characterization in the composition structure and antibacterial activity of fungal communities from healthy and diseased fruits of C. burmannii was investigated, the secondary metabolites in the fungus Medicopsis romeroi were reported for the first time. Results The fungal community in diseased fruits differed from that of healthy fruits at the Phylum, Class, Order or Genus levels, with important changes in the species and relative abundance of the dominant flora. Forty-one different strains (11 from healthy fruits and 30 from diseased fruits) were successfully identified by morphological and molecular biological methods which were classified into 8 groups and 23 genera using phylogenetic tree analysis, with Pleosporales, Glomerellales and Hypocreales were the dominant group at the phylum level and Colletotrichum was the dominant group at the genus level. The secondary metabolites of all strains had different degrees of antibacterial activity, while the secondary metabolites of diseased fruit symbiotic fungi were generally stronger than those of healthy fruits, with the active secondary metabolites dominated by small and medium polar compounds. Both the up-regulated and down-regulated flora in diseased fruit had strong antibacterial activity. Two new compounds, 5, 6-Dimethoxy-[1',1:4,1''-terphenyl]-2-ol (1), 5-(methoxycarbonyl)-2-methylbenzo[d][1, 3]dioxole-2-carboxylic acid (2) and three known compounds (3, 4, 5) were isolated and identified for the first time from the symbiotic fungus Medicopsis romeroi. Conclusion Although the diversity of fungal communities decreases after plant fruit disease, the antibacterial activity capacity of the fungi among them is generally enhanced, and the development of secondary metabolites of active strains from diseased fruits holds great promise. This study is significant for understanding the functional variation of bioactivity in fungal communities and developing a broader range of bioactive resources.

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Spirobisnaphthalenes from Fungi, Biological activities and Total Synthesis: A Research Review

et al. 2022

Asian J Org Chem

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Spirobisnaphthalenes are a family of natural products containing two naphthalenes connected with spiroketal or dinaphthalene spiroether. This family of natural compounds has a wide range of biological activities and complex structures, and has attracted great attention from synthetic chemists in recent years. Hundreds of novel spirobisnaphthalenes have been identified from the first separation in 1989. Despite the slow pace of research in recent years, since the latest review in 2017, 30 new natural products have been isolated from the endolichenic and endophytic fungus, including Type A, B, D and E, a new special skeleton with a spiro quaternary carbon center, as well as significant breakthroughs are made in the total synthesis. Here, the last research advances in the isolation of new spirobisnaphthalenes, their activities, mode of action and the total synthesis of spirobisnaphthalenes in the last five years were afforded based on our research in this field in this review.

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Enhancing the Discovery of Bioactive Secondary Metabolites From Fungal Endophytes Using Chemical Elicitation and Variation of Fermentation Media

Cited by 20 publications

References 80 publications

CRISPR/Cas9 RNP-assisted validation of palmarumycin biosynthetic gene cluster in Lophiotrema sp. F6932

CRISPR/Cas9 RNP-assisted validation of palmarumycin biosynthetic gene cluster in Lophiotrema sp. F6932

Diversity of fungal communities and changes in antibacterial activity of artificially culturable fungal secondary metabolites from diseased and healthy fruits of Cinnamomum burmannii

Spirobisnaphthalenes from Fungi, Biological activities and Total Synthesis: A Research Review

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