Isolation of endophytic fungi and screening of Huperzine A\u2013producing fungus from Huperzia serrata in Vietnam

Thanh Thi Minh Le,Anh Thi Hong Hoang,Thuy Thi Bich Vo,Ha Hoang Chu,Thuy Thi Bich Le,Dong Van Quyen

doi:10.1038/s41598-019-52481-2

Thanh Thi Minh Le, Anh Thi Hong Hoang + Show 4 more

Open Access

https://doi.org/10.1038/s41598-019-52481-2

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Abstract

Huperzine A (HupA), a natural Lycopodium alkaloid derived from Huperzia serrata (Thunb. ex Murray) Trev. plants, is a highly active acetylcholinesterase inhibitor and a key compound used for treating Alzheimer’s disease (AD). Recently, HupA has been reported in various endophytic fungi isolated from H. serrata. In the present study, 153 endophytic fungi were isolated from healthy tissues of H. serrata collected from natural populations in Lam Dong province of Central Vietnam. The endophytic fungi were identified based on morphological characteristics and Internal Transcribed Spacer sequences. Among them, 34 strains were classified into seven genera belonging to Ascomycota, including Alternaria, Fusarium, Trichoderma, Penicillium, Paecilomyces, and Phoma, and eight strains belonging to the genus Mucor (Zygomycota). The other strains remained unidentified. According to the results of thin-layer chromatography and high-performance liquid chromatography, only one of the 153 strains, Penicillium sp. LDL4.4, could produce HupA, with a yield 1.38 mg l−1 (168.9 µg g−1 dried mycelium) when cultured in potato dextrose broth, which was considerably higher than that of other reported endophytic fungi. Such a fungus is a promising candidate and alternative to presently available HupA production techniques for treating AD and preventing further memory decline.

Highlights

Huperzia serrata (Thunb. ex Murray) Trev. is a rare species, and it is listed in the research program on conservation and development of rare genes in medicinal plants[1]
Recent studies have demonstrated that in vitro cultures of H. serrata could provide higher quality Huperzine A (HupA) than in vivo cultures, the yields remain inadequate for the requirements of the pharmaceutical industry since plant tissues grow slowly in culture, and in vitro culture initiation requires the development of spores as starting material[7]
We explored and identified endophytic fungi associated with H. serrata in natural H. serrata populations in Lam Dong province in central Vietnam and determined their HupA production capacities

Summary

Introduction

Huperzia serrata (Thunb. ex Murray) Trev. is a rare species, and it is listed in the research program on conservation and development of rare genes in medicinal plants[1]. Is a rare species, and it is listed in the research program on conservation and development of rare genes in medicinal plants[1] It is considered a medicinal herb in Western countries and is used in a wide range of functional foods sold in the market. The isolation endophytic fungi in HupA-producing H. serrata has been investigated by scientists with the aim of finding an alternative and increasing HupA yields for pharmaceutical manufacturing processes. Several studies have attempted to identify HupA-producing endophytic fungi from plants of the family Huperziaceae, in China where H. serrata is widely distributed. We explored and identified endophytic fungi associated with H. serrata in natural H. serrata populations in Lam Dong province in central Vietnam and determined their HupA production capacities

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