Putative metabolic pathway for the bioproduction of bikaverin and intermediates thereof in the wild Fusarium oxysporum LCP531 strain

Juliana Lebeau,Yanis Caro,Thomas Petit,Laurent Dufossé

doi:10.1186/s13568-019-0912-4

Abstract

Fungal naphthoquinones, like red bikaverin, are of interest due to their growing applications in designing pharmaceutical products. Though considerable work has been done on the elucidation of bikaverin biosynthesis pathway in Fusarium fujikuroi, very few reports are available regarding its bioproduction in F. oxysporum. We are hereby proposing a putative metabolic pathway for bikaverin bioproduction in a wild F. oxysporum strain by cross-linking the pigment profiles we obtained under two different fermentation conditions with literature. Naphthoquinone pigments were extracted with a pressurized liquid extraction method, and characterized by HPLC–DAD and UHPLC-HRMS. The results led to the conclusions that the F. oxysporum LCP531 strain was able to produce bikaverin and its various intermediates, e.g., pre-bikaverin, oxo-pre-bikaverin, dinor-bikaverin, me-oxo-pre-bikaverin, and nor-bikaverin, in submerged cultures in various proportions. To our knowledge, this is the first report of the isolation of these five bikaverin intermediates from F. oxysporum cultures, providing us with steady clues for confirming a bikaverin metabolic pathway as well as some of its regulatory patterns in the F. oxysporum LCP531 strain, based on the previously reported model in F. fujikuroi. Interestingly, norbikaverin accumulated along with bikaverin in mycelial cells when the strain grew on simple carbon and nitrogen sources and additional cofactors. Along bikaverin production, we were able to describe the excretion of the toxin beauvericin as main extrolite exclusively in liquid medium containing complex nitrogen and carbon sources, as well as the isolation of ergosterol derivate in mycelial extracts, which have potential for pharmaceutical uses. Therefore, culture conditions were also concluded to trigger some specific biosynthetic route favoring various metabolites of interest. Such observation is of great significance for selective production of pigments and/or prevention of occurrence of others (aka mycotoxins).

Highlights

Bikaverin is classified as a mycotoxin, even if its occurrence was observed in non-virulent Fusarium sp
The Fusarium species, such as F. oxysporum, that produce bikaverin are commonly considered as phytopathogens with great economic and agricultural importance, the presence of the pigment has not been found to be related to the phytopathogenic activity
Despite its classification as a contaminant in food and feed, there are no reports to this date of harmful effects of products containing bikaverin on human or animal health, appropriate toxicological studies will still be required in order to ensure its complete safe use in any future applications (Norred et al 1992)

Summary

Introduction

Literature is abundantly reporting the significant application of metabolites from ascomycetous fungi in the industry, through the production of various bioactive compounds, such as plant hormones, enzymes, organic acids, mycotoxins as well as natural pigments (MapariLebeau et al AMB Expr (2019) 9:1862008), zearalenone (Gaffoor and Trail 2006; Lysøe et al 2006), beauvericin (Fotso et al 2002; Zhan et al 2007), fusaric acid (Bacon et al 1996; Son et al 2008; Niehaus et al 2014) and fusarin C (Wiebe and Bjeldanes 1981; Song et al 2004; Díaz-Sánchez et al 2012). Many Fusarium naphthoquinone pigments, like aurofusarin (Kim et al 2005; Frandsen et al 2006), fusarubin (Studt et al 2012) and bikaverin (Brewer et al 1973; review in: Limón et al 2010; Lale and Gadre 2016; Lebeau et al 2019) exhibit useful biological activities They are recognized as mycotoxins and this fact is important in safety concerns, mainly considering their possible applications in agrochemical, pharmacological and medical sectors (Caro et al 2017; Abdel-Azeem et al 2019; Lebeau et al 2019). Due to chemical and biological properties of naphthoquinones from Fusarium sp., these compounds may be applied in medical fields and as textile and material dyes

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