Chemically-Mediated Interactions Between Macroalgae, Their Fungal Endophytes, and Protistan Pathogens

Abstract

Filamentous fungi asymptomatically colonize the inner tissues of macroalgae, yet their ecological roles remain largely underexplored. Here, we tested if metabolites produced by fungal endophytes might protect their host against a phylogenetically broad spectrum of protistan pathogens. Accordingly, the cultivable fungal endophytes of four brown algal species were isolated and identified based on LSU and SSU sequencing. The fungal metabolomes were tested for their ability to reduce the infection by protistan pathogens in the algal model Ectocarpus siliculosus. The most active metabolomes effective against the oomycetes Eurychasma dicksonii and Anisolpidium ectocarpii, and the phytomixid Maullinia ectocarpii were further characterized chemically. Several pyrenocines isolated from Phaeosphaeria sp. AN596H efficiently inhibited the infection by all abovementioned pathogens. Strikingly, these compounds also inhibited the infection of nori (Pyropia yezoensis) against its two most devastating oomycete pathogens, Olpidiopsis pyropiae, and Pythium porphyrae. We thus demonstrate that fungal endophytes associated with brown algae produce bioactive metabolites which might confer protection against pathogen infection. These results highlight the potential of metabolites to finely-tune the outcome of molecular interactions between algae, their endophytes, and protistan pathogens. This also provide proof-of-concept toward the applicability of such metabolites in marine aquaculture to control otherwise untreatable diseases.

Keywords: fungal endophytes; macroalgae; metabolome; molecular interactions; protistan pathogens; pyrenocines; secondary metabolites.

Figure 1

Phylogenetic affinities based on LSU sequences of fungal endophytes isolates from the four brown algae sampled from French and Scottish sites. Isolate code are indicated in bold and accession numbers from reference sequences are marked in gray. Scale bar indicates 10% estimated sequence divergence.

Figure 2

Taxonomic assemblages of the fungal classes determined according to the different host-algae species and algal organs. Each class is displayed by a different color. Height of the bars represents the % of each fungal class according to the host-algae species or the algal organs. Width of the bars represents the total number of OTUs according to the host-algae species or the algal organs. Codes for host species and algal organs are SL, Saccharina latissima; PC, Pelvetia canaliculata; LD, Laminaria digitata; AN, Ascophyllum nodosum; F, Frond; H, Holdfast; R, Receptacle; T, Thallus.

Figure 3

Antipathogenic activities of the fungal extracts LD68H (Penicillium janczewskii), AN596H (Phaeosphaeria sp.), PC359H (Paradendryphiella arenaria), LD13H (Chaetomium globosum), SL469T (Chaetomium globosum), and SL333T (Phoma exigua) assessed on the infection of Ectocarpus siliculosus by Eurychasma dicksonii (CCAP 4018/1, CCAP 4018/3), Anisolpidium ectocarpii (CCAP 4001/1), and Maullinia ectocarpii (CCAP 1538/1). Controls consisted of algae alone (control uninfected), algae with parasite treatment (control infected), algae with parasite treatment, and 1% DMSO (control infected + DMSO 1%). Mean microscopy score values ± SE are displayed for biological triplicates.

Figure 4

Antipathogenic activities of the fungal extracts LD68H (Penicillium janczewskii), AN596H (Phaeosphaeria sp.), PC359H (Paradendryphiella arenaria), LD13H (Chaetomium globosum), SL469T (Chaetomium globosum), and SL333T (Phoma exigua) assessed by qPCR quantification of the infection of Ectocarpus siliculosus by Eurychasma dicksonii (CCAP 4018/1, CCAP 4018/3). Controls consisted of algae alone (control uninfected), algae with parasite treatment (control infected), algae with parasite treatment and 1% DMSO (control infected + DMSO 1%). Mean disease score ± SE are displayed for biological triplicates.

Figure 5

Key HMBC and COSY correlations of the new pyrenocine S.

Figure 6

Antipathogenic activities of the isolated assessed on the infection of Ectocarpus siliculosus by the pathogens Eurychasma dicksonii (CCAP4018/1, CCAP4018/3), Anisolpidium ectocarpii (CCAP 4001/1), and Maullinia ectocarpii (CCAP 1538/1). Controls consisted of algae alone (control uninfected), algae with parasite treatment (control infected), algae with parasite treatment and 1% DMSO (control infected + DMSO 1%). Mean microscopy score values ± SE are displayed for biological triplicates.