Associated bacterial microbiome responds opportunistic once algal host Scenedesmus vacuolatus is attacked by endoparasite Amoeboaphelidium protococcarum

Abstract

The interactions of microalgae and their associated microbiomes have come to the fore of applied phycological research in recent years. However, the functional mechanisms of microalgal interactions remain largely unknown. Here, we examine functional protein patterns of the microalgae Scenedesmus vacuolatus and its associated bacterial community during algal infection by the endoparasite Amoeboaphelidium protococcarum. We performed metaproteomics analyses of non-infected (NI) and aphelid-infected (AI) S. vacuolatus cultures to investigate underlying functional and physiological changes under infectious conditions. We observed an increase in bacterial protein abundance as well as a severe shift of bacterial functional patterns throughout aphelid-infection in comparison to NI treatment. Most of the bacterial proteins (about 55%) upregulated in AI were linked to metabolism and transport of amino acids, lipids, coenzymes, nucleotides and carbohydrates and to energy production. Several proteins associated with pathogenic bacterial-plant interactions showed higher protein abundance levels in AI treatment. These functional shifts indicate that associated bacteria involved in commensalistic or mutualistic interactions in NI switch to opportunistic lifestyles and facilitate pathogenic or saprotrophic traits in AI treatment. In summary, the native bacterial microbiome adapted its metabolism to algal host die off and is able to metabolize nutrients from injured cells or decompose dead cellular material.

Figure 1

(a) Dry weight biomass content, optical density (OD₇₅₀), normalized chlorophyll a fluorescence (OD₆₈₅) of aphelid-infected (AI, brown) und non infected (NI, green) S. vacuolatus cultures over days post inoculation (DPI) (mean of n = 3 ± SD). (b) Mean of infection status was revealed by fluorescence microscopic observations from AI treatment over time (n = 3).

Figure 2

Composition of protein groups (PGs) according to taxonomic affiliation to bacterial (blue) and eukaryotic (algae = dark green, fungi = green, various = light green) PGs in non-infected (NI) and aphelid-infected (AI) S. vacuolatus cultures over days post inoculation (DPI). Mean of three independent replicates per treatment and incubation time are indicated (mean of n = 3 ± SD).

Figure 3

Mean of relative abundances of algal, fungal and bacterial protein groups (PGs) over time. Red colors indicate higher, while green colors indicate lower protein abundances of aphelid-infected (AI) and non-infected (NI) S. vauolatus cultures 0, 4 and 7 days post inoculation (DPI). The hetamp was calculuated in the open-source platform R (v3.6.1) with the pheatmap package v1.0.12 (RRID: SCR_016418).The tree based on PG presence represents the clustering of euclidean PG distances.

Figure 4

Nonmetric dimensional scaling (NMDS) based on euclidean distances of bacterial and eucaryotic protein group (PG) abundances. S. vauolatus cultures with aphelid infection (AI, brown) and without aphelid infection (NI, green) are denoted at the start of incubation (filled circle), after 4 (filled triangle) and 7 (filled square) days post inoculation (DPI). Ordination stresses are indicated.

Figure 5

Volcano plot indicating differences in abundance of bacterial protein groups (PGs) between aphelid-infected (AI) and non-infected (NI) S. vauolatus cultures. PG fold changes (FCs) were calculated between AI and NI treatment after 4 days post inoculation (DPI) (light blue, left) and 7 DPI (dark blue, right). Log₂ fold change (Log₂FC) are plotted against − log₁₀ transformed p-values to determine significantly (p > 0.05) upregulated (FC > 1.5) and downregulated (FC < − 1.5) PGs.

Figure 6

Functional shifts in bacterial protein groups (PGs) between aphelid-infected (AI) and non-infected (NI) S. vauolatus cultures with significantly upregulated (Fold changes (FC) > 1.5, left) and downregulated (FC < − 1.5, right) PGs. PGs were categorized into functional groups (green: metabolism; blue: transcription/translation; red: posttranslational modification, for details see figure legend) based on EggNOG database by Prophane. Relative proportions of each functional role were determined to be more (197 proteins) or less abundant (226 proteins) in AI treatment compared to NI treatment.