Seed banking impacts native Acacia ulicifolia seed microbiome composition and function

Abstract

Background: Seed banks are a vital resource for preserving plant species diversity globally. However, seedling establishment and survival rates from banked seeds can be poor. Despite a growing appreciation for the role of seed-associated microbiota in supporting seed quality and plant health, our understanding of the effects of conventional seed banking processes on seed microbiomes remains limited. In this study we investigated the composition and functional potential of seed-associated bacterial epiphytes associated with stored and freshly collected seeds of a native plant, Acacia ulicifolia, using both 16S rRNA gene sequencing and culture-based approaches.

Results: Seeds obtained from seed banking facilities were found to host significantly less diverse bacterial populations, with substantial reductions in both low-abundance taxa and in community members commonly identified in freshly collected A. ulicifolia seeds. Bacteria with key plant growth promoting traits including IAA production, ACC deaminase activity, phosphate solubilisation, siderophore activity, and nitrogen fixation were identified in seed epiphytic communities, but these beneficial traits were less prevalent in stored seed compared to fresh seeds.

Conclusion: Overall, these results suggest that epiphytic seed microbiomes may undergo significant changes during the storage process, selecting for bacteria tolerant to storage conditions, and potentially reducing the population of plant-growth promoting bacteria on seeds.

Keywords: Bacterial traits; Conservation; Native seed microbiota; Plant growth-promoting traits; Seed epiphytes; Seed storage.

Fig. 1

Indicative image of A. ulicifolia (A) opened seed pod, (B) unripe unopened fruit and (C) map of locations from which seed samples were obtained in this study. Sample collection group indicated by colour. Yellow: Samples freshly collected from natural populations for this study. Green: sources of samples collected and later stored by Greening Australia. Green/Yellow: location where fresh collection and Greening Australia samples have shared geographic source. Blue: sources of samples collected and later stored by Australian PlantBank

Fig. 2

Acacia ulicifolia seed-associated epiphytic community alpha and beta diversity and number of shared OTUs observed in fresh and stored seed from different banking organisations. A Diversity of microbial communities was assessed based on Shannon diversity indices, Chao1 species richness, and Inverse Simpson evenness for fresh and stored seed (observed richness shown in Figure S5). B Principal coordinate analysis indicating differences in overall seed epiphyte bacterial community composition between fresh and stored seed (colour) and sampling location (points represented by different symbols). C Venn diagram indicating the number and proportion of unique and shared OTUs between the seed microbiomes of samples from different seed

Fig. 3

Composition of epiphytic bacterial communities from A. ulicifolia seeds at different taxonomic levels. A Relative abundance of the most predominant bacterial taxa at phylum and B family taxonomic levels for each sample location and stored seed lot, with less abundant taxa combined and entitled ‘others’. C Heatmap and dendrogram of the 75 most abundant genera in A. ulicifolia seed-associated bacterial communities showing clustering of stored (red) and fresh (aqua) samples

Fig. 4

Phylogenetic analysis of 16S rRNA gene sequences from bacterial isolates recovered from A. ulicifolia seeds. Phylogenetic tree inferred from maximum likelihood (RAxML) analysis and taxonomic assignments based on classification in the SILVA (v. 138) database using the Alignment, Classification and Tree (ACT) tool. Putative plant growth promoting functions for each isolate determined in vitro indicated by green circles for each trait tested (BNF = biological nitrogen fixation, Phos. = phosphate solubilisation, IAA = Indole-3-acetic acid production, Sid. = Siderophore production, ACC = 1-aminocyclopropane carboxylic acid deaminase activity). Isolates recovered from stored and fresh seeds are indicated by red and blue circles respectively

Fig. 5

Predicted set of differentially-abundant plant growth-promoting functions inferred using Tax4fun2 and applying linear discriminant analysis (LDA). The Functional classification are taken from the PLaBAse plant growth promoting trait ontology [53]

Fig. 6

Microbial cell counts of A. ulicifolia seed epiphytes determined by (A) flow cytometry and (B) colony forming unit counts. Counts are reported as mean (n = 12) values for each fresh seed sampling location (aqua) or stored sample group (red). Significant differences, as determined by Kruskal–Wallis tests, are denoted unshared letters