Foliose Ulva Species Show Considerable Inter-Specific Genetic Diversity, Low Intra-Specific Genetic Variation, and the Rare Occurrence of Inter-Specific Hybrids in the Wild

Abstract

Foliose Ulva spp. have become increasingly important worldwide for their environmental and financial impacts. A large number of such Ulva species have rapid reproduction and proliferation habits, which explains why they are responsible for Ulva blooms, known as "green tides", having dramatic negative effects on coastal ecosystems, but also making them attractive for aquaculture applications. Despite the increasing interest in the genus Ulva, particularly on the larger foliose species for aquaculture, their inter- and intra-specific genetic diversity is still poorly described. We compared the cytoplasmic genome (chloroplast and mitochondrion) of 110 strains of large distromatic foliose Ulva from Ireland, Brittany (France), the Netherlands and Portugal. We found six different species, with high levels of inter-specific genetic diversity, despite highly similar or overlapping morphologies. Genetic variation was as high as 82 SNPs/kb between Ulva pseudorotundata and U. laetevirens, indicating considerable genetic diversity. On the other hand, intra-specific genetic diversity was relatively low, with only 36 variant sites (0.03 SNPs/kb) in the mitochondrial genome of the 29 Ulva rigida individuals found in this study, despite different geographical origins. The use of next-generation sequencing allowed for the detection of a single inter-species hybrid between two genetically closely related species, U. laetevirens, and U. rigida, among the 110 strains analyzed in this study. Altogether, this study represents an important advance in our understanding of Ulva biology and provides genetic information for genomic selection of large foliose strains in aquaculture.

Keywords: Ulva; macroalgae; next-generation sequencing; phylogeny; sexual reproduction.

Fig. 1

Samples analyzed in this study. (A) Clustering of strains based on their chloroplast genome. Principal Component Analysis (PC1 and PC5) on SNP data for each strain against the chloroplast genome of strain U41’s nanopore assembly. Full PCA available in Figure S2. Shading represents the six clusters found. (B) Phylogram of the 110 sampled individuals based on rbcL and tufA concatenated sequences based on MCMC Bayesian analysis. Numbers represent support for clade clusters (first number: Bayesian posterior probability of clades, second number: GMYC support, third number: Bayesian Poisson Tree Processes support). Triangles represent the assigned species to the cluster. Scale bar = expected changes per site. (C) Sampling locations and relative abundance of each foliose Ulva species. Numbers represent the six clusters. Full list and detailed coordinates available in Table S1.

Fig. 2

Ulva organelles assembly and annotation. (A) Annotation of U. laetevirens chloroplast. B) Annotation of U. laetevirens mitochondrion. The annotation of the other species is available in Figures S3 and S4. (C) Detected inversion between U. laetevirens and U. fenestrata chloroplasts. Inversion is shaded in dark, syntenic blocs in the same orientation are shaded in gray. Ideogram represent coding sequence (genes), black shading indicates plus strand, gray shading the minus strand. Numbers indicate positions in kilobases (D) Variant sites (SNPs) detected within the chloroplast of the 11 U. fenestrata samples. Line represents variant sites, and their relative abundance between strains. Histogram represents GC content, ideogram same as (A).

Fig. 3

Agreement between barcoding and organelle coding sequence phylogenetic trees. Maximum Likelihood trees were constructed from the sequences of rbcL, tufA, and the entire coding sequence of the organelles. Triangles indicate species clusters, numbers indicate the bootstrap support (left) and Bayesian posterior probability (right) for the major nodes of the tree.

Fig. 4

Detection of a single hybrid among the 110 foliose Ulva strains. (A) Maximum Likelihood phylogenetic tree based on the 45S nuclear sequence. Triangles indicate the inferred species of the strains from the organellar (cytoplasmic) data. Numbers indicate the bootstrap support for the major nodes (left) and Bayesian posterior probability (right). (B) Principal Component Analysis of SNPs detected in the 45S sequence. Strain U99 is circled. Numbers represent species clusters. (C) Coverage visualization of illumina reads on the 45S locus between a single strain of U. laetevirens and U. rigida, as well as the hybrid strain U99. Bars represent SNPs between the two species and their relative ratios.