Sea lettuce (Ulva spp.), with its worldwide distribution and remarkable ability to grow rapidly under various conditions, represents an important natural resource that is still under-exploited. Its biomass can be used for a wide range of applications in the food/feed, pharmaceutical, nutraceutical, biofuel, and bioremediation industries. However, knowledge of the factors affecting Ulva biomass yield and composition is far from complete. Indeed, the respective contributions of the microbiome, natural genetic variation in Ulva species, environmental conditions and importantly, the interactions between these three factors on the Ulva biomass, have been only partially elucidated. Further investigation is important for the implementation of large-scale Ulva aquaculture, which requires stable and controlled biomass composition and yields. In this review, we document Ulva biomass composition, describe the uses of Ulva biomass and we propose different strategies for developing a sustainable and profitable Ulva aquaculture industry.

Green macroalgae of the genus Ulva play a key role in coastal ecosystems and are of increasing commercial importance. However, physiological differences between strains and species have yet to be described in detail. Furthermore, the strains of Ulva used in aquaculture usually originate from opportunistic collection in the wild without prior selection of best performing strains. Hence, efforts are required to detect the potential variability in growth and metabolic accumulation between Ulva strains and ultimately select the best performing strains under given environmental conditions. Here, the growth, physiological, and metabolic characteristics of 49 laminar Ulva spp. strains were investigated using a custom-made high-throughput phenotyping platform, enzymatic assays, and gas chromatography-mass spectrometry. We found large natural variation for a wide range of growth and metabolic characteristics, with growth rates varying from 0.09 to 0.37 mg.mg⁻¹.d⁻¹ among strains. Ulva spp. possess a unique diurnal growth pattern and primary metabolism compared with land plants, with higher growth rates during the night than during the light period. Starch and sucrose only contributed on average 35% of the carbon required to sustain Ulva’s night growth. Nitrates accumulated during the night in Ulva tissues, and nitrate accumulation and consumption was positively correlated with growth. In addition, we identified six amino acids as possible biomarkers for high growth in Ulva. The large variability in growth and metabolite accumulation recorded among morphologically similar Ulva strains justifies future efforts in strain selection for increasing biomass, metabolite yields, and nutrient removal in the growing aquaculture industry.

It is known that 22-nucleotide (nt) microRNAs (miRNAs) derived from asymmetric duplexes trigger phased small-interfering RNA (phasiRNA) production from complementary targets. Here we investigate the efficacy of 22-nt artificial miRNA (amiRNA)-mediated RNA silencing relative to conventional hairpin RNA (hpRNA) and 21-nt amiRNA-mediated RNA silencing. CHALCONE SYNTHASE (CHS) was selected as a target in Arabidopsis thaliana due to the obvious and non-lethal loss of anthocyanin accumulation upon widespread RNA silencing. Over-expression of CHS in the pap1-D background facilitated visual detection of both local and systemic RNA silencing. RNA silencing was initiated in leaf tissues from hpRNA and amiRNA plant expression vectors under the control of an Arabidopsis RuBisCo small subunit 1A promoter (SSU). In this system, hpRNA expression triggered CHS silencing in most leaf tissues but not in roots or seed coats. Similarly, 21-nt amiRNA expression from symmetric miRNA/miRNA* duplexes triggered CHS silencing in all leaf tissues but not in roots or seed coats. However, 22-nt amiRNA expression from an asymmetric duplex triggered CHS silencing in all tissues, including roots and seed coats, in the majority of plant lines. This widespread CHS silencing required RNA-DEPENDENT RNA POLYMERASE6-mediated accumulation of phasiRNAs from the endogenous CHS transcript. These results demonstrate the efficacy of asymmetric 22-nt amiRNA-directed RNA silencing and associated phasiRNA production and activity, in mediating widespread RNA silencing of an endogenous target gene. Asymmetric 22-nt amiRNA-directed RNA silencing requires little modification of existing amiRNA technology and is expected to be effective in suppressing other genes and/or members of gene families.

The 45S rRNA genes (rDNA) are among the largest repetitive elements in eukaryotic genomes. rDNA consists of tandem arrays of rRNA genes, many of which are transcriptionally silenced. Silent rDNA repeats may act as ‘back-up’ copies for ribosome biogenesis and have nuclear organization roles. Through Cas9-mediated genome editing in the Arabidopsis thaliana female gametophyte, we reduced 45S rDNA copy number (CN) to a plateau of ∼10%. Two independent lines had rDNA CNs reduced by up to 90% at the T7 generation, named low copy number (LCN) lines. Despite drastic reduction of rDNA copies, rRNA transcriptional rates, and steady-state levels remained the same as wild-type plants. Gene dosage compensation of rRNA transcript levels was associated with reduction of silencing histone marks at rDNA loci and altered Nucleolar Organiser Region 2 organization. Although overall genome integrity of LCN lines appears unaffected, a chromosome segmental duplication occurred in one of the lines. Transcriptome analysis of LCN seedlings identified several shared dysregulated genes and pathways in both independent lines. Cas9 genome editing of rRNA repeats to generate LCN lines provides a powerful technique to elucidate rDNA dosage compensation mechanisms and impacts of low rDNA CN on genome stability, development, and cellular processes.

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.