Establishment of a marine nematode model for animal functional genomics, environmental adaptation and developmental evolution

Abstract

Nematodes play key roles in marine ecosystem. Although oceans cover 71% of the Earth's surface, none of marine model nematode has been reported. Here, we constructed the first inbred line of free-living marine nematode Litoditis marina, sequenced and assembled its genome. Furthermore, we successfully applied CRISPR/Cas9 genome editing in L. marina.Comparative genomics revealed that immunity and oxygen regulation genes are expanded, which is probably central to its sediment adaptation. While L. marina exhibits massive gene contractions in NHRs, chemoreceptors, xenobiotics detoxification and core histones, which could explain the more defined marine environment. Our experiments showed that dozens of H4 genes in Caenorhabditis elegans might contribute to its adaptation to the complex terrestrial environments, while two H4 genes in L. marina are involved in salinity stress adaptation. Additionally, ninety-two conserved genes appear to be positively selected in L. marina, which may underpin its osmotic, neuronal and epigenetic changes in the sea. With short generation time, highly inbred lines, and genomic resources, our report brings L. marina a promising marine animal model, and a unique satellite marine model to the wellknown biomedical model nematode C. elegans. This study will underpin ongoing work on animal functional genomics, environmental adaptation and developmental evolution. 4 marine nematodes have had their genomes sequenced and analysed (International Helminth Genomes Consortium 2019; Smythe et al. 2019; Weinstein et al. 2019). Oceans cover 71% of the Earth's surface, and represents almost 99% of available habitat, which are a key element for the existence and evolution of life (Robert 1999; Peng et al. 2020). In marine sediments, free-living nematodes abound both in numbers and in local species diversity, comprising about 80% of the abundance of meiofauna (Heip et al. 1985; Danovaro et al. 2010; Nascimento et al. 2012), they play a key role in the benthic food web and ecosystem. However, the study of marine nematodes is largely limited to taxonomy and ecology (De Meester et al. 2016; Derycke et al. 2016). Molecular mechanisms underlying their evolution, development regulation and adaptation mechanisms are rarely reported, and none of marine model nematode has been reported. The bacterivorous marine nematode Litoditis marina (Bastian, 1865) Sudhaus, 2011, formerly known as Rhabditis marina or Pellioditis marina, is found widely distributed in the littoral zone of coasts and estuaries of European, American, African and Asian countries, and play an important role in these marine ecosystems (Tietjen et al. 1970; Kito 1981; Houthoofd et al. 2003; Derycke et al. 2016). In addition, the embryonic cell lineage of L. marina has been reported by Houthoofd et al. (2003). In comparison with C. elegans, the overall L. marina lineage homology is 95.5%, whereas the fate homology is only 76.4%, and most of the differences in fate homology concern nervous, epidermal, and pharyngeal tissues, suggesting that L. marina wil...

Genome-Wide Transcriptional Responses of Marine Nematode Litoditis marina to Hyposaline and Hypersaline Stresses

“…The wild strain of marine nematode L. marina , HQ1, was isolated from intertidal sediments (Huiquan Bay, Qingdao). Healthy worms were cultured on SW-NGM agar plates (prepared with seawater with a salinity of 3%) seeded with Escherichia coli OP50 as a food source, as reported previously ( Xie et al, 2020 ). Worms were maintained and propagated at 20°C in the laboratory for about 3 years till this study.…”

“…The minimum of base score Q20 was over 97.5% and Q30 was over 93.13%. Then, the clean data were aligned to the L. marina reference genome ( Xie et al, 2020 ) by Hisat2 (v2.0.5, with the default parameters) ( Kim et al, 2015 ), with mapping ratio from 66.91 to 69.72% ( Supplementary Table 1 ). New transcripts for novel genes were predicted and assembled by StringTie (v1.3.3b, with the default parameters) ( Pertea et al, 2015 ), then annotated with Pfam, SUPERFAMILY, Gene Ontology (GO) and KEGG databases.…”

“…Litoditis marina is a dioecious free-living marine nematode, which is widely distributed in the littoral zone of coasts and estuaries, and plays an important role in these marine ecosystems ( Derycke et al, 2016 ; Xie et al, 2020 ). It possesses some promising characteristics similar as C. elegans , such as short generation time, clear genetic background and a sequenced genome ( Xie et al, 2020 ), which facilitated its laboratory application for the in-depth study of molecular biology, cell biology, physiology and behavior regulation in this species. Generally, the habitat salinity for intertidal marine nematodes, including L. marina , is frequently changed due to the influence of many factors such as tides, sun exposure, rainfall, ocean currents and climate.…”

Genome-wide transcriptional responses of marine nematodeLitoditis marinato hyposaline and hypersaline stresses

“…The wild strain of marine nematode L. marina , HQ1, was isolated from intertidal sediments (Huiquan Bay, Qingdao). Healthy worms were cultured on SW-NGM agar plates (prepared with seawater with a salinity of 30‰) seeded with a lawn of Escherichia coli OP50 as a food source, as reported previously (Xie et al, 2020). Worms were maintained and propagated at 20°C in the laboratory for about 3 years till this study.…”

“…Clean data, with Q20 value higher than 97.5 for each sample, were first obtained by removing reads containing sequencing adaptors, reads having poly-N and low-quality ones from raw data. Then, they were aligned to the L. marina reference genome (Xie et al, 2020) by Hisat2 (v2.0.5) (Kim et al, 2015). New transcripts for novel genes were predicted and assembled by StringTie (v1.3.3b) (Pertea et al, 2015), then annotated with Pfam, SUPERFAMILY, GO and KEGG databases (Kanehisa and Goto, 2000;Young et al, 2010).…”