Periodic Reporting for period 1 - DeepSym (Understanding the drivers of the genetic and functional structure of deep-sea sponge symbiont communities) Reporting period: 2018-05-01 to 2020-04-30 Summary of the context and overall objectives of the project Sponges are known for hosting dense, diverse, and highly specific microbial communities that contribute appreciably to the host’s metabolism and biochemical repertoire. The sponge associated microbial community varies within and among species, and this variation can be attributed to biotic and abiotic factors. The major aims of DeepSym were: first, to determine whether the intra-species variability of the microbiome is driven by an environmental or a host genotype effect. And second, to characterise microbiome composition and functions of deep-sea sponges, and identify functional redundancy among different sponge species and locations. This is important for society in both general and specific ways: a) sponges are keystone species in marine environments, essential for the ocean’s health due to their role in both habitat formation and benthic-pelagic coupling. Maintenance of a healthy and balanced community of symbionts is critical for their survival, therefore it is essential to improve our understanding on the microbial composition and functioning if we aim at guaranteeing the survival of sponge communities in changing environments. b) The resources put in place by this project will be useful for a range of future work in sponges and more generally in the investigation of deep-sea sponge grounds. Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far WP 1: Functional roles of symbiotic community of deep-sea spongesWe sequenced the 16S rRNA gene and metatranscriptomes of 18 samples representing three species of sponge in one location, and one species in 4 different locations of the Cantabrian Sea (deep-sea water). The assembled reference metatranscriptome has been uploaded to the public repository IMG/MER. Functional pathways were identified using KEGG Mapper, and core taxa and functions have been identifies for host species, and for different locations. High expression of genes related with methane metabolism, purine metabolism, benzoate degradation, and carbon fixation pathways were found among the associated bacterial members. A large percentage of transcripts were assigned to ABC transporters and Two-component systems within the membrane transport pathway, pointing to the importance of metabolite exchange between symbionts and host. Interestingly, microbial composition (16S rRNA amplicon) was congruent with the host taxonomy, and therefore followed the well-known host-specificity of the sponge microbiome, however, transcriptome sequencing revealed that two species of the same genus showed a more different symbiotic functionality than two sponges of different genera.WP 2: Host genetic structure and environment effect on functional profilesTo disentangle the influence of the host genotype versus an environmental effect in the microbiome variation within sponge species, we sequenced the 16S rRNA gene of 393 shallow water sponges covering a range of different population structures, from weak structure (high gene flow) to high population structure. Our results showed that in sponge species with high population structure, the host genetic cluster influenced the microbial community more than the geographic location. WP 3: Functional equivalence of microbial taxa The taxonomic annotation of all microbial transcripts (1,2 million assembled transcripts) resulted in 537,117 different prokaryotic taxa (using UniProt database). Functional profiles of the communities of the 3 sponge species were quite similar, even though they hosted different bacterial communities, revealing that the active community perform equivalent functions. Groups of orthologous genes included 3.5% of the metatranscriptome sequences, which represented examples of functional redundancy. WP 4: Molecular signatures for symbiosis Eukaryotic-like protein were identified from the PFAM annotation of the translated transcripts. We identified 4,228 ELPs containing transcripts, with similar levels of expression among sponge species or locations. A total of 3,575 transcripts were identified as differentially expressed (DE) between sponge samples. WP5: Co-ordination and management of researchMeetings were held on a regular basis in the Riesgo lab, with informal discussion occurring on an almost daily basis. Risks were managed appropriately and the project delivered efficiently. The fellow attended a variety of training sessions held for postdoctoral fellows and mixed groups at the NHM. WP6: Outreach activities and the communication of results to a wider audienceNumerous opportunities arose to communicate work directly to the general public at events such as the European Researchers’ Night, Nature Live talks and more informally. To reach a more focused scientific community, a number of conferences and symposia were attended in the course of the project. Six papers have been published during the duration of the project, with a few more in process. Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far) The two year DeepSym project lead to the generation of a number of fundamental molecular resources from deep-sea and shallow sponges species, for the examination of the community composition and function of the sponge microbiome. These findings will be useful to understand the roles and evolution of the sponge microbiome, which is essential to interpret the impact of different climate change scenarios and the resilience of the sponge communities. This will be useful for policy makers, as well as of interest to the general public and scientific audiences. The project has led to 6 publications available at the time of writing, with several other publications under review or in process of finishing the manuscript. As part of the project, the researcher gave a invited talk at the Gordon Research Conference and few talks and courses within the Natural History Museum. It also allowed the fellow to present her results to the general public at a number informal events in the host institution, where she delivered ‘Nature Live’ talks and met the public at "European Research Night" events. This fellowship also led to a number of learning and professional advancement opportunities for the researcher employed on this project. These included additional training in bioinformatic analyses, increased ‘wet lab’ experience, opportunities to teach and communicate her science, and formal training in a number of “soft skills” that allowed her to continue her professional development in a manner that will aid her future prospects in research and assist her in her career path. Giving a talk in the ‘Nature Live’ event at the Natural History Museum with Dr. Riesgo.