Introduction
During the early post-settlement juvenile stages, the abalone Haliotis midae has a known ecological relationship with the Cape sea urchin Parechinus angulosus in nature. The urchins provide the juvenile abalone with shelter and protection, as well as with improved food supply and possibly provide beneficial microbial communities, all of which could act as drivers of the relationship between these species. These species are therefore strong candidate species for integrated multi-trophic aquaculture (IMTA), as they also have a similar preferred temperature range (12 - 20°C). Furthermore, hatchery-produced abalone could benefit from sea urchin faecal matter supplementation during early juvenile stages as the transfer of microbial communities could aid in digestion, growth and overall health. Therefore, the aim of this study was to evaluate the potential benefits for abalone growth, survival and subsequent changes in the microbiome associated with using sea urchin faecal matter as a supplementary feed or probiotic for juvenile abalone.
Materials and Methods
An 8 week experiment was conducted in a commercial hatchery in South Africa (Buffeljags Abalone) where two sets of three tanks, each tank (L x W x D: 0.68 m x 0.50 m x 0.12 m) with 25 000 juvenile abalone with starting shell lengths of ca. 3 mm that were fed a mixed diet of diatoms, formulated feed, Ulva lacinulata and Gracilaria. One set of tanks was only fed the mixed diet (control) and the other set was provided with a daily supplement of sea urchin faecal matter collected from 310 sea urchins per tank. Sea urchins were only fed U. lacinulata and were held in baskets in separate tanks (L x W x D: 0.68 m x 0.50 m x 0.12 m) as preliminary trials indicated that this simplifies animal husbandry practices for these species.
Juvenile abalone growth and survival was monitored for the duration of the trial. Abalone growth was assessed by measuring shell length and width (in cm) of 300 randomly chosen animals per tank at each timepoint (0 weeks, 1 week, 4 weeks and 8 weeks). Survival was assessed weekly by collecting and counting mortalities per tank. Using the Illumina MiSeq next-generation sequencing platform, a 16S rDNA and ITS2 metagenomics approach was used to characterise bacteria, fungi and oomycete communities. Metagenomic samples (n = 75) included incoming seawater (500 mL) that was collected on 0.22µm filters, sea urchin intestines and faecal matter, abalone intestines and faecal matter, as well as each of the feeds. Once raw sequence data is received, data will be processed using QIIME2 (Boylen et al., 2019). Bacterial 16S reads will be mapped against the SILVA 16S rRNA reference database (Quast et al., 2013) for taxonomic identification. Fungal and oomycete ITS2 reads will be mapped against a custom fungal and oomycete database that included the UNITE database (Abrenkov et al., 2021) and oomycete sequences downloaded from the National Center of Biotechnology Information (NCBI). MicrobiomeAnalyst will be used to assess within- and between sample diversity, as well as to quantify and visualise taxonomic abundance and differential abundance analyses (DESeq2). Lastly, the functional capabilities of the identified taxa will be assessed.
Results & Discussion
Results from the large-scale trial showed that the shell length of juvenile abalone were statistically significantly larger (P < 0.05; one-way ANOVA) at 1 week, 4 weeks and 8 weeks (Figure 1), with shell width being statistically significantly larger at 1 week and 4 weeks.
Although there were limited statistically significant differences in survival, the growth results suggest that the faecal matter and possibly the microbes transferred from the urchins to the abalone provides an advantage in the early post-weaning stages. This trophic transfer of microbial communities will be further assessed in this study. Enteric bacteria are thought to play roles in the synthesis of essential amino acids, provisioning of digestive enzymes, improvement of digestive efficiency, nitrogen metabolism, as well as access to essential micronutrients.
Conclusion
Supplementing juvenile abalone diets with sea urchin faecal matter is advantageous for abalone growth and health in their early juvenile stages, possibly as a result of microbial community transfer from the sea urchins. The intestinal microbiome is often established during the juvenile stages for aquatic animals and could have long-lasting effects on digestion, overall performance and health, possibly through the microbial provisioning of digestive enzymes. This study will provide insight on the potential benefits and effects of supplementing juvenile abalone diets with sea urchin faeces.
References
Abarenkov K., Zirk A., Piirmann T., Pöhönen R., Ivanov F., Nilsson R.H., Kõljalg U., 2021. UNITE QIIME release for Fungi. Version 10.05.2021.
Bolyen E., Rideout J.R., Dillon M.R., Bokulich N.A., Abnet C.C., Al-Ghalith G.A., Alexander H., et al. 2019. Reproducible, interactive, scalable and extensible microbiome data science using QIIME 2. Nature Biotechnology 37, 852–857.
Quast C., Pruesse E., Yilmaz P., Gerken J., Schweer T., Yarza P., Peplies J., Glöckner F.O., 2013. The SILVA ribosomal RNA gene database project: improved data processing and web-based tools. Nucleic Acids Research 41(D1), D590–D596.
This study received funding from the EU Horizon 2020 Research & Innovation Programme ASTRAL Project under Grant Agreement No. 863034