The Structure and Function of Gut Microbiomes of Two Species of Sea Urchins, Mesocentrotus nudus and Strongylocentrotus intermedius, in Japan

Sea urchin is an indicator of coastal environmental changes in the global warming era, and is also a model organism in developmental biology and evolution. Due to the depletion of wild resources, new aquaculture techniques for improving stocks have been well studied. The gut microbiome shapes various aspects of a host’s physiology. However, these microbiome structures and functions on sea urchins, particularly Mesocentrotus nudus and Strongylocentrotus intermedius which are important marine bioresources commonly found in Japan, have not been fully investigated yet. Using metagenomic approaches including meta16S and shotgun metagenome sequencings, the structures, functions, and dynamics of the gut microbiome of M. nudus and S. intermedius, related to both habitat environment and host growth, were studied. Firstly, a broad meta16S analysis revealed that at the family level, Psychromonadaceae and Flavobacteriaceae reads (38–71%) dominated in these sea urchins, which is a unique feature observed in species in Japan. Flavobacteriaceae reads were more abundant in individuals after rearing in an aquarium with circulating compared to one with running water. Campylobacteraceae and Vibrionaceae abundances increased in both kinds of laboratory-reared sea urchins in both types of experiments. 2-weeks feeding experiments of M. nudus and S. intermedius transplanted from the farm to laboratory revealed that these gut microbial structures were affected by diet rather than rearing environments and host species. Secondly, further meta16S analysis of microbial reads related to M. nudus growth revealed that at least four Amplicon Sequence Variant (ASV) affiliated to Saccharicrinis fermentans, which is known to be a nitrogen (N2) fixing bacterium, showed a significant positive correlation to the body weight and test diameter. Interestingly, gut microbiome comparisons using shotgun metagenome sequencing of individuals showing higher and lower growth rates revealed a significant abundance of “Nitrate and nitrite ammonification” genes in the higher-grown individuals under the circulating water rearing. These findings provide new insights on the structure-function relationship of sea urchin gut microbiomes beyond previously reported nitrogen fixation function in sea urchin in 1950s; we discovered a nitrate reduction function into ammonium for the growth promotion of sea urchin.

Application of SNP in Genetic Sex Identification and Effect of Estradiol on Gene Expression of Sex-Related Genes in Strongylocentrotus intermedius

Sea urchin (Strongylocentrotus intermedius) is an economically important mariculture species in Asia, and its gonads are the only edible part. The efficiency of genetic breeding in sea urchins is hampered due to the inability to distinguish gender by appearance. In this study, we first identified a sex-associated single nucleotide polymorphism (SNP) by combining type IIB endonuclease restriction site-associated DNA sequencing (2b-RAD-seq) and genome survey. Importantly, this SNP is located within spata4, a gene specifically expressed in male. Knocking down of spata4 by RNA interference (RNAi) in male individuals led to the downregulation of other conserved testis differentiation-related genes and germ cell marker genes. We also revealed that sex ratio in this validated culture population of S. intermedius is not 1:1. Moreover, after a 58-day feeding experiment with estradiol, the expression levels of several conserved genes that are related to testis differentiation, ovary differentiation, and estrogen metabolism were dynamically changed. Taken together, our results will contribute toward improving breeding efficiency, developing sex-controlled breeding, and providing a solid base for understanding sex determination mechanisms in sea urchins.