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Nitrogen cycle is a critical biogeochemical process of the oceans. The nitrogen fixation by sponge cyanobacteria was early observed. Until recently, sponges were found to be able to release nitrogen gas. However the gene-level evidence for the role of bacterial symbionts from different species sponges in nitrogen gas release is limited. And meanwhile, the quanitative analysis of nitrogen cycle-related genes of sponge microbial symbionts is relatively lacking. The  nirK  gene encoding nitrite reductase which catalyzes soluble nitrite into gas NO and  nosZ  gene encoding nitrous oxide reductase which catalyzes N 2 O into N 2  are two key functional genes in the complete denitrification pathway. In this study, using  nirK  and  nosZ  genes as markers, the potential of bacterial symbionts in six species of sponges in the release of N 2  was investigated by phylogenetic analysis and real-time qPCR. As a result, totally, 2 OTUs of  nirK  and 5 OTUs of  nosZ  genes were detected by gene library-based saturated sequencing. Difference phylogenetic diversity of  nirK  and  nosZ  genes were observed at OTU level in sponges. Meanwhile, real-time qPCR analysis showed that  Xestospongia testudinaria  had the highest abundance of  nosZ  gene, while  Cinachyrella  sp. had the greatest abundance of  nirK  gene. Phylogenetic analysis showed that the  nirK  and  nosZ  genes were probably of  Alpha-, Beta-,  and  Gammaproteobacteria  origin. The results from this study suggest that the denitrification potential of bacteria varies among sponges because of the different phylogenetic diversity and relative abundance of  nosZ  and  nirK  genes in sponges. Totally, both the qualitative and quantitative analyses of  nirK  and  nosZ  genes indicated the different potential of sponge bacterial symbionts in the release of nitrogen gas.

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The Different Potential of Sponge Bacterial Symbionts in N2 Release Indicated by the Phylogenetic Diversity and Abundance Analyses of Denitrification Genes, nirK and nosZ