Development of a multilocus-based approach for sponge (phylum Porifera) identification: refinement and limitations

Abstract

For sponges (phylum Porifera), there is no reliable molecular protocol available for species identification. To address this gap, we developed a multilocus-based Sponge Identification Protocol (SIP) validated by a sample of 37 sponge species belonging to 10 orders from South Australia. The universal barcode COI mtDNA, 28S rRNA gene (D3-D5), and the nuclear ITS1-5.8S-ITS2 region were evaluated for their suitability and capacity for sponge identification. The highest Bit Score was applied to infer the identity. The reliability of SIP was validated by phylogenetic analysis. The 28S rRNA gene and COI mtDNA performed better than the ITS region in classifying sponges at various taxonomic levels. A major limitation is that the databases are not well populated and possess low diversity, making it difficult to conduct the molecular identification protocol. The identification is also impacted by the accuracy of the morphological classification of the sponges whose sequences have been submitted to the database. Re-examination of the morphological identification further demonstrated and improved the reliability of sponge identification by SIP. Integrated with morphological identification, the multilocus-based SIP offers an improved protocol for more reliable and effective sponge identification, by coupling the accuracy of different DNA markers.

Figure 1. Sequence data processing flowchart.

Figure 2. The Sponge Identification Protocol (SIP) flow diagram used in this study.

Figure 3. Phylogenetic relationship of three representative sponge species using the Maximum Likelihood method based on 28S rRNA gene and COI mtDNA to validate the sponge identification by SIP.

(a) Phylogenetic relationship of sponge SAMA S1981 based on 28S rRNA gene. (b) Phylogenetic relationship of sponge SAMA S1981 based on COI mtDNA. (c) Phylogenetic relationship of sponge SAMA S1962 based on 28S rRNA gene. (d) Phylogenetic relationship of sponge SAMA S1962 based on COI mtDNA. (e) Phylogenetic relationship of sponge SAMA S1960 based on 28S rRNA gene. (f) Phylogenetic relationship of sponge SAMA S1960 based on COI mtDNA.

Figure 4. The sampling locations and the photos of the 37 sponge species.

(a) Four sampling locations (A–D), Map data: Google Earth. (b) Photos of the 37 sponge species (underwater photos courtesy of David Wiltshire).