DNA barcoding and phylogenetic analysis of five ascidians (Phlebobranchia) distributed in Gulf of Mannar, India.

DNA barcoding has played a significant role in biodiversity assessment as well as its conservation. This technique involves sequencing of mitochondrial marker gene including a short COI gene, known as barcode gene. It has proved its efficiency in identifying several species and resolving the limitations incurred during identification through conventional taxonomy. This study involves the use of DNA barcoding of ascidian species belonging to order Phlebobranchia. A total of 14 individuals, covering two families, three genera and five species, were barcoded. COI gene sequences of all the five species were deposited for the first time in NCBI as well as BOLD. The NJ tree revealed identical phylogenetic relationship among the individuals collected from three different stations. Mean Kimura 2-parameter (K2P) distances within-species, genus, family and order were 0.08%, 6.69%, 9.49% and 18.58%, respectively. This result concludes that COI gene sequencing is the efficient tool in identifying ascidians of the order Phlebobranchia. We report for the first time the COI gene sequences of four species of ascidians studied.

Identification of four Indian ascidians based on COI gene sequences.

DNA barcoding involving the sequencing of a short mitochondrial DNA segment, cytochrome c oxidase subunit 1 (COI) gene, is a specialized technique for the identification of species even at the early embryonic and larval stages, which is quite difficult in morphology-based taxonomy. Ascidians are sessile invertebrate chordates possessing numerous biochemical as well as pharmacological activities. In this study, a total of 36 ascidian samples belonging to the family Didemnidae were sequenced for a 650 bp region of the mitochondrial COI gene. All the species were represented by multiple specimens. The barcode sequences showed no stop-codons and indels in the alignments. The aligned sequences were submitted in Barcode submission tool, NCBI, and the accession numbers were obtained. The minimum intraspecific distance was found to be 0.00% and the maximum was 2.23%. Mean Kimura 2-parameter (K2P) distances within-species, genus, and family were 0.88, 5.98, and 20.03%, respectively. The mean interspecific distance is more than the mean intraspecific divergence, which indicates efficiency of DNA barcoding for identification of ascidians.

DNA barcoding of a colonial ascidian, Lissoclinum fragile (Van Name, 1902).

Ascidians (tunicates) are marine benthic organisms possessing various pharmacological activities, including anti-oxidant, anti-tumour, antimicrobial, etc. They also play a key role as model organisms to study various neurobehavioral disorders. Ascidian diversity is reportedly less in India due to lack of taxonomists as well as the limitations in morphology based taxonomy. Molecular taxonomy, comprising the sequencing of cytochrome c oxidase 1 gene (barcode region) otherwise known as DNA barcoding reduces these bottlenecks. Since several species of the family Didemnidae closely resemble in morphology, the present study was aimed to develop DNA barcodes of a colonial ascidian, Lissoclinum fragile belonging to the family Didemnidae. CO1 gene of L. fragile from Thoothukudi, Mandapam, and Vizhinjam waters were sequenced and submitted in GenBank, NCBI through Barcode submission tool. BLAST results showed maximum identity (97-100%) for L. fragile collected from different stations. The pairwise genetic distances within species and genera were calculated using Kimura two parameter (K2P) and the phylogenetic tree was constructed using Neighbour-Joining Tree.

New distributional data on ascidian fauna (Tunicata: Ascidiacea) from Mandapam coast, Gulf of Mannar, India.

BACKGROUND: Ascidians play a key role in the ecology and biodiversity of marine ecosystem. Ascidians can be transported in ship ballast water and while attached to ship and boat hulls. Heavy traffic by domestic and international ships as well as cargo vessels between the major and minor ports warrants continuous monitoring for new introductions of ascidians. The Mandapam coast is situated in the Gulf of Mannar, India, a marine hot spot area in the Indian Ocean which provides an environment suitable for the settlement of ascidians. NEW INFORMATION: A total of 30 species of ascidians were reported from Mandapam coastal waters, of which 26 species were new to the study area and five species: Ecteinascidia turbinata, Eudistoma carnosum, Trididemnum caelatum, T. vermiforme and Didemnum spadix, were new to India.