Phylogeographic and Morphological Analysis of Botrylloides niger Herdman, 1886 from the Northeastern Mediterranean Sea

Abstract

Botrylloides niger (class Ascidiacea) is an invasive marine filter-feeding invertebrate that is believed to originate from the West Atlantic region. This species of colonial tunicate has been observed in several locations along the coasts of Israel and around the Suez Canal, but it has not yet been reported on the coasts of the Northeastern Mediterranean Sea (NEMS), suggesting an ongoing Lessepsian migration. However, the extent of this invasion might be concealed by reports of other potentially misidentified species of Botrylloides, given that the strong morphological similarities within this genus renders taxonomical identification particularly challenging. In this study, we performed a phylogeographic and morphological analysis of B. niger in the NEMS. We collected 238 samples from 8 sampling stations covering 824 km of the coastlines of NEMS. We reported 14 different morphotypes, of which the orange-brown, orange, and brown-striped morphs were the most abundant. Using the mitochondrial cytochrome C oxidase I (COI) as a DNA barcode marker, we identified 4 haplotypes. The COI haplotypes clustered with the reference B. niger sequences from GenBank and differed significantly from the sister Botrylloides species. We confirmed our identification using three additional barcoding markers (Histone 3, 18S rRNA, and 28S rRNA), which all matched with over 99% similarity to reference sequences. In addition, we monitored a station for a year and conducted a temporal analysis of the collected colonies. The colonies were absent during the winter and spring, while new colonies were established in the summer and expanded during autumn. We performed demographic population analysis on our spatial data that identified a possible population subdivision at a sampling site, which might have been caused by local freshwater input. Herein, we present the first report on the presence of Botrylloides niger in the NEMS. This study represents a key step toward understanding the diversity and the propagation of this highly invasive species of colonial ascidians, both within the Mediterranean basin as well as globally.

Seven ascidian (Tunicata) species from the North Eastern Mediterranean

“…While the distribution and characteristics of Botryllus schlosseri (Pallas, 1766) clades are well-known (Reem et al, 2022), there are limited data available from the Turkish coastline, with the exception of a presence record from the Black Sea coast of Türkiye (Kayış, 2011). Moreover, with the exception of Botrylloides niger (Temiz et al, 2023), to our knowledge, no other records of botryllid ascidian have been reported from the Mediterranean coast of Türkiye. Although the occurrence of Symplegma brakenhielmi (Michaelsen, 1904) on the Mediterranean coast of Türkiye has been reported (Çınar et al, 2006), no genetic data about its taxonomy has been provided thus far.…”

“…The findings of this study disclose the occurrence and distribution patterns of seven colonial ascidian species along the coastline of the Northeastern Mediterranean region in Türkiye. Furthermore, a separate publication by Temiz et al (2023) presents information on B. niger, an additional species obtained from the sampling sites of this study, which exhibits a wide range of color morphs. It should be noted that due to the focus on shallow waters in the sampling process, not all colonial species may have been included, possibly because of factors like seasonal variations or the existence of deeper ascidian habitats.…”

“…Nowadays, alongside the traditional taxonomy, researchers utilize a diverse range of molecular tools to unravel the intricacies of biodiversity (Brunetti et al, 2017;Viard et al, 2019;Reem et al, 2017aReem et al, , b, 2022 and tackle emerging challenges in species delineation (Nydam et al, 2021;Temiz et al, 2023). One commonly used marker for this purpose is the cytochrome oxidase subunit 1 (COI) gene (Hebert et al, 2003).…”

DNA Barcodes for Evolution and Biodiversity

“…A combination of DNA barcoding, the sequencing of additional genes, and phenotypic data analysis was also used in another paper included in this Special Issue [26] to characterize population of the invasive colonial ascidian Botrylloides niger in the northeastern Mediterranean Sea. Several distinct morphotypes were found, but DNA-based species delimitation methods suggest that these all belong to B. niger.…”