Corresponding author: Kennet Lundin (
Academic editor: Nathalie Yonow
Using the nudibranch genus
Korshunova T, Malmberg K, Prkić J, Petani A, Fletcher K, Lundin K, Martynov A (2020) Fine-scale species delimitation: speciation in process and periodic patterns in nudibranch diversity. ZooKeys 917: 15–50.
Species delimitation, and hence the degree of separation between different groups of biological organisms, is a pivotal concept for modern biology, despite the fact that there is no universal agreement about the species concept itself (
Here we are using a complex case of nudibranch mollusc species of the genus
Material for this study was obtained by scuba diving at widely separate locations in Europe: in the, Croatia, France, Norway, Sweden, Spain, and the United Kingdom. The specimens were deposited in the Gothenburg Natural History Museum (
Specimens of
The molecular phylogenetic and delimitation methods were combined with morphological data (Figs
Phylogenetic analysis was performed using 46 specimens of the genus
Phylogenetic relationships of
Initially, Automatic Barcode Gap Discovery (ABGD) was used for species delimitation. ABGD analysis of the COI dataset run with two different models for species of the genus
Analysis of multi-locus genomic sequence data under the multispecies coalescent model was conducted. The sequences were divided into an eight-species scenario (Suppl. material
The haplotype network based on cytochrome c oxidase subunit I (COI) molecular data showing genetic mutations occurring within species of the genus
The molecularly and morphologically confirmed specimens of all species of the genus
Periodic-like presentation of colour variation patterns among all species of the genus
Ceratal rows not branched. Up to six anterior ceratal rows (commonly no more than four). Cerata without tubercles, usually considerably swollen. Rhinophores smooth. Pharynx and jaws moderately broad. Central teeth with central cusp adpressed by adjacent lateral denticles. Prostate thick, readily distinct from vas deferens, moderate in length to very long. Distal receptaculum seminis oval to elongate on a moderately long stalk. Supplementary gland inserts into penis commonly via a narrowing stalk. Penis conical, always with a relatively short, slightly curved, hollow stylet.
In this study, we confirm that genus
Body up to 20 mm; large dorsal pigment spots, if present, yellow-orange, bright; in specimens with bright yellow-orange spots on dorsal side and cerata, a distinct yellow-orange spot or stripe on the tail is always present; completely pale specimens lacking tail spot or stripe; no light pinkish subapical ring on cerata; absence of punctuated white line on edge of foot; cerata commonly moderate in width without distinctly attenuated apices; digestive gland in cerata relatively broad without distinct short branches; up to four anterior rows of cerata; radular formula 35–38 × 1.1.1, copulative stylet short and slightly bent at the top, receptaculum seminis pear-shaped without short distinct stalk between reservoir and short wide base.
Mediterranean Sea and all European Atlantic coasts to Norway, from very shallow water (0–0.5 m) to ca. 25 m. On the Swedish west coast, it lives below the halocline (15–25 m).
Morphologically
The species
Minimum uncorrected p-distances of the COI marker which separate
Body up to ca. 12 mm; large dorsal pigment spots, if present, yellow-orange, dull; in specimens with yellow-orange spots on body and cerata there is never any yellow-orange pigment spot or stripe on the tail, but there might be a median whitish line or broken line on the tail; completely pale specimens lack tail spot; light pinkish subapical ring on cerata present; absence of white punctuated line on external edge of foot; cerata commonly moderate in width without distinctly attenuated apices; digestive gland in cerata relatively broad without distinct short branches; up to four anterior rows of cerata; radular formula 31–47 × 1.1.1, copulative stylet relatively long and almost straight, at the top, receptaculum seminis pear-shaped with short distinct stalk between reservoir and long base.
Swedish northwest Skagerrak coast, in the south from the town of Lysekil at the Gullmar fjord, onwards to Smögen and the Väderö Island archipelago, to the Ide fjord in the north by the border with Norway. It is always found very shallow and above the halocline (situated at 6–7 m depth within the fjords and 15 m outside the fjords), most often from 0.1 to 6 metres depth, commonly on wharf pontoons in the marina. Inhabits exclusively the brackish water layer, salinity-range: ordinarily ca. 24–25‰ but may vary from 12 to 30‰.
Morphologically the brackish water-living
Minimum uncorrected p-distances of the COI marker which separate
Body up to at least 20 mm; large dorsal pigment spots, if present, bright yellow-orange or reddish orange; in specimens with yellow-orange or reddish spots on dorsal side and cerata, there is never any yellow-orange spot or stripe on the tail, but there could be a whitish median line on the tail; completely pale specimens lack tail stripe or spot; light pinkish subapical ring on cerata absent; absence of a punctuated white line or row of dots on the edge of foot; cerata commonly moderate in width without distinctly attenuated apices; digestive gland in cerata relatively broad without distinct short branches; up to four anterior rows of cerata; radular formula 30–37 × 1.1.1, copulative stylet very short and conical, receptaculum seminis subcircular with long distinct stalk between reservoir and rapidly widening base.
Mediterranean Sea and all European Atlantic coasts to Gulen at the mouth of Hardanger fjord, Norway, also possibly further north to the Trondheim fjord (Klas Malmberg, personal observation). Salinity-range: 33 to 35‰, ordinary oceanic salinity, or close to it. On the Swedish west coast, it lives below the halocline. In areas without a halocline and in more oceanic environments, it can be found closer to the surface or intertidally. In Croatia it is quite common from very shallow water (0–0.5 m) to ca. 20 m.
Morphologically this inhabitant of waters with normal to nearly normal ocean salinity,
Minimum uncorrected p-distances of the COI marker which separate
NE Atlantic, Skagerrak, Sweden, Västra Götalands län, Bohuslän, Väderöarna Islands (
Body up to 30 mm; dorsal spots, if present, reddish orange; in specimens with dorsal and ceratal spots distinct colouration of tail absent; completely pale specimens lack tail stripe or spot; light pinkish subapical ring on cerata absent; presence of distinct line of white pigment, sometimes punctuated, on the edge of the foot; cerata commonly broad with distinctly attenuated apices; digestive gland in cerata relatively thin without distinct short branches; up to six anterior rows of cerata; radular formula 38–61 × 1.1.1, copulative stylet relatively long, slightly bent at the middle, receptaculum seminis elongate oval with moderate distinct stalk between reservoir and rapidly widening base.
Mediterranean Sea and all European Atlantic coasts to Sweden and Southwest Norway. On the Swedish west coast, it lives below the halocline.
Morphologically
Minimum uncorrected p-distances of the COI marker which separate
NE Atlantic, Skagerrak, Sweden, Region Västra Götaland, Bohuslän county, town of Smögen, outermost skerries, Pesaskär (
Body up to 25 mm; dorsal pigment spots (if present), small and often rounded, forming an almost continuous orange-brownish covering; in specimens with dorsal pigment spots there is never any colouration of the tail; completely pale specimens likewise lack a tail spot; absence of light pinkish subapical ring on cerata; absence of punctuated white line on external edge of foot; cerata commonly moderate in width without distinctly attenuated apices; digestive gland in cerata relatively broad without distinct short branches; up to four anterior rows of cerata; radular formula 18–41 × 1.1.1, copulative stylet long and bent at the top, receptaculum seminis oval without stalk and widened base.
Western Mediterranean Sea and all European Atlantic coasts to northern Norway. On the Swedish west coast, it lives below the halocline.
Morphologically
Minimum uncorrected p-distances of the COI marker which separate
Reproductive systems, schemes.
The genus
The present case clearly differs from the situation when a reticulated molecular phylogenetic pattern of two closely related species was used for evidence of their synonymy (
Remarkably, both species,
Taking into consideration the population-to-species continuum (
Periodic-like patterns in application to biology, though discussed for a long time (e.g.,
The appearance of similar colour patterns across different species of the genus
Application of a periodic-like arrangement of vertical rows and horizontal periods helps to highlight subtle differences between apparently highly similar forms. For example, some white forms with distinct yellow-orange spots of
We would like to give special thanks to the team of Gulen Dive Centre (Christian Skauge, Ørjan Sandnes, Monica Bakkeli, and Guido Schmitz) and to Torkild Bakken (NTNU University Museum) for their generous help during fieldwork in Norway. Klas Malmberg and Kennet Lundin warmly thank the staff at the Smögen Dyk och Upplevelse Dive Centre for their enthusiasm and generous help during fieldwork in Sweden. Jakov Prkić and Alen Petani would like to thank Marko Lete and Đani Iglić for their great help during fieldwork in Croatia. David Fenwick (
In memory of Rolf Lundin.
Table S1. List of samples, localities, GenBank accession numbers, and voucher references
species data