Colour pattern measurements successfully differentiate two cryptic Onchidiidae Rafinesque, 1815 species
Cryptic species, by definition, appear very similar to each other. In the absence of obvious external morphological differences, quantitative measurements of fine-scale colour pattern differences may be used to distinguish between cryptic species. To demonstrate how this is accomplished, 30 specimens each of two cryptic onchidiid sea slug species in Singapore were collected and identified by sequencing a fragment of the mitochondrial cytochrome c oxidase subunit I gene. Sequences displayed a clear barcode gap: intraspecific distances (0–0.4%) and interspecific distances (4.8–5.5%) were distinct. To quantify colour patterns, eight pattern properties on the animals’ dorsal surface were measured using the PAT-GEOM software. Linear discriminant analysis and classification tree analysis were able to classify specimens with 80% and 81.7% accuracy respectively, and both identified proportion cover and randomness as the most important properties for differentiating the two species. Agreement between the genetic and pattern data is demonstrated by a significant correlation between the pairwise genetic and pattern distance matrices, as well as the significantly greater interspecific than intraspecific distances in both datasets. These results demonstrate that fine-scale pattern differences can be used to differentiate Peronia cryptic species. This approach has potential applications for a range of disciplines, including behaviour and ecology, and as an additional line of evidence for integrative taxonomy.
KeywordsPeronia Sensory ecology PAT-GEOM Integrative taxonomy Pattern quantification
The authors thank L. Roman Carrasco for his advice on statistics and the National Parks Board, Singapore for the permit (number NP/RP 15-088) under which the specimens were collected.
I.Z.W.C. conceived and designed the experiment, performed image and data analysis and wrote the manuscript, including the preparation of the figures and tables. J.J.M.C. and D.H. conceived and designed the experiment, collected the specimens and performed genetic and morphological analyses and contributed to the final draft of the manuscript. P.A.T. conceived and designed the experiment and contributed to the final draft of the manuscript.
This study was funded by a Singapore Ministry of Education Academic Research Fund (MOE AcRF) Tier 1 Grant (R154-000-660-112) and by the National Research Foundation, Prime Minister’s Office, Singapore under its Marine Science R&D Programme (MSRDP-P03).
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
All applicable international, national and/or institutional guidelines for the care and use of animals were followed by the authors.
Sampling and field studies
The specimens were collected under a permit issued by the National Parks Board, Singapore (number NP/RP 15-088).
The datasets and scripts used for all analyses are included in the “Electronic supplementary material” of the article.
- Awati PR, Karandikar KR (1948) Onchidium verraculatum, Cuv. (anatomy, embryology and bionomics). Zool Mem Univ Bombay 1:1–52Google Scholar
- Bergh R (1891) Die cryptobranchiaten Dorididen. Zool Jahrb Abt Anat Ontog Tiere 6:103–144Google Scholar
- Breiman L, Friedman JH, Stone CJ, Olshen RA (1984) Classification and regression trees. Taylor & Francis, Abingdon-on-ThamesGoogle Scholar
- Britton KM (1984) The Onchidiacea (Gastropoda: Pulmonata) of Hong Kong with a worldwide review of the genera. J Molluscan Stud 50:179–191Google Scholar
- Chan IZW, Stevens M, Todd PA (2018) PAT-GEOM: a software package for the analysis of animal patterns. Methods Ecol Evol, 0:1–10. https://doi.org/10.1111/2041-210X.13131
- Chang JJM, Tay YC, Ang HP, Tun KPP, Chou LM, Meier R, Huang D (2018) Molecular and anatomical analyses reveal that Peronia verruculata (Gastropoda: Onchidiidae) is a cryptic species complex. Contrib Zool 87(3):149–165Google Scholar
- Cuthill IC, Partridge JC, Bennett AT, Church SC, Hart NS, Hunt S (2000) Ultraviolet vision in birds. In: Slater P, Rosenblatt J, Snowdon C, Roper T (eds) Advances in the study of behavior, vol 29. Academic Press, Cambridge, pp 159–214Google Scholar
- Cuvier G, Latreille PA (1830) Le règne animal distribué d’après son organisation, pour servir de base à l’histoire naturelle des animaux et d’introduction à l’anatomie comparée, 2nd edn. Chez Déterville, ParisGoogle Scholar
- Golikov AN, Starobogatov YI (1972) Class Gastropoda. In: Mordukhai-Boltovskoi FD (ed) Guide to the fauna of the Black and Azov seas, vol 3. Naukova Dumka, Kiev, pp 65–166Google Scholar
- Joseph S, Poriya P, Vakani B, Singh SP, Kundu R (2014) Identification of a group of cryptic marine limpet species, Cellana karachiensis (Mollusca: Patellogastropoda) off Veraval Coast, India, using mtDNA COI sequencing. Mitochondrial DNA A DNA Mapp Seq Anal 27(2):1328–1331. https://doi.org/10.3109/19401736.2014.945577
- Linnaeus C (1758) Systema naturæ per regna tria naturæ, secundum classes, ordines, genera, species, cum characteribus, differentiis, synonymis, locis, 10th edn. Laurentius Salvius, StockholmGoogle Scholar
- Maechler M, Rousseeuw P, Struyf A, Hubert M, Hornik K (2018) cluster: cluster analysis basics and extensions. R package version 2.0.7–1. https://cran.r-project.org/web/packages/cluster/. Accessed 5 Jun 2018
- Mantel N (1967) The detection of disease clustering and a generalized regression approach. Cancer Res 27(2):209–220Google Scholar
- Oksanen J, Blanchet FG, Friendly M, Kindt R, Legendre P, McGlinn D, … Wagner H (2018) vegan: Community Ecology Package. R package version 2.5–1. https://CRAN.R-project.org/package=vegant. Accessed 5 Jun 2018
- R Development Core Team (2008) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. http://www.R-project.org/. Accessed 5 Jun 2018
- Rafinesque CS (1815) Analyse de la nature ou tableau de l’univers et des corps organisés. PalermeGoogle Scholar
- Ripley B (2018) tree: classification and regression trees. R package version 1.0–39. https://cran.r-project.org/web/packages/tree/index.html. Accessed 5 Jun 2018
- Smith MA, Rodriguez JJ, Whitfield JB, Deans AR, Janzen DH, Hallwachs W, Hebert PD (2008) Extreme diversity of tropical parasitoid wasps exposed by iterative integration of natural history, DNA barcoding, morphology, and collections. Proc Natl Acad Sci 105(34):12359–12364. https://doi.org/10.1073/pnas.0805319105 CrossRefGoogle Scholar