Evolving in the darkness: Phylogenomics of Sinocyclocheilus cavefishes highlights recent diversification and cryptic diversity

Abstract

Troglomorphism-any morphological adaptation enabling life to the constant darkness of caves, such as loss of pigment, reduced eyesight or blindness, over-developed tactile and olfactory organs-has long intrigued biologists. However, inferring the proximate and ultimate mechanisms driving the evolution of troglomorphism in freshwater fish requires a sound understanding of the evolutionary relationships between surface and troglomorphic lineages. We use Restriction Site Associated DNA Sequencing (RADseq) to better understand the evolution of the Sinocyclocheilus fishes of China. With a remarkable array of derived troglomorphic traits, they comprise the largest cavefish diversification in the world, emerging as a multi-species model system to study evolutionary novelty. We sequenced a total of 120 individuals throughout the Sinocyclocheilus distribution. The data comprised a total of 646,497Jbp per individual, including 4378 loci and 67,983 SNPs shared across a minimum of 114 individuals at a given locus. Phylogenetic analyses using either the concatenated RAD loci (RAxML) or the SNPs under a coalescent model (SVDquartets, SNAPP) showed a high degree of congruence with similar topologies and high node support (> 95 for most nodes in the phylogeny). The major clades recovered conform to a pattern previously established using Sanger-based mt-DNA sequences, with a few notable exceptions. We now recognize six major clades in this group, elevating the blind cavefish S. tianlinensis and the micro-eyed S. microphthalmus as two new distinct clades due to their deep divergence from other clades. PCA plots of the SNP data also supports the recognition of six major clusters of species congruent with the identified clades based on the spatial arrangement and overlap of the species in the PC space. A Bayes factor delimitation (BFD) analysis showed support for 21 species, recognizing 19 previously described species and two putative new cryptic ones.Two species whose identities were previously disputed, S. furcodorsalis and S. tianeensis, are supported here as distinct species. In addition, our multi-species calibrated tree in SNAPP suggests that the genus Sinocyclocheilus originated around 10.5 Mya, with most speciation events occurring in the last 2 Mya, likely favored by the uplift of the Qinghai-Tibetan Plateau and cave occupation induced by climate-driven aridification during this period. These results provide a firm basis for future comparative studies on the evolution of Sinocyclocheilus and its adaptations to cave life.

Sinocyclocheilus longicornus (Cypriniformes, Cyprinidae), a new species of microphthalmic hypogean fish from Guizhou, Southwest China

“…Initially, Sinocyclocheilus was divided into four species groups, S. jii, S. angularis, S. cyphotergous, and S. tingi, based on mitochondrial Cyt b and morphological differences . Phylogenetic trees reconstructed using mitochondrial ND4 and Cyt b, mitochondrial genome, and restriction site-associated DNA sequencing supported monophyly of the S. jii and S. tingi species groups and rejected monophyly of the S. angularis and S. cyphotergous species groups (Xiang 2014;Liu 2018;Mao et al 2021Mao et al , 2022Wen et al 2022;this study). These studies proposed new classification schemes, such as two new clades (Clades E and F) from Mao et al (2022) and a new species group (S. microphthalmus group) from Wen et al (2022).…”

“…Phylogenetic trees reconstructed using mitochondrial ND4 and Cyt b, mitochondrial genome, and restriction site-associated DNA sequencing supported monophyly of the S. jii and S. tingi species groups and rejected monophyly of the S. angularis and S. cyphotergous species groups (Xiang 2014;Liu 2018;Mao et al 2021Mao et al , 2022Wen et al 2022;this study). These studies proposed new classification schemes, such as two new clades (Clades E and F) from Mao et al (2022) and a new species group (S. microphthalmus group) from Wen et al (2022). Inconsistent topological differences may be related to molecular marker types, number of species and evolutionary models.…”

Sensory evolution in a cavefish radiation: patterns of neuromast distribution and associated behaviour inSinocyclocheilus(Cypriniformes: Cyprinidae)

“…These species are distributed across a vast karstic expanse spanning three provinces in southern China [22]. Prior work shows that one of the major drivers of speciation in this group was isolation over long periods of time, suggesting geographical speciation has dominated this radiation [19,66,67]. These divergences occurred during the geological uplift of the Yunnan–Kweichow Plateau alongside with the aridification of China, which occurred during the Pliocene and Pleistocene eras [66].…”

Expansion and contraction of lake basin shape the genetic structure of Sinocyclocheilus (Osteichthyes: Cypriniformes: Cyprinidae) populations in Central Yunnan, China

“…Based on the phylogenetic tree, in order to avoid having an impact on the multi-species tracing of ancestral patterns, one sample of Sinocyclocheilus grahami and four samples of Sinocyclocheilus tingi were censored here, which was then filtered with 'AC==0 || AC==AN || F_MISSING >0.2' and the distance between the locus and the set SNPs to be no less than 100 bp, and ultimately reassembled them (2,642,407 SNPs for 103 individuals). For time calibration, due to the absence of a specific fossil record provides a calibration for Sinocyclocheilus, we used the time inferred by Chen et al (2009) to the split between S. grahami (F) and S. grahami (M) at approximately 2.37~0.47 Ma (Chen et al, 2009), Mao, Liu, Vasconcellos, et al (2021) to the split between S. oxycephalus and S. qujingensis at approximately 4.0~2.0 Ma (Mao, Liu, Vasconcellos, et al, 2021) and Yang et al (2021) to the split between S. grahami and S. tingi at approximately 0.68 Ma (Yang et al, 2021), and set the differentiation rate of S. grahami at 3.51 × 10 −9 per year per nucleotide (Yang et al, 2016). We ran an independent SNAPP analysis with 20 million MCMC generations, sampling at every 2000 steps.…”