The high level of phenotypic diversity in southern African tent tortoises (Psammobates tentorius complex) has for decades prevented systematists from developing a stable taxonomy for the group. Here, we used a comprehensive DNA sequence dataset (mtDNA: Cytb, ND4, ND4 adjacent tRNA-His, and tRNA-Ser, 12S, 16S; and nDNA:PRLR gene) of 455 specimens, and the latest phylogenetic and species delimitation analytical procedures, to unravel the long-standing P. tentorius complex systematic puzzle. Our results for mtDNA and nDNA were incongruent, with the poorly supported nDNA phylogeny differentiating the three recognized subspecies, and showing potential hybridization in some regions. In contrast, the concatenated mtDNA phylogeny identified seven operational taxonomic units, with strong support. Clades 1, 4, 5, and 7 corresponded to tortoises identified as P. t. tentorius, clade 3 to P. t. trimeni, and clades 2 and 6 to P. t. verroxii. Our analyses showed conflicting topologies for the placement of C6 (P. t. verroxii north of the Orange River), with stronger support for it being sister to C2 + C3 than to the other clades. Clades 1, 2, and 6 had significantly higher genetic diversity than clades 3, 4, 5, and 7, perhaps because these clades inhabit substantially larger areas. The potential for future cladogenic radiations seems high in C1 and C6, particularly in C6 for which the within-clade diversification level was highest. Further research involving microsatellite DNA, phylogeographic evaluations, and morphological variation among clades is crucial for understanding the adaptive radiation of the P. tentorius complex and for modifying their taxonomy. K E Y W O R D SmtDNA, phylogeny, reptile, southern Africa, tent tortoise | 309 ZHAO et Al.

Background Climatic and topographic changes function as key drivers in shaping genetic structure and cladogenic radiation in many organisms. Southern Africa has an exceptionally diverse tortoise fauna, harbouring one-third of the world’s tortoise genera. The distribution of Psammobates tentorius (Kuhl, 1820) covers two of the 25 biodiversity hotspots in the world, the Succulent Karoo and Cape Floristic Region. The highly diverged P. tentorius represents an excellent model species for exploring biogeographic and radiation patterns of reptiles in Southern Africa. Results We investigated genetic structure and radiation patterns against temporal and spatial dimensions since the Miocene in the Psammobates tentorius species complex, using multiple types of DNA markers and niche modelling analyses. Cladogenesis in P. tentorius started in the late Miocene (11.63–5.33 Ma) when populations dispersed from north to south to form two geographically isolated groups. The northern group diverged into a clade north of the Orange River (OR), followed by the splitting of the group south of the OR into a western and an interior clade. The latter divergence corresponded to the intensification of the cold Benguela current, which caused western aridification and rainfall seasonality. In the south, tectonic uplift and subsequent exhumation, together with climatic fluctuations seemed responsible for radiations among the four southern clades since the late Miocene. We found that each clade occurred in a habitat shaped by different climatic parameters, and that the niches differed substantially among the clades of the northern group but were similar among clades of the southern group. Conclusion Climatic shifts, and biome and geographic changes were possibly the three major driving forces shaping cladogenesis and genetic structure in Southern African tortoise species. Our results revealed that the cladogenesis of the P. tentorius species complex was probably shaped by environmental cooling, biome shifts and topographic uplift in Southern Africa since the late Miocene. The Last Glacial Maximum (LGM) may have impacted the distribution of P. tentorius substantially. We found the taxonomic diversify of the P. tentorius species complex to be highest in the Greater Cape Floristic Region. All seven clades discovered warrant conservation attention, particularly Ptt-B–Ptr, Ptt-A and Pv-A.

We compared the eye anatomy of the scotopic fossorial Acontias orientalis, Acontias rieppeli and Typhlosaurus vermis with that of the photopic surface‐living Trachylepis punctatissima, with particular reference to the retina. The findings were compared with published data on gecko species (Röll, 2001), to determine whether similar trends existed. The vestigial eye of T. vermis was not comparable with that of the other three skink species. The findings in A. orientalis, A. rieppeli and T. punctatissima were as follows: (a) A. rieppeli lacked a conus papillaris, (b) A. orientalis, A. rieppeli and T. punctatissima were pure‐cone species but lacked a fovea, (c) estimated cone density in A. orientalis and A. rieppeli was lower than that in T. punctatissima, (d) the ellipsoid cone segment was smaller and the paraboloid segment larger in A. orientalis and A. rieppeli with the reverse in T. punctatissima, (e) VCL%, ONL%, OPL% and GCL% in A. orientalis and A. rieppeli were significantly greater than that of T. punctatissima, (f) INL% and IPL% in T. punctatissima was significantly greater, and (g) T. punctatissima had abundant Müller cells and fibres. Findings in the gecko species were congruent with those of the three skink species of the present study.

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Aim: Comparative phylogeographic studies provide important insights into the biogeographic processes shaping regional patterns of diversity. Yet, comparative studies are lacking for southern African herpetofauna, despite their high diversity. We statistically compare phylogeographic structure and divergence-time estimates among five co-distributed forest-living herpetofaunal taxa to assess rivers, climatic refugia and climatic gradients as congruent drivers of phylogeographic diversity.Location: Maputoland-Pondoland-Albany biodiversity hotspot, Southern Africa. Taxon: herpetofauna (reptiles and amphibians).Methods: Phylogeographic structure and divergence-times within species were estimated from mitochondrial and nuclear DNA sequence data. Phylogeographic concordance factors were used to estimate the degree of phylogeographic congruence among sympatric localities. Full-likelihood Bayesian comparisons were used to estimate synchronous divergence between phylogeographic regions and across a putative river barrier. Palaeoclimatic niche models were compared among taxa to identify congruent climatic refugia. Nonparametric statistics were used to identify climatic differences between regions and among populations within each species. Finally, redundancy analyses were used to assess geographic distance, climate and the putative river barrier as explanatory variables to genetic diversity. Results: There is comprehensive phylogeographic structuring within each species, comprising distinct northerly and southerly clades. Phylogeographic concordance factors generally support co-diversification in a north/south axis. Yet, analyses of the divergence-time estimates through the Mio/Plio/Pleistocene indicate asynchronous phylogeographic histories. Climatic niche models identified idiosyncratic responses to palaeoclimatic change. Climatic variables are significantly different among populations in all species and correlated with latitude. A combined model of distance, climate and rivers explained the greatest proportion of genetical diversity in most taxa, of which climate explained the highest variance. Main Conclusions: Ancient and recent species-specific responses to climatic and geological processes resulted in pseudo congruent phylogeographic histories among the five co-distributed species. The presence of a congruent north/south pattern in multiple taxonomic groups occupying different forested microhabitats, from fossorial to arboreal, supports latitudinal gradients as global drivers of phylogeographic diversity along the east coast of South Africa.