the evidence here. Molecular baraminological analysis of snakes and lizards showed that they are separate apobaramins, and that there are likely several snake and lizard baramins (Cserhati 2020). As such, putative elasmobranch holobaramins would be classified as type 3b Carter baramins. The first three groups are rays, whereas the fourth and fifth are sharks. Two of the three ray baramins have a normalized entropy value of less than 0.25, indicating that the species in these baramins are adapted to either brackish or saltwater conditions. This is no surprise since sharks and rays are mainly marine animals. These animals are less adapted to freshwater, as they must constantly swim to keep afloat; the less dense freshwater makes this more difficult for them. In contrast with the several putative baramins found within the Cyprinodontiformes apobaramin, it seems that the several Elasmobranchii groups have adapted to saltwater (see Figure 10C) after the initial euryhaline stage, as 74% of the species with FishBase annotation were oceanodromous. Nevertheless, five shark species from group #4 and three ray species from group #3 can also inhabit freshwater. These species are Carcharhinus leucas (bull shark), Glyphis fowlerae (Borneo river shark), Glyphis gangeticus (Ganges shark), Glyphis glyphis (Bizant river shark), Rhizoprionodon acutus (milk shark), Potamotrygon magdalenae (Magdalena River stingray), Potamotrygon motoro (ocellate river stingray), and Potamotrygon orbignyi (smooth back river stingray). Of these, C. leucas is known to inhabit freshwater lakes in Mozambique, KwaZulu-Natal, Nicaragua, and Sydney Harbour, Australia, a large temperate estuary (Smoothey et al. 2019). This species leads a catadromous lifestyle, using natural rivers and estuaries as nursery grounds before migrating out to the sea (Werry et al. 2012). It is possible that as the Flood waters receded, separate groups of bull sharks could have been entrapped in inland lakes in these three locations. The capability of male great white sharks to undertake transoceanic migrations observed by Pardini et al. (2001). These animals could have been entrapped in these freshwater lakes recently since they have not had much time to diverge morphologically. 8. Pleuronectiformes The order Pleuronectiformes consists of around 700 species such as flounders, turbots, and soles. These fish are characterized by the following synapomorphies: a flat body, with both eyes on one side of their head, with one of the eyes migrating to the other side of the head during development. Their dorsal fin is also positioned dorsal to their skull (Campbell et al. 2013). They also have a muscular sac in the eye called a recessus orbitalis, which can fill with fluid, thereby protruding the eyes above the plane of the fish’s body (Chapleau 1993). No extant flatfish have been discovered that have intermediate skull morphology. These fish apparently form an apobaramin, as they are unrelated to all other fish. Evolutionists such as Lamarck (1809) hypothesized that the ancestors of flatfish lay flatly on the seabed in extremely shallow water. This is exceedingly hard to imagine, because, as such, flatfish ancestors would become prey animals that would be very easy to capture. The mtDNA of 72 species of Pleuronectiformes was analyzed. The results can be seen in Figure 11 and are also summarized in Supplementary File 8. The heatmap in Figure 11A shows six clusters. The Hopkins statistic is 0.809. The Silhouette plot (Figure 11B) shows a maximum value at 8 groups, although there may be distortion in the data. Just as with sharks and rays, flatfish also may form multiple baramins, despite their general morphological similarity with one another. Besides the outlier group, there were four putative holobaramins with at least three species, each one of them with a statistically significant p-value. Groups #2 and #3, of 27 and 16 species respectively, both have a normalized water-type entropy value of just over 0.5. These fish live predominantly in saltwater. Group #4 with 12 species is comprised of species that exclusively inhabit saltwater environments (see Figure 11C); thus, their normalized entropy value is 0. The last group, with 15 species has a normalized water type entropy value of 0.896, with four, eight, and 14 species from the genera Cynoglossus and Paraplagusia, living in freshwater, brackish water, and saltwater respectively. This suggests that this putative baramin is still in the euryhaline stage. It is noteworthy that here also the gene order of the mtDNA differs between groups #4 (Arnoglossus, Asterorhombus, Bothus, Chascanopsetta, Crossorhombus, Grammatobothus, Laeops, Lophonectes, and Psettina) compared to groups #2, #3, and #5 (see Figure 12). The individual mitogenomes of all Pleuronectiformes species analyzed in this study can be seen in Figure 12. The 3’ end of the mtDNA in group #4 contains nine gene rearrangements compared to the mitogenomes of groups 2, 3, and 5. These genes are tRNA-Gln, tRNA-Ala, tRNA-Cys, tRNA-Tyr, tRNA-Ser, tRNA-Asp, NADH6, tRNA-Glu, and tRNA-Pro. Asterorhombus intermedius differs from the regular mtDNA gene order in that tRNA-Val has been inserted between the 16S rRNA and tRNA-Leu (Luo et al. 2019). Furthermore, it appears that the mtDNA of this archaebaramin had two control regions, CR1 and CR2, one of which was differentially lost in some species and the other in other species (Li et al. 2015). Group #6, made up of only two species (Samaris cristatus and Samariscus latus) also has a gene order configuration that is different from all the other groups. These fish live in deep-water benthic zones and inhabit only saltwater environments. Their mtDNA gene order signals discontinuity not only from other flatfishes but also from all other vertebrates, making them a truly unique group. For example, the mitogenome of S. latus has 39 genes (two rRNA genes, 24 tRNAs, 13 protein-coding genes), as well as a duplicated control region and a 376 bp non-coding region, inserted between tRNA-Phe and tRNA-Pro (Shi et al. 2014). 9. Salmoniformes These fish include species such as salmon, trout, chars, whitefishes, graylings, taimens, and lenoks. The mtDNA of 75 species was analyzed in this study. The results can be seen in Figure 13 and are also available in Supplementary File 9. The Hopkins clustering statistic is 0.885, which corresponds to very good clustering. The Silhouette plot in Figure 13B shows a maximum value at four clusters. There is a difference between the number of optimum clusters as shown in the elbow plot and the number of groups that seem to be present in the heatmap. The difference could be due to distortion in the data. However, besides the outlier group, six statistically significant putative CSERHATI Molecular baraminology of fish 2023 ICC 197
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