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For mtDNA HVS1 sequences, genetic distances were estimated using the Kimura 2P model for molecular distance and a gamma of 0.25 (Kimura, 1980).
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The SVM parameters were set to normalize data and use a gamma of 0.1 and C of 1.0, refined through experimentation.
Images were corrected with a gamma of 0.45 and spines were counted manually by scrolling through the z-stacks.
Similarly for the K2P model, a transition/transversion ratio of 9.17, a proportion of invariable sites of 29% and a gamma shape of 0.44 were used.
Phylogenetic trees were constructed for each dataset using PHYML, using the JTT model of substitution, and a gamma distribution of rates with four categories.
The identified program settings for all partitions, under the Akaike Information Criteria, included six character states (General Time Reversible model), a proportion of invariable sites, and a gamma distribution of rate variation across sites (GTR+I+G).
We used the general time reversible model allowing for a proportion of invariant sites and a gamma distribution of among-site variation with five categories.
A general time-reversible model of nucleotide substitution with estimated base frequencies, a proportion of invariant sites, and a gamma distribution of rates across sites was applied to conduct maximum likelihood analysis.
Phylogenetic reconstruction was performed by maximum-likelihood inference using RAxML v7.4.2 (Stamatakis 2006) with a general time reversible model of nucleotide substitution and a GAMMA model of rate heterogeneity, branch support values were determined using 1,000 bootstrap replicates.
The NJ tree was generated using the Tamura-Nei model of evolution [ 54] and a gamma parameter of 0.065.
The empirical nucleotide sequences and a gamma distribution of site heterogeneity with 5 categories of substitution rates were set as priors.
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