Five Complete Mitochondrial Genomes Of Crabs From Podotremata And The Phylogenetic Implications Of Brachyuran Crabs

Brachyura is one of the most species-rich taxa within the Decapoda,occurring in marine, freshwater, and terrestrial habitats. The phylogenetic relationships among Brachyura have been contentious for more than a century. Though Brachyura itself is widely regarded as a monophyletic grouping, the relationships among the various groups of the Brachyura are still controversial, in particular, the status of Podotremata. The phylogenetic position of Podotremata is important not only for uncovering the pattern of early brachyuran evolution, but also as a prerequisite to understanding eubrachyuran diversification.Due to maternal inheritance, lack of the genetic recombination, and relatively rapid evolutionary rate, the mitochondrial DNA sequence has been widely used for phylogenetic and population genetic studies of animals. Compared with a signle or a few genes, the complete mitogenome sequence can provide more genetic information and thus has gained increasing popularity in reconstruction of deeper phylogenetic relationships within metazoans.The complete mitochondrial genomes of Dynomene pilumnoides, Homola orientalis, Moloha majora, Umalia orientalis and Lyreidus brevifrons were determined by Long- PCR and primer walking sequencing. The date was combined with the other 28 species of brachyuran and six species in Anomura. Seven Thalassinidea species as outgroups were used to reconstruct the phylogenetic relationship of Meiura. Main results of the study were as follows:(1) The mitogenome sizes are 16,475 bp in D. pilumnoides, 15,903 bp in M. majora, 16,082 bp in H. orientalis, 15,466 bp in U. orientalis and 16,112 bp L. brevifrons, respectively. Each of mitogenomes contains 37 genes, including 13 PCGs, 2 rRNA genes and 22 tRNA genes.(2) The mitogenomes of H. orientalis, M. majora and U. orientalis have only one large non-coding region(NCR), which loact between rrnS and trnI. In L. brevifrons, there are two NCRs; one is locted between rrnS and trnI, and the other between trnA and nad3. Comparing to the Pancrustacean ground pattern, the NCR locating between rrnS and trnI is supposed to be putative control region. However, several NCRs are existence in D. pilumnoides mitogenome, and the largest NCR locating between trnL1 and trnQ is the putative control region.(3) The control regions of these three Homoloidea crabs are characterized by the large repeat region with different repeat length and copy number, accounting for the size variance. The highly levels of sequence identity among repeat units appear in two kinds of Homolidaes, H. orientalis and H. malayens.(4) Except for D. pilumnoides, the primitive crab gene order is identical to the published marine brachyurans and differ fom the pancrustacean ground pattern by only the position of trnH. D. pilumnoides mitogenome exhibits rearrangements and a novel pattern with positional translocation of two genes/regions(trnQ and CR). The novel gene order could have resulted from one-step occurrence of this process: tandem duplication occurring in the region between trnI and trnM, followed by deletions of redundant genes. Multiple deletions of redundant genes seem to be incomplete, because the short non-coding sequences remain at the deleted region.(5) In five primitive crabs, except for cox1, the other protein genes are all used the typical start codons ATN, the ATP8 and nad1 in U. orientalis and L. brevifrons with GTG. The start codon for cox1 is ATG in all reported Eubrachyura mitogenomes, while a common start codon ACG is found in the Podotremata. The Leu, Phe, Met and Ile are the most used amino acids, and the UUA, GUA, UCU, CCU and GCU were the most used codons.(6) The values of uncorrected nucleotide diversity(Pi), Ka/Ks and the similarities between the same gene indicated that nad5, nad4 and nad1 evolved faster while cox1, cox2, cytb, cox3 evolved much slowly. nad5, nad4 and nad1 are considered to be more reliable molecular marker for the further population studies among Podotremata.(7) Most(20 of 22) tRNAs are folded into a normal clover-leaf secondary structure, except for trnS-AGN and trnI. As in other brachyuran crabs, trnS-AGN lacks DHU stem arm in D. pilumnoides U. orientalis and M, majora mitogenome, while it is folded into a clover-leaf structure with short DHU arm in H. orientalis and L. brevifrons. Moreover, it is also worth noted in L. brevifrons that trnI of the mitogenome lacks a DHU stem arm while the other reported brachyuran species all have a standard cloverleaf structure for trnI.(8) In the primitive crab mitogenomes, lrRNA and srRNA are separated by the trnV and are both coded on the L-strand.(9) The phylogenetic relationships indicate that Homoloidea is a sister group to Dynomenidae locating the basal position of phylogenetic tree, and Raninidae is a sister group to Eubrachyura, indicating that Podotremata is paraphyletic and Dromiacea separates from a primitive brachyuran line at earlier stage than Raninoidea.