Computational protocol: Evolution of the connection patterns of the cephalic lateral line canal system and its use to diagnose opsariichthyin cyprinid fishes (Teleostei, Cyprinidae)

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Protocol publication

[…] The genus level classification of the Opsariichthyini follows , although that classification still needs to be confirmed (, cf. ). The cephalic lateral line canal system was observed in 12 species of opsariichthyins; data on the canal system in the out-group were compiled from previous studies (Tables –).Methods used for observation of the cephalic lateral line canal systems followed those of . The canals were stained using Cyanine suminol 5R. The canal terminology follows that of , with additional reference to that of . These are as follows: infraorbital canal (IO), preoperculomandibular canal (POM), supraorbital canal (SO), supratemporal canal (ST), and temporal canal (TC) (Fig. ).Furthermore, the canaliculi branching from each canal are defined as “bridges,” whereas the junctions connecting canals were termed “joints.” The three bridges and one joint were as follows: “frontal bridge” between SO and IO; “centroparietal bridge” recognizing that ST meets the opposite side ST; “infratemporal bridge” between POM and TC; and “anteropterotic joint” between IO and TC (Fig. ).In some species in cyprinid subfamilies such as Gobioninae and Leuciscinae, development of the cephalic lateral line canal system is generally completed when the fish is approximately 60 mm in total length (= TL) (, , ). Therefore, in the present study, specimens larger than 60 mm in TL were selected for examination. The pores on each canal were counted from end to end. Statistical tests were used to assess differences in the number of pores among the species. Tests for homogeneity of variance were carried out on the number of pores on each canal using Bartlett’s test in R 3.3.1 (). When the variances were homogeneous, the Tukey-Kramer test was used, whereas when variances were heterogeneous, the Steel-Dwass test in R 3. 3.1 was used.To obtain a hypothesis about the branching pattern of the opsariichthyin species, we analyzed mitochondrial cytochrome b (cyt b) gene sequences downloaded from GenBank. This is because molecular data for the cyt b gene sequence of all the species examined in the present study have been accumulated by previous studies (Table ). Cyt b sequence alignment of 1137 bp long sequences was performed using MEGA 7 () and checked manually for accuracy. Maximum likelihood (ML) analysis for phylogenetic reconstruction was applied using PAUP* v. 4.0b10 (). Models of molecular evolution were selected using the program MODELTEST v.3.7 (), with the best fitting model being determined by the Akaike information criteria (AIC) (= GTR+G+I model, in the present analysis). Three species of the subfamily Acheilognathinae, two species of the subfamily Gobioninae, two species of the subfamily Leuciscinae, and 12 species of the Xenocypridinae were chosen as out-groups (Table ). Polarity in the character evolutions of the connecting pattern of the cephalic lateral line canals was determined by character state reconstruction using Mesquite v.2.75 () with maximum parsimony methodology. Maximum parsimony character state reconstruction was performed on the ML tree.Specimens studied are deposited in the following institutions: Chonbuk National University, Jeollabuk-do, Korea (CNUC); Department of Fisheries, Faculty of Agriculture, Kyoto University, Kyoto, Japan (FAKU); Fisheries Research Laboratory, Mie University, Mie, Japan (FRLM); Lake Biwa Museum, Shiga, Japan (LBM); the National Museum of Nature and Science, Tsukuba, Japan (NSMT); Swedish Museum of Natural History, Stockholm, Sweden (NRM); Smithsonian Institution National Museum of Natural History, Washington DC, United States (USNM). The institutional code of the Faculty of Agriculture, Kindai University, was changed from FKUN (Department of Fisheries, Kindai University, Nara) to KUN-P (Kindai University, Nara, Pisces) with faculty reorganization in 2005. […]

Pipeline specifications

Software tools MEGA, ModelTest-NG, Mesquite
Application Phylogenetics
Organisms Nipponocypris sieboldii, Nipponocypris temminckii, Nipponocypris koreanus, Opsariichthys uncirostris, Opsariichthys bidens