Biogeography 17. 43–52. Sep. 20, 2015 Geographical variations in morphological characters of the fluvial eight-barbel loach, Nagare-hotoke-dojo (Cobitidae: Nemacheilinae) Taiki Ito*, Kazuhiro Tanaka and Kazumi Hosoya Program in Environmental Management, Graduate School of Agriculture, Kinki University, 3327-204 Nakamachi, Nara 631-8505, Japan Abstract. The morphological and color variations of Lefua sp. 1 Nagare-hotoke-dojo individuals from 13 river systems were examined. Analysis of variance revealed highly significant variations in Lefua sp. 1 mor- phology and coloration among the 13 populations examined, across all 19 measurements and counts. The 13 populations of Lefua sp. 1 were classified into two major clusters (I and II) by using UPGMA cluster analy- sis. Cluster I comprised fish from the Maruyama, Yura, Muko, Mihara, Yoshino, Hidaka, Kumano, Yoshii, Chikusa, and Ibo river systems. Cluster II comprised fish from the Yoshida, Saita, and Sumoto river systems. Cluster I was further subdivided into sub-clusters: I-i (the Maruyama, Yura, Muko, Mihara, Yoshino, Hidaka, Kumano, and Yoshii river systems) and I-ii (the Chikusa and Ibo river systems). Principal component analysis revealed that populations within cluster II clearly possessed longer caudal peduncles, while populations within cluster I possessed a longer anterior body on average and a deeper body. Populations within sub-cluster I-ii possessed a higher average dorsal fin and a longer average dorsal fin base than those of populations within sub-cluster I-i. A strong correlation was noted between the PC3 score and population latitude (r = 0.621). Observations of body color patterns revealed that individuals from the Yoshino, Mihara, Sumoto, and Hidaka river systems had dark brown mottling on both sides and the dorsal regions of their bodies and many small dark brown spots on the dorsal and caudal fins, while those from the Yura, Muko, and Kumano river systems possessed neither. Individuals from the Yoshii, Chikusa, Ibo, Maruyama, Yoshida, and Saita river systems, no dark brown mottling was present on both sides and the dorsal regions of their body and many small dark brown spots present on the dorsal and caudal fins. Overall, it was determined that Lefua sp. 1 exhibits a high degree of variability in its morphological characters among populations, which may be because of population isolation due to the specify preference for small freshwater bodies, such as streams. Key words: body color, endangered species, geographical cline, river capture, undescribed species Introduction were taxonomically separated from L. echigonia as an undescribed species sensu stricto, based on their The fluvial eight-barbel loach, known in Japanese morphological, ecological, and genetic features (Ho- common name as “Nagare-hotoke-dojo” (=Lefua soya, 1993; Hosoya, 2000; Hosoya, 2003; Sakai et sp. 1) is classified as a cobitid in the subfamily al., 2003). Lefua sp. 1 is distributed in freshwaters Nemacheilinae (Hosoya, 2013). Previously, this spe- along the coasts of the Seto Inland Sea, the Ise Bay cies was considered to represent several local popu- and the Sea of Japan that involves Kyoto, Fukui and lations of Lefua echigonia. However, the populations Tottori prefectures, Japan (Hosoya, 2013). ——————————————————————— Some studies have suggested that Lefua sp. 1 *Corresponding author: [email protected] exhibits several morphological and body color − 43 − Geographical variations of Lefua sp. 1 variations among populations. Takahashi (1999) imens, 38.10–44.91 mm SL, Tatsuno City, Hyogo showed variations in the number of vertebrae among Prefecture. the Lefua sp. 1 populations of the Kinsei, Chigiri, Maruyama River system: KUN-P 44173-44177, 5 and Yoshino river systems in the eastern part of the specimens, 32.25–55.88 mm SL, Toyooka City, Hy- Ehime Prefecture. In addition, Takahashi (1999) ogo Prefecture. showed that the position of the ventral-dorsal fin and Yura River system: KUN-P 44148-44151, 4 spec- the coloration of the dorsal and caudal fins differed imens, 36.53–52.2 mm SL, Tanba City, Hyogo Pre- among river systems. Furthermore, Hosoya (1994) fecture. and Nakatani & Yoshida (1996) reported the obser- Muko River system: KUN-P 44178-44181, 44246- vation of two body coloration morphs in Lefua sp. 1: 44247, 6 specimens, 35.21–47.29 mm SL, Takarazu- one with dark brown mottling on both sides and on ka City, Hyogo Prefecture. the dorsal regions of the body and the other without Kumano River system: KUN-P 44155, 44157- the mottle body coloration. 44160, 5 specimens, 31.33–43.24 mm SL, Yoshino As the studies described above only examined in- District, Nara Prefecture. dividuals from a few river systems, the entire range Saita River system: KUN-P 44596-44616, 21 of intraspecific variation within Lefua sp. 1 is still specimens, 33.71–58.98 mm SL, Mitoyo City, Kaga- unclear. Clarifying the level of variation within Lefua wa Prefecture. sp. 1 and the geographical patterns of this variation Yoshino River system: KUN-P 44562-44573, 12 will contribute not only to the taxonomic description specimens, 35.17–47.69 mm SL, Tokushima City, of Lefua sp. 1 but also to the species’ recognition Tokushima Prefecture. as a conservation unit. In this study, we collected Yoshida River system: KUN-P 42268-42272, specimens of Lefua sp. 1 from a wide distribution of 44165, 6 specimens, 48.2–62.3 mm SL, Shozu Dis- populations and described the geographical variation trict, Kagawa Prefecture. of its morphological characters. Mihara River system: KUN-P 44133-44140, 8 specimens, 37.46–45.26 mm SL, Minamiawaji City, Materials and methods Hyogo Prefecture. Materials examined We examined a total of 110 Lefua sp. 1 individu- als from 13 sites, each in one of 13 river systems (Fig. 1), during 2010–2014. The study materials were deposited at Kinki University, Nara, Japan (KUN-P). List of materials examined Yoshii River system: KUN-P 44152-4453, 44584- 44590, 44592-44593, 11 specimens, 36.57–55.72 mm SL, Mimasaka City, Okayama Prefecture. Chikusa River system: KUN-P 44161-44164, 4 Fig. 1. The locations of river systems in this study. C: Chikusa specimens, 43.79–49.31 mm SL, Tatsuno City, Hyo- River, H: Hidaka River, I: Ibo River, K: Kumano River, go Prefecture. Ma: Maruyama River, Mi: Mihara River, Mu: Muko River, Sa: Saita River, Su: Sumoto River, Yd: Yoshida Ibo River system: KUN-P 44129-44132, 4 spec- River, Yi: Yoshii River, Yn: Yoshino River, Yu: Yura River. − 44 − Taiki Ito, Kazuhiro Tanaka and Kazumi Hosoya Sumoto River system: KUN-P 44141-44147, Morphological examination 44166-44167, 44574-44583, 19 specimens, 32.27– Measurements and counts were carried out ac- 60.69 mm SL, Minamiawaji City, Hyogo Prefecture. cording to the methods described by Hubbs & Lagler Hidaka River system: KUN-P 44168-44172, 5 (2004), with the exception of the vertebrae counts, specimens, 28.16–47.90 mm SL, Hidaka District, which were conducted according to the methods Wakayama Prefecture. described by Hosoya (1983). The vertebrae were counted on radiographs: we also included the four vertebrae of the Weberian apparatus, and the hypural Table 1. Measurements and counts of Lefua sp. 1 from each river system River system Yoshii Chikusa Ibo Maruyama Yura Muko Counts and measurements range mean±SD range mean±SD range mean±SD range mean±SD range mean±SD range mean±SD Measurements Percentage of standard length Head length 20.0-22.3 21.3±0.700 20.1-21.2 20.7±0.415 20.2-21.6 20.8±0.494 19.7-22.3 21.0±1.084 20.0-21.1 20.4±0.426 19.9-21.4 20.6±0.482 Body width 10.9-12.5 11.6±0.561 12.7-13.6 13.2±0.334 11.5-13.7 12.4±0.894 11.6-14.1 12.6±0.884 11.6-13.6 12.8±0.861 11.4-14.2 12.6±0.975 Body depth 11.7-13.9 12.8±0.692 13.8-16.4 15.4±0.981 13.8-14.8 14.3±0.351 13.2-16.2 14.2±1.025 12.7-15.0 13.7±0.934 11.9-14.9 13.5±1.138 Length of caudal peduncle 11.8-16.5 14.3±1.190 11.5-13.6 12.3±0.784 10.8-13.0 11.7±0.787 12.1-15.0 12.8±1.094 11.3-13.3 12.7±0.815 11.6-15.1 12.9±1.248 Depth of caudal peduncle 9.8-11.7 10.8±0.557 11.6-13.1 12.3±0.561 12.0-13.8 12.9±0.769 10.6-12.1 11.3±0.590 10.7-13.0 11.7±0.844 12.0-13.9 12.5±0.650 Predorsal length 63.6-68.4 66.0±1.280 68.6-71.7 70.6±1.167 67.3-70.9 69.2±1.557 65.5-69.0 67.0±1.329 65.7-68.8 67.0±1.151 67.4-68.9 68.1±0.544 Preanal length 77.4-86.4 80.3±2.323 80.4-81.9 81.2±0.549 79.6-84.7 81.9±1.798 80.0-82.2 81.1±0.730 79.7-81.7 80.5±0.815 79.3-83.5 81.8±1.426 Prepelvic length 56.1-59.0 57.6±0.912 59.3-62.1 60.1±1.171 56.9-60.2 58.5±1.170 56.2-61.0 58.5±1.696 57.4-58.5 57.9±0.482 57.0-60.7 58.8±1.147 Pectoral fin to pelvic fin base 35.3-39.2 37.8±1.178 39.4-40.4 40.0±0.377 38.9-39.6 39.2±0.312 36.2-42.2 38.8±2.119 37.7-39.1 38.4±0.497 35.7-41.4 39.2±1.827 Height of dorsal fin 9.2-12.9 11.0±1.005 11.2-15.0 12.9±1.519 12.8-14.7 13.6±0.793 10.5-13.0 12.0±0.912 11.7-13.7 12.8±0.733 9.9-13.4 11.2±1.169 Length of dorsal fin base 6.5-8.8 7.7±0.611 7.9-9.4 8.8±0.561 8.4-9.7 9.1±0.496 7.3-9.1 8.1±0.709 8.0-9.3 8.7±0.461 7.1-9.1 8.1±0.660 Height of anal fin 9.6-12.2 11.2±0.837 11.2-14.1 12.0±1.219 12.1-13.3 12.7±0.570 9.8-13.9 11.8±1.482 11.4-12.8 11.9±0.552 9.7-12.6 11.0±0.898 Pectoral fin length 12.5-15.7 14.1±0.928 12.2-13.2 12.8±0.456 13.8-15.7 14.9±0.686 13.6-15.6 14.6±0.849 13.4-16.1 14.6±1.089 13.4-16.0 14.6±0.936 Percentage of head length Snout length 33.6-44.4 39.9±2.668 38.8-44.7 41.0±2.243 36.6-39.2 38.3±0.970 36.0-47.3 42.9±3.923 41.4-45.9 43.4±1.614 37.9-44.5 41.5±2.393 Orbit diameter 11.1-14.7 12.8±1.196 10.5-13.8 12.3±1.350 8.8-13.9 12.0±1.932 8.8-14.4 12.4±1.915 11.0-13.7 12.1±0.983 11.4-13.0 12.1±0.506 Interorbital width 27.8-38.4 34.3±2.550 33.7-40.1 37.6±2.643 35.2-38.9 37.1±1.400 32.9-41.0 35.3±2.958 33.9-38.2 35.2±1.751 34.2-39.2 36.5±1.472 Counts Abdominal vertebral number 20-22 21.0±0.603 21-22 21.5±0.500 22 22.0±0 21-22 21.4±0.490 21 21.0±0 20-23 21.7±1.374 Caudal vertebral number 16-20 18.4±1.226 18-19 18.3±0.433 16-18 17.0±0.707 17-21 19.0±1.265 18-19 18.3±0.433 15-19 17.2±1.462 Total vertebral number 38-41 39.4±1.150 39-41 39.8±0.829 38-40 39.0±0.707 39-42 40.4±1.020 39-40 39.3±0.433 38-40 38.8±0.687 Table 1.