Neozoarces spp.

Neozoarces spp. began spawning in the Peter the Great Bay usually in the second part of September under the water temperature below 18°C. Spawning period proceed to the middle of December (temperature 1– 5°C). Pairs of spawning fish find and occupy shelters on rocky bottom in the empty cavities of dead mollusks’ shells Crenomytilus grayanus, Mizuhopecten yessoensis, Azumapecten farreri (Jones & Preston 1904) (Bivalvia: Pectinidae), and barnacles Balanus crenatus Bruguière, 1789 (Arthropoda: Thecostraca: Balanidae) (Fig 1). Morphological description. Meristic counts: males—D XXXXII–XXXXV 50–52, A I 71–73, P 1 9, vert. 98– 100, females—D XXXXII 52, A I 72, P 1 9, vert. 98. All specimens of both sexes have one strong spine in the origin of anal fin and a small (2–6 mm length) and narrow (1–2 mm) tentacle which is in the middle of head top, between and ahead of the eyes. All reproductive males have good visible urogenital papilla. Total length of males (mean 99.1 ± 8.2 mm, range 87.5–113.5) is noticeably more than same of females (mean 89.7 ± 7.8 mm, range 78–102) (Fig. 2A). Some females are bigger than males but usually male is bigger than female in the spawning pairs (Fig. 3A, B, Table 1). In total, difference in TL among sexes is authentic (Mann–Whitney U–test, p= 0.002337). Remark. *—CAS-SU 7025, additional original material, n=3, females; **—USNM 71471, n=6, sex not identified; ***— USNM 105238, n=1, sex not identified. Proportional measurements c/ TL, % of males (mean 19.3 ± 1.6, range 16.7–21.9) is noticeably more than same of females (mean 16.8 ± 1.6, range 12.2–18.4), too (Fig. 2B). Usually, than bigger TL fish than higher proportion c/TL, small fishes both sexes have lowest proportion. Difference c/TL among sexes is authentic (Mann–Whitney U–test, p= 0.002337), it is some higher than difference in TL. Proportional measurements lmx/c, % of males (mean 54.8 ± 5.1, range 47.9–67.44) is noticeably more than same of females (mean 45.4 ± 5.4, range 35.2–52.49), too (Fig. 2C). Difference is authentic (Mann–Whitney U–test, p= 0.000018). Visually, males usually have much more large mouth than females, and maxilla at male usually extends far beyond eye (Fig. 3 A, B). TL, proportion c/TL and lmx/c at spawning pairs of fish have the same tendency but with others meanings (Table 1). Both males and females have similar coloration: brownish background of body, up to 18 white club-shaped spots on dorsal and anal fins, row of 8–10 whitish spots laterally on the mid of body, row of 5–7 whitish spots on the belly (Fig. 3A, B). These spots are larger than eye diameter. Numerous fine whitish spots are on the body sides. Some females have reddish body background (Fig. 3B). Coloration of males is more brightly than one at females, dark colors look like as sharp contrast with white spots. Coloration of upper part of head (above of eye) have the same background as body—brownish at both sexes. Coloration of lower part of male head consists from 5–6 thin black twisting oblique lines with short vertical ones, which together form net-like figure. Such figure on the lower part of head of females absent, only 1–2 black thin twisting oblique lines are visible there (Fig. 3A, B). The form of males head looks a little wider, than at females, due to more massive upper jaw, a greater mouth and slightly ventricose cheeks. The head of females looks more pointed and accurate. Fish head coloration changes during growth from “ N. steindachneri ” form (1) to “ N. pulcher ” form (2), so all young fish and mature females have coloration form 1 (Fig. 4), and mature males have coloration form 2 (Fig. 3). Redescription of Neozoarces sp. type series specimens. First description of N. pulcher by Steindachner (1880) incomplete: number of fish type series and holotype were not determined, TL and other measurements (in mm) absent, sex of 3 syntypes not identified.Therefore, we made a brief morphological description of N. pulcher syntypes. TL of syntypes 107.0, 104.2, 90.5 mm (here and later—measurements of syntypes 1–3, respectively), head length 19.0, 16.5, 13.7 (Table 2); body depth 9.6, 10.2, 9.0; snout length 2.8, 2.9, 2.6; upper jaw length 8.0, 7.5, 6.6; orbit length 3.2, 3.1, 3.0; pectoral fin length 8.1, 8.7, 6.1 mm. Proportional measurements: c/TL 17.8, 15.8, 15.1 % (Table 2); H/TL 9.0, 9.8, 9.8; ao/c 14.7, 17.6, 19.0; lmx/c 42.1, 45.5, 48.2; o/c 16.8, 18.8, 21.9; hP/c 42.6, 52.7, 44.5 %, counts: D XLV–XLVI 45–50, A I 72–75, P 1 9–11, vert. 93–97. Body coloration of syntypes (in alcohol) looks like a pale brown color version of our specimens described above, net-like figure on the lower part of heads not visible. F. Steindachner (1880) gives next proportional measurements: c/TL 15.79–19.05 %; H/TL 10.5–11.1; ao/c 20–22.2; o/c 16.7–21.05; hP/c 40.0–50.0 %; counts: D XL 50, A I 75, P 1 10. Since Steindachner (1880) did not determine which of type series specimens is the holotype, we propose to define syntype 3, due to its better preservation, as the lectotype N. pulcher, and the syntypes 1, 2 as paralectotypes. First description of N. steindachneri by Jordan & Snyder (1902) is some more full, authors noted TL collected five specimens — 60 mm, and proportional measurements: c/TL 15.79%; H/TL 11.1; o/c 20.0; ao/c 20.0; io/c 13.3%; hP/c 43.75; counts D XXXVIII 49, A I 72. Sex not identified. Our measurements as follows (holotype, paratype, additional materials): TL 60.1, 63.0, 51.0–62.7 (Table 2); head length 9.6, 8.9, 7.8–10.7; c/TL 16.0, 14.1, 12.5– 19.8%; upper jaw length 4.4, 4.5, 3.2–5.9; lmx/c 45.8, 50.8, 43.6–52.0; ao 18.4, 19.0, 18.4; hP/c 52.3, 54.3–62.5%. Sex identified for additional material* (type series), all specimens are females. Counts (holotype CAS-SU 7025, paratype USNM 50227): D XLIII 53, A I 72, vert. 93, D XLII 49, A I 75, vert. 92. Coloration of fish in alcohol monotonously brown, bars and spots poorly visible. Genetic description. Two fragments of mt DNA (COI and cytb) from 6 specimens having some morphological differences have been analyzed. In the fragment of a gene cytochrome oxidase I with a length 655 base pairs (bp) two samples (N. pulcher- form (B and W) (voucher MIMB-28445-7– KJ427778 and voucher MIMB-28445- 6– KJ427779 respectively) had on one nucleotide replacement, distinguishing them from other sequences. Protein sequences at all individuals were identical. When comparing the nucleotide sequences of the COI gene of our fish with the data available in the GenBank for N. pulcher (length of alignment 576 bp), variability was found in the FJ932603 sequence (1 nucleotide). In the KP237941 sequence, one nucleotide is not defined (“N”). Figure 5 displays a phylogenetic tree showing the location of the species we are studying on the same branch with bootstrap supports of 100/100 for NJ and ML trees. It is necessary to note, that the sequence of gene COI at one N. pulcher—form (B) (voucher MIMB-28445-9) to sequencing directly was not possible. After its cloning in T/A Clone (Thermo Fisher Scientific), it has appeared, that it is presented by two fragments: one is a functional copy of a gene and the second, having deletion in 11 nucleotides, a framework of reading leading breakage. Sequences of a gene cytochrome b (1066 bp) at all individuals were identical. We found only two nucleotide sequences of the Neozoarces pulcher cytb gene in the GenBank. After aligning them with the sequences we obtained, a difference of 1 nucleotide with the KP237952 sequence was found. We did not build phylogenetic trees based on the nucleotide sequences of the cytb gene due to the small amount of data in the GenBank on this mtDNA fragment for the Neozoarcidae family.