Euprymna parva ( Sasaki, 1913 ) Sanchez & Jolly & Reid & Sugimoto & Azama & Marlétaz & Simakov & Rokhsar, 2019

Sanchez, Gustavo, Jolly, Jeffrey, Reid, Amanda, Sugimoto, Chikatoshi, Azama, Chika, Marlétaz, Ferdinand, Simakov, Oleg & Rokhsar, Daniel S., 2019, New bobtail squid (Sepiolidae: Sepiolinae) from the Ryukyu islands revealed by molecular and morphological analysis, Communications Biology 2 (465), pp. 1-15 : 6-7

Euprymna parva ( Sasaki, 1913) , comb. nov.

( Figure 1c left, Table 1 , Supplementary Figs. 1 , 2c, d , Supplementary Table 2).

Sepiola parva Sasaki, 1913: 252 , Fig. 4 . — Sasaki, 1929: 136–137 Pl. XV, Figs. 4 and 5 , text Fig. 80; Takayama and Okutani, 1992: 203–214, fig, 2, Figs. 4–6 . — Okutani, 1995: 45, fig. 43. Reid and Norman, 1998: 717. — Reid and Jereb, 2005: 165–166, fig. 239.

Inioteuthis parva Sasaki, 1914: 595 , pl. 11 Figs. 9 and 10.

Type locality: Japan, Tokyo Bay .

Material examined. 3 ♂ (8.5–10.7 mm ML) , 3 ♀ (7.0–8.0 mm ML), East China Sea , Okinawa, Diamond Beach in Seragaki, 26.51N, 127.88E, <2 m, 15 June 2016, coll. J. Jolly, G. Sanchez, A. Masunaga & K. Asada ( AM C.574777, Hap 3, and GenBank accession number: LC417215 ) .

Remarks. The correct generic placement of this taxon has been equivocal since it was first described. It was first placed in Sepiola , transferred to Inioteuthis Sasaki, 1914 , then later retained in the genus Sepiola . Sasaki does not explain why parva was placed in Sepiola in his 1929 treatise, nor why it was referred to Inioteuthis in his 1914 work. Clearly, however, the features noted by Sasaki,

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“the nipple-like protruberance” near the base of the hectocotylus and the “peculiar and unstalked cylindrical suckers” (Sasaki48: 137) are characteristic of Euprymna . Mature males have two rows of suckers on their arms, in contrast to the four rows typically (but not always) found among Euprymna . This has, no doubt, resulted in its misplacement in the genus Sepiola , that has continued largely without question until now. However, two rows of arm suckers are also found in E. pardalota and E. phenax . The inclusion of the E. pardalota COI sequence in our analyses ( Fig. 2 ) confirms its position, and that of “ Sepiola ” parva in the monophyletic genus Euprymna . Optic lobe transcriptome data ( Fig. 3 ) clearly places S. parva in the monophyletic Euprymna clade, and the pairwise synonymous substitution rate (Ks) between clades (Supplementary Fig. 1 ) highlights the disjunction between Euprymna and Sepiola , providing strong support for their distinct generic status. The genetic distance data fully supports the placement of S. parva in the genus Euprymna and this evidence now permits a more robust definition of the genus based on morphological characters.

The hectocotylus (dorsal left arm) of Euprymna is unique among the Sepiolinae . In all genera but Euprymna , the hectocotylus is clearly tripartite, with a morphologically distinct basal part, copulatory apparatus, and distal part49,50. In contrast, in Euprymna , the hectocotylized arm has a bipartite form, with a proximal portion and a distal modified part. In Euprymna there is no distinct copulatory apparatus, instead the pedicels of the ventral suckers in the third to fourth proximal rows are modified to form 1–2 papillae in most species, sometimes bearing a vestigial sucker, whereas in all other genera the sucker pedicels forming the copulatory apparatus are more conspicuously modified (mostly horn- or hook-like). More importantly, the distal-most portion of the Euprymna hectocotylus bears deeply modified sucker-stalk elements: the stalks are columnar, i.e., thickened and lengthened, and appressed to each other to form palisades, and the sucker proper is reduced to a small opening surrounded by a chitinous rim, often covered by a fleshy cap and embedded in the columnar pedicel. On the contrary, in all other genera, the hectocotylus distal suckers are normal (in some cases some of them may be enlarged and/or their stalks slightly c Male arm crown, dorsal view, holotype 14.9 mm ML (NSMT Mo 85885), scale bar 5 mm. d Female right side (of animal) arm crown, oral view, paratype, 15.3 mm ML (NSMT Mo 85893), scale bar 2 mm. c, d Numbers 1 – 4 indicate Arms 1 – 4. e SEM Arm 4 sucker rim, paratype female, 19.5 mm ML (NSMT Mo 85889), scale bar 20 µm. f SEM enlargement of sucker rim shown in (d), scale bar 10 µm.

lengthened) (Bello, submitted). The very simple copulatory apparatus of Euprymna is considered a plesiomorphic character state 51 in the Sepiolinae , placing this genus in a basal position within the subfamily36,52.

Figures

Fig. 1 Ryukyuan bobtail collection sites, eggs masses, and adult morphotypes. a Ryukyu Archipelago collection sites. b Ryukyuan bobtail squid egg clutches from Types 1–3; scale bars 5, 4 and 12 mm, respectively. c Live adult Type 2 and Type 3 collected during surveys, scale bars 5 and 11 mm, respectively.

Fig. 2 Maximum likelihood tree estimated for COI sequences. Possible misidentifications of original taxa: *suspected E. morsei (Verrill, 1881); **suspected E. hyllebergi Nateewathana, 1997; ***suspected misidentification, likely sepiadariid; Sepiadariidae; +, likely S. birostrata (see Supplementary Table 1). Nodes with bootstrap support <95% are shown. #We have reassigned this species to Euprymna below.

Fig. 4 Euprymna brenneri sp. nov. Holotype male, 14.9 mm ML (NSMT Mo 85885): a Dorsal view, scale bar 5 mm and b Ventral view, scale bar 5 mm.

Fig. 5 Euprymna brenneri sp. nov. a Ventral mantle dissected to show funnellocking cartilage, fc, and mantle-locking cartilage, mc, paratype male, 22 mm ML (NSMT Mo 85891), scale bar 3 mm. b Funnel dissected from ventral side to show funnel organ (fo), same specimen, scale bar 2 mm.

Fig. 9 Euprymna brenneri sp. nov. developmental stages. a Developing embryo inside egg attached to yolk sac (note attachment threads on egg). b Laboratory-reared hatchling, ventral view. c hatchling, dorsal view. d, e Captive reared adults. Note arm banding clearly visible in (a and d) and concentration of blue reflective iridophores in (e).

Fig. 3 Maximum likelihood and Bayesian tree based on 452 orthologous genes using their coding sequences. Nodes with bootstrap support <95% and posterior probability <0.9 are shown. *Members of the Type 2 clade are morphologically indistinguishable from Sepiola parva. Based on the COI analyses, this phylogeny and our morphological analysis, however, we have redefined the genus Euprymna and reassigned this species to Euprymna.