Phoxinusseptimaniae Kottelat, 2007

Denys, Gaël P. J., Dettai, Agnès, Persat, Henri, Daszkiewicz, Piotr, Hautecoeur, Mélyne & Keith, Philippe, 2020, Revision of Phoxinus in France with the description of two new species (Teleostei, Leuciscidae), Cybium 44 (3), pp. 205-238 : 216-220

Phoxinusseptimaniae Kottelat, 2007

( Fig. 7 )

Pisciculus varius Dralet, 1821: 35 (type locality: Luech Stream, Chamborigaud, France, see below).

Phoxynus montanus Ogérien, 1863: 362 (type locality: Ain River near Champagnole; Queue de Cheval waterfall, near Saint-Claude, France).

Phoxinus laevis var. montanus Blanchard, 1896: 155 (type locality: Ain River near Champagnole; Queue de Cheval waterfall, near Saint-Claude, France).

Phoxinusseptimaniae Kottelat, 2007: 148 , fig. 3 (type locality: River Agly, bridge downstream of Latour-de- France, 42°46’09”N, 2°39’39”E, Dépt. Pyrénées-Orientales, France).

Material examined

MNHN 2006-1701 , holotype, m, 46.1 mm SL; France: Agly River , Latour-de-France, 42°46.1’N, 2°38.5’E, alt. 92 m. MNHN 2006-1702 , paratypes, 2 m, 1 f, 41.6-46.4 mm SL; France: Agly River , Latour-de-France, 42°46.1’N, 2°38.5’E, alt. 92 m. MNHN 2014-2754 , 5, 46.1-56.2 mm SL; France: Agly River , Latour-de-France, 42°46.1’N, 2°38.5’E, alt. 92 m. MNHN 2019-0264 , m, 64.8 mm SL; France: Luech Stream, Chamborigaud, 44°19.7’N, 3°57.5’E, alt. 335 m. MNHN 2014-2804 , 2 m, 3 f, 50.7-61.4 mm SL; France: Luech Stream, Chamborigaud, 44°19.7’N, 3°57.5’E, alt. 335 m. MNHN 2014-2746 , 4 m, 3 f, 36.6-51.3 mm SL; France: Séran Stream, Talissieu, 45°51.3’N, 5°43.2’E, alt. 239 m. MNHN 2014-2730 , 7 m, 11 f, 35.3-63.1 mm SL; France: Suran Stream, Neuville-sur-Ain, 46°04.9’N, 5°20.4’E, alt. 268 m. MNHN 2014-2742 , 2 f, 46.8-50.1 mm SL; France: Galaure Stream, Chateauneuf-de-Galaure, 45°13.4’N, 5°57.6’E, alt. 246 m. MNHN 2014-2726 , 3 m, 3 f, 53.1-61.4 mm SL; France: Bourne Stream, Saint-Thomasen-Royan, 45°03.7’N, 5°16.8’E, alt. 173 m. MNHN 2014- 2727 , 2 m, 3 f, 51.1-65.3 mm SL; France: Vannon Stream, Brotte-lès-Ray, 47°36.2’N, 5°44.1’E, alt. 204 m. MNHN 2014-2744 , 1 m, 3 f, 50.6-71.2 mm SL; France: Lez Stream, Montjoux, 44°30.1’N, 5°05.2’E, alt. 444 m. MNHN 2014- 2802 , 1 f, 46.4 mm SL; France: Linotte Stream, Loulans- Verchamp, 47°26’N, 6°12.4’E, alt. 234 m. MNHN 2014- 2752 , 2 m, 3 f, 50.3-55.8 mm SL; France: Calavon Stream, Viens, 43°51.7’N, 5°34.3’E, alt. 342 m. MNHN 2014-2737 , 4 m, 3 f, 55.4-72.3 mm SL; France: Ilouvre Stream, Babeau- Bouldoux, 43°26.8’N, 2°54.7’E, alt. 222 m. MNHN 2014- 2758 , 2 f, 54.9-57.3 mm SL; France: Orbieu Stream, Ribau- te, 43°06.1’N, 2°37.3’E, alt. 107 m. MNHN 2014-2736 , 4 m, 2 f, 45.1-61.9 mm SL; France: Hérault River , Valleraugue, 43°05.2’N, 3°36.2’E, alt. 430 m. MNHN 2014-2763 , 2 m, 3 f, 46.1-56.2 mm SL; Lez River , Prades-le-Lez, 43°41.9’N, 3°51.3’E, alt. 61 m .

Material used for molecular analysis

MNHN 2014-2754 , 1, tag 4089; France: Agly River , Latour-de-France, 42°46.1’N, 2°38.5’E, alt. 92 m. (COI: GenBankaccessionnumber: MT975754 ). MNHN 2019 - 0264, tag 1994; France: Luech Stream, Chamborigaud, 44°19.7’N, 3°57.5’E, alt. 335 m. (COI: GenBank acces- sion number: MT975748 , 12S: GenBank accession number: MT975373 ). MNHN 2011-0388 , 1, tag 4416; France: Orain Stream, Saint-Baraing, 46°53.8’N, 5°26.8’E, alt. 496 m. (COI: GenBankaccessionnumber: MT975792 ). MNHN 2011-0407 , 1, tag 4393; France: Ouche Stream, Fleurey-sur- Ouche, 47°18.5’N, 4°50.6’E, alt. 293 m. (COI: GenBank accession number: MT975789 ). MNHN 2014-2727 , 1, tag 12454; France: Vannon Stream, Brotte-lès-Ray, 47°36.2’N, 5°44.1’E, alt. 204 m. (COI: GenBankaccessionnumber: MT975728 , 12S: GenBankaccessionnumber: MT975360 ). MNHN 2011-0946 , 1, tag 10820; France: Rhône River , Jons, 45°48.7’N, 5°05.5’E, alt. 180 m. (COI: GenBank accession number: MT975712 ). MNHN 2010-0519 , 1, tag 4131; France: Boulzane Stream, Puilaurens, 42°47.3’N, 2°18.5’E, alt. 514 m. (COI: GenBankaccessionnumber: MT975764 , 12S: GenBankaccessionnumber: MT975379 ). MNHN 2011-0838 , 2, tag 10701 and 10702; France: Boul- zane Stream, Saint-Paul-de-Fenouillet, 42°48.3’N, 2°29’E, alt. 252 m. (COI: GenBank accession numbers: MT975700 , MT975701 , 12S: GenBankaccessionnumber: MT975346 ). MNHN 2010-0495 , 1, tag 4107; France: Desix Stream, Ansignan, 42°45.3’N, 2°31.3’E, alt. 197 m. (COI: GenBank accession number: MT975758 ). MNHN 2014-2737 , 3, tag 12344-12346; France: Ilouvre Stream, Babeau-Bouldoux, 43°26.8’N, 2°54.7’E, alt. 222 m. (COI: GenBank accession numbers: MT975721 - MT975723 , 12S: GenBank acces- sion numbers: MT975355 - MT975357 ). MNHN 2011-0244 , 2, tag 4349 and 4350; France: Orbieu Stream, Ribaute, 43°06.1’N, 2°37.3’E, alt. 107 m. (COI: GenBank accession numbers: MT975782 , MT975783 ). MNHN 2011-0275 , 3, tag 4311-4313; France: Sals Stream, Coustaussa, 42°56.3’N, 2°16.7’E, alt. 242 m. (COI: GenBank accession numbers: MT975773 - MT975775 , 12S: GenBank accession num- bers: MT975385 ). MNHN 2010-1069 , 1, tag 4272; France: Buèges River , Pégairolles-de-Buèges, 43°48.8’N, 3°35.5’E, alt. 176 m. (COI: GenBankaccessionnumber: MT975771 , 12S: GenBankaccessionnumber: MT975384 ). MNHN 2010-1071 , 1, tag 4273; Garel Stream, Saint-Jean-de-Buèg- es, 43°50’N, 3°37’E, alt. 181 m. (COI: GenBank acces- sion number: MT975772 ). MNHN 2011-0834 , 2, tag 4440 and 4441; France: Hérault River , Valleraugue, 43°05.2’N, 3°36.2’E, alt. 430 m. (COI: GenBank accession numbers: MT975794 , MT975795 ). MNHN 2014-2736 , 2, tag 12449 and 12451; France: Hérault River , Valleraugue, 43°05.2’N, 3°36.2’E, alt. 430 m. (COI: GenBank accession numbers: MT975726 , MT975727 , 12S: GenBankaccessionnumber: MT975359 ). MNHN 2010-1066 , 1, tag 4271; France: Lez River , Prades-le-Lez, 43°41.9’N, 3°51.3’E, alt. 61 m. 12S: GenBank accession numbers: MT975383 ). MNHN 2011- 0243 , 2, tag 4347 and 4348; France: Lez River , Prades-le- Lez, 43°41.9’N, 3°51.3’E, alt. 61 m. (COI: GenBank acces- sion numbers: MT975780 , MT975781 , 12S: GenBank accession number: MT975387 ). MNHN 2010-0511 , 1, tag 4127; France: Massane River , Argelès-sur-Mer, 42°31.9’N, 2°59.3’E, alt. 68 m. (COI: GenBankaccessionnumber: MT975781 , 12S: GenBankaccessionnumber: MT975387 ). MNHN 2010-0502 , 1, tag 4114; France: Tanyari Stream, Saint-Génis-des-Fontaines, 42°32.3’N, 2°56.1’E, alt. 57 m. (COI: GenBankaccessionnumber: MT975759 , 12S: GenBankaccessionnumber: MT975377 ). MNHN 2013- 0682 , 1, tag 16630; France: Maureillas Stream, Le Bou- lou, 42°31.1’N, 2°49.7’E, alt. 80 m. (12S: GenBank acces- sion number: MT975364 ). MNHN 2010-0995 , 2, tag 4445 and 4446; France: Tech River , Elne, 42°35’N, 2°58.2’E, alt. 8 m. (COI: GenBank accession numbers: MT975797 , MT975798 , 12S: GenBankaccessionnumber: MT975390 ). MNHN 2010-1000 , 1, tag 4177; France: Têt River , Eus, 42°38.6’N, 2°28.9’E, alt. 275 m. (COI: GenBank accession number: MT975765 ). MNHN 2011-0245 , 2, tag 4331 and 4332; France: Crieulon Stream, Orthoux-Serignac-Quilhan, 43°53.3’N, 4°02.1’E, alt. 59 m. (COI: GenBank accession numbers: MT975776 , MT975777 ). MNHN 2011-0251 , 2, tag 4333 and 4334; France: Crieulon Stream, Orthoux- Serignac-Quilhan, 43°53.3’N, 4°02.1’E, alt. 59 m. (COI: GenBank accession numbers: MT975386 , MT975779 , 12S: GenBank accession numbers: MT975386 ) .

Diagnosis

Phoxinus septimaniae is distinguished from the other French Phoxinus species by its unique nuptial colour pattern. The male has a blackish stripe along Z3 sometimes fractioned in blotches (vs. green stripe or black / green bars), Z4 greyish (vs. greenish or yellowish), pectoral fin bases yellow-orange with a red disc on the inner side (vs. yellow or orange without disc), a yellow spot between the eye and the mouth (vs. absence) ( Fig. 7A, C ). In the nuptial female, the Z4 is white (vs. greyish, yellowish or greenish) ( Fig. 7B, D ).

Phoxinus septimaniae is further distinguished from P. bigerri by having 75-94 scales along the lateral series (vs. 68-87), a slightly pointed snout (vs. very stout), a subterminal mouth (vs. terminal or slightly subterminal), a straight to slightly concave anal fin margin (vs. straight to convex), a deep caudal peduncle (caudal peduncle depth 2.7-4.6 times in its length vs. 2.1-3.4), and some populations have continuous breast scale patches across the breast (vs. separated or rarely connected anteriorly by 1-2 rows of scales).

Phoxinus septimaniae is further distinguished from P. csikii by having a slightly pointed snout (vs. very stout and blunt), a straight to slightly concave anal fin margin (vs. straight to slightly convex), and in some populations the breast is fully covered by scales (vs. two scale patches separated by an unscaled area).

The nuptial male P. septimaniae is further distinguished by having, black bars crossing Z2-Z3 (vs. Z1-Z5 in P. bigerri ), Z4 greyish (vs. golden-yellow in P. bigerri ; green in P. csikii ; green-yellowish in P. phoxinus ), belly white (vs. greyish to black in P. bigerri ; shiny red in P. csikii ), lips red (vs. pinkish in P. bigerri and P. phoxinus ), pectoral, pelvic and anal fin bases red (vs. pinkish in P. bigerri and P. phoxinus ). The nuptial female is further distinguished by having a black lateral stripe along Z3 (vs. no stripe), and no bars (vs. bold black bars crossing Z2 to Z4 in P. bigerri ).

Nuptial colouration

Male. – Snout and top of head dark brown. Ablack bar on operculum reaching from top of head to branchiostegal rays. Cheeks white. Yellow spot between eye and mouth, a second one covering suboperculum and posterior part of operculum. Lips red. Z1 brown to black, Z2 pale-brown to golden. A blackish stripe along Z3 fractioned in blotches in some individuals and at some places. Z4 greyish, without green colouration, except rarely around pectoral fin. Z5 blackish. Pectoral fin bases usually yellowish-orange or red with a red disc at inners side ( Fig. 7A, C ).

Female. – No or an indistinct yellow spot on operculum. Z1 brownish. Z2 pale-brown. Stripe black along Z3. Z4 silvery white and Z5 black. ( Fig. 7B, D ).

Molecular characterization

On the COI marker, P. septimaniae has 10 diagnostic sites, including three synapomorphies (Tab. II). For the 12S rDNA marker, three diagnostic sites characterise this species: A (vs. G) in position 304, T (vs. Aor G) in position 390 and T (vs. C) in position 394.

Distribution

Phoxinus septimaniae occurs in all the Rhône drainage as well as the French coastal Mediterranean catchments ( Fig. 1 ) until northeastern Spain ( Palandačić et al., 2017; Corral-Lou et al., 2019). It was introduced in the Roya, the Rhine catch- ments as well as in the Geneva (France/Switzerland) and the Ceresole (Italy) Lakes ( Kottelat, 2007; Palandačić et al., 2017).

Vernacular name

Languedoc minnow (English), Vairon du Languedoc (French).

Remarks

Phoxinus septimaniae was originally diagnosed having a continuous patch of breast scales ( Kottelat, 2007). The specimens examined by Kottelat (2007) originate from the Tech River, the stream Lez, the Gardon (Rhône drainage), the River Hérault and the stream Vidourle. We examined materials from all these populations and identified these morpho- logically as P. septimaniae according to Kottelat (2007). However in our analysis, these “scaled breast” minnows form a paraphyletic group within the Mediterranean clade which includes also fish having two separated scaled patches on the breast from the Rhône drainage and some Mediter- ranean rivers as the Vidourle and the Buège ( Fig. 3B ). While the “scaled breast” minnows have very different haplotypes, each of these haplotypes is present in other minnows from Mediterranean catchments (stream Lez, Aude catchment) and in the Rhône drainage without the connection between the patches of breast scales. Kottelat (2007) had observed that some specimens from the Vidourle and the Aude catchment had two separate patches of breast scales and hypothesized the existence of introgressions with P. phoxinus . This hypothesis is plausible as there were many stocking operations of minnows in Europe ( Palandačić et al., 2020). However, an alternative explanation would be that the continuous patches of breast scales are a variable character state in P. septimaniae . Then, P. septimaniae should include populations from the Rhône drainage, which have separated breast scaled patches in order to be a monophyletic group. Therefore, we identify the fish from the Rhone as P. septimaniae .

Nomenclatural notes

The name Pisciculus varius , first introduced by Rondelet (1555: 205), is made available by Dralet (1821: 35). No type specimens are known and no type locality is mentioned, as this was not the standard at these times. Dralet (1821) diagnosed P. varius by having a gold shining back, a silver belly and crimson at its sides (“l’or brille sur son dos, l’argent sur son ventre et le pourpre à ses côtés” in the original French description). This is the translation of Rondelet’s description: “ Dorsum aurei splendoris est, venter argentei, latera purpurascut ” ( Rondelet, 1555). The minnow drawn by Ron- delet (1555: 205) is one of the syntypes mentioned by Dralet. Moreover, Dralet describes the taste and the flesh texture of the species. These minnows, which were eaten, also belong to the syntype series, but as Dralet lived in southwestern France, they might have come from another drainage (Garonne or Adour) and in fact might belong to another species ( P. dragarum or P. bigerri ). Thus, we designate the specimen illustrated by Rondelet as lectotype of Pisciculus varius Dralet, 1821 . The type locality of Pisciculus varius is restricted to the locality of the lectotype. Rondelet (1555) gave no information on the origin locality of the specimen drawn. Rondelet was born and lived in Montpellier, however he also travelled to Paris, Maringues, Florence, the Netherlands and Belgium. So, while the lectotype could have come from the French Mediterranean, it could also have originated from the Seine, Loire, Meuse or Rhine catchments and could be attributed to P. septimaniae , P. phoxinus or P. fayollarum . Furthermore, the illustration of the lectotype does not allow us to identify a minnow ( Cuvier and Valenciennes, 1844), much less the Phoxinus species to which it may correspond. Rondelet’s collection, if it has ever existed, has disappeared. The lectotype is thus not extant. On the different colouration patterns described in these areas (e.g. Fournel, 1836; Selys- Longchamps, 1842; Villatte des Prûgnes, 1897; Moreau, 1899; Piton, 1931), only the description from Crespon (1844: 296-297) is concordant. Crespon lived in Nîmes (about 50 km to the Northeast of Montpellier) and described the minnows living in the Fountain basins as having “…a gold- en line on each side of the body, crimson and purplish glints with gold-coloured stripes on the sides; silver belly…” (“… une lignée dorée sur chaque côté du corps, des reflets pour- prés et violâtres avec des lames couleur d’or sur les côtés; ventre argentin…” in the original French description). Both descriptions given by Rondelet (1555) and Crespon (1844) are very similar, so we consider that the lectotype of Pisciculus varius came from the French Mediterranean catchments where Rondelet worked extensively. We need to stabilise the type locality (art. 75.3.1) choosing a population adjacent to Montpellier and Nîmes for which we already have molecular and morphological data allowing the identification among all other French Phoxinus species (art. 75.3.2; see above). We designate as neotype the specimen MNHN 2019-0264 (art. 75.3.3) (64.8 mm SL), as the lectotype is not extant (art. 75.3.4) andcorresponds to what we know of the former types (art. 75.3.5). The neotype came from a locality close to the original type locality (art. 75.3.6) in the lower Rhône drain- age close to Montpellier and Nîmes (France: Luech stream, Chamborigaud, 44°19.7’N, 3°57.5’E), which now becomes the type locality of Pisciculus varius . This specimen is now deposited in the MNHN collections (art. 75.3.7). Conse- quently, we consider Pisciculus varius Dralet, 1821 and Phoxinus septimaniae Kottelat, 2007 as the same species. However, Pisciculus varius was used as a valid name only in 1821 to our knowledge, so before 1899 (art. 23.9.1.1). More- over, since its description in 2007, P. septimaniae was used as a presumed valid name in at least 31 works by 28 different authors ( Kottelat and Freyhof, 2007; Hanel et al., 2009; de Sostoa et al., 2010; Collin and Fumagalli, 2011; Keith et al., 2011; Agence Gaïadomo, 2012; Almeida et al., 2013; Mace- da-Veiga, 2013; Witté et al., 2013; Geiger et al., 2014; Beh- rens-Chapuis et al., 2015; Denys, 2015; FernÁndez Cortés, 2015; Ibrahim, 2015; Knebelsberger et al., 2015; Miró and Ventura, 2015; Palandačić et al., 2015, 2017, 2020; Aparicio et al., 2016; Civade, 2016; Leunda et al., 2017; Maire et al., 2017; Aparicio, 2018; Arsento et al., 2018; Schönhuth et al., 2018; Vucić et al., 2018; Bogutskaya et al., 2019; Corral-Lou et al., 2019; Gauliard et al., 2019; Vinyoles et al., 2019) (art. 23.9.1.2). Consequently, we state that, to our knowledge, the condition in Article 23.9.1 applies. From the date of publica- tion of that act P. septimaniae Kottelat, 2007 has precedence over Pisciculus varius Dralet, 1821 . Phoxinus septimaniae is a nomen protectum and Pisciculus varius a nomen oblitum. Then, Pisciculus varius Dralet, 1821 is a senior synonym of Phoxinusseptimaniae Kottelat, 2007 .

Ogérien (1863) described Phoxynus montanus from the Ain River near Champagnole and the Queue de Cheval waterfall near Saint-Claude (Eastern France, Rhône drain- age). This species is characterised by having 10 pectoral fin rays (confirmed by Blanchard (1896)) (vs. 13-16 in P. septimaniae ) and a nuptial colouration greyish black spattered with brown or olive colour and the flank with blue, yellow and greenish spots (vs. without blue or greenish spots in P. septimaniae ). Lunel and Lunel (1874) stated the population at the type locality is distinguished from the minnows observed in the Lake Geneva basin where characteristic haplotypes of both P. csikii and P. septimaniae were found suggesting that these minnows are hybrids ( Palandačić et al., 2017). With the absence of a syntype series and waiting for further investigations, we consider Phoxynus montanus Ogérien, 1863 and P. septimaniae Kottelat, 2007 as the same species. However, Phoxynus montanus was not used as a valid name after 1899 (art. 23.9.1.1). As discussed for P. varius , the article 23.9.1.2 is also applied for P. septimaniae . By consequence, we state that, to our knowledge, the condition in Article 23.9.1 applies here too. This makes P. montanus a nomen oblitum. Therefore, Phoxynus montanus Ogérien, 1863 becomes a senior synonym of P. septimaniae Kottelat, 2007 .

Phoxinus laevis var. montanus Blanchard, 1896 isalready an objective synonym of Phoxynus montanus Ogérien, 1863 . Phoxinus laevis var. montanus Blanchard, 1896 is a senior synonym of P. septimaniae Kottelat, 2007 .

Figures

Figure 1. – Records of Phoxinus used in this study. Coloured squares represent the six species delineated in this study: P. phoxinus (blue), P. bigerri (red), P. csikii (green), P. septimaniae (yellow), P. dragarum (purple), and P. fayollarum (orange).

Figure 3. – Phylogenetic tree by Bayesian inference with the COI marker (592 bp) on 482 sequences of Phoxinus spp. (A) with a focus on the P. septimaniae clade (B). Coloured boxes represent evolutionary lineages containing French specimens. Bold vertical bars represent ABGD delineations. Red asterisks represent specimen with a completely scaled breast. Posterior probabilities values are indicated above the nodes.

Figure 7. – Phoxinus septimaniae in the Rhône and Mediterranean catchments: m from the Luech Stream, Chamborigaud 44°19.7’N, 3°57.5’E, MNHN 2014-2804 (A); f from the Hérault River, Valleraugue 44°05.2’N, 3°36.3’E, MNHN 2014-2736 (B); nuptial col- ouration pattern for male (C) and female (D).