Paracapoeta, a new genus of the Cyprinidae from Mesopotamia, Cilicia and Levant (Teleostei, Cypriniformes)

The molecular and morphological studies carried out within the scope of this study revealed that the scrapers, known as the Mesopotamian group, belong to a different genus. The Paracapoeta gen. nov., from the Mesopotomia and Levant, is distinguished from Capoeta and Luciobarbus species by the presence of a strong ligament between the base of the last simple and the first branched rays of the dorsal-fin (vs. no or a very weak ligament). The Paracapoeta further differs from Capoeta by the last simple dorsal-fin ray strongly ossified in adult specimens (more than 75%, vs. less than 75%). The Paracapoeta further differs from Luciobarbus by the lower lip with horny layer (vs. fleshy lips). The molecular phylogeny based on the combined dataset (COI + Cytb, 1312 bp.) showed that the genus Paracapoeta was recovered from the other groups in the subfamily Barbinae with high bootstrap and posterior probability values (BP: 94%, PP: 0.96). Also, Paracapoeta and Capoeta are well differentiated by an average genetic distance of 8.02±0.78%. The morphological and molecular findings have largely overlapped each other. Besides, Capoeta turani is treated as a synonym of Capoeta erhani.


Introduction
Cyprinid genus Capoeta Valenciennes, 1842 has a wide distribution in the Mediterranean, Middle East, Caucasus and South-West Asia. Even though the members of the genus occur in lakes and spring waters, they generally prefer fast-flowing streams (Kaya 2019). The genus has attracted the attention of various fish taxonomists and they have described a number of new species over the last fifteen years (Turan et al. 2006a(Turan et al. , 2006bÖzuluğ and Freyhof 2008;Alwan 2010;Zareian et al. 2016;Turan et al. 2017;Elp et al. 2018). In parallel with this, many genetic studies have been performed (Turan 2008;Zareian et al. 2016;Bektaş et al. 2017;Bektaş et al. 2019). The Mesopotamian Capoeta group, which is proposed as a new genus in this study, appears to be in a different branch from the Capoeta genus in many genetic studies (Levin et al. 2012;Berrebi et al. 2014;Ghanavi et al. 2016;Jouladeh-Roudbar et al. 2017;Zareian et al. 2018;Bektaş et al. 2017Bektaş et al. , 2019. Molecular phylogenies based on nuclear and mitochondrial molecular markers unambiguously demonstrated that Capoeta is clustered together with the Western Palaearctic barbels of the genera Barbus Daudin, 1805and Luciobarbus Heckel, 1843(Durand et al. 2002Gante 2011;Levin et al. 2012;Buonerba et al. 2015;Yang et al. 2015). However, hybridization-based polyploidy has likely played a major role in the evolution of the hexaploid genus Capoeta (Yang et al. 2015;Levin et al. 2019), which emerged through intergeneric hybridization of Luciobarbus (2n = 100) and Cyprinion Heckel, 1843 (2n = 50) (Yang et al. 2015). In fact, the genus Capoeta (6n; Turan 2008), a monophyletic unit and well-defined genus, probably evolved from the ancestors of a tetraploid Luciobarbus as it is closely related to Luciobarbus mursa (Güldenstädt, 1773) and L. subquincunciatus (Günther, 1868) (Levin et al. 2012;Berrebi et al. 2014;Yang et al. 2015).
In recent years, various ideas have been put forward among researchers about the validity of Mesopotamian group species. Erk'akan and Özdemir (2011) compared C. turani (Seyhan) and C. erhani (Ceyhan) morphologically, based on data provided from literature and claimed that both are the synonyms of the C. barroisi. Later, Özdemir (2013) developed this claim further and stated that C. turani (Seyhan), C. erhani (Ceyhan), C. barroisi (Orontes) and C. trutta (Persian Gulf basin) are conspecific and all belong to C. trutta. Recent studies supported the close relationship between C. turani and C. erhani; however, C. erhani, C. barroisi and C. trutta are molecularly well distinguished (Bektaş et al. , 2019. Detailed morphological comparisons among these species confirmed these genetic results (Kaya 2019).
Here (i) we discussed the validity of the Capoeta turani and (ii) proposed a new generic name, Paracapoeta, for the scrapers, formerly known as the Mesopotamian Capoeta group based on morphological and molecular analysis.

Morphological analyses
The care of experimental animals was consistent with the Republic of Turkey's animal welfare laws, guidelines and policies approved by Recep Tayyip Erdogan University Local Ethics Committee for Animal Experiments (permit reference number 2014/77).
Samples were collected by electro-shocker. After anaesthesia, fish were fixed in 4% formaldehyde. Methods for counting followed Kottelat and Freyhof (2007). The later-al line scales were counted from the first scale touching the shoulder girdle to the posterior-most scale at the end of the hypural complex. The last two branched rays articulating on a single pterygiophore in the dorsal and anal fins were counted as "1½." The simple dorsal-and anal-fin rays were not counted because the anteriormost rays are deeply embedded.
For osteological preparation (last simple dorsal-fin ray), one specimen of each selected species of Paracaopeta and Capoeta ( (Pietschmann, 1933) [195 mm SL]) were cleared and stained with alizarin red S, according to the protocol of Taylor and Van Dyke (1985). The specimens were examined using a stereo microscope (Nikon SMZ1500), and photos were taken using a digital machine with a glycerol bath.

Molecular data analyses
The COI (577 bp.) and Cytb (735 bp.) fragments of the seventy-one samples for Capoeta, ( Table S1) were combined with the Clustal W method (Thompson et al. 1994) in the Bioedit 7.2.5 (Hall 1999) as a dataset with a total length of 1312 bp.

FFR
Recep Tayyip  In the original description, C. erhani (Ceyhan River) is distinguished from C. turani (Seyhan River) by having numerous spots (vs. few) and a brown back, lateral head and body (vs. silvery). Besides, the spots in C. erhani are large and often fused into blotches giving the fish a mottled appearance in individuals smaller than 120 mm SL (vs. spots are always small and never fused into blotches in C. turani. Also, caudal peduncle and operculum of C. erhani are always densely spotted (vs. few isolated spots or no spots). Indeed, the silvery body color and the shape, form and number of the spots used in the original description were useful to distinguish both species. However, the fact is that Çakıt (type locality) is a constantly turbid stream (confirmed by Jörg Freyhof, pers. comm., 2019) which has possibly caused these differences in body color and pattern. These changes have also been observed in other species (Capoeta damascina, Oxynoemacheilus sarus, Garra turcica, Squalius adanaensis, Chondrostoma ceyhanensis, Salariopsis sp.) co-occurring with Capoeta turani.
On the other hand, recent molecular studies have stated that there is no difference (0.35%) between these two species at species level (Bektaş et al. , 2019. In the light of this information, we treated C. turani as a synonym of C. erhani.

Rediagnosis of Capoeta
Capoeta Valenciennes, 1842 Rediagnosis. The body fusiform and slightly compressed laterally. In adult individuals, the general body color is brownish, and without dark brown or blackish spots (except C. pestai). The head plain brownish, and no black spots on head and cheek. The mouth inferior, mouth transversely slit or horseshoe-shaped. Lips not developed and lower lip with keratinize edge. One or two pairs of barbel around the mouth. The last simple dorsal-fin slightly or moderately ossified (less than %75) and its posterior edge serrated (except C. antalyensis). No or very weak ligament between the base of the last simple and the first branched rays of the dorsal-fin. There are melanophore rows on the posterior edge of the flank scales. There is no keel in predorsal area, in front of dorsal-fin. Type species. Cyprinus capoeta Güldenstädt, 1773 [actual status of the type species is Capoeta capoeta (Güldenstädt, 1773)].
Distribution. Afghanistan, Armenia, Azerbaijan, Georgia, Iran, Iraq, Jordan, Syria, Pakistan, Kazakhstan, Palestine, Tajikistan, Turkey, Turkmenistan and Uzbekistan: The genus Capoeta has a wide distribution in the Mediterranean, Middle East, Caucasus and South-West Asia.
Diagnosis. The new genus Paracapoeta is distinguished from other genus of Capoeta and Luciobarbus by having a strong ligament between the base of the last simple and the first branched rays of the dorsal-fin (Fig. 1a, b) (vs. no or a very weak ligament in Capoeta and Luciobarbus (Fig. 1c, d)). The new genus is further distinguished from Capoeta and Luciobarbus by the distribution of melanophores on the flank scales (Fig. 2). In Paracapoeta, the posterior part of the scales is covered by more or less melanophores that are irregularly scattered. In Luciobarbus and Capoeta, there are melanophore rows on the posterior edge of the flank scales, and there are no or numerous irregularly scattered melanophores pigments behind the melanophore rows (Fig. 2). It further differs from the genus Capoeta by the last simple dorsal-fin ray strongly ossified in adult specimens (more than % 75, vs. less than % 75) (Fig. 3), a well-developed naked keel in front of dorsal-fin (except P. anamisensis, vs. absent in Capoeta) and the body with numerous irregular-shaped small black spots on the back and flank (except P. anamisensis, vs. absent in Capoeta, except C. pestai) (Figs 4, 5). It further differs from Luciobarbus by having the lower lip with horny layer (vs. with fleshy lips) and lips without papillae (vs. lips with papillae). Table 1. Nucleotide positions for some genera within the subfamily Barbinae. Diagnostic and distinctive nucleotide positions are represented in bold font and gray background, respectively.

Barbus T G C/T T A/G C G G T/C T/C T T T C C C C A/G T C C C G/A A T T C C C A C A/G C C/T
Distribution. Turkey, Iran, Iraq and Syria: Seyhan, Ceyhan and Orontes rivers, Levant drainages; Tigris, Euphrates, Mond and Minab River, Persian Gulf drainages.
Etymology. The name of the new genus is formed by combining the words "Para" and "Capoeta". "Para" means "beside" or "near", and "Capoeta" is the available name of the closest genus of Paracapoeta, deriving from the local vernacular name "kapwaeti" used in Georgia and Azerbaijan.

Discussion
In an effort to re-evaluate the generic structure of the scrapers, Paracapoeta gen. nov., formerly known as the Mesopotamian Capoeta group, was assessed based on morphological and molecular data.
For a combined dataset of Cytb (735 bp) and COI (577 bp), the intergeneric genetic distances for Paracapoeta and Capoeta lineages (mean 8.02±0.78%) represent the lowest limit of the predicted intergeneric genetic distances estimated for especially the western Palearctic Barbinae genera, while it corresponds to the lower limits of intergeneric distance (8-10%; Ward et al. 2005;Hubert et al. 2008;Nguyen et al. 2008;Lara et al. 2010;Perea et al. 2010;Schönhuth et al. 2012) for some closely related Cyprinid genera. On the other hand, Luciobarbus lineage exhibited the highest levels of intrageneric genetic diversity (7.1%, Table 2) for the combined dataset because of the presence of two distinct subclades. Since the newly uncovered lineage of Paracapoeta gen. nov. from genus Capoeta exhibited significant genetic distance in comparison with Capoeta species, the classification of the Mesopotamian Capoeta group as a subgroup of Capoeta is unjustifiable; therefore reclassification as a genus is suggested in the present study.
Consistent with previous studies (Levin et al. 2012;Doadrio et al. 2016;Šimková et al. 2017), our phylogenetic trees revealed some inconsistencies in the current taxonomy of the tribe Barbini, such as the paraphyletic status of the genus Luciobarbus and L. subquincunciatus having a more recent common ancestor with Capoeta and Paracapoeta than the other Luciobarbus.  Note: Values on the diagonal indicate the mean genetic distances within clades.