First record of Paraschistura alta

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1 Iran. J. Ichthyol. (December 2015), 2(4): Received: September 21, Iranian Society of Ichthyology Accepted: Novembers 17, 2015 P-ISSN: ; E-ISSN: doi: First record of Paraschistura alta (Nalbant and Bianco, 1998) from Eastern Iran and providing its COI barcode region sequences (Teleostei: Nemacheilidae) Arash JOULADEH-ROUDBAR *1, Soheil EAGDERI 2, Saber VATANDOUST 3 1 Department of Fisheries, Sari University of Agriculture Sciences and Natural Resources, Mazandran, Iran. 2 Department of Fisheries, Faculty of Natural Resources, University of Tehran, Karaj, Iran. 3 Department of Fisheries, Babol Branch, Islamic Azad University, Babol, Iran. * arash.aarshaan@yahoo.com Abstract: Paraschistura alta, formerly reported from the eastern part of Sistan basin in Afghanistan, is reported for the first time from the Zahak River in Iranian part of this basin and its morphological characteristics, COI barcode and its phylogenetic relationship within other members of the genus Paraschistura from Iran are provided. All the measured meristic and morphometric characters of seven collected specimens of P. alta (SL= mm) from the Zahak River were in the range of its original description. The results of phylogenetic analysis based on the COI barcode region places the Iranian members of the genus Paraschistura into 11 groups, which show between 3.05% and 14.10% K2P sequence divergence. Paraschistura alta from Zahak River corresponds to a distinct clade, sister to P. bampurensis + P. hurmozensis. Keywords: Mitochondrial gene, Phylogenetic analysis, Helmand River, Sistan basin, Iran. Introduction Nemacheilid loaches with about 67 genera and more than 652 valid species are found across Eurasia with one species in northeast Africa (Eschmeyer & Fong 2011). This family has a great diversity in Iranian Inland waters (Coad 2015; Freyhof et al. 2015; Jouladeh-Roudbar et al., 2015a,b; Mafakheri et al., 2015; Azimi et al., 2015a,b; Mousavi-Sabet et al., 2015a,b; Ghasemi et al., 2015). Among the member of family Nemacheilidae, Paraschistura Prokofiev, 2009 is a newly described genus, and therefore, not all of its species are fully examined and ascribed (Coad 2014) till Freyhof et al. (2015) reviewed the genus Paraschistura from Iran and described six new species and provided new information on this taxon. They also provided an identification key for all 11 recognized species. However, it seems that still members of this genus are poorly known from some River drainages in Iran, Turkey, Turkmenistan, Pakistan and Afghanistan. The members of this genus with a dark black spot or strip at the base of the anterior dorsal fin rays (Coad 2015; Freyhof et al. 2015). They are small fishes and characterised by an elongated body with dorsal and ventral profiles almost parallel. The body is compressed anteriorly and the head is compressed or depressed. The caudal peduncle is short and moderately deep. The snout is usually blunt. The dorsal and anal fins usually have 7 and 5 branched rays, respectively. The caudal fin is slightly to deeply emarginated. The lateral line is incomplete, may be very short and rarely reaches the end of the anal fin level. Scales arc absent or rarely present but weakly developed on the caudal peduncle (Coad 2015). Kottelat (2012) listed 14 species in Paraschistura, based mostly on the proposals of Prokofiev (2009) which are described from Afghanistan (P. lindbergi), Pakistan (P. alepidota, 235

2 Iranian Journal of Ichthyology (December 2015), 2(4): P. kessleri, P. lepidocaulis, P. microlabra, P. naseeri, P. pakistanica, P. prashari, and P. punjabensis), Turkey (P. chrysicristinae), and Turkmenistan (P. turcomana = P. turcomanus), and the three species from Iran, including P. bampurensis, P. nielseni and P. sargadensis. Vatandoust & Eagderi (2015) described P. ilamensis from Tigris River drainage as first species of the genus of Paraschistura from Iranian part of this basin. Freyhof et al. (2015) reviewed the genus Paraschistura from Iran and described of six new species including P. abdolii (from the Sirjan basin and the western tributaries of the Hamun-e Jaz Murian basin), P. aredvii (from the Zohreh drainage), P. hormuzensis (from the Minab drainage), P. naumanni (from the Kol drainage, the Mond drainage and the Lake Maharlo basin), P. pasatigris (from the Karun and Karkheh drainages), and P. susiani (from the Jarahi drainage) based on the morphological and molecular (the mtdna COI barcode region) data set. Recently Mousavi-Sabet & Eagderi (2015) also described P. delvari from Mond River drainage (Persis basin). Based on the described characteristics of Paraschistura pasatigris, this species can be suggested as synonym of P. ilamensis. In addition, Freyhof et al. (2015) treated Metaschistura Prokofiev, 2009 as a synonym of Paraschistura Prokofiev, 2009, P. sargadensis as synonym of P. kessleri and P. turcomanus as synonym of P. turcmenica. Mousavi-Sabat et al. (2015) reviewed the genus Paraschistura in the Hari River basin, and revalidated P. turcomana (= P. turcomanus). In addition, P. baluchiorum described from Pakistan is treated as the synonym of P. bampurensis according to Nalbant and Bianco (1998). Also, P. alta described from Afghanistan is retained in the genus Schistora by Kottelat (2012), whereas Coad (2014) considered it as P. alta. The type locality of P. alta is Helmand River basin in Afghanistan at the Kajaki, north east of Girisk and it was originally described as Schistura alta Nalbant & Bianco, This species is characterised by a deep body, a complete lateral line, 236 Fig.1. Map of Eastern Iran showing sampling station (, Zahak River) in the Sistan basin. deeply forked caudal fin and emarginated dorsal finfree margin. Paraschistura alta has been reported only from Helmand River basin in Afghanistan and now found in the Zahak River of the Sistan basin of Iran which may be due to recent floods caused entered this species to the Sistan basin. Therefore, this study is aimed to report first record of this species in Iran by providing its morphological characteristics, diagnostic nucleotide substitutions in the mtdna COI barcode region and its phylogentic relationship within the members of the genus Paraschistura in Iran. Materials and Methods Seven specimens of P. alta were collected from the Zahak River, Sistan and Baluchestan Province, near Zabol 30 49'32"N, 61 45'36"E, in September 2014 during fieldwork on the ichthyofauna of Sistan basin by electro-fishing (Fig. 1). After anesthesia with 1% clove solution, three specimens were fixed in 96% ethanol and the other four specimens were preserved in 4% buffered formaldehyde and transferred to the laboratory for further investigations. The taxonomic key given by Coad (2014) were used to identify the specimens. Meristic characteristics of the specimens were counted using a stereomicroscope. A total of 26

3 Jouladeh-Roudbar et al.-first record of Paraschistura alta from Iran Table 1. Morphaometric data of Paraschistura alta (n=7). range mean SD Standard length (mm) In percent of standard length Head length Body depth at dorsal-fin origin Body width at dorsal-fin origin Predorsal length Postdorsal length Prepelvic length Preanal length Distance between pectoral and pelvic-fin origins Distance between pelvic and anal-fin origins Depth of caudal peduncle Length of caudal peduncle Dorsal-fin depth Pectoral-fin length Pelvic-fin length Distance between anus and anal fin origin In percent of head length Head depth at nape Head depth at eye Snout length Eye diameter Postorbital distance Maximum head width Interorbital width Length of inner rostral barbell Length of outer rostral barbell Length of maxillary barbell morphometric features were measured by a caliper to the nearest 0.01mm (Table 1). All measurements are made point to point based on Kottelat & Freyhof (2007). The percentage ratios of morphometric characters in relations to SL and HL were calculated. The last two branched rays articulating on a single pterygiophore in the dorsal and anal fins are noted as 1½. Unbranched rays of dorsal and anal fins are not counted as they are deeply. DNA extraction and PCR: DNA was extracted from muscle tissue at base of dorsal fin using a Genomic DNA Purification Kit (#K0512; Thermo Scientific Corporation, Lithuania) following the manufacturer s protocol. The COI gene was amplified using primers FCOI20 (5 -AACCTCTGTCTTCGGGGCTA-3 ) and RCOI20III (5 -TTGAGCCTCCGTGAAGTGT G-3 ), designed by (Hashemzadeh-Segherloo et al. 2014). Polymerase chain reaction (PCR) conditions were as follows: a 50μl final reaction volume 237 containing 5μl of 10X Taq polymerase buffer, 1μl of (50mM) MgCl2,1μl of (10mM) deoxynucleotide triphosphate (dntp), 1μl (10μm) of each primer, 1μl of Taq polymerase (5Uμl-1), 7μl of total DNA and 33μl of H2O. Amplification cycles were as follows: denaturation for 10 min at 94 C; 30 cycles at 94 C for 1 min, 58.5 C for 1 min, 72 C for 1 min and a final extension for 5min at 72 C. PCR products were purified using purification Kit (Expin Combo GP mini; Macrogen incorporation, Korea). The PCR products were sequenced using Sanger method by a robotic ABI-3130xl sequencer using manufacturer s protocol. The forward primer FCOI20 was used to single strand sequencing. Molecular data analysis: The sequences were compared to the published Paraschistura sequences using (BLASTn) basic local alignment search tool (Altschul et al. 1990). All sequence data were aligned using MEGA6 software (Tamura et al. 2013). To

4 Iranian Journal of Ichthyology (December 2015), 2(4): Table 1. Accession numbers of the new materials and materials from NCBI (mainly from Freyhof et al. 2015). Number Gen Bank No. Species Number Gen Bank No. Species 1 KM P. abdolii 20 KM P. naumanni 2 KM P. abdolii 21 KM P. naumanni 3 KM P. abdolii 22 KM P. naumanni 4 KU P. alta 23 KM P. nielseni 5 KU P. alta 24 KM P. nielseni 6 KU P. alta 25 KM P. nielseni 7 KM P. aredvii 26 KM P. pasatigris 8 KM P. aredvii 27 KM P. pasatigris 9 KM P. aredvii 28 KM P. pasatigris 10 KM P. bampurensis 29 KM P. susiani 11 KM P. bampurensis 30 KM P. susiani 12 KM P. bampurensis 31 KM P. susiani 13 KM P. bampurensis 32 KM P. turcmenica 14 KM P. cristata 33 KM P. turcmenica 15 KM P. cristata 34 KM P. turcmenica 16 KM P. cristata 35 KJ Paracobitis malapterura 17 KJ P. hormuzensis 36 KJ T. hafezi 18 KM P. hormuzensis 37 KJ Turcinoemacheilus hafezi 19 KM P. malapterura unify the length of the sequences, the common 650bp length segments were selected and used for phylogenetic analysis. Maximum likelihood reconstructions were performed using RAxML (Stamatakis 2006) under the GTR+G+I model of nucleotide substitution, with CAT approximation of rate heterogeneity and bootstrap (10000 bootstrap replicates). Bayesian analysis was performed using MrBayes Starting from a random tree, Metropolis-coupled Markov chain Monte Carlo sampling was performed with four chains run for generations for COI data set of with a sampling frequency of 100. Bayesian posterior probabilities were obtained from the 50% majority rule consensus trees. The resulting BI topology was applied for both presented trees. (Huelsenbeck et al. 2004). All constructed phylograms were displayed and edited in the programmes FigTree, v1.2.1 (Rambaut, 2009). The evolutionary divergence over sequence pairs between groups of the studied species were estimated using the Kimura 2-parameter model (Kimura 1980). The retrieved sequences of the other members of the genus Paraschistura from GenBank database (NCBI) following Blast search is shown in Table 2. As appropriate outgroup to root the 238 constructed phylogenetic hypothesis, the geographically adjacent loach genera i.e. Turcinoemacheilus, Paracobitis and Oxynoemacheilus were included. Results Morphological analysis: Seven specimens of P. alta ranged mm of SL (Fig. 2) were collected from the Zahak River (Sistan and Baluchestan Province, near Zabol) in September The general body shape of this species is displayed in Figure 2. Morphometric characteristics are provided in Table 2. All the measured meristic and morphometric characters of Iranian population of P. alta were in the range of its description provided by Coad (2014). The other morphological features and its coloration are provided as following: Description: Medium sized, slender species with large and depressed head. Body deepest at dorsal-fin origin, depth slightly decreasing towards caudal-fin base. Greatest body width at middle between pectoral- and pelvic fin bases. Section of the head roundish, flattened on ventral surface. Caudal peduncle compressed laterally and short, (mean 1.1) times longer than deep. Pectoral fin reaching approximately 50-55% of distance from

5 Jouladeh-Roudbar et al.-first record of Paraschistura alta from Iran Fig.2. Lateral view of Paraschistura alta collected from the Zahak River (Sistan basin of Iran). Fig.3. Lateral view of the cleared and stained specimen of Paraschistura alta. pectoral-fin origin to pelvic-fin origin. Pelvic axillary lobe ovoid, fully attached to body or tip of axillary lobe free. Pelvic fin origin below second or third branched dorsal fin ray. Pelvic fin not reaching to anus. Anal fin origin about one eye diameter behind anus. Anal-fin origin is at vertical of middle between dorsal- and caudal fin origins and reaches to caudal fin origin. A shallow dorsal adipose keel on caudal peduncle, without procurrent rays. Margin of dorsal fin concave. Caudal fin forked with pointed lobe. Usually upper lob longer than lower lobe. Dorsal fin with 3 unbranched and 7½ and 8½ branched rays. Anal fin with 3 unbranched and 5½ branched rays. Pectoral fin with 9-10 (usually 9) and pelvic fin with 6-7 (usually 7) branched rays (Fig. 3.). Small, rounded, deeply embedded scales on caudal peduncle and flanks. Lateral line incomplete, reaching to dorsal-fin origin. Mouth small, strongly arched. Lips fleshy, with many deep furrows. A median interruption in lower lip. Upper lip with median incision. There is no Processus dentiformis. Anterior nostril opening at tip of a pointed and flap-like tube. Barbels long. Largest known specimen 112mm SL. Coloration: Body olive-green or beige in life and yellowish in preserved individuals. Body with 9-10 regularly shaped dark bars (7-10 based on Coad (2014)), including 4 predorsal, 1 at dorsal-fin origin, 1 at middle of dorsal-fin base, 1 at posterior margin of dorsal fin-base and 3 (in one specimen 2) bars on caudal peduncle. Bars often dissociate on flank but usually meeting their homologues on back. Bars on flank in front of the dorsal-fin origin sometimes faded or absent in some individuals. A large black spot at base of unbranched and first branched dorsalfin rays. Head olive-green on top and side with cheek pale-olive. Dorsal and caudal fins with sporadic black spots on rays and other fins hyaline. Molecular analysis: We generated COI barcodes for 3 specimens of P. alta collected from Zahak River. The estimation of the phylogenetic relationships based on the COI barcode region places the used sequenced of the members of the genus Paraschistura into 11 groups (Fig. 4), which show between 3.05% and 14.1% K2P sequence divergence in their COI barcode region. Paraschistura alta from Zahak River corresponds a distinct clade, forming a monophyletic 239

6 Iranian Journal of Ichthyology (December 2015), 2(4): Fig.4. Bayesian consensus tree inferred from COI data. Bayesian posterior probabilities followed by Maximum likelihood bootstrap values are listed above the nodes. group with sister group of P. bampurensis + P. hurmozensis (Fig. 4). All analysed species of the genus Paraschistura corresponds to distinct clades (Fig. 4). Table 3 lists the nucleotide substitutions found in the 650 base pairs long mtdna COI barcode region. Table 4 lists the average estimates of the evolutionary divergence between the Paraschistura species. Discussion This study reports the presence of Paraschistura alta for first time from Iranian inland waters, Zahak River (Sistan basin), showing its range extension further to the western part of the Sistan basin. In addition, the 240 current study provided the morphometric and meristic characteristics, COI barcoding of the Iranian population of this species as well as its phylogenetic relationship with other members of the genus Paraschistura from Iran. The Sistan basin consists of several lakes of varied extent i.e. Hamune puzak, Hamun-e Saberi and Hamun-e hirmand and connections straddling the Iran-Afghanistan border. Paraschistura alta has been described and reported only from Helmand River basin (the eastern part of the Sistan basin) in Afghanistan at the Kajaki, north east of Girisk. The Helmand River basin consists about 36800km 2 and originate from Baba Mountain (east of Farakhulum)

7 Jouladeh-Roudbar et al.-first record of Paraschistura alta from Iran Table 3. Nucleotide substitutions found in mtdna COI barcode region some of Paraschistura species in Iran. Nucleotide position relative to Oryzias latipes complete mitochondrial genome (AP004421) P. alta T C T G C T C G T A T A T G A G A T T T A G A T A T A A T A C A G A G T G G A T A G C C C P.abdolii T C A G C C T G A A T A A A A A A T T A A A G C C C G A T G C G A A A C G G G T A G C C T P.aredvii T T T G C T T C T A T G A G A A A T T T G A G C A C G G T A T A A A A T G A A C A A C C C P.bampurensis T C T G C T C G T T C A T G A A A T C T A G A T G C G A T A T A G G A C G G A T A A C C C P.cristata T C T A C T T G T A T A A A G G G T T T A A G C C C A G T A C G A A A C G G A C G G T C C P.hormuzensis T C T G C T C G T T C G T A A A A T T T A A A T G C G A T A T A A A A C A G A T A A C C C P.naumanni T C T A C C T G A A T A A G A A A T T A A A G C C C G A T G C G A A A C A G G T A G C C T P.nielseni C T T G C T T G T A T G A G A A A T T T A A G C A C G G T A T A G A A C G A A T A A C T C P.pasatigris T T T G T T T G T A T A A G A A A C C T A A A C A C G G C A T A A A A C A A A C A A C C C P.susiani T T T A C T T G T A T A A G A A A T T T A A G C A C G A T A T G A A A T G A A C A A C C C P.turcmenica T C T G C T T G T A T A A A A A G T T T A A G C C T A G C A C G G G A C G G A C G G C C C P. alta A C G A C C T C G A T A A A T C A G T C G C A G G A T A C T A T A T A A T G A T G C A C T P.abdolii A C C C T T C C G A T G G A T A G G T T C T A G A A C G T T A G A T G G T G A T G C A C C P.aredvii A C G T C C T T A A T A A A T A G A T T A T G G A T C A C T A T G T A A C A A T G T A T T P.bampurensis A T G A C C C C G A T A A G T A G A T C G C A G A A T A C T A T A C G A C A A T G C A C T P.cristata A C A C T C T C A A T G G G T A G G T T C T A G A A C A A T G G A T G A C A A T G C G C T P.hormuzensis A C G A C C C C G A T A G G T A A G T C G C A G A A T A C C A T A C G A C A A C G C A C T P.naumanni A C C C T T C C G A T G A G T A G G T T C T A G A A C G C T A A A T G A T G A T G C A C T P.nielseni A C G T C C T C G A G A G G T A A G T T G T G G A T C A C T A T A T G A C A A T G T A T T P.pasatigris A C A A C C T C A G T A T A T A A G T C A T A G A G C A C T A T A T G A C A T T G T A C T P.susiani G C G A C C T C G A T A T G T A A A T C A T G G A A C A C T G T A T G A C A T T G T A C T P.turcmenica A C A C T T T C A G T G G G C A A A C T C T A G A A C A A T A G A T G A C A A T G C G C T P. alta G G C T A T T T C A C A C C A A T T C A T C G C C A G T C C C A C G G C T A C C C T A C P.abdolii A G T C A C C C T A C G C T G G C C C G C C A C C A G C T C C A T G A C C A C C C T A C P.aredvii A A C T A T T C C A C A C C A A T T T A C C G T C A A T C C C A C G A C T A T C G T A T P.bampurensis A G C T A T T T C A C A C T A A T T C A T C A T C A G T C T T A C A A C T A C C C T A C P.cristata G G T T A C C T C G C A C T G A C C C G C C G C T A A T C C C G T A A C C A C C C T G C P.hormuzensis G G C T A T T C C A C G C T A A T T C A T C G T C A A C C C C A C A A T T A C T C T A C P.naumanni G A T C A C C C C G C G T T G G C C C G C C A C C A G C T C C A T G A C C A C C C T G C P.nielseni A A T T A T T C C A T A C T A A T T T A C C G C C G A T C C C A C G A C C A C C G T A T P.pasatigris A G C C G T T C C A C G C C G A T T T A C T G C C A A T C C C A C G A C C G C T A T A C P.susiani A G C T A T T C C A C A C C G A T T T A T C A C C A G T C C C G C G A C T A C C G T A T P.turcmenica G G T T A C C T C A C A C T G A C C C G C C G C C A A T C C C A T G A C C A C C C C G C Table 3. Estimates of the average evolutionary divergence between the Iranian Paraschistura species, expressed as number of base substitutions per site. All positions with less than 95 % site coverage were eliminated before analysis, leading to a total of 650 nucleotide positions. No. Species P. alta 2 P. abdolii P. aredvii P. bampurensis P. cristata P. hormuzensis P. naumanni P. nielseni P. pasatigris P. susiani P. turcmenica

8 Iranian Journal of Ichthyology (December 2015), 2(4): and drain south-west for about 1300km (Dupree 1973; Coad 2014) before Sistan (Hamoun) lakes. The previous studies have reported little relationship between the western (Iran) and eastern (Afghanistan) parts of the Sistan basin (Coad 2014). Our results also confirm the absence of 10 species (Garra wanae, Schizothorax esocinus, S. labiatus, S. plagiostomus, Paracobitis boutanensis, P. gazniensis, Paraschistura lindbergi, Triplophysa farwelli, and T. griffithii, T. kullmanni) out of 22 reported species from eastern part of the Sistan basin (Afghanistan) and vice versa absence of Capoeta fusca in eastern part of this basin that presents in Iranian part (Coad 2014; Jouladeh- Roudbar et al. 2015a). The Iranian part of Sistan basin is a network basin and the flooding of the lake could be led to fish movement toward western part of this basin, where the most of lakes lies in Iran. The result of the phylogenetic relationships of the Iranian members of the genus Paraschistura (except P. delvarii) based on COI barcode region revealed that P. alta is distinct clade sister group to P. hurmuzensis (from Makran basin, southeast of Iran) + P. bampurensis (from Jaz-e Murian and Makran basins, southeast of Iran). The nucleotide substitution of studied 650bp mtdna COI barcode region showed 5.92% and 12.91% differences between P. alta with P. bampurensis and P. abdolii, receptivity. The Sistan, Jaz-e Murian and Makran basins are connected and therefore their close relationships can be verified based on close geographical distance. Acknowledgement We would like to thank University of Tehran for financial support. We are pleased to thank H. Gohari Moghadam, A. Bahalkeh, and P. Jalili for helping in fish collection and clearing and staining of the specimens. References Altschul, S.F.; Gish, W.; Miller, W.; Myers, E.W. & Lipman, D.J Basic local alignment search tool. Journal of Molecular Biology 215(3): Azimi, H.; Mousavi-Sabet, H. & Eagderi, S. 2015a. Osteological characteristics of Paraschistura nielseni (Cypriniformes, Nemacheilidae). Iranian Journal of Ichthyology 2(3): Azimi, H.; Mousavi-Sabet, H.; Eagderi, S. & Vatandoust, S. 2015b. Osteological characteristics of Turkmenian stone loach, Paraschistura cristata (Cypriniformes: Nemacheilidae). International Journal of Aquatic Biology 3(5): Coad, B.W Fishes of Afghanistan. Pensoft Publishers, Sofia-Moscow. 393 pp. Coad, B.W Freshwater Fishes of Iran (Available at (accessed on 10 October 2015). Dupree, L Afghanistan in 1983: and still no solution. Asian Survey Eschmeyer, W.N. & Fong, J.D Pisces. In: Zhang, Z.-Q. (Ed.). Animal biodiversity: An outline of higher level classification and survey of taxonomic richness. Zootaxa 3148: Freyhof, J.; Sayyadzadeh, G.; Esmaeili, H.R. & Geiger, M Review of the genus Paraschistura from Iran with description of six new species (Teleostei: Nemacheilidae). Ichthyological Exploration Freshwaters 26(1): Ghasemi, H.; Jouladeh-Roudbar, A.J.; Eagderi, S.; Abbasi, K.; Vatandoust, S. & Esmaeili, H.R. (2015). Ichthyofauna of Urmia basin: Taxonomic diversity, distribution and conservation. Iranian Journal of Ichthyology 2(3): Hashemzadeh-Segherloo, I.; Abdoli, A.; Purahmad, R.; Puria, M. & Golzarianpour, K Genetic barcoding of Capoeta species in Karoon and Tigris tributaries. New Genetics 9(2): (In Farsi) Huelsenbeck, J.P.; Larget, B. & Alfaro, M.E Bayesian phylogenetic model selection using reversible jump Markov chain Monte Carlo. Molecular Biology and Evolution 21(6): Jouladeh-Roudbar, A.; Eagderi, S. & Esmaeili, H.R. 2015a. Fishes of the Dasht-e Kavir basin of Iran: an updated checklist. International Journal of Aquatic Biology 3(4): Jouladeh-Roudbar, A.; Vatandoust, S.; Eagderi, S.; Jafari- Kenari, S. & Mousavi-Sabet, H. 2015b. Freshwater fishes of Iran; an updated checklist. AACL Bioflux

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