Iranian Journal of Fisheries Sciences 20(2) 333-342 2021 DOI: 10.22092/ijfs.2021.123784 Research Article Otolith shape analysis of three mudskipper species of Persian Gulf

Ghanbarifardi M.1*; Zarei R.2

Received: August 2020 Accepted: January 2021

Abstract The variation in otolith shape of three syntopic species of mudskippers Boleophthalmus dussumieri, Periophthalmus waltoni, and Scartelaos tenuis from the tidal flats of Qeshm Island in Persian Gulf was studied based on Elliptic Fourier Analysis. Principal component analysis and Discriminant analysis could separate the specimens of every species with 100% classification success, which proves the variability of otolith among the three fish species. Cluster analysis produced two main clusters, one cluster encompasses only specimens from P. waltoni and the other contains S. tenuis and B. dussumieri. The result of the present study revealed the power of geometric morphometrics in discriminating the three mudskippers. It is also concluded that otolith traits especially its geometric morphometric could be regarded as an invaluable source of information for paleontology, phylogeny, taxonomy and ecology of fishes.

Keywords: Mudskippers, Persian Gulf, Otolith shape, Elliptic Fourier Analysis

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1-Department of Biology, Faculty of Sciences, University of Sistan and Baluchestan, Zahedan, Iran 2-Department of plant sciences, Faculty of biological sciences, Alzahra University, Tehran, Iran *Corresponding author's Email: [email protected]

334 Ghanbarifardi and Zarei, Otolith shape analysis of three mudskipper species of Persian Gulf

Introduction rockfish species (Sebastes spp.) (Tuset Fish otoliths are located in the inner ear et al., 2016). of bony fishes, where they operate as Moreover, using otolith integral pieces of the hearing and morphometric traits has shown equilibrium organs (Popper and Fay, geographical variation between 2011). They consist of aragonite and populations of Aphaniops stoliczkanus develop independently from the skeletal from Southeastern Arabian Peninsula system (Maisey, 1987). Three pairs of (Reichenbacher et al., 2009) and otoliths exist - the saccular otoliths or Southern Iran (Teimori et al., 2012a). sagittae, the utricular otoliths or lapilli, These characteristics have been used on and the lagenar otoliths or asterisci genus Aphanius to differentiate between (Nolf, 1985). In most teleosts, the species and identify new species morphology of sagitta represents a (Teimori et al., 2012b; Gholami et al., meaningful taxonomic tool for species 2014) which revealed its importance in identification (Tuset et al., 2008; Nolf, taxonomic and phylogenetic studies. 2013), although considerable Some studies investigated otolith intraspecific variation can occur, shape analyses in the family Gobiidae especially when connectivity between (Lord et al., 2012; Yu et al., 2014; populations is restricted (Teimori et al., Bănaru et al., 2017; Lombarte et al., 2012a; Gholami et al., 2015; Teimori et 2018; Gut et al., 2020). Wang et al. al., 2018). Furthermore, environmental (2011) used this method for stock factors and ecological or behavioral discrimination of spotted tail goby characteristics can influence otolith Synechogobius ommaturus from the morphology (Torres et al., 2000; Yellow Sea. Five lineages of the Volpedo and Echeverria, 2003; gobionelline-like Gobiidae (sensu

Downloaded from jifro.ir at 3:24 +0330 on Thursday September 30th 2021 Lombarte et al., 2010; Gierl et al., Agorreta et al., 2013) were recognized 2018; Doustdar et al., 2019). using otolith shape analysis; the Elliptical Fourier-shape descriptors outcome revealed that Pomatoschistus have been used to discriminate between and Periophthalmus lineages were species of species-rich genera such as discriminated better than other lineages Sebastes (Stransky and MacLellan, (Gierl et al., 2018). 2005). Otolith shape analysis of fishes Mudskippers (Gobioidei, Gobiidae, has also been found effective to Oxudercinae) reside in intertidal and discriminate stocks of some species supratidal mudflats and mangroves of such as Gadus morhua (Campana and Africa, Indian Ocean, and whole Indo- Casselman, 1993), two anchovy West Pacific region (Murdy,1989; species, silver and blue anchovies Jaafar and Murdy, 2017). These fishes (Karahan et al., 2014), three encompass ten genera, but only four, populations of Lutjanus kasmira namely Boleophthalmus, (Vignon and Morat, 2010), and 42 Periophthalmodon, Periophthalmus and Iranian Journal of Fisheries Sciences 20(2) 2021 335

Scartelaos, spend time on land as part their otolith shape relationships with of their daily life cycle (Murdy 1989, their phylogenetic relations as revealed 2011). In Persian Gulf and Gulf of by molecular trees of the past studies. Oman, three species of mudskippers live in mangroves and tidal flats: Materials and methods Boleophthalmus dussumieri A total of 34 mudskipper specimens (B. Valenciennes, 1837, Periophthalmus dussumieri, 10 specimens; P. waltoni, waltoni Koumans, 1955, and Scartelaos 16; S. tenuis, 8) were captured from a tenuis (Day, 1876) (Polgar et al., 2017). station located in Qeshm Island The aims of this study were A) to (Dokuhak, 26º59'58"N, 56º09'23"E) present and compare otolith shape during June 2017, Qeshm is the largest characteristics of the three mudskippers Iranian Persian Gulf Island (Fig. 1). from Persian Gulf based on Elliptic Fourier analysis, and B) to compare Downloaded from jifro.ir at 3:24 +0330 on Thursday September 30th 2021

Figure 1: Sampling site for the three mudskipper species in Qeshm Island (Dokuhak, 26º59'58"N, 56º09'23"E).

Fish samples were collected using a subsequently put in distilled water for hand-net during low-tide and 4-5 hours and then rinsed several times. transported to the laboratory for further Left otoliths were photographed using a examination. Specimens were identified stereomicroscope and scanning electron using morphological characters microscopy (SEM LEO 1430 VP at the according to Murdy (1989). Otoliths Zoological State Collection Munich, were removed by dissecting the fish Germany and VEGA3 TESCAN at skulls. Otoliths were immersed in 1% Alzahra University, Tehran, Iran). A KOH solution for 3-4 hours to remove representative image of sagitta (termed organic debris, which were otolith in the following) of each

336 Ghanbarifardi and Zarei, Otolith shape analysis of three mudskipper species of Persian Gulf

mudskipper species is shown in Figure Fourier harmonics were made invariant 2. The outline of the otolith was traced to otolith size, rotation and starting using the soft-wares tpsUtil v. 1.38, position of the tracing of the outline tpsDig v. 2.16 and GMTP (Rohlf, 2006, (Ferson et al., 1985; Rohlf and Slice, 2013; Taravati and Darvish, 2010). The 1990). Variation in otolith shape was x- and y- coordinates of 300 harmonics assessed using Elliptical Fourier were obtained from points equally analysis (Ferson et al., 1985). Principal spaced along the otolith's outline using component analysis, Discriminant tpsDig v. 2.16, and the TPS files analysis and Cluster analysis were run opened in PAST 3.20 (Hammer et al., using EFA PC scores with PAST 3.20. 2001). The x- and y- coordinates of

Boleophthalmus dussumieri Scartelaos tenuis Periophthalmus waltoni

Figure 2: SEM photographs of otoliths from the three mudskipper species.

Results scores could recognize every species as Thirty-four SEM pictures of sagittae a separate group, with 100%

Downloaded from jifro.ir at 3:24 +0330 on Thursday September 30th 2021 otoliths of three mudskippers from classification success (Fig. 4). The first Qeshm Island were used to conduct the two canonical variates (CVs) accounted analyses. The outcome of the principal for 87.25% and 12.75% of total component analysis (PCA) based on variance, respectively. CV1 separates EFA PC scores assigned the specimens P. waltoni from B. dussumieri and S. of each species to a separate group (Fig. tenuis. Cluster analysis based on PC 3). The first four principal components scores showed that there were two main (PCs) accounted for 40.9%, 18.8%, clusters, one cluster included of only 9.6%, and 5.7% of total variance, specimens from P. waltoni and the respectively. PC1 separated P. waltoni other cluster contained S. tenuis and B. from B. dussumieri, but with some dussumieri (Figure 5). Cluster analysis overlap. PC2 clearly isolated S. tenuis recognized B. dussumieri and S. tenuis from the other two species. as more similar taxa based on otolith Discriminant analysis based on PC shape analysis (yellow line in Fig. 5). Iranian Journal of Fisheries Sciences ( ) 2021 337

Figure 3: Principal component scores for the otolith shapes of the three mudskipper species, B. dussumieri, P. waltoni and S. tenuis.

Figure 4: Discriminant function scores for the otolith shapes of the three mudskipper species, B. dussumieri, P. waltoni and S. tenuis.

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Figure 5: Cluster analysis for the otolith shapes of the three mudskipper species, B. dussumieri, P. waltoni and S. tenuis.

Discussion and Scartelaos tenuis are identified as In the present study, three mudskipper more similar taxa while Periophthalmus species from three genera are compared waltoni is recognized as a separate using otolith shape analysis (outline clade (Figs. 3 to 5). The body shape of method). Boleophthalmus dussumieri the examined species has been 338 Ghanbarifardi and Zarei, Otolith shape analysis of three mudskipper species of Persian Gulf

evaluated by geometric morphometric examinations (Agorreta et al., 2013; approach (14 landmarks) in a previous Ghanbarifardi et al., 2016; Polgar et al., study (Polgar et al., 2017); the resulting 2017). Therefore, the otolith characters PCA scatter plot is similar to the differentiate mudskippers classification of the otoliths of our phylogenetically better than the other study; Periophthalmus waltoni was osteological characters. The posterior separated from Boleophthalmus vertebral column and the caudal dussumieri and Scartelaos tenuis with skeleton of ten mudskipper species PC1 (89% of variance). were examined and showed isolation of Molecular studies define Periophthalmus and Periophthalmodon Periophthalmus lineage within from other mudskipper genera to some Gobiidae family, which constitutes two extent (Ghanbarifardi et al., 2020a). traditional subfamilies Amblyopinae Teimori et al. (2012b) have concluded and Oxudercinae, including that otolith morphology of Aphanius Periophthalmus, Boleophthalmus, species probably has a higher rate of Scartelaos and some other genera divergence compared to other (Agorreta et al., 2013; Ghanbarifardi et morphological traits. Similarly, it seems al., 2016; Polgar et al., 2017). The mudskippers' otoliths could mentioned studies are in consistent with discriminate mudskippers better than our result and recognize Scartelaos and other morphological characters (Murdy, Boleophthalmus as sister taxa and 1989; Ghanbarifardi et al., 2020a). Periophthalmus as an isolated genus for Otolith characters (the present study), all other genera within Periophthalmus skull bones (Murdy, 1989), the lineage. Otolith morphometry of posterior vertebral column and the mudskippers from Persian Gulf were caudal skeleton of mudskippers

Downloaded from jifro.ir at 3:24 +0330 on Thursday September 30th 2021 compared and it is documented that (Ghanbarifardi et al., 2020a) have good Otolith morphometry were more clearly fossilization potential and could separated P. waltoni from B. therefore facilitate recognition of fossil dussumieri, than S. tenuis from either P. species of mudskippers, which are waltoni or B. dussumieri (Ghanbarifardi currently unknown. Moreover, these et al., 2020b). Murdy (1989) used 39 examinations proved that otolith shape morphological (mostly osteological) analysis along with body shape analysis apomorphies to conduct a cladistic could be regarded as informative analysis on Oxudercines. That study phylogenetic traits that are useful for puts Periophthalmus, taxonomic and systematic surveys. Periophthalmodon, Boleophthalmus, Scartelaos in one clade, isolated from Acknowledgments other genera of Oxudercinae, which is The authors are really thankful to Dr. in contradiction with the result of the Bettina Reichenbacher and Dr. Zeinab present study and molecular Gholami for their help during lab work Iranian Journal of Fisheries Sciences 20(2) 2021 339

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