Introduction
Estuaries and protected inshore habitats are well known for their ecosystem services to the aquatic biota, especially in that they provide suitable habitats for early life stages. The nursery function of these coastal habitats to a wide range of marine fish species is well-documented (Beck et al., Reference Beck, Heck, Able, Childers, Eggleston, Gillanders, Halpern, Hays, Hoshino, Minello and Orth2001; McLusky and Elliott, Reference McLusky and Elliott2004; Dahlgren et al., Reference Dahlgren, Kellison, Adams, Gillanders, Kendall, Layman, Ley, Nagelkerken and Serafy2006; Igulu et al., Reference Igulu, Nagelkerken, Dorenbosch, Grol, Harborne, Kimirei, Mumby, Olds and Mgaya2014; Seitz et al., Reference Seitz, Wennhage, Bergström, Lipcius and Ysebaert2014). Fish recruitment into estuaries is known to be influenced by environmental cues and behavioural responses of their larvae (Boehlert and Mundy, Reference Boehlert and Mundy1988). However, larvae and juveniles face significant threats from large-scale predation and starvation in inshore waters, which often drives them to seek refuge in estuarine habitats (Kapsenberg and Cyronak, Reference Kapsenberg and Cyronak2019). Estuarine environments offer relatively lower predation risks and more abundant food resources, making them critical for the survival and growth of these early life stages (Wasserman and Strydom, Reference Wasserman and Strydom2011; Lima et al., Reference Lima, Barletta and Costa2015; Strydom, Reference Strydom2015; Ferreira et al., Reference Ferreira, Barletta, Lima, Dantas, Justino and Costa2016). Furthermore, larval congregation and survival in estuarine habitats are also influenced by seasonal variation in environmental parameters like pH, temperature, turbidity, and salinity (Dando, Reference Dando1984; Takahashi, Reference Takahashi2005; Rodrigues et al., Reference Rodrigues, Almeida, Silva, Cunha, Antunes, Freitas and Ramos2019; Molina et al., Reference Molina, Duque and Cogua2020). The fluctuations in biotic and abiotic properties of the ecosystem in which they live, are primarily responsible for larval dynamic growth. Hydrological characteristics, especially temperature, largely influence migration of marine nektonic groups into estuaries. Estuaries usually retain warm temperature that increases metabolism and feeding success and thereby increases larval growth. Further, it also increases swimming capability. It is also known that larvae and juveniles of many marine fish species prefer estuarine salinity for improved growth (Boeuf and Payan, Reference Boeuf and Payan2001).
Diverse habitat utilization during ontogeny is believed to benefit organisms at distinct life stages (Sutherland, Reference Sutherland1996; Stephens and Krebs, Reference Stephens and Krebs2019). Almost all bony reef fish species have a pelagic early life history stage that commences with a weak planktonic stage and completes with a nektonic stage. The nektonic stage has effective swimming capability that helps their spatial dispersal. Pelagic stages of coral reef fishes can sometimes last for extended periods, which allows larvae to better identify suitable settlement habitats (Ogden and Quinn, Reference Ogden and Quinn1984; McCormick, Reference McCormick1999; Leis and McCormick, Reference Leis and McCormick2002). However, despite the importance of this long pelagic phase, there is still limited information on the larval biology of tropical coral reef fishes, largely due to challenges in larval taxonomy.
Among the fishes primarily associated with coral reefs and deep lagoons, family Acanthuridae, comprises 85 species belonging to six genera popularly known as ‘surgeon fishes’ (Kuiter and Tonozuka, Reference Kuiter and Tonozuka2001; Fricke et al., Reference Fricke, Eschmeyer and van der Laan2022). They constitute the largest tropical inshore marine fish groups and have ecological importance in the dynamics of reefs, but little is known about their biology (Randall, Reference Randall1961). The genus Acanthurus alone comprises 39 species (Devi and Rao, Reference Devi and Rao2003). Acanthurids are known for their vagrant nature (Feary et al., Reference Feary, Pratchett, Emslie, Fowler, Figueira, Luiz, Nakamura and Booth2014) and many of them have been reported from very wide geographic ranges (Cambra et al., Reference Cambra, Cubero, Chinchilla and Azofeifa-Solano2018). Most species in this family begin their life cycle with a pelagic larval stage, followed by an extended post-larval ‘acronurus’ stage. During this phase, they settle in shallow coastal environments, such as bays and lagoons, before metamorphosing to juveniles inhabiting deeper areas. Acanthurus species, like Acanthurus xanthopterus, are ecologically adaptable, thriving in both sheltered zones and exposed reef areas. After settling, they start feeding on benthic algae, a crucial part of their herbivorous diet. Their gizzard-like stomach helps in digesting food mixed with sand and other debris. In some species, the larval stage can be prolonged by delaying metamorphosis, aiding dispersal across regions. As adults, these surgeonfish form a significant part of the algivorous biomass, contributing to coral reef resilience. Acronurus larvae of some acanthurids are reported to have significantly longer pelagic larval durations (Robertson, et al., Reference Robertson, Grove and McCosker2004). Larval forms of acanthurids in the Indo-Pacific region have been described by Leis and Carson-Ewart (Reference Leis and Carson-Ewart2000). Yellowfin surgeon fish, Acanthurus xanthopterus is the largest of the genus Acanthurus, widely distributed across the Indo-Pacific region (Randall, Reference Randall2001).
The present paper is the first report of the occurrence of A. xanthopterus larvae in a tropical estuary, Vembanad Lake, South west coast of India.
Materials and methods
Study area and sample collection
The Vembanad-kol wetland system, which encompasses mangroves, mudflats, swamps, and marshes, is the largest of its kind on India's south west coast and is home to a diverse range of aquatic flora and fauna. It is a tropical positive microtidal estuary system recognized as a Ramsar site that stretches from 9°28′ to 10°10′ N and 76°13′ to 76°31′ E. The lake has a permanent opening to the Arabian Sea at Cochin barmouth, through which seawater enters the estuary during flood tide and flows back during ebb tide. Monsoon (June–September), post-monsoon (October–January) and pre-monsoon (February–May) are the three different seasons on India's west coast. In contrast to post-monsoon and pre-monsoon seasons, the lake receives increased river discharges during monsoon (Jacob et al., Reference Jacob, Revichandran and Naveen Kumar2013).
Ichthyoplankton specimens were collected using a conical plankton net (300 μm mesh size) with a 60 cm mouth diameter. The net was equipped with a water flowmeter (General Oceanics) to quantify the volume of water filtered during each sampling event. Upon collection, fish larvae were transferred to labelled containers and photographed. Specimens were preserved in absolute alcohol for further analysis. Morphological identification was conducted using standard literature (Leis and Carson-Ewart, Reference Leis and Carson-Ewart2000; Miller and Kendall, Reference Miller and Kendall2009), with larvae classified up to the family level. Voucher specimens were stored in the laboratory of the School of Industrial Fisheries, Cochin University of Science and Technology, Kochi, Kerala, India.
The sorting and examination of larvae were carried out using a stereo microscope (Leica, EZ4D). Fish larvae were initially categorized as morphologically similar taxonomic units (MSTUs) to facilitate identification and analysis. The morphometric and meristic characters of 20 acanthurid larval specimens were recorded, following the methods outlined by Leis and Carson-Ewart (Reference Leis and Carson-Ewart2000). The colour descriptions were made shortly after preservation. Illustrations of the larvae were generated using Inkscape graphic software (version 1.1.1; Tran et al., Reference Tran, Trinh-Dang and Brancelj2021). Statistical analyses were performed using R software (Figure 1).
Map showing Vembanad lake's downstream region and sampling stations (S1–S5).
DNA barcoding
Acanthurid larval samples were also identified using DNA barcoding after morphological examinations of a total of 20 specimens, of which two specimens (randomly selected on a 1:10 ratio) were selected (Jose et al., Reference Jose, Mahadevan, Rozario, Pradeep, Maitra and Shamsudheen2020). Total genomic DNA was isolated from tissues of randomly selected fish larvae using spin column protocol (proposed in Qiagen DNeasy Blood and Tissue Kit). Quality and quantity of DNA isolates were estimated using spectrophotometer (Nanodrop 2000) at a wavelength range of 260/280 nm. DNA isolates exhibiting optical density (OD) value between 1.7 and 2.0 were selected for thermal amplification in a gradient PCR.
For PCR amplification, a reaction mix of 50 μl was prepared using 25 μl Sigma Aldrich ReadyMix™ Taq PCR Reaction Mix with MgCl2, 8 μl template (isolated total genomic DNA), 1 μl of forward and reverse primers (0.01 mM) and 15 μl demineralized water. Partial mitochondrial COI gene sequences from total genomic DNA was amplified using primer pair Fish F1 (5′-TCAACCAACCACAAAGACATTGGCAC-3′) and Fish R1 (5′-TAGACTTCTGGGTGGCCAAAGAATCA-3′) (Ward et al., Reference Ward, Zemlak, Innes, Last and Hebert2005). Amplification was performed in a Corbett gradient thermal cycler and a thermal profile was selected modified from Lakra et al. (Reference Lakra, Verma, Goswami, Lal, Mohindra, Punia, Gopalakrishnan, Singh, Ward and Hebert2011) as follows. Initial denaturation of 95°C for 2 min (one cycle) followed by 35 cycles of denaturation for 40 s at 94°C, annealing for 40 s at 45–55°C, extension for 70 s at 72°C and a single cycle of final extension at 72°C (for 10 min). PCR amplicons, after electrophoresis were visualized on 1.2% agarose gel and amplicons with intense bands after visualization and documentation were outsourced for purification and sequencing.
Nucleotide sequence analyses
Manual editing was done on chromatogram of COI sequences using BioEdit 7.0.9 (Hall, Reference Hall1999) and evenly spaced peak regions without baselines were selected. Initial screening of nucleotide sequences was done using Basic Local Alignment Search Tool (BLASTn) for identifying homologous sequences in public database like NCBI (Deepak and Harikrishnan, Reference Deepak and Harikrishnan2017). COI sequences were aligned and compiled using Clustal X (Thompson et al., Reference Thompson, Gibson, Plewniak, Jeanmougin and Higgins1997) and phylogram based on neighbour joining (NJ) and minimum evolution (ME) analyses with 1000 bootstraps were generated using MEGA X (Kumar et al., Reference Kumar, Stecher, Li, Knyaz and Tamura2018). In order to confirm the findings of phylogram, pair-wise sequence distance data based on Kimura 2 Parameter model was generated using the same software.
Results
In the present study, fish larvae belonging to 18 families could be collected from five stations in downstream part of Vembanad lake. The percentage abundance of these larval forms is depicted in Figure 2 and it could be noted that larvae belonging to families Ambassidae (51.7%), Mugilidae (17.5%), Acanthuridae (10.0%), Scatophagidae (5.87%), Aplocheilidae (5.07%) represented the predominant forms. The acanthurid larvae were characterized by their deep orbicular bodies and folding spine. These larvae were collected consistently from all five stations, however, no significant difference was detected in their occurrence with respect to stations and year (P > 0.05).
Percentage composition of larval fish families in downstream part of Vembanad Lake.
Larval morphology
Body is orbicular, deep (BD 59% BL) and laterally compressed, scaleless with narrow vertical ridges which later develop into scales. Head moderate in size (HL 28% BL). Eyes large (ED 37% HL) and rounded. Mouth very small (1.1 mm width), terminal, does not reach eyes and has serrated small teeth with denticulation (Figure 5A). 14 teeth present in upper jaw, each with 5 or 6 denticulations, with a wide gap (0.2 mm) at the symphysis of the maxilla. Lower jaw has 12 teeth with 5 or 6 denticulations each without a gap at the symphysis of the mandible. Lateral line complete. Reduced head serrations present on premaxilla, nasal, supra ocular, and supra occipital area (Figure 5B). Serrations also present on lower jaw (dentary and angular) and along the throat. A distinct small folding spine (0.3 mm in length) on both sides of caudal peduncle. First spine short while second spine being long, stout and serrated in dorsal and anal. Acronurus' spines have T-shaped cross section. Body is lightly pigmented, generally transparent except for gut and brain region. Gut silvery and brain region golden yellow. Distal part of pectoral fin is pale yellow. Melanophores are concentrated on the posterior part of the caudal peduncle forming a dark band. Melanophores are also present on the margins of the body. Details of meristics and morphometrics of the larvae are given in Table 1.
Micrograph of A. xanthopterus.
Diagrammatic representation of A. xanthopterus.
(A) Denticulation of teeth in A. xanthopterus; (B) serrations present on the snout and frontal region of A. xanthopterus.
Details of meristic and morphometric characters of A. xanthopterus (n = 20)
BL, body length; HL, head length; PAL, pre-anal length; BD, body depth; ED, eye diameter.
DNA barcoding, phylogram and genetic distance data
COI sequences were successfully generated for larval samples of Acanthurus sp. collected from downstream part of Vembanad lake using the mentioned primer pair and thermal profile. Homology and speciation of these nucleotide sequences were evaluated using BLASTN annotations of National Centre for Biotechnology Information (NCBI) (Deepak and Harikrishnan, Reference Deepak and Harikrishnan2017) which showed more than 97% identity score towards homologous sequences of Acanthurus xanthopterus in public database. Generated COI sequences were 627 base pairs in length and these sequences were submitted in National Center for Biotechnology Information (NCBI) database with accession numbers MW714349 and MW714395.
Mitochondrial DNA COI sequences of A. xanthopterus were obtained as a single haplotype, devoid of insertions, deletions, and stop codons. In order to perform molecular analysis, additional homologous mtCOI sequence of congeners of genus Acanthurus (detailed in Table 2) and Parastromateus niger (with accession number MF737196; Outgroup) from NCBI database were acquired. Phylogram (Figure 6) and genetic distance data (Supplementary file) were generated using these sequences.
Details regarding COI sequences acquired from NCBI for molecular analyses
Phylogram based on 1000 bootstrap replications and Kimura-2-parameter for selected species. Bootstrap values less than 70 were omitted from the phylogram.
The phylogram demonstrated genetic congruency among the Acanthurus species, with all sequences clustering within a single major monophyletic clade. As expected, the outgroup Parastromateus niger exhibited the greatest genetic diversity. Within the monophyletic clade, Acanthurus xanthopterus was the first species to appear, represented by both generated and acquired sequences, and clustered within a major clade with strong bootstrap support (100%). The closest related species was A. albimento, which also had two sequence representations, supported by a bootstrap value of 100%. The genetic relationship between A. xanthopterus and A. albimento was further confirmed by a high bootstrap value (98%) linking their clades. The next closest species were A. nigricauda and A. olivaceus, though their lower bootstrap values were insufficient for strong support. A second major clade included A. guttatus, A. achilles, and A. nigricans, but their interspecific relationships lacked significant bootstrap support. Similarly, A. pyroferus, A. nigrofuscus, and A. lineatus were grouped together, though again without substantial bootstrap support. Finally, A. triostegus and A. thompsoni formed a separate clade, also with low bootstrap support. Despite high interspecific divergence among the Acanthurus species, intraspecific divergence was very low, as indicated by the consistent 100% bootstrap support for each species. To further validate the phylogram, a distance matrix (Supplementary file) was developed and analysed.
Genetic distance data also confirmed the persistence of lower intraspecific divergence with the selected species. For A. xanthopterus, there was no genetic distance within generated sequences as well as the acquired one. But the level of genetic difference between A. xanthopterus and other congeners (interspecific divergence) was very high (refer Supplementary material).
Water quality parameters
The mean seasonal surface water quality parameters recorded in five sampling stations are given in Figure 7A–D. It was noted that mean values of salinity (F = 47.99, P < 0.01), surface water temperature (F = 132.9, P < 0.01), pH (F = 4.78, P < 0.05) and dissolved oxygen (F = 13.29, P < 0.01) varied significantly between seasons in all stations. Highest mean salinity was recorded in pre monsoon season (28.11 ± 4.56 psu) while a mean salinity of 14.85 ± 3.63 psu was recorded during post monsoon, followed by 5.2 ± 2.27 psu in monsoon. The highest surface water temperature was recorded in the pre monsoon season (29.95 ± 0.11°C) in contrast to post monsoon (28.56 ± 0.13°C) and monsoon (28.54 ± 0.21°C) seasons. The pH of surface water varied significantly between 7.68 ± 0.11 (pre monsoon) and 7.48 ± 0.11 (post monsoon). Highest mean dissolved oxygen was measured in post monsoon season (5.17 ± 0.26 mg l−1) followed by monsoon (4.85 ± 0.34 mg l−1) and pre monsoon (4.22 ± 0.29 mg l−1).
Mean ± SD of (A) salinity, (B) temperature, (C) pH and (D) dissolved oxygen in surface water at sampling locations during three seasons (Abbreviations: POM- post monsoon, PRM – premonsoon and MON – monsoon).
Temporal larval abundances
Seasonal variations in mean abundances of acanthurid larvae and other fish larvae in the downstream part of the lake are depicted in Figure 8. Highest fish larval abundance could be recorded in monsoon season (3719 ± 284 numbers/100 m3). However, no acanthurid larvae could be recorded during this season. During post monsoon season, a mean larval abundance of 354 ± 180 numbers/100 m3 was recorded in acanthurid larvae whereas the mean abundance for other fish larvae was 1099 ± 206 numbers/100 m3. Acanthurid larvae could also be encountered in premonsoon period (mean 217 ± 120 numbers/100 m3) when the other fish larvae recorded a mean of 956 ± 142 numbers/100 m3. It could also be noted that monsoon season period was also marked by low species diversity, in contrast to high diversity values recorded in post monsoon and pre monsoon periods.
Seasonal variations (mean ± SE) in the abundance of Acanthurid larvae (AL) and other fish larvae (OL) in the Vembanad lake's downstream region.
Discussion
Reef fishes have a bipartite life cycle, consisting of a pelagic dispersive larval stage and demersal juvenile/adult stages. The pelagic stage is primarily responsible for the distribution of adult fish (Leis, Reference Leis1991; Bonhomme and Planes, Reference Bonhomme and Planes2000). Several reef fishes have different habitat preferences during their ontogenetic stages; larvae and juveniles prefer shallow bay environments such as mangroves and seagrasses, whereas adults return to reefs (Nagelkerken et al., Reference Nagelkerken, Dorenbosch, Verberk, De La Morinière and Van Der Velde2000a, Reference Nagelkerken, Dorenbosch, Verberk, De La Morinière and van Der Velde2000b, Reference Nagelkerken, Kleijnen, Klop, Van Den Brand, de La Moriniere and Van der Velde2001, Reference Nagelkerken, Roberts, Van Der Velde, Dorenbosch, Van Riel, De La Moriniere and Nienhuis2002; De La Moriniere et al., Reference De La Moriniere, Pollux, Nagelkerken and Van der Velde2002). For many tropical reef fishes, active habitat selection and behavioural characteristics have been attributed to be the major drivers of larval settlement (Leis, Reference Leis2010).
The morphological traits observed in the larval samples from the present study align closely with the descriptions of acanthurid larvae provided by Leis and Carson-Ewart (Reference Leis and Carson-Ewart2000), with one notable exception: the body shape. According to their findings, the kite-shaped body profile is a distinguishing feature that sets acanthurid larvae apart from leognathid larvae. However, in our study, we found that the body shape of Acanthurus xanthopterus larvae is more ovoid rather than kite-shaped, which contrasts with the established descriptions. This discrepancy highlights the variability that can occur within larval morphology and suggests that additional research may be necessary to further understand these differences. Therefore, DNA barcoding, a viable tool used for avoiding taxonomic ambiguity (Panprommin et al., Reference Panprommin, Soontornprasit, Tuncharoen and Iamchuen2020) was performed to generate COI sequences from the collected fish larvae using aforementioned primer pair and thermal regime. Panprommin et al. (Reference Panprommin, Soontornprasit, Tuncharoen and Iamchuen2020) performed DNA barcoding of larval fish species collected from tributaries of Ing River and identified 35 species belonging to 31 genera contained within 19 families and nine orders. In the present study, molecular sequences developed were compared with homologous COI sequences of species, already reported in public databases (NCBI) and BLAST search confirmed the speciation of collected larvae as A. xanthopterus. Additional analyses using acquired COI sequences also supported the preliminary results, being evident from higher bootstrap support in phylogram and lower intraspecific genetic divergence. This study is the first authentic report of morphological and molecular characterization of A. xanthopterus in Vembanad Lake.
Adult stages of reef fishes have previously been reported to enter estuaries from marine habitats all over the world (de Paiva et al., Reference de Paiva, Lima, de Souza and Araujo2009) and A. xanthopterus has been reported along India's west coast (Kurup and Samuel, Reference Kurup, Samuel, Ravindran, Nair, Perigreen, Pillai, Panicker and Thomas1985; Prakash et al., Reference Prakash, Balamurugan, Kumar and Balasubramanian2012). Kurup and Samuel (Reference Kurup, Samuel, Ravindran, Nair, Perigreen, Pillai, Panicker and Thomas1985) had previously reported the rare occurrence of adult A. matoides in Vembanad lake, however, no records on the early stages of Acanthurus have so far been reported from the lake. Recent studies on fishery production in the lake (Asha et al., Reference Asha, Suson, Retina and Nandan2014; Roshni et al., Reference Roshni, Renjithkumar, Raghavan and Ranjeet2021) also did not reveal the occurrence of acanthurids in this lake. However, rare occurrences of adult A. xanthopterus have been reported from rocky reefs along southern Kerala coast (Baiju et al., Reference Baiju, Prabhakaran, Benno Pereira and Jayaprakas2016). Juveniles of this reef fish usually prefer shallow, protected, turbid inshore waters while adults prefer deeper areas of protected bays and lagoons (Ajith Kumar et al., Reference Ajith Kumar, Vinoth, Prakash and Balasubramanian2012). Recruitment of post-larval stages of acanthurids into protected inshore locations for prolong metamorphosis has also been documented (Devi and Rao, Reference Devi and Rao2003).
In the present study, A. xanthopterus larvae were collected from the downstream regions of Vembanad lake, close to its opening in to the Arabian Sea. Further, these larval forms were encountered in the lake only in high saline months (from December to April) during when salinity exceeded 20 psu. Transient fish larvae are known to utilize estuaries with oyster shell beds as the water turbulence and flow disturbance over oyster shellbeds favoured their settlement (Breitburg et al., Reference Breitburg, Palmer and Loher1995; Lehnert and Allen, Reference Lehnert and Allen2002). It may be pointed out that oyster shell beds have also been reported in the downstream regions of Vembanad lake (Unnikrishnan and Balakrishnan, Reference Unnikrishnan Nair and Balakrishnan Nair1984; Kripa et al., Reference Kripa, Jenni, Chinnadurai, Khambadkar, Prema and Mohamed2014) and these shell rubble areas might be providing congenial habitats for these acronurus larvae. Cross-habitat dispersal during ontogenetic development stages for ensuring potential growth and reducing predation risks has previously been documented and such ontogenetic niche shifts are known to explain seasonal migrations in many fish species (Grol et al., Reference Grol, Rypel and Nagelkerken2014; Fokkema et al., Reference Fokkema, van der Jeugd, Lameris, Dokter, Ebbinge, de Roos, Nolet, Piersma and Olff2020). A. bahianus is known to spread across the Amazon–Orinoco barrier owing to environmental changes (Castellanos-Gell et al., Reference Castellanos-Gell, Robainas-Barcia, Casane, Chevalier-Monteagudo, Pina-Amargos and García-Machado2012).
Although the genus Acanthurus is commonly found in coral reefs and associated habitats, its late pelagic larval stages have been reported in open water habitats (Leis et al., Reference Leis, Sweatman and Reader1996). The reef fish larvae swim actively over long distances and are highly selective in identifying their settlement sites. Acanthurids have a long pelagic larval period and have the capacity for self-recruitment (Leis and McCormick, Reference Leis and McCormick2002; Fisher et al., Reference Fisher, Leis, Clark and Wilson2005). They are more abundant in near-shore areas and enter very shallow waters to develop into juveniles (Reintjes and King, Reference Reintjes and King1953; Randall, Reference Randall1961). A. xanthopterus has been found in the Lakshadweep coral reefs of the Arabian Sea (Ajith Kumar et al., Reference Ajith Kumar, Vinoth, Prakash and Balasubramanian2012), which is about 225–450 km off the coast of Kerala (Muley et al., Reference Muley, Venkataraman, Alfred and Wafar2002). Lakshadweep sea is known for yellow fin tuna resources which largely predate on reef fish larvae (Randall, Reference Randall1961) and therefore, these larvae may seek congenial environments for survival and for escaping predation.
Several studies have found that fish larvae migrate to nursery areas for enhanced survival rather than for high food resources (Grol et al., Reference Grol, Dorenbosch, Kokkelmans and Nagelkerken2008, Reference Grol, Nagelkerken, Rypel and Layman2011, Reference Grol, Rypel and Nagelkerken2014; Kimirei et al., Reference Kimirei, Nagelkerken, Trommelen, Blankers, Van Hoytema, Hoeijmakers, Huijbers, Mgaya and Rypel2013). It may be pointed out that the guts of larval samples collected in the present study were empty. Low feeding rates and empty guts in acronurus larvae have previously been reported (Randall, Reference Randall1961; McCormick, Reference McCormick1999). Therefore, it could be inferred that the occurrences of acanthurid larvae in the downstream part of estuary as reported in the present study were not accidental and could be attributed to their effective utilization of these habitats as nursery areas favouring their survival and settlement.
Supplementary Material
The supplementary material for this article can be found at https://doi.org/10.1017/S0025315425000165
Author Contributions
Conceptualization: N. B., H. M. Methodology: N. B., D. J., S. K. Sample collection: N. B., R. R. M. Data Curation: N.B., S. K. Writing – Original Draft: N. B., D. J., S. K. Writing – Review and Editing: N. B., H. M. Visualization: N. B., H. M., S. K. All authors read and approved the final version of the manuscript.
Financial Support
The authors acknowledge KSCSTE for financial support and thank the Director, School of Industrial Fisheries, Cochin University of Science and Technology, for providing the necessary research facilities.
Competing interest
The authors declare no conflicts of interest relevant to this study.
Ethical Standards
This study was conducted following all applicable ethical guidelines and standards. No ethical approval was required.
Data Availability
The authors confirm that the data supporting the findings of this study are included within the article. Additional information can be obtained from the corresponding author upon reasonable request.
