Bamboo Shark Research Articles

Three new species of 'tetraphyllidean' cestodes from an undescribed bamboo shark (Orectolobiformes: Hemiscylliidae) in Sri Lanka.

Examination of seven specimens of an undescribed species of bamboo shark (Orectolobiformes: Hemiscylliidae), currently referred to as Chiloscyllium sp. 1, from Sri Lanka yielded three new species of 'tetraphyllidean' cestodes, one each in the genera Carpobothrium Shipley et Hornell, 1906, Spiniloculus Southwell, 1925, and Yorkeria Southwell, 1927. Carpobothrium marjorieae Caira, Otto et Jensen sp. n. differs from its three valid congeners in total length, number of testes, and size of the apical sucker of its anterior bothridial flap. Like Carpobothrium eleanorae Koontz et Caira, 2016 it possesses spherical eggs with highly elongate bipolar filaments, but its eggs are conspicuously larger than those of the latter species. Spiniloculus akshayi Caira, Otto et Jensen sp. n. can be distinguished from its five congeners in total length and number of proglottids. Yorkeria sachiniae Caira, Otto et Jensen sp. n. differs from its 16 congeners in number of proglottids and testes, total length, terminal proglottid, cirrus sac, and cephalic peduncle length, hook size, genital pore position, and bothridial shape. The eggs of Y. sachiniae Caira, Otto et Jensen sp. n. are spindle-shaped and bear a single long, polar filament. All three species were found to exhibit microthrix patterns on their scolices like those of the majority of their congeners that have been examined to date with scanning electron microscopy. Microtriches on the distal surfaces of the bothridial pouches of a species of Carpobothrium were characterised for the first time; C. marjorieae Caira, Otto et Jensen sp. n. was found to possess a modified form of gladiate spinithrix on this surface. Among the seven members of the genus, this is the first species of Chiloscyllium Müller et Henle that has been found to host representatives of all three of these 'tetraphyllidean' genera. However, unlike most of its congeners, a representative of the non-hooked genus Scyphophyllidium Woodland, 1927 was not found infecting Chiloscyllium sp. 1.

Developmental hourglass and heterochronic shifts in fin and limb development.

How genetic changes are linked to morphological novelties and developmental constraints remains elusive. Here, we investigate genetic apparatuses that distinguish fish fins from tetrapod limbs by analyzing transcriptomes and open-chromatin regions (OCRs). Specifically, we compared mouse forelimb buds with the pectoral fin buds of an elasmobranch, the brown-banded bamboo shark (Chiloscyllium punctatum). A transcriptomic comparison with an accurate orthology map revealed both a mass heterochrony and hourglass-shaped conservation of gene expression between fins and limbs. Furthermore, open-chromatin analysis suggested that access to conserved regulatory sequences is transiently increased during mid-stage limb development. During this stage, stage-specific and tissue-specific OCRs were also enriched. Together, early and late stages of fin/limb development are more permissive to mutations than middle stages, which may have contributed to major morphological changes during the fin-to-limb evolution. We hypothesize that the middle stages are constrained by regulatory complexity that results from dynamic and tissue-specific transcriptional controls.

Reproductive biology of a bamboo shark as a framework for better fisheries management

The brown-banded bamboo shark, Chiloscyllium punctatum, is one of the most common sharks caught in coastal fisheries across Southeast Asia; however, its small size and low economic value typically exclude it from the attention of fisheries managers. As part of an assessment of a potential role for this shark in improving the sustainability of coastal shark fisheries in Indonesia, we investigated its reproductive biology from waters around the Seribu Islands on the basis of monthly sampling from a fish landing in Tanjung Pasir, Banten. Females have fully functional uteri, but asymmetric ovaries, with only the right ovary being functional and the left ovary being atrophied. The species is reproductively active throughout the year, with the optimum period for males to copulate being from June to August, and the optimum period for egg-laying in females from July to January on the basis of their physiological condition. Females mature at a larger size than do males, 704- and 653-mm total length respectively. To allow a continuous recruitment flow to the population, and to facilitate compliance and monitoring, we recommend that fishers release all brown-banded bamboo sharks greater than 700mm in total length.

Sexual development and maturity of the male brown banded bamboo shark,

The present study aimed to assess the reproductive biology of male brown banded bamboo shark (Chiloscyllium punctatum). Morphological and histological changes in the reproductive tract and changes in gonad steroid testosterone were assessed in 110 male C. punctatum collected from Bahrakan Creek between January 2018 and March 2019. The reproductive tract in male C. punctatum consisted of a pair of testes, efferent ducts, epididymidis, ductus deferens, seminal vesicles, Leydig gland and claspers. Four sexual development stages were determined: (1) immature, no calcified claspers (2) maturing, partially calcified claspers; (3) active and resting adults. Both adult resting and active stages had calcified claspers. In this study, immature male C. punctatum without calcified claspers were found from August to October, maturing C. punctatum with partially calcified claspers were found from November to January and adult C. punctatum caught from February to April were active with calcified claspers, whereas adult fish collected from May to July and during the rest of the year (until mid-January in the following year) were in the resting stage with inactive testes. The testis had a lobular structure and the spermatocysts were arranged diametrically (i.e. along the cross-sectional width of the testis) in each lobule. Plasma testosterone concentrations were associated with histomorphological changes in the reproductive tract. Testosterone concentrations peaked in March in active males and then decreased significantly in resting males. Overall, the variations in the reproductive organs during different sexual maturity stages were consistent with their roles in the production, movement and release of spermatozoa.

Identification of circRNAs in the Liver of Whitespotted Bamboo Shark (Chiloscyllium plagiosum).

Whitespotted bamboo shark (Chiloscyllium plagiosum), a member of the cartilaginous fish family, has an extremely large liver and demonstrates a strong regeneration ability and immune regulation. Circular RNAs (circRNAs) is an important class of non-coding RNAs. Increasing evidences suggest that circRNAs are a kind of potential regulators. Recently, researchers have isolated and identified different circRNAs from various species, while few reports were on the circRNAs of C. plagiosum. In this study, we have identified a total of 4,558 circRNAs in the liver of C. plagiosum. This finding suggests that circRNAs are not evenly distributed in the chromosomes and follow the GT-AG rule during cyclization. Alternative back-splicing might exist in shark circRNAs as shown by the authenticity identification of predicted circRNAs. The binding strength of circRNAs (<2,000 bp) and the detected miRNAs in shark liver were simultaneously analyzed to construct an mRNA–miRNA–circRNA network for the Glutathione S-transferase P1 gene, and the circRNA authenticity was simultaneously verified. Our data provide not only novel insights into the rich existence of circRNAs in marine animals, but also a basis for characterizing functions of identified circRNAs in the liver homeostasis of C. plagiosum.

Multimodal Imaging and Analysis of the Neuroanatomical Organization of the Primary Olfactory Inputs in the Brownbanded Bamboo Shark, Chiloscyllium punctatum.

There is currently a limited understanding of the morphological and functional organization of the olfactory system in cartilaginous fishes, particularly when compared to bony fishes and terrestrial vertebrates. In this fish group, there is a clear paucity of information on the characterization, density, and distribution of olfactory receptor neurons (ORNs) within the sensory olfactory epithelium lining the paired olfactory rosettes, and their functional implications with respect to the hydrodynamics of incurrent water flow into the nares. This imaging study examines the brownbanded bamboo shark Chiloscyllium punctatum (Elasmobranchii) and combines immunohistochemical labeling using antisera raised against five G-protein α-subunits (Gαs/olf, Gαq/11/14, Gαi–1/2/3, Gαi–3, Gαo) with light and electron microscopy, to characterize the morphological ORN types present. Three main ORNs (“long”, “microvillous” and “crypt-like”) are confirmed and up to three additional microvilli-bearing types are also described; “Kappe-like” (potential or homologous “Kappe” as in teleosts), “pear-shaped” and “teardrop-shaped” cells. These morphotypes will need to be confirmed molecularly in the future. Using X-ray diffusible iodine-based contrast-enhanced computed tomography (diceCT), high-resolution scans of the olfactory rosettes, olfactory bulbs (OBs), peduncles, and telencephalon reveal a lateral segregation of primary olfactory inputs within the OBs, with distinct medial and lateral clusters of glomeruli, suggesting a potential somatotopic organization. However, most ORN morphotypes are found to be ubiquitously distributed within the medial and lateral regions of the olfactory rosette, with at least three microvilli-bearing ORNs labeled with anti-Gαo found in significantly higher densities in lateral lamellae [in lateral lamellae] and on the anterior portion of lamellae (facing the olfactory cavity). These microvilli-bearing ORN morphotypes (microvillous, “Kappe-like,” “pear-shaped,” and “teardrop-shaped”) are the most abundant across the olfactory rosette of this species, while ciliated ORNs are less common and crypt cells are rare. Spatial simulations of the fluid dynamics of the incurrent water flow into the nares and within the olfactory cavities indicate that the high densities of microvilli-bearing ORNs located within the lateral region of the rosette are important for sampling incoming odorants during swimming and may determine subsequent tracking behavior.

Convergence of Olfactory Inputs within the Central Nervous System of a Cartilaginous and a Bony Fish: An Anatomical Indicator of Olfactory Sensitivity

The volume of the olfactory bulbs (OBs) relative to the brain has been used previously as a proxy for olfactory capabilities in many vertebrate taxa, including fishes. Although this gross approach has predictive power, a more accurate assessment of the number of afferent olfactory inputs and the convergence of this information at the level of the telencephalon is critical to our understanding of the role of olfaction in the behaviour of fishes. In this study, we used transmission electron microscopy to assess the number of first-order axons within the olfactory nerve (ON) and the number of second-order axons in the olfactory peduncle (OP) in established model species within cartilaginous (brownbanded bamboo shark, Chiloscyllium punctatum [CP]) and bony (common goldfish, Carassius auratus [CA]) fishes. The total number of axons varied from a mean of 18.12 ± 7.50 million in the ON to a mean of 0.38 ± 0.21 million in the OP of CP, versus 0.48 ± 0.16 million in the ON and 0.09 ± 0.02 million in the OP of CA. This resulted in a convergence ratio of approximately 50:1 and 5:1, respectively, for these two species. Based on astroglial ensheathing, axon type (unmyelinated [UM] and myelinated [M]) and axon size, we found no differentiated tracts in the OP of CP, whereas a lateral and a medial tract (both of which could be subdivided into two bundles or areas) were identified for CA, as previously described. Linear regression analyses revealed significant differences not only in axon density between species and locations (nerves and peduncles), but also in axon type and axon diameter (p < 0.05). However, UM axon diameter was larger in the OPs than in the nerve in both species (p = 0.005), with no significant differences in UM axon diameter in the ON (p = 0.06) between species. This study provides an in-depth analysis of the neuroanatomical organisation of the ascending olfactory pathway in two fish taxa and a quantitative anatomical comparison of the summation of olfactory information. Our results support the assertion that relative OB volume is a good indicator of the level of olfactory input and thereby a proxy for olfactory capabilities.

Cell culture-based karyotyping of orectolobiform sharks for chromosome-scale genome analysis

Karyotyping, traditionally performed using cytogenetic techniques, is indispensable for validating genome assemblies whose sequence lengths can be scaled up to chromosome sizes using modern methods. Karyotype reports of chondrichthyans are scarce because of the difficulty in cell culture. Here, we focused on carpet shark species and the culture conditions for fibroblasts and lymphocytes. The utility of the cultured cells enabled the high-fidelity characterization of their karyotypes, namely 2n = 102 for the whale shark (Rhincodon typus) and zebra shark (Stegostoma fasciatum), and 2n = 106 for the brownbanded bamboo shark (Chiloscyllium punctatum) and whitespotted bamboo shark (C. plagiosum). We identified heteromorphic XX/XY sex chromosomes for the two latter species and demonstrated the first-ever fluorescence in situ hybridization of shark chromosomes prepared from cultured cells. Our protocols are applicable to diverse chondrichthyan species and will deepen the understanding of early vertebrate evolution at the molecular level.

The White-Spotted Bamboo Shark Genome Reveals Chromosome Rearrangements and Fast-Evolving Immune Genes of Cartilaginous Fish.

SummaryChondrichthyan (cartilaginous fish) occupies a key phylogenetic position and is important for investigating evolutionary processes of vertebrates. However, limited whole genomes impede our in-depth knowledge of important issues such as chromosome evolution and immunity. Here, we report the chromosome-level genome of white-spotted bamboo shark. Combing it with other shark genomes, we reconstructed 16 ancestral chromosomes of bamboo shark and illustrate a dynamic chromosome rearrangement process. We found that genes on 13 fast-evolving chromosomes can be enriched in immune-related pathways. And two chromosomes contain important genes that can be used to develop single-chain antibodies, which were shown to have high affinity to human disease markers by using enzyme-linked immunosorbent assay. We also found three bone formation-related genes were lost due to chromosome rearrangements. Our study highlights the importance of chromosome rearrangements, providing resources for understanding of cartilaginous fish diversification and potential application of single-chain antibodies.

Volumetric analysis and morphological assessment of the ascending olfactory pathway in an elasmobranch and a teleost using diceCT.

The size (volume or mass) of the olfactory bulbs in relation to the whole brain has been used as a neuroanatomical proxy for olfactory capability in a range of vertebrates, including fishes. Here, we use diffusible iodine-based contrast-enhanced computed tomography (diceCT) to test the value of this novel bioimaging technique for generating accurate measurements of the relative volume of the main olfactory brain areas (olfactory bulbs, peduncles, and telencephalon) and to describe the morphological organisation of the ascending olfactory pathway in model fish species from two taxa, the brownbanded bamboo shark Chiloscyllium punctatum and the common goldfish Carassius auratus. We also describe the arrangement of primary projections to the olfactory bulb and secondary projections to the telencephalon in both species. Our results identified substantially larger olfactory bulbs and telencephalon in C. punctatum compared to C. auratus (comprising approximately 5.2% vs. 1.8%, and 51.8% vs. 11.8% of the total brain volume, respectively), reflecting differences between taxa, but also possibly in the role of olfaction in the sensory ecology of these species. We identified segregated primary projections to the bulbs, associated with a compartmentalised olfactory bulb in C. punctatum, which supports previous findings in elasmobranch fishes. DiceCT imaging has been crucial for visualising differences in the morphological organisation of the olfactory system of both model species. We consider comparative neuroanatomical studies between representative species of both elasmobranch and teleost fish groups are fundamental to further our understanding of the evolution of the olfactory system in early vertebrates and the neural basis of olfactory abilities.

New Distributional Record of Hasselt’s Bamboo Shark Chiloscyllium hasseltii (Orectolobiformes: Hemiscylliidae) from Bangladesh Waters

A new record of Hasselt’s bamboo shark Chiloscyllium hasseltii (Orectolobiformes: Hemiscylliidae) has been confirmed from the St. Martin’s Island, the northeastern part of the Bay of Bengal of Bangladesh. Two male specimens were collected on 19th December 2018 from the coast of St. Martin’s Island as a bycatch of the local fishermen. The traditional morphometric method was applied for taxonomic identification which was further validated by a molecular approach based on partial cytochrome c oxidase subunit I (COI) sequences (DNA barcodes). The key characteristics of this species are the distance from first to second dorsal fin less than 9.3% of TL; first dorsal fin height less than 6.6% of TL and second dorsal fin height less than 5.8% of TL. The morphometric, meristic and DNA barcoding data thus confirmed the presence of C. hasseltii in Bangladesh. With this report a new shark species has been added in the treasure of the marine ichthyofaunal diversity of Bangladesh and also updates the geographical distribution range of this species.

Morphometric and DNA Barcoding of endemic Halmaheran walking shark (Hemiscyllium halmahera, Allen, 2013) in North Maluku, Indonesia

Abstract. Madduppa H, Putri ASP, Wicaksono RZ, Subhan B, Akbar N, Ismail F, Arafat D, Prabuning D, Sani LMI, Srimariana ES, Baksir A, Bengen DG. 2020. Morphometric and DNA Barcoding of endemic Halmaheran walking shark (Hemiscyllium halmahera, Allen, 2013) in North Maluku, Indonesia. Biodiversitas 21: 3331-3343. The walking shark (Hemiscyllium halmahera) commonly referred to as bamboo shark, is a shark species that belongs to the Hemiscyllidae family which is endemic in North Maluku, and Indonesian waters. These Hemiscyllium species are generally confined to the Australian-New Guinea region, save a specie which originates from Halmahera, an Indonesian island. Ever since its discovery, sufficient information on its existence has been largely unavailable. Hence, the purpose of this study is (1) to assess the morphology, using meristic measurements and length-weight relationship, and (2) to determine the molecular characteristics through DNA barcoding, phylogenetic reconstruction, and genetic diversity. The samples used in this research were obtained from Halmahera Archipelago, North Maluku, including Halmahera, Tidore, Ternate, Maitara, and Guraici in several expeditions between March 2017 and January 2018. 36 individuals in total were measured for morphometric parameters including Total Length (TL), Standard Length (SL), Head Length (HL), Body Circumference (BC), and Body Weight (BW). Out of a total of 36 sharks, tissue samples were obtained from 32 individuals for molecular identification. After the required measurements were obtained, all individuals sharks were released back to their habitat safely. Equations of linear regression were used to analyze the relationship between length and weight. Morphometric measurements indicated that the average total length of the walking shark samples obtained, were not more than 75± cm for both male and female specimens. The length-weight analysis revealed a positive allometric growth pattern in H. halmahera species, where weight was gained faster than an increase in length. The results of the DNA fragment length analysis from all samples were 793-bp. All 32 tissue samples identified as H. halmahera consisted of 3 samples from Ternate Island, 14 samples from Maitara Island, 9 samples from Guraici Island (South Halmahera), 10 samples from Central Halmahera, and 2 samples from Tidore Island. These results confirmed that DNA subsequence analysis indicates that H. halmahera is different from those sequences extracted from Genbank in the Australia-New Guinea region. This study successfully identified H. halmahera based on their morphological and molecular characters, and has therefore contributed to international genetic database (Barcoding Of Life Database-BOLD) for further studies.

RADIOGRAPHIC DETERMINATION OF GASTRIC EMPTYING AND GASTROINTESTINAL TRANSIT TIME IN COWNOSE RAYS (RHINOPTERA BONASUS) AND WHITESPOTTED BAMBOO SHARKS (CHILOSCYLLIUM PLAGIOSUM) AND THE EFFECT OF METOCLOPRAMIDE ON GASTROINTESTINAL MOTILITY.

Gastrointestinal (GI) pathology is common in elasmobranchs; however, information regarding normal GI transit time and the effect of therapeutics on GI motility is lacking. The objective of this study was to determine baseline gastric emptying and GI transit times in cownose rays (Rhinoptera bonasus) and whitespotted bamboo sharks (Chiloscyllium plagiosum) via radiographic barium sulfate contrast studies. Additionally, a pilot study was undertaken to determine the effect of metoclopramide on GI transit time in whitespotted bamboo sharks. Eight cownose rays and eight whitespotted bamboo sharks were administered a 98% w/w barium sulfate suspension at 8 ml/kg via orogastric tube. Post-contrast radiographs were obtained at 2 min, 3, 6, 12, and 23 hr for rays; and 2 min, 3, 6, 9, 12, 16, 25, 30, 36, and every 12 hr until complete gastric emptying occurred for sharks. In cownose rays, the mean and standard error were established for time of initial spiral colon filling (3.4 ± 0.4 hr), complete spiral colon opacification (12 ± 0 hr), initial spiral colon emptying (21.6 ± 1.4 hr), and complete gastric emptying (23 ± 0 hr). In bamboo sharks, the mean and standard error were established for time of initial spiral colon filling (5.3 ± 0.5 hr), complete spiral colon opacification (12.4 ± 1.3 hr), initial spiral colon emptying (22.5 ± 2.7 hr), and complete gastric emptying (39.9 ± 3.3 hr). Cownose rays had a significantly shorter time to spiral colon filling and complete gastric emptying compared with bamboo sharks (P < 0.05). Whitespotted bamboo sharks (n = 8) were administered metoclopramide (0.4 mg/kg orally once daily for 10 days) and the barium series was repeated. Complete gastric emptying time was significantly shorter in treated sharks compared with control (P < 0.05), suggesting that metoclopramide may be a useful therapeutic for GI motility disorders in elasmobranchs.

DiceCT: A Valuable Technique to Study the Nervous System of Fish

Contrast-enhanced X-ray imaging provides a non-destructive and flexible approach to optimizing contrast in soft tissues, especially when incorporated with Lugol’s solution (aqueous I2KI), a technique currently referred to as diffusible iodine-based contrast-enhanced computed tomography (diceCT). This stain exhibits high rates of penetration and results in excellent contrast between and within soft tissues, including the central nervous system. Here, we present a staining method for optimizing contrast in the brain of a cartilaginous fish, the brownbanded bamboo shark, Chiloscyllium punctatum, and a bony fish, the common goldfish, Carassius auratus, using diceCT. The aim of this optimization procedure is to provide suitable contrast between neural tissue and background tissue(s) of the head, thereby facilitating digital segmentation and volumetric analysis of the central nervous system. Both species were scanned before staining and were rescanned at time (T) intervals, either every 48 h (C. punctatum) or every 24 h (C. auratus), to assess stain penetration and contrast enhancement. To compare stain intensities, raw X-ray CT data were reconstructed using air and water calibration phantoms that were scanned under identical conditions to the samples. Optimal contrast across the brain was achieved at T = 240 h for C. punctatum and T = 96 h for C. auratus. Higher resolution scans of the whole brain were obtained at the two optimized staining times for all the corresponding specimens. The use of diceCT provides a new and valuable tool for visualizing differences in the anatomic organization of both the central and peripheral nervous systems of fish.

Sequence structure character of IgNAR Sec in whitespotted bamboo shark (Chiloscyllium plagiosum)

Whitespotted bamboo shark (Chiloscyllium plagiosum) is a demersal cartilaginous fish with an adaptive immune system founded upon immunoglobulins. In this manuscript, we characterize the IgNAR of the whitespotted bamboo shark. A newly discovered alternative splicing form of IgNAR Sec (IgNARshort (ΔC2-C3) Sec) was identified, in which the C1 domain was spliced directly to the C4 domain, the process resulted in a molecule containing three constant domains. However, a single unpaired cysteine remains in the highly flexible hinge region, contributing in the formation of an interchain disulfide bond. Two types of C1 domain were found, and the one lacking a short α-helix showed lower proportion. This finding suggests that short α-helices might be important to the stability of IgNAR. High-throughput sequencing revealed that the percentage of VNAR types significantly vary between the diverse species of sharks. The variable region of IgNAR (the VNAR) with small size and stabilization is a potential candidate for immunotherapeutic agents. The structure and stability analysis in this manuscript may be useful in future biomedical applications.

Examining an ontogenetic shift in the diet of the whitespotted bamboo shark Chiloscyllium plagiosum in northern Taiwanese waters

The diet and feeding habits of the whitespotted bamboo shark (Chiloscyllium plagiosum) were described using stomach content analysis and stable isotope analysis based on 327 and 78 specimens, respectively, that were collected in northern Taiwanese waters from October 2013 to March 2015. The index of relative importance indicated that both females and males mainly fed on Demospongiae, unidentified organisms, Annelida, and crustaceans. Juveniles (< 65 cm in total length, TL) preferred Annelida and crustaceans, while adults (> 65 cm in TL) fed more on teleost fishes. Similar standardized diet breadths (Ba) were found for sharks caught by different fishing gear and for sharks at different maturity stages; however, seasonal variation in Ba was noted. These results suggest that the whitespotted bamboo shark is a prey specialist. No significant differences in prey composition among fishing gear, sexes, maturity stages, or seasons were found by the multi-variate statistical analysis. Permutational multi-variate analysis of variance and a global test also indicated that there were no significant differences in diets between the maturity stages and sexes. The stable isotope analysis indicated that the mean values of δ15N and δ13C for the females and males were similar, suggesting the existence of high overlap in terms of feeding for both sexes. A multi-variate analysis of variance indicated that significant differences were found for the mean values of δ13C and δ15N among the different seasons, maturity stages and season–maturity stage interactions. These results suggested that an ontogenetic shift in the diet of C. plagiosum may occur when the fish approach the size of maturity.

Evaluation of the Antibacterial Material Production in the Fermentation of Bacillus amyloliquefaciens-9 from Whitespotted Bamboo Shark (Chiloscyllium plagiosum).

Bacillus amyloliquefaciens-9 (GBacillus-9), which is isolated from the intestinal tract of the white-spotted bamboo shark (Chiloscyllium plagiosum), can secrete potential antibacterial materials, such as β-1,3-1,4-glucanase and some antimicrobial peptides. However, the low fermentation production has hindered the development of GBacillus-9 as biological additives. In this study, the Plackett–Burman design and response surface methodology were used to optimize the fermentation conditions in a shake flask to obtain a higher yield and antibacterial activity of GBacillus-9. On the basis of the data from medium screening, M9 medium was selected as the basic medium for fermentation. The data from the single-factor experiment showed that sucrose had the highest antibacterial activity among the 10 carbon sources. The Plackett–Burman design identified sucrose, NH4Cl, and MgSO4 as the major variables altering antibacterial activity. The optimal concentrations of these compounds to enhance antibacterial activity were assessed using the central composite design. Data showed that sucrose, NH4Cl, and MgSO4 had the highest antibacterial activities at concentrations of 64.8, 1.84, and 0.08 g L−1, respectively. The data also showed that the optimal fermentation conditions for the antibacterial material production of GBacillus-9 were as follows: Inoculum volume of 5%, initial pH of 7.0, temperature of 36 °C, rotating speed of 180 rpm, and fermentation time of 10 h. The optimal fermentation medium and conditions achieved to improve the yield of antibacterial materials for GBacillus-9 can enhance the process of developing biological additives derived from GBacillus-9.

Everything is subjective under water surface, too: visual illusions in fish.

The study of visual illusions has captured the attention of comparative psychologists since the last century, given the unquestionable advantage of investigating complex perceptual mechanisms with relatively simple visual patterns. To date, the observation of animal behavior in the presence of visual illusions has been largely confined to mammal and bird studies. Recently, there has been increasing interest in investigating fish, too. The attention has been particularly focused on guppies, redtail splitfin and bamboo sharks. Overall, the tested species were shown to experience a human-like perception of different illusory phenomena involving size, number, motion, brightness estimation and illusory contours. However, in some cases, no illusory effects, or evidence for a reverse illusion, were also reported. Here, we review the current state of the art in this field. We conclude that a wider investigation of visual illusions in fish is fundamental to form a broader comprehension of perceptual systems of vertebrates. Furthermore, we believe that this type of investigation could help us to address general important issues in perceptual studies, such as the role of ecology in shaping perceptual systems, the existence of interindividual variability in the visual perception of nonhuman species and the role of cortical activity in the emergence of visual illusions.

Squalene from Fish Livers Extracted by Ultrasound‐Assisted Direct In Situ Saponification: Purification and Molecular Characteristics

AbstractAn ultrasound‐assisted direct in situ saponification (U‐DS) process was developed to extract unsaponifiable matter (rich in squalene) from the livers of four fish species, sea bass, skipjack tuna, gray bamboo shark, and spot‐tail shark. The conventional solvent extraction method was used for comparison. Box–Behnken experimental design (BBD) was adopted to optimize the independent variables including the biomass/methanol ratio (1:3 to 1:9 w/v), 50% KOH volume (1–12 mL), and sonication time (0–30 min) over the unsaponifiable yield. With the conventional process, the yield of squalene was 0.10 ± 0.02 to 5.52 ± 0.06 g (100 g)−1, whereas the U‐DS process rendered the yield of 0.13 ± 0.03 to 6.86 ± 0.05 g (100 g)−1, depending on the fish species. After extraction, squalene was further concentrated (purity ≥60%) from all fish species via fractional crystallization (yield of squalene concentrate ranged from 48.35% to 74.49%) and purified using a silica gel column with a maximum recovery up to 98%. For all the fractions, components were examined using thin layer chromatography (TLC). Squalene was qualitatively and quantitatively analyzed using reversed‐phase high‐performance liquid chromatography (RP‐HPLC). Entire extraction processes yielded squalene with a purity of ≥94%. Fourier transform infrared (FTIR) analysis confirmed the native structure of squalene with six nonconjugated bonds, suggesting no degradation of squalene taken place during the U‐DS process. Thus, the U‐DS process along with fractionation and purification could be used for recovering the squalene from fish livers.

New Geographical Record of the Burmese Bamboo Shark, Chiloscyllium burmensis (Orectolobiformes: Hemiscylliidae), from Bangladesh Waters

We report the first geographical record of the Burmese bamboo shark, Chiloscyllium burmensis (Orectolobiformes: Hemiscylliidae) from the coast of Bay of Bengal, Bangladesh. Three specimens (two males and one female) of C. burmensis were collected on 22nd March 2018, from the Bangladesh Fisheries Development Corporation Fish Landing Center of Cox’s Bazar. Morphometric and molecular approach were applied for taxonomic identification. The shark showed a body slender, snout blunt, nostrils subterminal and well separated from snout tip. Mouth slightly closer to eyes than snout tip, labial folds connected across chin by a dermal fold. Pectoral and pelvic fins are relatively thin. The key characteristics of this species are small eyes, less than 1.25% of total length (TL); distance from vent to tail tip more than 67% of TL; head length less than 15.8% of TL; distance from nose tip to vent less than 31% of TL. The morphometric and DNA barcoding data confirmed the presence of C. burmensis in Bangladesh. This report updates the geographical distribution for this species confirming its presence in the coastal region of Bangladesh.