Blue Mussel Larvae Research Articles

The impact of the multichannel quorum sensing systems of Vibrio tasmaniensis and Vibrio crassostreae on virulence towards blue mussel (Mytilus edulis) larvae

Mussel culture in Europe is currently suffering from mass mortality events with mortality rates up to 100%. Vibrios belonging to the Splendidus clade have recently been implicated in these events. In this study, we demonstrate that V. tasmaniensis LGP32 and V. crassostreae J2–9, two model pathogens of oysters, are also pathogenic to blue mussel larvae in an immersion challenge test. After five days, LGP32 and J2–9 had killed 93 and 73% of challenged mussel larvae, respectively. Because quorum sensing, bacterial cell-to-cell communication with small signal molecules, has been demonstrated to control the virulence of various vibrios, we further investigated whether it had an impact on the virulence of V. tasmaniensis and V. crassostreae. We identified the components of a multichannel quorum sensing system (as also found in many other vibrios) in the genomes of both species. Knock out of selected components of this pathway revealed that in general quorum sensing in V. tasmaniensis LGP32 and V. crassostreae J2–9 has no impact on motility, protease activity, biofilm formation and virulence towards mussel larvae. Finally, the quorum sensing inhibitor cinnamaldehyde did not protect mussel larvae from these pathogens. Together, these data indicate that the multichannel quorum sensing systems of V. tasmaniensis LGP32 and V. crassostreae J2–9 have no impact on virulence of the bacteria towards blue mussel larvae. Hence, quorum sensing controlling virulence is not a general feature in vibrios as it has different outcomes on virulence in different species.

Factors affecting settlement, growth and metamorphosis of hatchery-produced Australian blue mussel larvae

Factors affecting settlement, growth and metamorphosis of hatchery-produced Australian blue mussel larvae

Pathogenesis of experimental vibriosis in blue mussel (Mytilus edulis) larvae based on accurate positioning of GFP-tagged Vibrio strains and histopathological and ultrastructural changes of the host

Bacteria, especially Vibrio spp., are largely responsible for larval diseases in shellfish aquaculture. Despite the severe impact of bacterial diseases on larval health, only a few direct studies have characterized the pathogenesis of vibriosis in bivalve larvae. To clarify the onset and advancement of vibriosis in blue mussel Mytilus edulis larvae, two known larval pathogens Vibrio splendidus ME9 and Vibrio anguillarum NB10 were tagged with green fluorescence protein (GFP), and combined with synchronous histopathological analyses, we identified a new type of pathogenesis of experimental vibriosis in bivalve larvae. The GFP-tagged Vibrio strains were first filtered by larvae through the velum and entered into the larval stomach through the esophagus. Later, they proliferated rapidly in the style sac and digestive glands, inducing necrosis of the digestive organs in the dorsal region during the first 24 h post challenge. Necrosis continued with esophagus disruption and velar deformation. Ultimately, the loss of distinguishable internal structures further disrupted the associated regulatory functions and rapidly induced larval death. It was confirmed later by ultrastructural observations that the Vibrio strains grew along the digestive tract and disrupted the digestive epithelium as a portal of entry. Moreover, we obtained the first evidence that the microvilli and cilia of the gastrointestinal epithelial cells were most sensitive to the vibriosis since the earliest disruptions of the larvae occurred in these positions, as shown by ultrastructural observations. The progression of the infection pathway as described in this study will facilitate further research that aims at the analysis of the gastrointestinal morphology after the introduction of anti-Vibrio therapy in the culture system.

Toxicity and histopathological effects of toxic dinoflagellate, Alexandrium catenella exudates on larvae of blue mussel, Mytilus galloprovincialis, and Pacific oyster, Crassostrea gigas

HighlightToxicity and pathological effects of A. catenella were investigated on shellfish larvaeUnfiltered exudates of A. catenella caused significant mortality of blue mussel larvaeApplication of 0.22 mm filtration on A. fundyense exudates potentially decrease the toxicity effectsPathological effects of A. catenella occurred as early as 3 h after exposureThe prevalence and intensity of necrosis increased with exposure duration to A. catenella exudatesAbstractBlooms of the toxic dinoflagellate Alexandrium catenella have affected shellfish industries globally due to their capacity to produce paralytic shellfish toxins(PST). This study aimed to investigate the toxicity effect of exudate A. catenella on larvae of blue mussel Mytilus galloprovincialis and Pacific oyster Crassostrea gigas and filtration methods to reduce the toxic effect. Blue mussel and Pacific oyster larvae were assessed their survival and histopathological changes after exposure to extracellular exudates of A. catenella ranging from 100 to 1,000 cells ml-1 . The results showed that exposure to exudate A. catenella caused significantly higher larval mortality (39 to 52%) than exposure to an equivalent biovolume of the nontoxic species, Tisochrysis lutea (33%) or unfed controls (17%). Filter-sterilization (0.22 µm) of exudates and activated carbon filtration decreased the mortality of Pacific oyster larvae to a level similar to controls (unfed), with the exception of the highest concentrations (600 and 1,000 cells ml-1 ) and mortality of bluemussel larvae mortality by 32% respectively. Blue mussel larvae exposed to exudate A. catenella showed pathological changes mainly in the stomach (digestive gland and style sac) as early as three hours after onset of exposure. The findings of this study suggest that early detection of blooms in the vicinity of mussel and Pacific oyster hatcheries and taking steps to mitigate their effects, is important to reduce the effects of A. catenella blooms on shellfish larval rearing.

Does Ralstonia eutropha, rich in poly-β hydroxybutyrate (PHB), protect blue mussel larvae against pathogenic vibrios?

The natural amorphous polymer poly-β-hydroxybutyrate (PHB-A: lyophilized Ralstonia eutropha containing 75% PHB) was used as a biological agent to control bacterial pathogens of blue mussel (Mytilus edulis) larvae. The larvae were supplied with PHB-A at a concentration of 1 or 10mg/L for 6 or 24hr, followed by exposure to either the rifampicin-resistant pathogen Vibrio splendidus or Vibrio coralliilyticus at a concentration of 105 CFU/ml. Larvae pretreated 6hr with PHB-A (1mg/L) survived a Vibrio challenge better relative to 24hr pretreatment. After 96hr of pathogen exposure, the survival of PHB-A-treated mussel larvae was 1.41- and 1.76-fold higher than the non-treated larvae when challenged with V.splendidus and V.coralliilyticus, respectively. Growth inhibition of the two pathogens at four concentrations of the monomer β-HB (1, 5, 25 and 125mM) was tested in vitro in LB35 medium, buffered at two different pH values (pH 7 and pH 8). The highest concentration of 125mM significantly inhibited the pathogen growth in comparison to the lower levels. The effect of β-HB on the production of virulence factors in the tested pathogenic Vibrios revealed a variable pattern of responses.

Ingestion and effects of micro- and nanoplastics in blue mussel (Mytilus edulis) larvae

It is well known that mussels are exposed to microplastics but ingestion and potential effects on mussel larvae are not well understood. We quantified ingestion and egestion of 100 nm and 2 μm polystyrene beads in blue mussel larvae after 4 h exposure and 16 h depuration using different plastic-to-microalgae ratios. Effects on growth and development of mussel larvae were investigated at 0.42, 28.2 and 282 μgL−1 within 15 days of exposure. We found that, on a mass basis, larvae ingested a higher amount of 2 μm than 100 nm beads, while egestion was independent of particle size and the plastics-to-algae ratio. Although particle egestion occurred readily, microplastics remained inside the larvae. Larval growth was not affected but abnormally developed larvae increased after exposure to polystyrene beads. Malformations were more pronounced for 100 nm beads, at higher concentration and after longer exposure time.

Ralstonia eutropha, containing high poly-β-hydroxybutyrate levels, regulates the immune response in mussel larvae challenged with Vibrio coralliilyticus

Marine invertebrates rely mainly on innate immune mechanisms that include both humoral and cellular responses. Antimicrobial peptides (AMPs), lysozyme and phenoloxidase activity, are important components of the innate immune defense system in marine invertebrates. They provide an immediate and rapid response to invading microorganisms. The impact of amorphous poly-β-hydroxybutyrate (PHB-A) (1 mg PHB-A L−1) on gene expression of the AMPs mytimycin, mytilinB, defensin and the hydrolytic enzyme lysozyme in infected blue mussel larvae was investigated during “in vivo” challenge tests with Vibrio coralliilyticus (105 CFU mL−1). RNAs were isolated from mussel larvae tissue, and AMPs were quantified by q-PCR using the 18srRNA gene as a housekeeping gene. Our data demonstrated that AMPs genes had a tendency to be upregulated in challenged mussel larvae, and the strongest expression was observed from 24 h post-exposure onwards. The presence of both PHB-A and the pathogen stimulated the APMs gene expression, however no significant differences were noticed between treatments or between exposure time to the pathogen V. coralliilyticus. Looking at the phenoloxidase activity in the infected mussels, it was observed that the addition of PHB-A significantly increased the activity.

Isolation of Vibrionaceae from wild blue mussel (Mytilus edulis) adults and their impact on blue mussel larviculture.

The blue mussel (Mytilus edulis) is known as a robust bivalve species, although its larviculture appears to be highly susceptible to diseases. In this study, we isolated 17 strains from induced mortality events in healthy wild-caught blue mussel adults and demonstrated that they caused between 17% and 98% mortality in blue mussel larvae in a newly developed, highly controlled immersion challenge test model. Eight of the isolates belong to the Splendidus clade of vibrios, while the other isolates belong to the genus Photobacterium. The genomes of the most virulent Vibrio isolate and the most virulent Photobacterium isolate were sequenced and contained several genes encoding factors that have previously been linked to virulence towards bivalves. In vitro tests confirmed that all 17 isolates were positive for these virulence factors. The sequenced genomes also contained a remarkably high number of multidrug resistance genes. We therefore assessed the sensitivity of all isolates to a broad range of antibiotics and found that there were indeed many strong positive correlations between the sensitivities of the isolates to different antibiotics. Our data provide an ecological insight into mass mortality in blue mussels as they indicate that wild mussels contain a reservoir of pathogenic bacteria.

Sole and Combined Usage of Ultra-sonication and Hydrogen Peroxide as Mitigation Techniques of Bio-fouling

Mussels are stubborn organism attached to solid substrate by byssus threads and caused operational problems in utility of power generating stations. Sole and combined usage of ultrasonic (28 kHz- and 42 kHz- frequencies) and hydrogen peroxide (H2O2) has studied for control of blue mussel larvae and adult stage in seawater condition. A theoretical wo rking model using disinfection (Chick and Watson type) approaches is presented based on helpful results of experiments. This study also demonstrate that the combined treatment (ultra-sonication with H2O2) is overall highly efficient than individual treatment would, but on the basis of exposure time, the ultra-sonication was the most efficient among them. Therefore the development of sole and combined technique might be effective practical mitigation strategy against mussel attachment for water handling facilities.

Validation of trophic and anthropic underwater noise as settlement trigger in blue mussels.

Like the majority of benthic invertebrates, the blue mussel Mytilus edulis has a bentho-pelagic cycle with its larval settlement being a complex phenomenon involving numerous factors. Among these factors, underwater noise and pelagic trophic conditions have been weakly studied in previous researches. Under laboratory conditions, we tested the hypothesis that picoplankton assimilation by the pediveliger blue mussel larvae acts as a food cue that interacts with anthropic underwater sound to stimulate settlement. We used 13C-labeling microalgae to validate the assimilation of different picoplankton species in the tissues of pediveliger larvae. Our results clearly confirm our hypothesis with a significant synergic effect of these two factors. However, only the picoeukaryotes strains assimilated by larvae stimulated the settlement, whereas the non-ingested picocyanobacteria did not. Similar positive responses were observed with underwater sound characterized by low frequency vessel noises. The combination of both factors (trophic and vessel noise) drastically increased the mussel settlement by an order of 4 compared to the control (without picoplankton and noise). Settlement levels ranged from 16.5 to 67% in 67 h.

Response of competent blue mussel (Mytilus edulis) larvae to positive and negative settlement cues

Recent work on larval settlement cues has emphasized mechanisms by which larvae exploit individual, positive cues (cues that larvae move toward), often in complex flow fields. Yet in natural systems, larvae of habitat generalists probably respond to multiple settlement cues, including a mixture of positive and negative cues. First, a simple test chamber in which cue dispersal was dominated by diffusion was used to assess whether competent blue mussel (Mytilus edulis) larvae responded negatively or positively to cues from a variety of intertidal species. Second, choice experiments tested the responses of larvae offered a mixture of conflicting (positive and negative) cues from the same direction and conflicting cues from different directions. Responses to individual cues were predictable from established ecological interactions. Larvae were attracted to odors from conspecifics, tended to move toward odors from a filamentous alga, avoided odors from two predators of post-settlement mussels, and exhibited little response to odors from an herbivorous gastropod. Negative and positive cues offered from the same direction produced movement both toward and away from the mixture, while offering the combined cues from different directions resulted in net movement that was largely consistent with predictions from the individual cue responses. Larvae presented with a choice between two negative cues mainly remained where they started, in the middle of the test apparatus, and exhibited no choice. These results highlight the need to consider the combined effects of a broad range of cues when evaluating the net effect of cues on settlement in the field.

Maintained larval growth in mussel larvae exposed to acidified under-saturated seawater.

Ocean acidification (OA) is known to affect bivalve early life-stages. We tested responses of blue mussel larvae to a wide range of pH in order to identify their tolerance threshold. Our results confirmed that decreasing seawater pH and decreasing saturation state increases larval mortality rate and the percentage of abnormally developing larvae. Virtually no larvae reared at average pHT 7.16 were able to feed or reach the D-shell stage and their development appeared to be arrested at the trochophore stage. However larvae were capable of reaching the D-shell stage under milder acidification (pHT ≈ 7.35, 7.6, 7.85) including in under-saturated seawater with Ωa as low as 0.54 ± 0.01 (mean ± s. e. m.), with a tipping point for normal development identified at pHT 7.765. Additionally growth rate of normally developing larvae was not affected by lower pHT despite potential increased energy costs associated with compensatory calcification in response to increased shell dissolution. Overall, our results on OA impacts on mussel larvae suggest an average pHT of 7.16 is beyond their physiological tolerance threshold and indicate a shift in energy allocation towards growth in some individuals revealing potential OA resilience.

Family effects on the growth and survival of congeneric blue mussel larvae (Mytilus edulis and M. trossulus)

Intraspecific genetic variation is widely recognized to affect how bivalve larvae respond to variation in environmental conditions, but has been largely ignored in comparisons of larval performance between closely related sibling species. Replicates of five different full-sib families of larval blue mussels (Mytilus edulis and its more northerly congener, M. trossulus) were reared under different temperature (10, 13, and 17 °C) and food conditions in a full factorial design. Growth and survival were strongly affected by temperature, family, the interaction of family and temperature, and the three-way interaction of family, temperature, and food. Family means for days to 20 % survival ranged from 10.2 to 20.0 and did not cluster by species. The two M. trossulus families were intermediate in survival and ranked first and third for growth. The temperature by family interaction effect for survival was very strong, and the two M. trossulus families responded very differently to temperature manipulations. One M. trossulus family exhibited highest survival at 13 °C, while the other M. trossulus family and all three M. edulis families exhibited highest survival at 10 °C. By contrast, greatest growth consistently occurred at 17 °C, indicating that higher mortality in warmer water may be at least partially offset (at a population level) by more rapid growth in body size. The results illustrate the importance of assaying both growth and survival when evaluating environmental effects on larvae and the need to ensure a high level of genetic diversity when pools of larvae are used to study genetically similar species.

Across-shelf distribution of blue mussel larvae in the northern Gulf of Maine: consequences for population connectivity and a species range boundary.

Studies of population connectivity have largely focused on along-shelf, as opposed to across-shelf, processes. We hypothesized that a discontinuity in across-shelf mixing caused by the divergence of the Eastern Maine Coastal Current (EMCC) from shore acts as an ecological barrier to the supply of mussel larvae to the coast. Existing data on the relative abundance of two congeneric blue mussels, Mytilus edulis and M. trossulus, were analysed to quantify the association of M. trossulus with the colder temperature signal of the EMCC and generate larval distribution predictions. We then sampled the across-shelf distribution of larvae along two transects during 2011. Larvae were identified using restriction digests of PCR amplicons from the mitochondrial 16S rDNA. Mytilus edulis larvae were consistently abundant on either the inshore and offshore transect ends, but not homogeneously distributed across the shelf, while M. trossulus larvae were less common throughout the study area. The divergence of the EMCC from shore appears to create a break in the connectivity of M. edulis populations by isolating those inshore of the EMCC from upstream larval sources. Across-shelf transport processes can thus produce connectivity patterns that would not be predicted solely on the basis of along-shelf processes.

Early development of the blue mussel Mytilus edulis (Linnaeus, 1758) cultured in potassium-fortified inland saline water

The low potassium concentration in inland saline water (ISW) restrains the normal development of cultured marine organisms, and thus, possesses challenges for the development of ISW aquaculture. Therefore, assessing the effects of potassium fortification in ISW on the performance of cultured marine species is an important step to determine the feasibility of their culture in ISW. The aim of this research was to investigate the effects of potassium fortification in ISW on the performance of early life stages of the blue mussel Mytilus edulis including fertilised eggs, trochophore, veliger and pediveliger larvae. These stages were reared in five different levels of potassium-fortified ISW, namely 20, 40, 60, 80 and 100% of potassium levels equivalent to the potassium level in ocean water (OW) and two controls namely, ISW at 27ppt (ISW27) and OW at 25ppt (OW25). The results showed that the higher levels of potassium in ISW, particularly with 100% K+ fortification (ISW100K+), invariably improved the survival and size, and reduced the developmental stage interval and deformities of blue mussel larvae. Deformities, such as faulty cell cleavage, abnormal formation of trochophore larvae, protruding mantle in veliger larvae, and indented shell margin in veliger and in pediveliger, were observed when reared in any ISW. However, rearing in ISW did not result in any deformities in settlement larvae. The number of deformities was reduced at higher K+ fortification levels, and there were no deformities in pediveliger larvae reared in ISW100K+ and in OW. These results showed that K+ fortification in ISW improves the performance of the rearing of the larval stages of the blue mussel. Statement of relevanceThis study is the first attempt to culture larval stages of blue mussel Mytilus edulis in potassium-fortified inland saline water. The results have shown that that the larval development is feasible only if ISW is fortified with K+. By initiating larval culture in inland saline water, we expect better results will be reflected by adult stages.

Application of poly-β-hydroxybutyrate (PHB) in mussel larviculture

Abstract In this study, the effect of poly-β-hydroxybutyrate (PHB) delivery on the performance (survival, growth and metamorphosis) of blue mussel ( Mytilus edulis ) larvae was studied for the first time. Upon addition of PHB in either crystalline (i.e. extracted from the bacterial cell) or amorphous (i.e. still contained in the bacterial cell) forms to a standard algal diet at concentrations of 0.1 mg L − 1 , 1.0 mg L − 1 and 10.0 mg L − 1 , no significant improvement in growth performance or metamorphosis was observed. However, a concentration of 1.0 mg L − 1 amorphous PHB almost doubled the survival of the larvae in several independent experiments. A total of sixteen PHB-degrading bacterial strains belonging to genus Pseudoalteromonas could be isolated from the PHB treated mussel larvae. The application of these in combination with PHB however, did not further increase the culture performance of the mussel larvae. Where previous studies suggested that the PHB effect could be related to a change in the intestinal microbial community of the treated animals, molecular fingerprinting analysis of the mussel larvae associated microbiota did not show a relationship between changes in the microbiota composition and the improved survival following PHB treatment. Overall, this study showed for the first time that PHB can be applied in blue mussel larviculture to increase the survival of the cultured animals. Statement of relevance Novel approach to increase survival in mussel larviculture.

Stimulation of heterotrophic bacteria associated with wild-caught blue mussel (Mytilus edulis) adults results in mass mortality

Abstract Although hatchery technology is available and although hatcheries have several advantages over collection of natural spat, hatchery production of blue mussel ( Mytilus edulis ) larvae currently is not economically feasible. In contrast to other bivalves such as oysters and clams, no mortality episodes due to pathogenic bacteria have been described for adult blue mussel thus far. In this study, we aimed at investigating whether opportunistic pathogens are associated with wild-caught adult blue mussel, as we reasoned that environmental conditions that are beneficial to opportunistic pathogens might be responsible for mass mortalities of mussel larvae under hatchery conditions. The growth of heterotrophic bacteria associated with wild-caught blue mussel adults was stimulated by the addition of organic matter (tryptone and yeast extract) to the rearing water. The addition of organic matter resulted in a 3 log increase in total heterotrophic bacterial counts in the rearing water, and resulted in complete mortality of the animals after 6 days, and this could be prevented by the addition of antibiotics. Water quality parameters (TAN, nitrite, oxygen and pH) were monitored regularly, and were all within the acceptable range throughout the experiment, excluding water quality deterioration as the cause of mortality. These data might give a first insight into the reason why blue mussel larviculture still is problematic.

Effects of different dietary microalgae on survival, growth, settlement and fatty acid composition of blue mussel ( Mytilus galloprovincialis) larvae

The diatom Chaetoceros calcitrans is a major component of many bivalve hatcheries, yet it is expensive and notoriously difficult to culture on a commercial scale. In an attempt to reduce dependence on the diatom C. calcitrans, mussel larvae ( Mytilus galloprovincialis) were subjected to feeding experiments which altered levels of the diatom under controlled hatchery conditions. Growth, survival and settlement success of mussel larvae were determined in response to five mixed algal diets in which the relative contributions of C. calcitrans was varied over the experimental period (30 days). Fatty acid profiles of the larvae and algal diets were also assessed. The exclusion of C. calcitrans from the diet had no significant differences on larval growth and only minor differences in total fatty acid content were found between treatments. Fatty acid analysis revealed that larval survival was strongly influenced by the proportions of dietary docosahexaenoic acid (DHA), while settlement was positively correlated with higher ratios of the n-3 long-chained polyunsaturated fatty acid (n-3 LC-PUFA) (namely, DHA and eicosapentaenoic acid, EPA), to the n-6 LC-PUFA (arachidonic acid, ARA). Despite similar relative and absolute n-3 LC-PUFA levels in the larvae under different dietary treatments, the larvae receiving high levels of C. calcitrans performed significantly better in terms of survival and settlement success. These results indicate that the (DHA + EPA)/ARA ratio is a key factor in determining larval performance, rather than the total amount of these fatty acids.

Effect of ocean acidification on the early life stages of the blue mussel <i>Mytilus edulis</i>

Abstract. Several experiments have shown a decrease of growth and calcification of organisms at decreased pH levels. There is a growing interest to focus on early life stages that are believed to be more sensitive to environmental disturbances such as hypercapnia. Here, we present experimental data, acquired in a commercial hatchery, demonstrating that the growth of planktonic mussel (Mytilus edulis) larvae is significantly affected by a decrease of pH to a level expected for the end of the century. Even though there was no significant effect of a 0.25–0.34 pH unit decrease on hatching and mortality rates during the first 2 days of development nor during the following 13-day period prior to settlement, final shells were respectively 4.5±1.3 and 6.0±2.3% smaller at pHNBS~7.8 (pCO2~1100–1200 μatm) than at a control pHNBS of ~8.1 (pCO2~460–640 μatm). Moreover, a decrease of 12.0±5.4% of shell thickness was observed after 15d of development. More severe impacts were found with a decrease of ~0.5 pHNBS unit during the first 2 days of development which could be attributed to a decrease of calcification due to a slight undersaturation of seawater with respect to aragonite. Indeed, important effects on both hatching and D-veliger shell growth were found. Hatching rates were 24±4% lower while D-veliger shells were 12.7±0.9% smaller at pHNBS~7.6 (pCO2~1900 μatm) than at a control pHNBS of ~8.1 (pCO2~540 μatm). Although these results show that blue mussel larvae are still able to develop a shell in seawater undersaturated with respect to aragonite, the observed decreases of hatching rates and shell growth could lead to a significant decrease of the settlement success. As the environmental conditions considered in this study do not necessarily reflect the natural conditions experienced by this species at the time of spawning, future studies will need to consider the whole larval cycle (from fertilization to settlement) under environmentally relevant conditions in order to investigate the potential ecological and economical losses of a decrease of this species fitness in the field.

Spawning Behavior and Early Life History of the Sharpnose Puffer, Canthigaster rivulata, in the Aquarium

Specimens ofCanthigaster rivulata (Temminck et Schlegel) were collected from Kominato and Hayama, central Japan, from May, 1985 to October, 1986. On the basis of the gonadosomatic index, gonadal histology and results of artificial fertilization of these specimens, the spawning season is considered to extend from late June to mid-September. The specimens exhibited the following dimorphic differences associated with sex: 1) The male is larger than the female. 2) Ventral side of the body is brownish orange in the male with vermiculated or reticulated patterns of bright violet, while it is white in the female. 3) The male has a well-developed skin fold along the mid-dorsal and mid-ventral lines, which is greatly elevated during courtship; whereas the female’s skin folds are not or slightly developed and conspicuous only during courtship. In an aquarium with the water temperatures of 22 to 26°C, a pair of fish spawned every four days late in the morning for three consecutive months. Courtship and spawning occurred in a pair. The male swam in front of the female, and elevated the skin folds both dorsally and ventrally, fully spreading the unpaired fins, with the ventral side of the body flashing bright blue and the dorsal side turning dark. Both fish swam in a circular fashion, elevating the skin folds. The male followed the female nudging her abdomen with his snout. Both fish turned upward, and released gametes. The eggs are spherical, 0.53–0.73 mm in diameter, demersal, adhesive, transparent, and pale yellowish orange in color, and contain a cross-shaped or asteroid cluster of oil globules. The egg membrane was thick and consisted of about 14 concentric layers. The incubation period ranged from 73.5 hours at 28.2–28.5°C to 145.0 hours at 22.1–22.4°C. The newly hatched larvae were 1.38–1.98 mm in total length (TL) with 84-11-13 = 19–21 myomeres. The yolk was absorbed when the larvae attained 1.49–2.22 mm TL, three days after hatching. The larvae were fed on oyster larvae, blue mussel larvae, sea-urchin larvae and rotifers, but all of them died in 16 days. During the embryonic and early larval stages, the only pigment cells that appeared on the body were the black chromatophores.