Zooplankton-based adverse outcome pathways: A tool for assessing endocrine disrupting compounds in aquatic environments.

Endocrine disrupting compounds (EDCs) pose a significant ecological risk, particularly in aquatic ecosystems. EDCs have become a focal point in ecotoxicology, and their identification and regulation have become a priority. Zooplankton have gained global recognition as bioindicators, benefiting from rigorous standardization and regulatory validation processes. This review aims to provide a comprehensive summary of zooplankton-based adverse outcome pathways (AOPs) with a focus on EDCs as toxicants and the utilisation of freshwater zooplankton as bioindicators in ecotoxicological assessments. This review presents case studies in which zooplankton have been used in the development of AOPs, emphasizing the identification of molecular initiating events (MIEs) and key events (KEs) specific to zooplankton exposed to EDCs. Zooplankton-based AOPs may become an important resource for understanding the intricate processes by which EDCs impair the endocrine system. Furthermore, the data sources, experimental approaches, advantages, and challenges associated with zooplankton-based AOPs are discussed. Zooplankton-based AOPs framework can provide vital tools for consolidating toxicological knowledge into a structured toxicity pathway of EDCs, offering a transformative platform for facilitating enhanced risk assessment and chemical regulation.

Public perception on human exposure risk: A case study on endocrine disrupting compounds in the environment.

Humans are exposed to environmental risks owing to the broad usage of endocrine disrupting compounds (EDCs). However, the subjective evaluation of risk levels and characteristics, as well as the variation in risk processing, have not been thoroughly examined. The objective was to understand the public's perception of the risk associated with human exposure to environmental EDCs and identify any variations in risk perception. In this pioneering study conducted within the distinctive social and cultural context of Malaysia, a developing nation, a quantitative analysis approach was employed to assess the subjective evaluation of risk levels and characteristics among the public while developing a risk perception model. Data gathered from surveys and questionnaires were analyzed to gather information on the public's perception of environmental and health issues pertaining to pesticides, hormones, plastics, medicines, and cosmetics. The analysis revealed that the majority of the public assessed the level of human exposure to environmental risks based on experiential processing, which was influenced by cognitive and affective variables. Interestingly, a higher proportion of individuals in the community had a low risk perception of environmental EDCs, surpassing the overall risk perception by 19.3%. Furthermore, the public showed significant awareness of environmental and health issues related to pesticides, hormones, and plastics but had a lesser inclination to acknowledge the vulnerability of humans to risks associated with medicines and cosmetics. These findings suggest that the public is likely to be exposed to environmental EDCs based on their current perceived risks, and that sociopsychological factors play a significant role in shaping perceptions and judgments. This understanding can inform the development of targeted risk management strategies and interventions to mitigate the potential harm caused by environmental EDCs.

Dietary supplementation of Polygonum chinense improves the immunity of Asian seabass, Lates calcarifer (Bloch, 1790) against Vibrio harveyi infection.

Aquaculture plays a significant role in the overall fish production in Malaysia, contributing a substantial quantity of food-fish amounting to roughly 573,683 tonnes with an estimated economic value of US$860 million in 2022. However, diseases have become a significant limitation for aquaculture production. Therefore, herbal immunostimulant has been considered a natural and practical approach of preventing disease infection in fish. The ability of Polygonum chinense extract (PCE) on haemato-biochemistry parameters, immunomodulatory properties, and disease resistance of Lates calcarifer (Asian seabass) under Vibrio harveyi challenge was evaluated in this study, with a focus on dose-response associations and variability over various exposure durations (0-, 7- and 14-day post-infection). A total of 480 Asian seabass (9.5 ± 0.2 g) were distributed in 12 aquaria and fed four diets supplemented with 0 (control), 2, 5 and 10 g/kg diet for 60 days before being challenged with V. harveyi. Dietary PCE significantly improved (P < 0.05) survival, with the dose of 10 g/kg showing the highest survival rate (90 %) when compared to the control (60 %). Additionally, hematological (red and white blood cell counts, hemoglobulin, packed cell volume, and mean corpuscular volume) and immunological (activities of lysozyme, phagocytic activity and respiratory burst, and serum total immunoglobulin) properties were significantly increased (P < 0.05) in comparison to the control group. In contrast, serum aspartate aminotransferase and alanine aminotransferase levels, as well as glucose level were significantly reduced (P < 0.05) in PCE-fed fish compared to the control group. Conclusively, the current study discovered that supplementing fish feed with P. chinense extract improves fish haemato-biochemical profile, immunocompetence and disease resistance to V. harveyi infection.

De Novo Transcriptomic and Life-History Responses of Moina Micrura Under Stress Environment Conditions.

Moina micrura represents a promising model species for ecological and ecotoxicological investigations in tropical freshwater ecosystems. Illumina NovaSeq™ 6000 sequencing was employed in this study to analyze M. micrura across three distinct developmental stages: juvenile, adult, and male. Current study successfully annotated 51,547 unigenes (73.11%) derived from seven (7) different databases. A total of 554 genes were found to be significantly upregulated, while 452 genes showed significant downregulation between juvenile and male. Moreover, 1001 genes were upregulated, whereas 830 genes exhibited downregulation between the adult and male. Analysis of differentially expressed genes revealed upregulation of chitin, cuticle, myosin (MYO), mitogen-activated protein kinases (MAPK), fibrillin (FBN), cytochrome (CYP), glutathione s-transferase (GST), vitellogenin (VTG), acetylcholinesterase (AChE), and transforming growth factor beta (TGFB) under unfavorable environmental conditions (male), as compared to favorable environmental conditions (juveniles and adults). These alterations in gene expression significantly impact the phenological and life-history traits of M. micrura. Furthermore, the upregulation of hemoglobin (HMB), doublesex (DSX), juvenile hormone analogs (JHA), heat shock protein (HSP), and methyltransferase (METT) genes in males initiates the sex-switching effects observed in M. micrura. These findings hold substantial value for researchers interested in determining M. micrura sequences for future investigations of gene expression and comparative reproductive genome analysis within the Moina genus and cladoceran families.

Occurrence, distribution, and ecological risk of bisphenol analogues in marine ecosystem of urbanized coast and estuary.

Bisphenol analogues are prevalent globally because of rampant usage and imprecise processing techniques, prompting alerts about environmental and health hazards. The method employed in this study by solid phase extraction (SPE) and liquid chromatography-tandem quadrupole mass spectrometer (LC-MS/MS) for both quantification and qualitative analysis of the bisphenol compounds in the surface water samples. The coastal and estuarine surface water of Port Dickson and Lukut ranges from 1.32 ng/L to 1890.51 ng/L of bisphenol analogues. BPF mean concentration at 1143.88 ng/L is the highest, followed by BPA and BPS at 59.01 ng/L and 10.96 ng/L, respectively. Based on RQm for bisphenol analogues, the highest for BPF at 2.49 (RQ > 1, high risk), followed by BPS at 0.12 (0.1 < RQ < 1, medium risk) and BPA at 0.09 (0.1 < RQ < 1, medium risk). The presence and current risk of bisphenols analogues should alert the possible water quality degradation soon.

Risk assessment of bisphenol analogues towards mortality, heart rate and stress-mediated gene expression in cladocerans Moina micrura.

Bisphenol A (BPA) is a well-known endocrine-disrupting compound that causes several toxic effects on human and aquatic organisms. The restriction of BPA in several applications has increased the substituted toxic chemicals such as bisphenol F (BPF) and bisphenol S (BPS). A native tropical freshwater cladoceran, Moina micrura, was used as a bioindicator to assess the adverse effects of bisphenol analogues at molecular, organ, individual and population levels. Bisphenol analogues significantly upregulated the expressions of stress-related genes, which are the haemoglobin and glutathione S-transferase genes, but the sex determination genes such as doublesex and juvenile hormone analogue genes were not significantly different. The results show that bisphenol analogues affect the heart rate and mortality rate of M. micrura. The 48-h lethal concentration (LC50) values based on acute toxicity for BPA, BPF and BPS were 611.6 µg L-1, 632.0 µg L-1 and 819.1 µg L-1, respectively. The order of toxicity based on the LC50 and predictive non-effect concentration values were as follows: BPA > BPF > BPS. Furthermore, the incorporated method combining the responses throughout the organisation levels can comprehensively interpret the toxic effects of bisphenol analogues, thus providing further understanding of the toxicity mechanisms. Moreover, the output of this study produces a comprehensive ecotoxicity assessment, which provides insights for the legislators regarding exposure management and mitigation of bisphenol analogues in riverine ecosystems.

Acute toxicity and risk assessment of perfluorooctanoic acid (PFOA) and perfluorooctanesulfonate (PFOS) in tropical cladocerans Moina micrura.

Per- and polyfluoroalkyl substances (PFAS) are gaining worldwide attention because of their toxicity, bioaccumulative and resistance to biological degradation in the environment. PFAS can be categorised into endocrine disrupting chemicals (EDCs) and identified as possible carcinogenic agents for the aquatic ecosystem and humans. Despite this, only a few studies have been conducted on the aquatic toxicity of PFAS, particularly in invertebrate species such as zooplankton. This study evaluated the acute toxicity of two main PFAS, perfluorooctanoic acid (PFOA) and perfluorooctanesulfonate (PFOS), by using freshwater cladocerans (Moina micrura) as bioindicators. This study aimed to assess the adverse effects at different levels of organisations such as organ (heart size and heart rate), individual (individual size and mortality) and population (lethal concentration, LC50). PFOA was shown to be more hazardous than PFOS, with the LC50 values (confidence interval) of 474.7 (350.4-644.5) µg L-1 and 549.6 (407.2-743.9) µg L-1, respectively. As the concentrations of PFOS and PFOA increased, there were declines in individual size and heart rate as compared to the control group. The values of PNECs acquired by using the AF method (PNECAF) for PFOA and PFOS were 0.4747 and 0.5496 µg L-1, respectively. Meanwhile, the PNEC values obtained using the SSD method (PNECSSD) were 1077.0 µg L-1 (PFOA) and 172.5 µg L-1 (PFOS). PNECAF is more protective and conservative compared to PNECSSD. The findings of this study have significant implications for PFOS and PFOA risk assessment in aquatic environments. Thus, it will aid freshwater sustainability and safeguard the human dependency on water resources.

Development of a single-run liquid chromatography-mass spectrometry analysis for the detection of 11 multiclass contaminants of emerging concern using a direct filtration method.

In toxicological analysis, the analytical validation method is important to assess the exact risk of contaminants of emerging concern in the environment. Syringe filters are mainly used to remove impurities from sample solutions. However, the loss of analyte to the syringe filter could be considerable, causing an underestimate of the analyte concentrations. The current study develops and validates simultaneous liquid chromatography-mass spectrometry analysis using a direct filtration method to detect four groups of contaminants of emerging concern. The adsorption of the analyte onto three different matrices and six types of syringe filters is reported. The lowest adsorption of analytes was observed in methanol (16.72%), followed by deionized water (48.19%) and filtered surface lake water (48.94%). Irrespective of the type of the matrices, the lowest average adsorption by the syringe filter was observed in the 0.45 µm polypropylene membrane (15.15%), followed by the 0.20 µm polypropylene membrane (16.10%), the 0.20 µm regenerated cellulose (16.15%), the 0.20 µm polytetrafluoroethylene membrane (47.38%), the 0.45 µm nylon membrane (64.87%) and the 0.20 µm nylon membrane (71.30%). In conclusion, the recommended syringe filter membranes for contaminants of emerging concern analysis are polypropylene membranes and regenerated cellulose, regardless of the matrix used.

Dominant phytoplankton groups as the major source of polyunsaturated fatty acids for hilsa (Tenualosa ilisha) in the Meghna estuary Bangladesh.

The tropical estuarine ecosystem is fascinating for studying the dynamics of water quality and phytoplankton diversity due to its frequently changing hydrological conditions. Most importantly, phytoplankton is the main supplier of ω3 polyunsaturated fatty acids (PUFA) in the coastal food web for fish as they could not synthesize PUFA. This study evaluated seasonal variations of water quality parameters in the Meghna River estuary (MRE), explored how phytoplankton diversity changes according to hydro-chemical parameters, and identified the major phytoplankton groups as the main source of PUFA for hilsa fish. Ten water quality indicators including temperature, dissolved oxygen, pH, salinity, dissolved inorganic nitrogen (DIN = nitrate, nitrite, ammonia) and phosphorus, dissolved silica and chlorophyll-a were evaluated. In addition, phytoplankton diversity was assessed in the water and hilsa fish gut. Principal component analysis (PCA) was used to analyze the spatio-temporal changes in the water quality conditions, and the driving factors in the MRE. Four main components were extracted and explained 75.4% variability of water quality parameters. The most relevant driving factors were dissolved oxygen, salinity, temperature, and DIN (nitrate, nitrite and ammonia). These variabilities in physicochemical parameters and dissolved inorganic nutrients caused seasonal variations in two major groups of phytoplankton. Peak abundance of Chlorophyta (green algae) occurred in water in nutrient-rich environments (nitrogen and phosphorus) during the wet (36%) season, while Bacillariophyta (diatoms) were dominant during the dry (32%) season that depleted dissolved silica. Thus, the decrease of green algae and the increase of diatoms in the dry season indicated the potential link to seasonal changes of hydro-chemical parameters. The green algae (53.7%) were the dominant phytoplankton group in the hilsa gut content followed by diatoms (22.6%) and both are contributing as the major source of PUFAs for hilsa fish according to the electivity index as they contain the highest amounts of PUFAs (60 and 28% respectively).

Use of Taguchi design for optimization of diesel-oil biodegradation using consortium of Pseudomonas stutzeri, Cellulosimicrobium cellulans, Acinetobacter baumannii and Pseudomonas balearica isolated from tarball in Terengganu Beach, Malaysia.

A consortium of bacteria capable of decomposing oily hydrocarbons was isolated from tarballs on the beaches of Terengganu, Malaysia, and classified as Pseudomonas stutzeri, Cellulosimicrobium cellulans, Acinetobacter baumannii and Pseudomonas balearica. The Taguchi design was used to optimize the biodegradation of diesel using these bacteria as a consortium. The highest biodegradation of diesel-oil in the experimental tests was 93.6%, and the individual n-alkanes decomposed 87.6-97.6% over 30 days. Optimal settings were inoculum size of 2.5 mL (1.248 OD600nm); 12% (v/v) the initial diesel-oil in a minimal salt medium of pH 7.0, 30.0 gL-1 NaCl and 2.0 gL-1 NH4NO3 concentration, incubated at 42 °C temperature and 150 rpm agitation speed. Parameters significantly improved diesel-oil removal by consortium as shown by the model determination coefficient (R2 = 90.89%; P < 0.001) with a synergistic effect of agitation speed significantly contributing 81.03%. Taguchi design determined the optimal settings for the parameters under study, which significantly improved diesel-oil removal by consortium. This can be used to design a novel bioremediation strategy that can achieve optimal decontamination of oil pollution in a shorter time. Supplementary Information: The online version contains supplementary material available at 10.1007/s40201-022-00812-3.

Fucoxanthin Production of Microalgae under Different Culture Factors: A Systematic Review.

Fucoxanthin is one of the light-harvesting pigments in brown microalgae, which is increasingly gaining attention due to its numerous health-promoting properties. Currently, the production of microalgal fucoxanthin is not yet feasible from an economic perspective. However, the cultivation of microalgae at favourable conditions holds great potential to increase the viability of this fucoxanthin source. Hence, this study aimed to review the fucoxanthin production of microalgae under different conditions systematically. A literature search was performed using the Web of Science, Scopus and PubMed databases. A total of 188 articles were downloaded and 28 articles were selected for the current review by two independent authors. Microalgae appeared to be a more reliable fucoxanthin source compared to macroalgae. Overall, a consensus fucoxanthin production condition was obtained and proposed: light intensity ranging from 10 to 100 µmol/m2/s could achieve a higher fucoxanthin content. However, the optimal light condition in producing fucoxanthin is species-specific. The current review serves as an antecedent by offering insights into the fucoxanthin-producing microalgae response to different culture factors via a systematic analysis. With the current findings and recommendations, the feasibility of producing fucoxanthin commercially could be enhanced and possibly achieve practical and sustainable fucoxanthin production.

Occurrence, potential sources and ecological risk estimation of microplastic towards coastal and estuarine zones in Malaysia.

Extensive global plastic production has led to microplastic (MP) pollution of marine ecosystems. This study analysed the abundance of MPs in the surface water of tropical coastal and estuarine zones in Malaysia affected by rapid urbanisation and intense human activity. It also estimated the risk posed by MPs to the marine environment. Mean MP abundance ranged from 2.10 to 6.80 particles/L. Fourier-transform infrared spectroscopic analysis found that the MP polymers were dominated by cellophane (54%), followed by polyester (33%) and polyethylene (2%). The risk posed by MPs was estimated with the risk quotient (RQ) method which found no potential ecological risk to both coastal and estuarine areas (RQ < 1). This study will serve as a baseline for future monitoring of MP pollution of marine water to assess the impact of heavily urbanised coastal and estuarine zones.

Pharmaceuticals, hormones, plasticizers, and pesticides in drinking water.

Humans are exposed to endocrine disrupting compounds (EDCs) in tap water via drinking water. Currently, most of the analytical methods used to assess a long list of EDCs in drinking water have been made available only for a single group of EDCs and their metabolites, in contrast with other environmental matrices (e.g., surface water, sediments, and biota) for which more robust methods have been developed that allow detection of multiple groups. This study reveals an analytical method of one-step solid phase extraction, incorporated together with liquid chromatography-tandem mass spectrometry for the quantification of multiclass EDCs (i.e., pharmaceuticals, hormones, plasticizers, and pesticides) in drinking water. Fifteen multiclass EDCs significantly varied in amount between field samples (p < 0.05), with a maximum concentration of 17.63 ng/L observed. Daily exposure via drinking water is unlikely to pose a health risk (risk quotient < 1). This method serves as an analytical protocol for tracing multiclass EDC contamination in tap water as part of a multibarrier approach to ensure safe drinking water for good health and well-being. It represents a simpler one-step alternative tool for drinking water analysis, thereby avoiding the time-consuming and expensive multi-extraction steps that are generally needed for analyzing multiclass EDCs.

A Review on a Hidden Gem: Phycoerythrin from Blue-Green Algae.

Phycoerythrin (PE) is a pink/red-colored pigment found in rhodophytes, cryptophytes, and blue-green algae (cyanobacteria). The interest in PE is emerging from its role in delivering health benefits. Unfortunately, the current cyanobacterial-PE (C-PE) knowledge is still in the infant stage. It is essential to acquire a more comprehensive understanding of C-PE. This study aimed to review the C-PE structure, up and downstream processes of C-PE, application of C-PE, and strategies to enhance its stability and market value. In addition, this study also presented a strengths, weaknesses, opportunities, and threats (SWOT) analysis on C-PE. Cyanobacteria appeared to be the more promising PE producers compared to rhodophytes, cryptophytes, and macroalgae. Green/blue light is preferred to accumulate higher PE content in cyanobacteria. Currently, the prominent C-PE extraction method is repeated freezing-thawing. A combination of precipitation and chromatography approaches is proposed to obtain greater purity of C-PE. C-PE has been widely exploited in various fields, such as nutraceuticals, pharmaceuticals, therapeutics, cosmetics, biotechnology, food, and feed, owing to its bioactivities and fluorescent properties. This review provides insight into the state-of-art nature of C-PE and advances a step further in commercializing this prospective pigment.

Effects of Probiotics on Growth, Survival, Water Quality and Disease Resistance of Red Hybrid Tilapia (Oreochromis spp.) Fingerlings in a Biofloc System.

Biofloc technology has shown positive effects in aquaculture, especially on the growth performance of cultured animals. The aims of this study were to evaluate the effects of adding different probiotic strains in a biofloc system on the growth performance and disease resistance of red hybrid tilapia (Oreochromis spp.). Three different probiotics (Lysinibacillus fusiformis SPS11, Bacillus amyloliquefaciens L9, and Enterococcus hirae LAB3), commercial probiotics (MG1) and a mixed probiotics (MP) combining all three strains were used in this study. The in vitro assay results showed that the mixed probiotic (MP) was able to inhibit the growth of Streptococcus agalactiae and Streptococcus iniae significantly compared to the single and commercial probiotic. The efficacy of MP was further tested in in vivo tilapia culture challenged with S. agalactiae. The best specific growth rate (3.73 ± 0.23% day-1) and feed conversion ratio (0.76 ± 0.04) were recorded in the group of biofloc with addition of MP. After being challenged with S. agalactiae, the group of biofloc with MP had significantly higher survival (83 ± 1.43%) compared to the other groups. Furthermore, the nitrogen concentration (NO2-N and NH4-N) was significantly lower in all the biofloc groups compared to the control. Hence, the addition of probiotics was able to provide beneficial effects to red hybrid tilapia culture in the biofloc system.

Uncovering Research Trends of Phycobiliproteins Using Bibliometric Approach.

Phycobiliproteins are gaining popularity as long-term, high-value natural products which can be alternatives to synthetic products. This study analyzed research trends of phycobiliproteins from 1909 to 2020 using a bibliometric approach based on the Scopus database. The current findings showed that phycobiliprotein is a burgeoning field in terms of publications outputs with "biochemistry, genetics, and molecular biology" as the most related and focused subject. The Journal of Applied Phycology was the most productive journal in publishing articles on phycobiliproteins. Although the United States of America (U.S.A.) contributed the most publications on phycobiliproteins, the Chinese Academy of Sciences (China) is the institution with the largest number of publications. The most productive author on phycobiliproteins was Glazer, Alexander N. (U.S.A.). The U.S.A. and Germany were at the forefront of international collaboration in this field. According to the keyword analysis, the most explored theme was the optimization of microalgae culture parameters and phycobiliproteins extraction methods. The bioactivity properties and extraction of phycobiliproteins were identified as future research priorities. Synechococcus and Arthrospira were the most cited genera. This study serves as an initial step in fortifying the phycobiliproteins market, which is expected to exponentially expand in the future. Moreover, further research and global collaboration are necessary to commercialize phycobiliproteins and increase the consumer acceptability of the pigments and their products.

The Critical Studies of Fucoxanthin Research Trends from 1928 to June 2021: A Bibliometric Review.

Fucoxanthin is a major carotenoid in brown macroalgae and diatoms that possesses a broad spectrum of health benefits. This review evaluated the research trends of the fucoxanthin field from 1928 to June 2021 using the bibliometric method. The present findings unraveled that the fucoxanthin field has grown quickly in recent years with a total of 2080 publications. Japan was the most active country in producing fucoxanthin publications. Three Japan institutes were listed in the top ten productive institutions, with Hokkaido University being the most prominent institutional contributor in publishing fucoxanthin articles. The most relevant subject area on fucoxanthin was the agricultural and biological sciences category, while most fucoxanthin articles were published in Marine Drugs. A total of four research concepts emerged based on the bibliometric keywords analysis: "bioactivities", "photosynthesis", "optimization of process'', and "environment". The "bioactivities" of fucoxanthin was identified as the priority in future research. The current analysis highlighted the importance of collaboration and suggested that global collaboration could be the key to valorizing and efficiently boosting the consumer acceptability of fucoxanthin. The present bibliometric analysis offers valuable insights into the research trends of fucoxanthin to construct a better future development of this treasurable carotenoid.

Life Cycle of Edible Jellyfish Acromitus hardenbergi Stiasny, 1934 (Scyphozoa: Rhizostomeae) Inhabiting a Brackish-Water Environment.

The edible jellyfish Acromitus hardenbergi Stiasny, 1934 is harvested throughout the year at the mouth of the Perak River, Malaysia. Although this species is an important fishery resource in the local area, limited biological studies have been carried out on it. The aim of the present study was to elucidate the life cycle of this unique brackish-water jellyfish in order to conserve the species and develop sustainable jellyfish fisheries. Mature medusae were collected at the mouth of the Perak River. Embryonic and larval development after fertilization was completed within 24 h until the planula stage and within 48 h until the polyp stage. Primary polyps had a long stalk with a small stolon at the base of the calyx. Fully developed polyps were bowl-or goblet-shaped but became an elongated stalk under starved conditions. Asexual reproduction was accomplished only by means of budding, and no podocysts were produced. Strobilation was mono-disc type. These characteristics may be adaptations to the dynamic environmental conditions in the estuary of the Perak River, where salinity fluctuates widely due to strong inflows of highly turbid freshwater coupled with tidal changes. This study suggests that polyps of A. hardenbergi expand their population not by podocysts, but by budding as quickly as possible and forming one large ephyra by mono-disc strobilation without the residuum, because the polyp cannot remain for a long time at its settlement place in the sediment-rich environment with drastic salinity change.

The Efficacy of Moina micrura Enriched with Probiotic Bacillus pocheonensis in Enhancing Survival and Disease Resistance of Red Hybrid Tilapia (Oreochromis spp.) Larvae.

The administration of probiotics via live feeds, such as Artemia and rotifers, has gained significant attention. Moreover, indiscriminate use of antibiotics in conventional aquaculture practices in order to prevent or control disease outbreaks has resulted in the occurrence of residues and antimicrobial resistance. Thus, the application of eco-friendly feed additives, such as probiotics, as a safer alternative has received increasing attention in recent years. However, only minimal information on the administration of probiotics via freshwater cladoceran Moina micrura is available despite being commonly used for larval and post-larval feeding of freshwater crustaceans and fish. Thus, this study aimed to evaluate the application of Bacillus pocheonensis strain S2 administered via M. micrura to red hybrid tilapia (Oreochromis spp.) larvae. Bacillus pocheonensis that has been previously isolated from Spirulina sp. was subjected to preliminary in vitro evaluation of antagonistic properties. The agar well-diffusion assay revealed that this probiont could inhibit the growth of Streptococcus agalactiae and Aeromonas hydrophila. The size of inhibition zones ranged from 8.8 ± 0.2 to 18.2 ± 0.4 mm. Moina micrura was later used as a biological model in preliminary in vivo bacterial challenge assays to evaluate the efficacy of B. pocheonensis in protecting the host from diseases. Moina micrura was pre-enriched with B. pocheonensis at 104 and 106 CFU mL-1 before S. agalactiae and A. hydrophila were introduced into the culture. The study revealed that B. pocheonensis at 104 CFU mL-1 was able to significantly enhance the survival of M. micrura after being challenged with both pathogens (63 ± 3%) in comparison to the control group. The relative percentage survival (RPS) of M. micrura was highest (p < 0.05) when treated with B. pocheonensis at both concentrations 104 and 106 CFU mL-1 (38.33) after being challenged against S. agalactiae. To assess the efficacy of B. pocheonensis in protecting red hybrid tilapia against streptococcosis, the larvae were fed with either unenriched (control) Moina or probiont-enriched Moina daily for 10 days. A significantly (p < 0.05) higher survival rate (77 ± 3%) was observed in larvae fed with probiont-enriched M. micrura compared to other treatments, and the RPS was recorded at 62.90. In addition, the S. agalactiae load was suppressed in larvae fed probiont-enriched M. micrura (6.84±0.39 CFU mL-1) in comparison to the control group (7.78±0.09 CFU mL-1), indicating that the probiont might have contributed to the improvement of tilapia health and survival. This study illustrated that M. micrura was suitable to be used as a vector for probiotics in freshwater fish larvae as an alternative to hazardous antibiotics for disease control.

A Recommendation for a Pre-Standardized Marine Microalgal Dry Weight Determination Protocol for Laboratory Scale Culture Using Ammonium Formate as a Washing Agent.

Microalgal biomass is one of the crucial criteria in microalgal studies. Many reported methods, even the well-established protocol on microalgal dry weight (DW) determination, vary greatly, and reliable comparative assessment amongst published results could be problematic. This study aimed to determine the best condition of critical parameters in marine microalgal DW determination for laboratory-scale culture using four different marine microalgal species. These parameters included the washing process, grades of glass microfiber filter (GMF), GMF pretreatment conditions, washing agent (ammonium formate) concentrations, culture: washing agent ratios (v:v) and washing cycles. GMF grade GF/A with precombustion at 450 °C provided the most satisfactory DW and the highest ash-free dry weight (AFDW)/DW ratio. Furthermore, 0.05 M ammonium formate with 1:2 culture: washing agent ratio and a minimum of two washing cycles appeared to be the best settings of microalgal DW determination. The present treatment increased the AFDW/DW ratio of the four respective microalgae by a minimum of 19%. The findings of this study could serve as a pivotal reference in developing a standardized protocol of marine microalgal DW determination to obtain veracious and reliable marine microalgal DW.