Influence of waterborne butachlor (BUC), a commonly used pesticide, on morphometric, biochemical, and molecular biomarkers was evaluated in juvenile, full sibling, diploid and triploid African catfish (Clarias gariepinus). Fish were exposed for 21 days to one of three concentrations of BUC [mean measured µg/L: 22, 44 or 60]. Unexposed (control) triploids were heavier and longer and had higher visceral-somatic index (VSI) than diploids. Also, they had lighter liver weight (HSI) and showed lower transcript levels of brain gonadotropin-releasing hormone (GnRH), aromatase (cyp191b) and fushi tarazu-factor (ftz-f1), and plasma testosterone levels than diploids. Butachlor treatments had no effects, in either diploid or triploid fish, on VSI, HSI, weight or length changes, condition factor (CF), levels of plasma testosterone, 17-β estradiol (E2), cortisol, cholesterol, or mRNA levels of brain tryptophan hydroxylase (tph2), forkhead box L2 (foxl2), and 11 β-hydroxysteroid dehydrogenase type 2 (11β-hsd2). Expressions of cyp191b and ftz-f1 in triploids were upregulated by the two highest concentrations of BUC. In diploid fish, however, exposures to all BUC concentrations decreased GnRH transcription and the medium BUC concentration decreased ftz-f1 transcription. Substantial differences between ploidies in basal biomarker responses are consistent with the reported impaired reproductive axis in triploid C. gariepinus. Furthermore, the present study showed the low impact of short term exposure to BUC on reproductive axis in C. gariepinus.
So far, several classes of digesting solutions have been employed to extract microplastics (MPs) from biological matrices. However, the performance of digesting solutions across different temperatures has never been systematically investigated. In the first phase of the present study, we measured the efficiency of different oxidative agents (NaClO or H2O2), bases (NaOH or KOH), and acids [HCl or HNO3; concentrated and diluted (5%)] in digesting fish tissues at room temperature (RT, 25°C), 40, 50, or 60°C. In the second phase, the treatments that were efficient in digesting the biological materials (>95%) were evaluated for their compatibility with eight major plastic polymers (assessed through recovery rate, Raman spectroscopy analysis, and morphological changes). Among the tested solutions, NaClO, NaOH, and diluted acids did not result in a satisfactory digestion efficiency at any of the temperatures. The H2O2 treatment at 50°C efficiently digested the biological materials, although it decreased the recovery rate of nylon-6 (NY6) and nylon-66 (NY66) and altered the colour of polyethylene terephthalate (PET) fragments. Similarly, concentrated HCl and HNO3 treatments at RT fully digested the fish tissues, but also fully dissolved NY6 and NY66, and reduced the recovery rate of most or all of the polymers, respectively. Potassium hydroxide solution fully eliminated the biological matrices at all temperatures. However, at 50 and 60°C, it degraded PET, reduced the recovery rate of PET and polyvinyl chloride (PVC), and changed the colour of NY66. According to our results, treating biological materials with a 10% KOH solution and incubating at 40°C was both time and cost-effective, efficient in digesting biological materials, and had no impact on the integrity of the plastic polymers. Furthermore, coupling this treatment with NaI extraction created a promising protocol to isolate MPs from whole fish samples.
Plastic debris is widespread and ubiquitous in the marine environment and ingestion of plastic debris by marine organisms is well-documented. Viscera and gills of 110 individual marine fish from 11 commercial fish species collected from the marine fish market were examined for presence of plastic debris. Isolated particles were characterized by Raman spectroscopy, and elemental analysis was assessed using energy-dispersive X-ray spectroscopy (EDX). Nine (of 11) species contained plastic debris. Out of 56 isolated particles, 76.8% were plastic polymers, 5.4% were pigments, and 17.8% were unidentified. Extracted plastic particle sizes ranged from 200 to 34,900 μm (mean = 2600 μm ±7.0 SD). Hazardous material was undetected using inorganic elemental analysis of extracted plastic debris and pigment particles. The highest number of ingested microplastics was measured in Eleutheronema tridactylum and Clarias gariepinus, suggesting their potential as indicator species to monitor and study trends of ingested marine litter.
Information on the biological responses of polyploid animals towards environmental contaminants is scarce. This study aimed to compare reproductive axis-related gene expressions in the brain, plasma biochemical responses, and the liver and gill histopathological alterations in diploid and triploid full-sibling juvenile African catfish (Clarias gariepinus). Fish were exposed for 96 h to one of the two waterborne phenanthrene (Phe) concentrations [mean measured (SD): 6.2 (2.4) and 76 (4.2) μg/L]. In triploids, exposure to 76 μg/L Phe increased mRNA level of fushi tarazu-factor 1 (ftz-f1). Expression of tryptophan hydroxylase2 (tph2) was also elevated in both ploidies following the exposure to 76 μg/L Phe compared to the solvent control. In triploids, 76 μg/L Phe increased plasma alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) levels compared to the other Phe-exposed group. It also elevated lactate and glucose contents relative to the other groups. In diploids, however, biochemical biomarkers did not change. Phenanthrene exposures elevated glycogen contents and the prevalence of histopathological lesions in the liver and gills of both ploidies. This study showed substantial differences between diploids and triploids on biochemical and molecular biomarker responses, but similar histopathological alterations following acute Phe exposures.
Skin is a major by-product of the fisheries and aquaculture industries and is a valuable source of gelatin. This study examined the effect of triploidization on gelatin yield and proximate composition of the skin of African catfish (Clarias gariepinus). We further investigated the effects of two commonly used pesticides, chlorpyrifos (CPF) and butachlor (BUC), on the skin gelatin yield and amino acid composition in juvenile full-sibling diploid and triploid African catfish. In two separate experiments, diploid and triploid C. gariepinus were exposed for 21 days to graded CPF [mean measured: 10, 16, or 31 μg/L] or BUC concentrations [Mean measured: 22, 44, or 60 μg/L]. No differences in skin gelatin yield, amino acid or proximate compositions were observed between diploid and triploid control groups. None of the pesticide treatments affected the measured parameters in diploid fish. In triploids, however, gelatin yield was affected by CPF treatments while amino acid composition remained unchanged. Butachlor treatments did not alter any of the measured variables in triploid fish. To our knowledge, this study is the first to investigate changes in the skin gelatin yield and amino acid composition in any animal as a response to polyploidization and/or contaminant exposure.
Presence of microplastics (MPs) in a broad range of wild and cultured marine organisms is well-documented, but transfer mechanisms by which cultured organisms are contaminated with MPs is poorly understood. MP loads in three Malaysian commercial brands of fish meal were investigated. Chemical composition of extracted MP-like particles was confirmed using micro-Raman spectroscopy. Inorganic composition of MPs and pigment particles were assessed through energy-dispersive X-ray spectroscopy (EDX). Out of 336 extracted particles, 64.3% were plastic polymers, 25% pigment particles, 4.2% non-plastic items, and 6.5% were unidentified. Fragments were the dominant form of MPs (78.2%) followed by filaments (13.4%) and films (8.4%). This study demonstrates that cultured organisms could be exposed to high levels of MPs via MP contaminated fish/shellfish used in fish meal production. Fish meal replacement with other sources of protein including meat meals and plant-based meals may mitigate MP exposure to cultured or farmed organisms.
This study examined the potential of artificial neural network (ANN) modeling to infer timing, route and dose of contaminant exposure from biomarkers in a freshwater fish. Hepatic glutathione S-transferase (GST) activity and biliary concentrations of BaP, 1-OH BaP, 3-OH BaP and 7,8D BaP were quantified in juvenile Clarias gariepinus injected intramuscularly or intraperitoneally with 10-50 mg/kg benzo[a]pyrene (BaP) 1-3 d earlier. A feedforward multilayer perceptron (MLP) ANN resulted in more accurate prediction of timing, route and exposure dose than a linear neural network or a radial basis function (RBF) ANN. MLP sensitivity analyses revealed contribution of all five biomarkers to predicting route of exposure but no contribution of hepatic GST activity or one of the two hydroxylated BaP metabolites to predicting time of exposure and dose of exposure. We conclude that information content of biomarkers collected from fish can be extended by judicious use of ANNs.
There are serious concerns over the adverse impacts of microplastics (MPs) on living organisms. The main objective of this study was to test the effects of MPs on the total length, weight, condition factor (CF), transcriptional level of antioxidant, anti and pro-apoptotic, and neurotransmitter genes, and the histopathology of the gill, liver, brain, kidney, and intestine in the larvae of zebrafish (Danio rerio). Fish were exposed to one of three levels of pristine low-density polyethylene (LDPE) fragments (5, 50, or 500 μg/L) for 10 or 20 days. No significant changes were observed in any of the selected biomarkers across MP concentrations at days 10 or 20. The expression of casp9 (caspase 9, apoptosis-related cysteine protease), casp3a (caspase 3, apoptosis-related cysteine protease a) and cat (catalase), however, were significantly lower in the larvae sampled at day 20 than day 10. We provide evidence that virgin short-term exposure to LDPE fragments has minimal impact on biomarker responses in D. rerio larvae.
Dietary organic acids are increasingly being investigated as a potential means of improving growth and nutrient utilization in aquatic animals. A 9-week study was performed to compare equal amounts (2%) of different organic acids (sodium butyrate, acetate, propionate, or formate) on the growth, muscle proximate composition, fatty acid composition, cholesterol and lipid peroxidation, differential cell counts, plasma biochemistry, intestinal short-chain fatty acid (SCFA) level, and liver histopathology to red hybrid tilapia (Oreochromis sp.) (initial mean weight of 2.87 g). A second experiment was performed to determine their effects on lipid peroxidation and trimethylamine (TMA) when added at 1% to tilapia meat and left out for 24 h. The results of the first experiment showed no treatment effect to growth, feeding efficiencies, or muscle fatty acid composition, but all dietary organic acids significantly decreased intestinal SCFA. Dietary butyrate and propionate significantly decreased muscle lipid peroxidation compared to the control group, but the dietary formate treatment had the lowest lipid peroxidation compared to all treatments. Muscle crude protein and lipid in tilapia fed the formate diet were significantly lower and higher, respectively, and showed evidence of stress based on the differential cell counts, significantly higher plasma glucose and liver glycogen, as well as inflammatory responses in the liver. Although a potential benefit of dietary organic acids was a reduction to lipid peroxidation, this could be accomplished post-harvest by direct additions to the meat. In addition, inclusions of butyrate and propionate to tilapia meat significantly decreased TMA, which might be a more cost-effective option to improve the shelf life of tilapia products.
The impacts of environmental stressors on polyploid organisms are largely unknown. This study investigated changes in morphometric, molecular, and biochemical parameters in full-sibling diploid and triploid African catfish (Clarias gariepinus) in response to chlorpyrifos (CPF) exposures. Juvenile fish were exposed to three concentrations of CPF (mean measured μg/L (SD): 9.71 (2.27), 15.7 (3.69), 31.21 (5.04)) under a static-renewal condition for 21days. Diploid control groups had higher hepatosomatic index (HSI), plasma testosterone (T), and brain GnRH and cyp19a2 expression levels than triploids. In CPF-exposed groups, changes in HSI, total weight and length were different between the diploid and triploid fish. In contrast, condition factor did not alter in any of the treatments, while visceral-somatic index (VSI) changed only in diploids. In diploid fish, exposure to CPF did not change brain 11β-hsd2, ftz-f1, foxl2, GnRH or cyp19a2 mRNA levels, while reduced tph2 transcript levels compared to the control group. In contrast, 11β-hsd2 and foxl2 expression levels were changed in triploids following CPF exposures. In diploids, plasma T levels showed a linear dose-response reduction across CPF treatments correlating with liver weight and plasma total cholesterol concentrations. In contrast, no changes in plasma cholesterol and T concentrations were observed in triploids. Plasma cortisol and 17-β estradiol (E2) showed no response to CPF exposure in either ploidy. Results of this first comparison of biomarker responses to pesticide exposure in diploid and polyploid animals showed substantial differences between diploid and triploid C. gariepinus.
The present study aimed to assess the effects of anthropogenic activities on the heavy metal levels in the Langat River by transplantation of Corbicula javanica. In addition, potential ecological risk indexes (PERI) of heavy metals in the surface sediments of the river were also investigated. The correlation analysis revealed that eight metals (As, Co, Cr, Fe, Mn, Ni, Pb and Zn) in total soft tissue (TST) while five metals (As, Cd, Cr, Fe and Mn) in shell have positively and significantly correlation with respective metal concentration in sediment, indicating the clams is a good biomonitor of the metal levels. Based on clustering patterns, the discharge of dam impoundment, agricultural activities and urban domestic waste were identified as three major contributors of the metals in Pangsun, Semenyih and Dusun Tua, and Kajang, respectively. Various geochemical indexes for a single metal pollutant (geoaccumulation index (I geo), enrichment factors (EF), contamination factor (C f) and ecological risk (Er)) all agreed that Cd, Co, Cr, Cu, Fe, Mn, Ni and Zn are not likely to cause adverse effect to the river ecosystem, but As and Pb could pose a potential ecological risk to the river ecosystem. All indexes (degree of contamination (C d), combined pollution index (CPI) and PERI) showed that overall metal concentrations in the tropical river are still within safe limit. River metal pollution was investigated. Anthropogenic activities were contributors of the metal pollution. Geochemical indexes showed that metals are within the safe limit.
Silver barb Barbodes gonionotus fry were exposed to polyvinyl chloride (PVC) fragments at increasing concentrations of 0.2, 0.5 and 1.0 mg/L for 96 h, following which whole body histological evaluation and analysis of the digestive enzymes trypsin and chymotrypsin were performed. Whole body trypsin and chymotrypsin activities increased significantly in fish exposed to 0.5 and 1.0 mg/L PVC as compared those exposed to zero or 0.2 mg/L PVC. In fish exposed to all tested concentrations, PVCs were observed in both the proximal and distal intestine, and fish exposed to 0.5-1.0 and 1.0 mg/L PVC, respectively, and these particles were associated with localized thickening of the mucosal epithelium. No tissue damage was evident in any other internal organs or gills. This lack of damage may be attributed to the absence of contaminants associated with the PVC fragments and their relatively smooth surface. The increased whole body trypsin and chymotrypsin activities may indicate an attempt to enhance digestion to compensate for epithelial thickening of the intestine and/or to digest the plastics.
This study represents a first attempt at applying a fuzzy inference system (FIS) and an adaptive neuro-fuzzy inference system (ANFIS) to the field of aquatic biomonitoring for classification of the dosage and time of benzo[a]pyrene (BaP) injection through selected biomarkers in African catfish (Clarias gariepinus). Fish were injected either intramuscularly (i.m.) or intraperitoneally (i.p.) with BaP. Hepatic glutathione S-transferase (GST) activities, relative visceral fat weights (LSI), and four biliary fluorescent aromatic compounds (FACs) concentrations were used as the inputs in the modeling study. Contradictory rules in FIS and ANFIS models appeared after conversion of bioassay results into human language (rule-based system). A "data trimming" approach was proposed to eliminate the conflicts prior to fuzzification. However, the model produced was relevant only to relatively low exposures to BaP, especially through the i.m. route of exposure. Furthermore, sensitivity analysis was unable to raise the classification rate to an acceptable level. In conclusion, FIS and ANFIS models have limited applications in the field of fish biomarker studies.
The contamination of aquatic environments with microplastics (MPs) has spurred an unprecedented interest among scientific communities to investigate their impacts on biota. Despite the rapid growth in the number of studies on the aquatic toxicology of MPs, controversy over the fate and biological impacts of MPs is increasingly growing mainly due to the absence of standardized laboratory bioassays. Given the complex features of MPs, such as the diversity of constituent polymers, additives, shapes and sizes, as well as continuous changes in the particle buoyancy as a result of fouling and defouling processes, it is necessary to modify conventional bioassay protocols before employing them for MP toxicity testings. Moreover, several considerations including quantification of chemicals on/in the MP particles, choice of test organisms, approaches for renewing the test solution, aggregation prevention, stock solution preparation, and units used to report MP concentration in the test solution should be taken into account. This critical review suggests some important strategies to help conduct environmentally-relevant MP bioassays.
Glutathione S-transferases (GST) are considered among the most controversial biomarkers of water pollutants in fish with little known about factors influencing their activities. The objective of this study was to investigate how gender, dose, ploidy, and sampling time alter hepatic GST activities in African catfish (Clarias gariepinus) following β-naphthoflavone (β-NF) injection. Newly matured male and female diploid and triploid fish were intraperitoneally (i.p.) injected with 0, 15, or 75 mg/kg of β-NF, and livers were excised 24, 48, and 72 h post-injection. Results showed that hepatic GST activities were significantly inhibited by both doses of β-NF. Inhibition was greater in females than males, but no significant differences were observed between diploid and triploid fish. Enzymatic activities differed over time with lowest levels 72 h post-injection. These results extend our understanding of GST activity in fish and highlight the necessity of considering confounding factors when comparing different studies.
A total of 40 marine mussel Perna viridis populations collected (2002-2009) from 20 geographical sites located in two busy shipping lanes namely the Straits of Malacca (10 sites; 16 populations) and the Straits of Johore (8 sites; 21 populations) and three populations (2 sites) on the east coast of Peninsular Malaysia, was determined for Cd, Cu, Fe, Ni, Pb and Zn concentrations. In comparison with the maximum permissible limits (MPLs) set by existing food safety guidelines, all metal concentrations found in all the mussel populations were lower than the prescribed MPLs. In terms of the provisional tolerable weekly intake prescribed by the Joint FAO/WHO Expert Committee on Food Additives (JECFA) and oral reference doses (ORDs) by the USEPA, all the studied metals (except for Pb) were unlikely to become the limiting factors or unlikely to pose a risk for the consumption of the mussel populations. The estimated daily intake (EDI) for average level mussel (ALM) and high level mussel (HLM) consumers of mussels was found to be lower than the ORD guidelines for Cd, Cu, Fe, Ni and Zn. Furthermore, the target hazard quotient (THQ) was found to be less than 1 for ALM consumers but higher than 1 for HLM consumers in some sites. Therefore, there were no potential human health risks to the ALM consumers of the mussels. However, for Pb THQ values, the Pb levels in some mussel populations could create a health risk problem. Present results indicate that the consumption amounts of mussels should be limited for minimizing potential health risks of heavy metals to the HLM consumers.
A 9-week study was conducted to compare dietary corn starch (CS) or tapioca starch (TS), with or without being pre-gelatinized (PG), on the growth, feeding efficiencies, plasma and muscle biochemistry, intestinal short chain fatty acids (SCFA), and liver glycogen of triplicate groups of 20 red hybrid tilapia (Orecohromis sp.). Various pellet characteristics were evaluated, along with their surface and cross sectional microstructure. The PG diets had significantly higher water stability, bulk density, and protein solubility, along with a smoother surface. Tilapia fed the TS diet had lower growth than had all other tilapia, but were significantly improved when diet was pre-gelatinized. In the PG treatments, intestinal SCFA significantly decreased while plasma glucose, cholesterol and triglycerides, as well as liver glycogen, significantly increased. Fish fed the CS diet had significantly more long chain polyunsaturated fatty acid than had those fed by other treatments. Pre-gelatinization may improve fish productivity and offer greater flexibility during aquafeed production.
No report was found on the occurrence of microplastics in processed seafood products that are manufactured for direct human consumption. This study investigates the potential presence of micro- and mesoplastics in 20 brands of canned sardines and sprats originating from 13 countries over 4 continents followed by their chemical composition determination using micro-Raman spectroscopy. The particles were further inspected for their inorganic composition through energy-dispersive X-ray spectroscopy (EDX). Plastic particles were absent in 16 brands while between 1 and 3 plastic particles per brand were found in the other 4 brands. The most abundant plastic polymers were polypropylene (PP) and polyethylene terephthalate (PET). The presence of micro- and mesoplastics in the canned sardines and sprats might be due to the translocation of these particles into the edible tissues, improper gutting, or the result of contamination from the canneries. The low prevalence of micro- and mesoplastics sized >149μm, and the absence of potentially hazardous inorganic elements on them, might indicate the limited health risks associated with their presence in canned sardines and sprats. Due to the possible increase in micro- and mesoplastic loads in seafood products over time, the findings of this study suggest their quantification to be included as one of the components of food safety management systems.
There is a paucity of information about the occurrence of microplastics (MPs) in edible fish tissues. Here, we investigated the potential presence of MPs in the excised organs (viscera and gills) and eviscerated flesh (whole fish excluding the viscera and gills) of four commonly consumed dried fish species (n = 30 per species). The MP chemical composition was then determined using micro-Raman spectroscopy and elemental analysis with energy-dispersive X-ray spectroscopy (EDX). Out of 61 isolated particles, 59.0% were plastic polymers, 21.3% were pigment particles, 6.55% were non-plastic items (i.e. cellulose or actinolite), while 13.1% remained unidentified. The level of heavy metals on MPs or pigment particles were below the detection limit. Surprisingly, in two species, the eviscerated flesh contained higher MP loads than the excised organs, which highlights that evisceration does not necessarily eliminate the risk of MP intake by consumers. Future studies are encouraged to quantify anthropogenic particle loads in edible fish tissues.
Cancer therapy in Malaysia primarily focuses on the clinical management of patients with cancer and malnutrition continues to be one of the major causes of death in these patients. There is a dearth of information on the nutrient intake and status of newly diagnosed patients with cancer prior to the initiation of treatment. The present study aims to assess the nutrient intake and status of newly diagnosed patients with cancer from the East Coast of Peninsular Malaysia.