Metal contents in house geckos (Squamata: Gekkonidae) from industrial and urban areas of southern Tamaulipas, Mexico and western Andalucía, Spain, may reflect airborne metal pollution.

House geckos share living quarters with humans in the tropical and subtropical regions inhabited by these reptiles. Gecko behavior, biological traits, continuous exposure to suspended particulate matter 0 µm in diameter (PM10) and dust, as well as status as exotic species, motivated the choice of these species to examine environmental exposure to ambient air pollutants, in particular metals, and subsequent accumulation in these organisms. One part of the study was conducted in Tamaulipas (Mexico) where Hemydactylus frenatus is abundant in urban and industrial environments, the other part was conducted in Andalucia (Spain) where Tarentola mauritanica is found in similar environments. Adult geckos were collected on buildings in locations affected by various air pollution sources. For both species, higher metal contents were observed in whole-body (including digestive tracts) analysis and were markedly different between collection sites. Contents in tails, digestive tracts, and carcasses without digestive tracts were not correlated. Based on contamination factor values, bioaccumulation in H. frenatus tissues occurred for 12 of the 15 metals analyzed. Data suggest that H. frenatus might serve as a biomonitor for Cu, Ni, Pb, Cr, Li, and V, whereas T. mauritanica might be a biomonitor for Cu, Ni, Pb, and Cr. To our knowledge, metal contents for H. frenatus are reported here for the first time. House gecko data could be integrated into a highly representative monitoring system and health risk assessments related to air quality in residential areas.

Combined effects of high temperature and pesticide mixture exposure on free-swimming behaviors and hepatic cytochrome P450 1A expression in goldfish, Carassius auratus.

The synergy between multiple compounds and other stressors, including heat, creates volatility and greater unpredictability than standard single-chemical toxicity testing, especially in the case of pesticides and metabolites which might contain several noxious ingredients resulting in adverse ecological effects. To address this, the aim of this study was to examine the dose- and time-dependent effects of low- and high-dose pesticide mixture (metalachlor, linuron, isoproturon, tebucanazole, aclonifen, atrazine, pendimethalin, azinphos-methyl) and heat stress co-exposure (22°C control/32°C treatment for 4-week) on free-swimming behaviors and cumulative actionless time (CAT) of goldfish. Behavioral analysis showed a dose- and time-dependent decrease in distance swam, as well as a subsequent increase in CAT. Vertical and horizontal spatial behavioral use were affected under heat and pesticides co-exposure conditions. In 3- and 4-week(s) exposure groups, horizontal spatial behavioral use demonstrated elevated time spent in the lower third of the aquarium. Similarly, during 3- and 4-week(s) exposure (32°C control and 32°C high doses) vertical spatial behavioral use was found to increase time spent in the outermost edges of the aquarium. In all treatment groups, the final condition factor (KM) showed significant attenuation when compared to the initial KM. However, there was an unclear relationship between heat/pesticide co-exposure and growth most notably in 32°C high-dose groups. In addition, the expression of hepatic cytochrome P450 1A mRNA was significantly higher in pesticide-exposed groups. Taken together, data demonstrated that co-exposure with low- or high-dose pesticide mixture and heat stress significantly impacted natural swimming patterns, which over time might result in the broader population and ecological effects.

Interactive effects of high temperature and pesticide exposure on oxidative status, apoptosis, and renin expression in kidney of goldfish: Molecular and cellular mechanisms of widespread kidney damage and renin attenuation.

One of many noteworthy consequences of increasing societal reliance on pesticides is their predominance in aquatic environments. These pernicious chemicals interact with high temperatures from global climate change, heat waves, and natural variations to create unstable environments that negatively impact organisms' health. To understand these conditions, we examined the dose-dependent effects of environmentally relevant pesticide mixtures (metolachlor, linuron, isoproturon, tebuconazole, aclonifen, atrazine, pendimethalin, and azinphos-methyl) combined with elevated temperatures (22 control vs. 32°C for 4-week exposure) on renin, dinitrophenyl protein (DNP, an indicator of reactive oxygen species, ROS), 3-nitrotyrosine protein (NTP, an indicator of reactive nitrogen species, RNS), superoxidase dismutase (SOD, an antioxidant), and catalase (CAT, an antioxidant) expressions in the kidneys of goldfish (Carassius auratus). Histopathological analysis showed widespread damage to kidney tissues in high temperature and pesticide co-exposure groups, including rupture of the epithelial layer, hemorrhaging, and degeneration of tubular epithelium. Quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemical analyses demonstrated significant declines in renin receptor-like mRNA and protein expressions in kidney tissues under combined exposure to high temperature and pesticides compared with controls; conversely, expression of DNP, NTP, SOD, and CAT increased in kidney tissues under the same conditions. Apoptotic cells were also increased in co-exposure groups as assessed by in situ terminal deoxynucleotidyl transferase dUTP nick labeling (TUNEL) assay. The enhanced apoptosis in kidneys of heat and pesticides co-exposed fish was associated with increased caspase-3 (a protease enzyme) mRNA levels. Our results demonstrated that high temperature and pesticides induced oxidative/nitrative stress (i.e., ROS/RNS), damaged tissues, increased cellular apoptosis, and suppressed renin expression in kidneys of goldfish.

Effects of elevated temperature on prooxidant-antioxidant homeostasis and redox status in the American oyster: Signaling pathways of cellular apoptosis during heat stress.

Increasing seawater temperature affects growth, reproduction, development, and various other physiological processes in aquatic organisms, such as marine invertebrates, which are especially susceptible to high temperatures. In this study, we examined the effects of short-term heat stress (16, 22, 26, and 30 °C for 1-week exposure) on prooxidant-antioxidant homeostasis and redox status in the American oyster (Crassostrea virginica, an edible and commercially cultivated bivalve mollusk) under controlled laboratory conditions. Immunohistochemical and real-time quantitative PCR (qRT-PCR) analyses were performed to examine the expression of heat shock protein-70 (HSP70, a biomarker of heat stress), catalase (CAT, an antioxidant), superoxide dismutase (SOD, an antioxidant), dinitrophenyl protein (DNP, a biomarker of reactive oxygen species, ROS), and 3-nitrotyrosine protein (NTP, an indicator of reactive nitrogen species, RNS), in the gills and digestive glands of oysters. In situ TUNEL assay was performed to detect cellular apoptosis in tissues. Histological analysis showed an increase in mucus secretion in the gills and digestive glands of oysters exposed to higher temperatures (22, 26, and 30 °C) compared to control (16 °C). Immunohistochemical and qRT-PCR analyses showed significant increases in HSP70, DNP and NTP protein, and mRNA expressions in tissues at higher temperatures. Cellular apoptosis was also significantly increased at higher temperatures. Thus, heat-induced oxidative and nitrative stress likely occur due to overproduction of ROS and RNS. Interestingly, expression of CAT and SOD increased in oysters exposed to 22 and 26 °C, but was at or below control levels in the highest temperature exposure (30 °C). Collectively, these results suggest that elevated seawater temperatures cause oxidative/nitrative stress and induce cellular apoptosis through excessive ROS and RNS production, leading to inhibition of the antioxidant defense system in marine mollusks.

Molecular characterization and expression of arginine vasotocin V1a2 receptor in Atlantic croaker brain: Potential mechanisms of its downregulation by PCB77.

The arginine vasotocin (AVT)-V1a receptor mediates critical reproductive behaviors of the nonapeptide vasotocin in the teleost brain. In this study, we report the molecular characterization of the AVT-V1a2 receptor and its messenger RNA (mRNA) and protein expressions in the Atlantic croaker brain after exposure to the planar polychlorinated biphenyl congener 3,3',4,4'-tetrachlorobiphenyl (PCB77). The full-length sequence of croaker AVT-V1a2 receptor complementary DNA (cDNA) is highly homologous to other teleost AVT-V1a2 receptor cDNAs. Double-labeled immunohistochemistry showed coexpression of AVT-V1a2 receptor and gonadotropin-releasing hormone-I (GnRH-I, a neuropeptide that regulates gonadotropin secretion) in hypothalamic neurons, thereby providing the anatomical basis for possible AVT modulation of croaker reproduction through alterations in GnRH-I secretion. AVT-V1a2 receptor mRNA and protein levels as well as GnRH-I mRNA levels were markedly decreased in hypothalamic tissues of croaker exposed to PCB77 (dose: 2 and 8 µg/g body weight for 4 weeks) compared with levels in untreated (control) fish. In contrast, hypothalamic cytochrome P450 1A (CYP1A, a monooxygenase enzyme) and interleukin-1ß (IL-1ß, a cytokine indicator of inflammation and response to neuronal damage) mRNA levels, and plasma protein carbonyl (PC, an indicator of reactive oxygen species) contents, important biomarkers of neural stress, were increased in PCB77-exposed fish compared with controls. Collectively, these results suggest that the downregulation of hypothalamic AVT-V1a2 receptor and GnRH-I transcripts due to PCB77 exposure is associated with induction of CYP1A, cellular inflammation and oxidative stress in Atlantic croaker, a marine teleost that inhabits estuaries along the US Atlantic coast and the Gulf of Mexico that are often contaminated with persistent organic pollutants such as PCBs.

Short-term heat stress impairs testicular functions in the American oyster, Crassostrea virginica: Molecular mechanisms and induction of oxidative stress and apoptosis in spermatogenic cells.

Global temperature is increasing due to anthropogenic activities. Abnormal temperature has devastating effects on growth, reproduction, and development of aquatic organisms. In this study, we examined the effects of short-term exposure to elevated temperatures (28 and 32°C for 1 week) on testicular functions, heat shock protein-70 (HSP70), dinitrophenyl protein (DNP, a biomarker of reactive oxygen species [ROS]), and nitrotyrosine protein (NTP, an indicator of reactive nitrogen species [RNS]) expressions, protein carbonyl (PC, a measure of ROS) contents, nitrates/nitrites (NOx, a metabolite of nitric oxide) levels, extrapallial fluid (EPF) conditions, and cellular apoptosis in American oyster (Crassostrea virginica). Higher temperatures significantly decreased (~26%) sperm production in oysters compared with controls (24°C). HSP70, NTP, and DNP expressions were increased after heat exposure, consistent with increased EPF pH, and cellular apoptosis. The enhanced apoptosis in spermatogenic cells is associated with increased caspase 3/7 activity, PC contents, and NOx levels in testicular tissues. Together these results suggest that elevated temperature drastically increases oxidative stress and cellular apoptosis which in turn leads to decreased testicular functions in oysters. To the best of our knowledge, this study reports the first findings on the impacts of elevated temperatures on testicular functions in oysters.

Short-term exposure to 12‰ brackish water has significant effects on the endocrine physiology of juvenile American alligator (Alligator mississippiensis).

American alligators (Alligator mississippiensis) mainly inhabit freshwater habitats but can be exposed to a wide range of salinities during storm surges, droughts or from alterations in freshwater flows. Although some salinization events last weeks, others only last a few days. This study assessed changes in the endocrine function of the renin-angiotensin-aldosterone system (RAAS) and steroid hormone production (steroidogenesis) in juvenile alligators exposed to brackish water (12‰) for 7 days. We quantified plasma levels of angiotensin II and the corticosteroids (aldosterone, corticosterone and 11-deoxycortisol). Various progestogens, androgens, and estrogens were further assessed. The protein expression for the RAAS enzymes, renin and angiotensin converting enzyme (ACE), was quantified immunohistochemically in kidney and lung tissue, respectively, and histology was performed on kidney, lung and gonad tissues. Finally, blood biochemistry parameters such as electrolyte levels and diagnostic indicators for dehydration, renal, and hepatic function were measured. Corticosterone, 11-deoxycortisol, Na+, Cl-, total protein, albumin, uric acid, and cholesterol levels were all significantly elevated in alligators exposed to brackish water compared with alligators in freshwater. The levels of 17ß-estradiol and estrone were significantly lowered while histology showed alterations in gonad tissue in the brackish water exposed group. In contrast, while there were no effects of exposure on aldosterone levels, angiotensin II was significantly reduced in brackish water exposed alligators. These results correlated with significantly decreased expressions for both renin and ACE in kidney and lung tissue. Overall, this study showed that short-term exposure of alligators to 12‰ brackish water has significant endocrine effects on juvenile alligators.

Molecular cloning and characterization of two ARNT (ARNT-1 and ARNT-2) genes in Atlantic croaker and their expression during coexposure to hypoxia and PCB77.

Aryl hydrocarbon receptor nuclear translocator (ARNT) is an important transcriptions factor that binds/coactivates drug-metabolizing genes in vertebrates. In this study, we report the cloning and characterization of two ARNT (ARNT-1 and ARNT-2) genes and their mRNA and protein expression in liver tissues of Atlantic croaker after co-exposure to hypoxia and 3,3',4,4'-tetrachlorobiphenyl (PCB77). The full-length croaker ARNT-1 and ARNT-2 genes encode proteins of 537 and 530 amino acids, respectively, and are highly homologous to ARNT-1 and ARNT-2 genes of other vertebrates. ARNT mRNAs are ubiquitously expressed in all tissues. Hypoxia (dissolved oxygen: 1.7 mg/L) exposure (1-4 weeks) did not affect hepatic ARNTs mRNA levels. Dietary PCB77 treatment (2 and 8 µg/g body weight/day for 4 weeks) caused marked increases in ARNTs mRNA and protein levels in normoxic fish. However, coexposure to hypoxia and PCB77 for 4 weeks significantly blunted the increase in ARNTs mRNA and protein levels in response to PCB77 exposure. These results suggest that ARNT activity and functions induced by exposure to PCB aryl hydrocarbon receptor (AhR) agonists could be compromised in croaker inhabiting hypoxic coastal regions.

Interactive effects of hypoxia and PCB co-exposure on expression of CYP1A and its potential regulators in Atlantic croaker liver.

Although marine and coastal environments which are contaminated with xenobiotic organic compounds often become hypoxic during the summer, the interactive effects of hypoxia and xenobiotic exposure on marine species such as teleost fishes remain poorly understood. The expression and activity of monooxygenase enzyme cytochrome P450-1A (CYP1A) in fishes are upregulated by exposure to polychlorinated biphenyls (PCBs), whereas they are down-regulated during hypoxia exposure. We investigated the interactive effects of hypoxia and PCB co-exposure on hepatic CYP1A expression in Atlantic croaker and on potential regulators of CYP1A. Croaker were exposed to hypoxia (1.7 mg/L dissolved oxygen), 3,3',4,4'-tetrachlorobiphenyl (PCB 77, dose: 2 and 8 µg/g body weight), and Aroclor 1254 (a common PCB mixture, dose: 0.5 and 1 µg/g body weight), alone and in combination for 4 weeks. PCB 77 exposure markedly increased hepatic CYP1A mRNA and protein expression, and ethoxyresorufin-O-deethylase (EROD, an indicator of CYP1A enzyme) activity and increased endothelial nitric oxide synthase (eNOS) protein expression. PCB 77 treatment also increased interleukin-1ß (IL-1ß, a cytokine) mRNA levels and protein carbonyl (PC, an indicator of reactive oxygen species, ROS) contents. These marked PCB 77- and Aroclor 1254-induced increases in CYP1A mRNA levels and EROD activity were significantly attenuated by co-exposure to hypoxia, whereas the increases in hepatic eNOS protein and IL-1ß mRNA expression, and PC contents were augmented by hypoxia co-exposure. The results suggest that biotransformation of organic xenobiotics by CYP1A is reduced in fish during co-exposure to hypoxia and is accompanied by alterations in eNOS, ROS, and IL-1ß levels.

Molecular characterization and hypoxia-induced upregulation of neuronal nitric oxide synthase in Atlantic croaker: Reversal by antioxidant and estrogen treatments.

Neuronal nitric oxide synthase (nNOS) catalyzes production of nitric oxide in vertebrate brains. Recent findings indicate that endothelial NOS and reactive oxygen species (ROS) are significantly increased during hypoxic stress and are modulated by antioxidants. However, the influence of antioxidants and steroids on nNOS upregulation by hypoxia is largely unknown. In this study, we characterized nNOS cDNA and examined the effects of hypoxia and antioxidant and steroid treatments on nNOS expression in Atlantic croaker hypothalamus. Hypoxia exposure (dissolved oxygen, DO: 1.7 mg/L for 2 and/or 4weeks) caused significant increases in hypothalamic nNOS mRNA, protein and its neuronal expression. Hypothalamic nNOS expression and superoxide radical (O2(·-), an index of ROS) production were increased by pharmacological treatment of fish exposed to normoxic conditions with N-ethylmaleimide, an alkene drug which covalently modifies sulfhydryl groups and inhibits aromatase activity. In contrast, treatments with Nω-nitro-L-arginine methyl ester, a competitive NOS-inhibitor, or vitamin E, an antioxidant, prevented the upregulation of O2(·-) production and nNOS expression in hypoxia-exposed (DO: 1.7 mg/L for 4 weeks) fish. Moreover, treatment with 1,4,6-androstatrien-3,17-dione, an aromatase inhibitor, increased hypothalamic O2(·-) production and nNOS expression in normoxic control fish; whereas estradiol-17ß treatment significantly reduced O2(·-) production and nNOS expression in hypoxia-exposed fish. Double-labeled immunohistochemical results showed that nNOS and aromatase proteins are co-expressed in the hypothalamus. Taken together, the results suggest that upregulation of nNOS and ROS in the croaker hypothalamus in response to hypoxia is influenced by antioxidant and overall estrogen status.

Histological, biochemical and immunohistochemical assessments of Roundup®, atrazine, and 2,4-D mixtures on tissue architecture, body fluid conditions, nitrotyrosine protein and Na+/K+-ATPase expressions in the American oyster, Crassostera virginica.

Pesticides are widely used to control weeds and pests in agricultural settings but harm non-target aquatic organisms. In this study, our objective was to evaluate the effect of short-term exposure (one week) to environmentally relevant concentrations of pesticides mixture (low concentration: 0.4 µg/l atrazine, 0.5 µg/l Roundup®, and 0.5 µg/l 2,4-D; high concentration: 0.8 µg/l atrazine, 1 µg/l Roundup®, and 1 µg/l 2,4-D) on tissue architecture, body fluid conditions, and 3-nitrotyrosine protein (NTP) and Na+/K+-ATPase, expressions in tissues of American oyster (Crassostrea virginica) under controlled laboratory conditions. Histological analysis demonstrated the atrophy in the gills and digestive glands of oysters exposed to pesticides mixture. Periodic acid-Schiff (PAS) staining showed the number of hemocytes in connective tissue increased in low- and high-concentration pesticides exposure groups. However, pesticides treatment significantly (P < 0.05) decreased the amount of mucous secretion in the gills and digestive glands of oysters. The extrapallial fluid (i.e., body fluid) protein concentrations and glucose levels were dropped significantly (P < 0.05) in oysters exposed to high-concentration pesticides exposure groups. Moreover, immunohistochemical analysis showed significant upregulations of NTP and Na+/K+-ATPase expressions in the gills and digestive glands in pesticides exposure groups. Our results suggest that exposure to environmentally relevant pesticides mixture causes morphological changes in tissues and alters body fluid conditions and NTP and Na+/K+-ATPase expressions in tissues, which may lead to impaired physiological functions in oysters.

New prospects of environmental RNA metabarcoding research in biological diversity, ecotoxicological monitoring, and detection of COVID-19: a critical review.

Ecosystems are multifaceted and complex systems and understanding their composition is crucial for the implementation of efficient conservation and management. Conventional approaches to biodiversity surveys can have limitations in detecting the complete range of species present. In contrast, the study of environmental RNA (eRNA) offers a non-invasive and comprehensive method for monitoring and evaluating biodiversity across different ecosystems. Similar to eDNA, the examination of genetic material found in environmental samples can identify and measure many species, including ones that pose challenges to traditional methods. However, eRNA is degraded quickly and therefore shows promise in detection of living organisms closer to their actual location than eDNA methods. This method provides a comprehensive perspective on the well-being of ecosystems, facilitating the development of focused conservation approaches to save at-risk species and uphold ecological equilibrium. Furthermore, eRNA has been recognized as a valuable method for the identification of COVID-19 in the environment, besides its established uses in biodiversity protection. The SARS-CoV-2 virus, which is accountable for the worldwide epidemic, releases RNA particles into the surrounding environment via human waste, providing insights into the feasibility of detecting it in wastewater and other samples taken from the environment. In this article, we critically reviewed the recent research activities that use the eRNA method, including its utilization in biodiversity conservation, ecological surveillance, and ecotoxicological monitoring as well as its innovative potential in identifying COVID-19. Through this review, the reader can understand the recent developments, prospects, and challenges of eRNA research in ecosystem management and biodiversity conservation.

Exploitation of environmental DNA (eDNA) for ecotoxicological research: A critical review on eDNA metabarcoding in assessing marine pollution.

The rise in worldwide population has led to a noticeable spike in the production, consumption, and transportation of energy and food, contributing to elevated environmental pollution. Marine pollution is a significant global environmental issue with ongoing challenges, including plastic waste, oil spills, chemical pollutants, and nutrient runoff, threatening marine ecosystems, biodiversity, and human health. Pollution detection and assessment are crucial to understanding the state of marine ecosystems. Conventional approaches to pollution evaluation usually represent laborious and prolonged physical and chemical assessments, constraining their efficacy and expansion. The latest advances in environmental DNA (eDNA) are valuable methods for the detection and surveillance of pollution in the environment, offering enhanced sensibility, efficacy, and involvement. Molecular approaches allow genetic information extraction from natural resources like water, soil, or air. The application of eDNA enables an expanded evaluation of the environmental condition by detecting both identified and unidentified organisms and contaminants. eDNA methods are valuable for assessing community compositions, providing indirect insights into the intensity and quality of marine pollution through their effects on ecological communities. While eDNA itself is not direct evidence of pollution, its analysis offers a sensitive tool for monitoring changes in biodiversity, serving as an indicator of environmental health and allowing for the indirect estimation of the impact and extent of marine pollution on ecosystems. This review explores the potential of eDNA metabarcoding techniques for detecting and identifying marine pollutants. This review also provides evidence for the efficacy of eDNA assessment in identifying a diverse array of marine pollution caused by oil spills, harmful algal blooms, heavy metals, ballast water, and microplastics. In this report, scientists can expand their knowledge and incorporate eDNA methodologies into ecotoxicological research.

Road mitigation structures designed for Texas ocelots: Influence of structural characteristics and environmental factors on non-target wildlife usage.

Roads negatively impact wildlife through habitat fragmentation, loss of habitat connectivity, and wildlife-vehicle collisions, thus road mitigation structures, such as wildlife crossing structures (WCS), wildlife guards (WG), and fencing are commonly used to address this issue all over the world, including in the United States. In South Texas, such structures were built or modified along a State Highway in an effort to address road mortality for the endangered ocelot (Leopardus pardalis) and non-target wildlife species. The goal of this study was to examine temporal changes in wildlife interactions with WCS and WG during and after their construction and modification along a South Texas highway and to determine whether environmental factors influenced use of WCS. Using camera traps deployed to monitor the road mitigation structures, we compared crossing rates, repel rates, and species richness of all species that interacted with the structures, and we examined whether differential wildlife use of WCS and WG was affected by one or more structural dimensions, distance to nearby vegetation, and water presence. Crossings through WCS by wildlife decreased following the completion of construction of mitigation structures; however, repel interactions at WG increased. Overall, crossings decreased at WCS that had higher openness ratios and during periods of precipitation and higher daily temperatures, but distance to vegetation had minimal influence. These factors were shown to influence crossings of each of the five most frequently observed species differently. Lastly, the presence of pooled water at one WCS caused a decrease in crossings when the water level was highest but was not a barrier at lower water levels. By examining influences on wildlife interaction with road mitigation structures, we conclude that a variety of structures, including different WCS configurations, can be beneficial in facilitating movement and restricting entry into the right-of-way for a diversity of wildlife species beyond the target species.

Paternò-Büchi Reaction Mass Spectrometry Enables Positional Assignment of Polymethylene-Interrupted Double Bonds in Food-Derived Lipids.

Fatty acids (FAs) containing polymethylene-interrupted (PMI) double bonds are a component of human foods; however, they present a significant analytical challenge for de novo identification. Covalent adduct chemical ionization and ozone-induced dissociation mass spectrometry (MS) methods enable unambiguous assignment of PMI-FA double bond positions, however, no method has been reported with electrospray ionization (ESI) platform using off-the-shelf systems. In the current work, we studied the Paternò-Büchi (PB) fragmentation patterns of PMI-FA and triacylglycerol (TG) by analyzing several known food sources. PB-MS/MS and MS3 enabled complete double bond location assignments, including the isolated double bond in PMI-FA and triacylglycerols. Sea urchin ("uni"), oyster, pine nut, and ginkgo nut were characterized for their signature PMI-FA, 20:2(5Z,11Z), 22:2(7Z, 15Z), 18:3(5Z,9Z,12Z), and 20:3(5Z,11Z,14Z), respectively. Quantitative analyses of the relative abundance of these PMI-FA led to results similar to reference methods. 18:3(5Z,9Z,12Z) was enriched at the sn-1/sn-3 position in pine nut major TG.

Interactive effects of hypoxia with estradiol-17ß on tryptophan hydroxylase activity and serotonin levels in the Atlantic croaker hypothalamus.

Hypoxia causes a marked decline in reproductive neuroendocrine function in Atlantic croaker due to decreases in the hypothalamic expression and activities of tryptophan hydroxylase (TPH, the rate limiting enzyme in serotonin synthesis) and aromatase. In the present study, the influence of the estrogen status on hypothalamic TPH and serotonin (5-HT) regulation by hypoxia (dissolved oxygen: 1.7 mg/L for 4 weeks) was investigated in croaker. Treatment in vivo with the aromatase inhibitor, ATD (1,4,6-androstatrien-3,17-dione), significantly decreased TPH activity, TPHs (TPH-1 and TPH-2) mRNAs expression, and 5-hydroxytryptophan (5-HTP, an immediate precursor of 5-HT) and 5-HT contents in croaker hypothalamus. Treatment with estradiol-17ß partially restored hypothalamic TPH activity, TPHs mRNA expression, and 5-HTP and 5-HT contents in hypoxia-exposed fish. These results suggest that the hypoxia-induced inhibition of TPH and 5-HT synthesis is dependent on the estrogen status. To our knowledge, this is the first report of a role for estrogens in modulating neural TPH and 5-HT responses to hypoxia in aquatic vertebrates.

Molecular and biochemical biomarkers in the American oyster Crassostrea virginica exposed to herbicide Roundup® at high temperature.

Aquatic organisms are frequently exposed to various environmental stressors. Thus, the effects of high temperatures and herbicides on aquatic organisms are a major subject of interest. In this study, we studied the effects of short-term exposure (1 week) to Roundup®, a glyphosate-based herbicide (concentrations: 0.5 and 5 µg/L), on the morphology of gills, digestive glands, and connective tissues, and the expression of heat shock protein-70 (HSP70, a chaperone protein), cytochrome P450 (CYP450, a biomarker of environmental contaminants), dinitrophenyl protein (DNP, a biomarker of protein oxidation), nitrotyrosine protein (NTP, a biomarker of protein nitration), antioxidant enzymes such as superoxidase dismutase (SOD) and catalase (CAT) in tissues of American oyster, Crassostrea virginica (Gmelin, 1791) maintained at high temperature (30 °C). Histological analyses showed an increase in mucous production in the gills and digestive glands, and in hemocyte aggregation in the connective tissues as well as a structural change of lumen in the digestive glands of oysters exposed to Roundup. Immunohistochemical and quantitative RT-PCR analyses showed significant (P < 0.05) increases in HSP70, CYP450, DNP, NTP, CAT, and SOD mRNA and protein expressions in the tissues of oysters exposed to Roundup. Taken together, these results suggest that exposure to Roundup at high temperature induces overproduction of reactive oxygen species/reactive nitrogen species which in turn leads to altered prooxidant-antioxidant activity in oyster tissues. Moreover, our results provide new information on protein oxidation/nitration and antioxidant-dependent mechanisms for HSP70 and CYP450 regulations in oysters exposed to Roundup at high temperature.

Roundup® disrupts tissue architecture, attenuates Na+/K+-ATPase expression, and induces protein oxidation/nitration, cellular apoptosis, and antioxidant enzyme expressions in the gills of goldfish, Carassius auratus.

Extensive agricultural activities to feed the growing population are one major driving force behind aquatic pollution. Different types of pesticides are used in farmlands to increase crop production and wash up into water bodies. Glyphosate-based herbicide Roundup® is one of the most used pesticides in the United States; however, its effects on teleost species are still poorly understood. This study focused on the effects of environmentally relevant concentrations of Roundup exposure (low- and high-dose: 0.5 and 5 µg/L for 2-week) on Na+/K+-ATPase (NKA, a biomarker for sodium­potassium ion pump efficacy), cytochrome P450-1A (CYP1A, a monooxygenase enzyme), 2,4-dinitrophenyl protein (DNP, a biomarker for protein oxidation), 3-nitrotyrosine protein (NTP, a biomarker for protein nitration), superoxidase dismutase (SOD, an antioxidant enzyme), catalase (CAT, an antioxidant enzyme) expressions, and cellular apoptosis in the gills of goldfish. Histopathological and in situ TUNEL analyses showed widespread tissue damage, including lamellar fusion, loss of gill architecture, club shape of primary lamellae, mucous formation, and distortion in the epithelium layer, as well as apoptotic nuclei in gills. Immunohistochemical and qRT-PCR analyses provided insights into the expressions of molecular indicators in gills. Fish exposed to Roundup exhibited a significant (P < 0.05) downregulation of NKA expression in gills. Additionally, we observed upregulation of CYP1A, DNP, NTP, SOD, and CAT expressions in the gills of goldfish. Overall, our results suggest that exposure to Roundup causes disruption of gill architecture, induces protein oxidation/nitration and cellular apoptosis, and alters prooxidant-antioxidant homeostasis in tissues, which may lead to reduced fitness and survivability of teleost species.

Molecular and biochemical responses to tributyltin (TBT) exposure in the American oyster: Triggers of stress-induced oxidative DNA damage and prooxidant-antioxidant imbalance in tissues by TBT.

Environmental pollution increases due to anthropogenic activities. Toxic chemicals in the environment affect the health of aquatic organisms. Tributyltin (TBT) is a toxic chemical widely used as an antifouling paint on boats, hulls, and ships. The toxic effect of TBT is well documented in aquatic organisms; however, little is known about the effects of TBT on DNA lesions in shellfish. The American oyster (Crassostrea virginica, an edible and commercially important species) is an ideal marine mollusk to examine the effects of TBT exposure on DNA lesions and oxidative/nitrative stress. In this study, we investigated the effects of TBT on 8'-hydroxy-2'-deoxyguanosine (8-OHdG, a biomarker of pro-mutagenic DNA lesion), double-stranded DNA (dsDNA), dinitrophenyl protein (DNP, a biomarker on reactive oxygen species, ROS), 3-nitrotyrosine protein (NTP, a biomarker of reactive nitrogen species, RNS), catalase (CAT, an antioxidant), and acetylcholinesterase (AChE, a cholinergic enzyme) expressions in the gills and digestive glands of oysters. We also analyzed extrapallial (EF) fluid conditions. Immunohistochemical and qRT-PCR results showed that TBT exposure significantly increased 8-OHdG, dsDNA, DNP, NTP, and CAT mRNA and/or protein expressions in the gills and digestive glands. However, AChE mRNA and protein expressions, and EP fluid pH and protein concentrations were decreased in TBT-exposed oysters. Taken together, these results suggest that antifouling biocide-induced production of ROS/RNS results in DNA damage, which may lead to decreased cellular functions in oysters. To the best of our knowledge, the present study provides the first molecular/biochemical evidence that TBT exposure results in oxidative/nitrative stress and DNA lesions in oysters.

Effects of elevated temperature on 8-OHdG expression in the American oyster (Crassostrea virginica): Induction of oxidative stress biomarkers, cellular apoptosis, DNA damage and γH2AX signaling pathways.

Global temperature is increasing due to anthropogenic activities and the effects of elevated temperature on DNA lesions are not well documented in marine organisms. The American oyster (Crassostrea virginica, an edible and commercially important marine mollusk) is an ideal shellfish species to study oxidative DNA lesions during heat stress. In this study, we examined the effects of elevated temperatures (24, 28, and 32 °C for one-week exposure) on heat shock protein-70 (HSP70, a biomarker of heat stress), 8­hydroxy-2'-deoxyguanosine (8-OHdG, a biomarker of pro-mutagenic DNA lesion), double-stranded DNA (dsDNA), γ-histone family member X (γH2AX, a molecular biomarker of DNA damage), caspase-3 (CAS-3, a key enzyme of apoptotic pathway) and Bcl-2-associated X (BAX, an apoptosis regulator) protein and/or mRNA expressions in the gills of American oysters. Immunohistochemical and qRT-PCR results showed that HSP70, 8-OHdG, dsDNA, and γH2AX expressions in gills were significantly increased at high temperatures (28 and 32 °C) compared with control (24°C). In situ TUNEL analysis showed that the apoptotic cells in gill tissues were increased in heat-exposed oysters. Interestingly, the enhanced apoptotic cells were associated with increased CAS-3 and BAX mRNA and/or protein expressions, along with 8-OHdG levels in gills after heat exposure. Moreover, the extrapallial (EP) fluid (i.e., extracellular body fluid) protein concentrations were lower; however, the EP glucose levels were higher in heat-exposed oysters. Taken together, these results suggest that heat shock-driven oxidative stress alters extracellular body fluid conditions and induces cellular apoptosis and DNA damage, which may lead to increased 8-OHdG levels in cells/tissues in oysters.