Growth of the harmful alga, Prymnesium parvum (Prymnesiophyceae), after gradual and abrupt increases in salinity.

Prymnesium parvum is a euryhaline, toxin-producing microalga. Although its abundance in inland waters and growth potential in the laboratory is reduced at high salinity (>20), the ability of inland strains to adjust their growth after long-term residence in high salinity is uncertain. An inland strain of P. parvum maintained at salinity of 5 in modified artificial seawater medium (ASM-5) was subjected to the following treatments over five sequential batch culture rounds: ASM-5 (control); modified ASM at salinity of 30, raised with NaCl; modified ASM at salinity incrementally increased to 30 with NaCl; and Instant Ocean® at salinity of 30 (IO-30). Exponential growth rate (r) was reduced when salinity was increased from 5 to 30 in ASM but returned to control values during the second round. When salinity was incrementally increased, a reduction in r still occurred when salinity reached 25-30. Maximum density was reduced at salinity of 30 in ASM upon abrupt transfer or incremental increase, and compensation did not occur. Growth performance in IO-30 was comparable to control values. In conclusion, (i) long-term compensation for acute inhibitory effects of high salinity occurred for r but not maximum density, (ii) incremental increases in salinity did not prevent growth inhibition, suggesting the existence of a salinity threshold of 25-30 for onset of salinity stress, and (iii) the presence of a seawater-like salt mixture prevented growth inhibition by high salinity. These findings provide new insights on P. parvum's long-term ability to adjust its growth in environments of different salinity and ionic composition.

Effects of thyroid endocrine manipulation on sex-related gene expression and population sex ratios in Zebrafish.

Thyroid hormone reportedly induces masculinization of genetic females and goitrogen treatment delays testicular differentiation (ovary-to-testis transformation) in genetic males of Zebrafish. This study explored potential molecular mechanisms of these phenomena. Zebrafish were treated with thyroxine (T4, 2nM), goitrogen [methimazole (MZ), 0.15mM], MZ (0.15mM) and T4 (2nM) (rescue treatment), or reconstituted water (control) from 3 to 33days postfertilization (dpf) and maintained in control water until 45dpf. Whole fish were collected during early (25dpf) and late (45dpf) testicular differentiation for transcript abundance analysis of selected male (dmrt1, amh, ar) and female (cyp19a1a, esr1, esr2a, esr2b) sex-related genes by quantitative RT-PCR, and fold-changes relative to control values were determined. Additional fish were sampled at 45dpf for histological assessment of gonadal sex. The T4 and rescue treatments caused male-biased populations, and T4 alone induced precocious puberty in ∼50% of males. Male-biased sex ratios were accompanied by increased expression of amh and ar and reduced expression of cyp19a1a, esr1, esr2a, and esr2b at 25 and 45dpf and, unexpectedly, reduced expression of dmrt1 at 45dpf. Goitrogen exposure increased the proportion of individuals with ovaries (per previous studies interpreted as delay in testicular differentiation of genetic males), and at 25 and 45dpf reduced the expression of amh and ar and increased the expression of esr1 (only at 25dpf), esr2a, and esr2b. Notably, cyp19a1a transcript was reduced but via non-thyroidal pathways (not restored by rescue treatment). In conclusion, the masculinizing activity of T4 at the population level may be due to its ability to inhibit female and stimulate male sex-related genes in larvae, while the inability of MZ to induce cyp19a1a, which is necessary for ovarian differentiation, may explain why its "feminizing" activity on gonadal sex is not permanent.

Thyroid endocrine disruption and external body morphology of Zebrafish.

This study examined the effects thyroid-active compounds during early development on body morphology of Zebrafish (Danio rerio). Three-day postfertilization (dpf) larvae were exposed to goitrogen [methimazole (MZ, 0.15mM)], combination of MZ (0.15mM) and thyroxine (T4, 2nM), T4 (2nM), or control (reconstituted water) treatments until 33dpf and subsequently maintained in reconstituted water until 45dpf. Samples were taken at 33 and 45dpf for multivariate analysis of geometric distances between selected homologous landmarks placed on digital images of fish, and for histological assessment of thyrocytes. Body mass, standard length, and pectoral fin length were separately measured on remaining fish at 45dpf. Histological analysis confirmed the hypothyroid effect (increased thyrocyte height) of MZ and rescue effect of T4 co-administration. Geometric distance analysis showed that pectoral and pelvic fins shifted backward along the rostrocaudal axis under hypothyroid conditions at 45dpf and that T4 co-treatment prevented this shift. Pectoral fin length at 45dpf was reduced by exposure to MZ and rescued by co-administration of T4, but it was not associated with standard length. Methimazole caused a reduction in body mass and length at 45dpf that could not be rescued by T4 co-administration, and non-thyroidal effects of MZ on body shape were also recognized at 33 and 45dpf. Alterations in the length and position of paired fins caused by exposure to thyroid-disrupting chemicals during early development, as shown here for Zebrafish, could affect physical aspects of locomotion and consequently other important organismal functions such as foraging, predator avoidance, and ultimately survival and recruitment into the adult population. Results of this study also suggest the need to include rescue treatments in endocrine disruption studies that rely on goitrogens as reference for thyroid-mediated effects.

Are endocrine and reproductive biomarkers altered in contaminant-exposed wild male Largemouth Bass (Micropterus salmoides) of Lake Mead, Nevada/Arizona, USA?

Male Largemouth Bass were sampled from two locations in Lake Mead (USA), a site influenced by treated municipal wastewater effluent and urban runoff (Las Vegas Bay), and a reference site (Overton Arm). Samples were collected in summer (July '07) and spring (March '08) to assess general health, endocrine and reproductive biomarkers, and compare contaminant body burdens by analyzing 252 organic chemicals. Sperm count and motility were measured in spring. Contaminants were detected at much higher frequencies and concentrations in fish from Las Vegas Bay than Overton Arm. Those with the highest concentrations included PCBs, DDTs, PBDEs, galaxolide, and methyl triclosan. Fish from Las Vegas Bay also had higher Fulton condition factor, hepatosomatic index, and hematocrit, and lower plasma 11-ketotestosterone concentration (KT). Gonadosomatic index (GSI) and sperm motility did not differ between sites, but sperm count was lower by nearly 50% in fish from Las Vegas Bay. A positive association between KT and GSI was identified, but this association was nonlinear. On average, maximal GSI was reached at sub-maximal KT concentrations. In conclusion, the higher concentration of contaminant body burdens coupled with reduced levels of KT and sperm count in fish from Las Vegas Bay suggest that male reproductive condition was influenced by contaminant exposures. Also, the nonlinear KT-GSI association provided a framework to understand why GSI was similar between male bass from both sites despite their large difference in KT, and also suggested the existence of post-gonadal growth functions of KT at high concentrations.

Novel associations between contaminant body burdens and biomarkers of reproductive condition in male Common Carp along multiple gradients of contaminant exposure in Lake Mead National Recreation Area, USA.

Adult male Common Carp were sampled in 2007/08 over a full reproductive cycle at Lake Mead National Recreation Area. Sites sampled included a stream dominated by treated wastewater effluent, a lake basin receiving the streamflow, an upstream lake basin (reference), and a site below Hoover Dam. Individual body burdens for 252 contaminants were measured, and biological variables assessed included physiological [plasma vitellogenin (VTG), estradiol-17ß (E2), 11-ketotestosterone (11KT)] and organ [gonadosomatic index (GSI)] endpoints. Patterns in contaminant composition and biological condition were determined by Principal Component Analysis, and their associations modeled by Principal Component Regression. Three spatially distinct but temporally stable gradients of contaminant distribution were recognized: a contaminant mixture typical of wastewaters (PBDEs, methyl triclosan, galaxolide), PCBs, and DDTs. Two spatiotemporally variable patterns of biological condition were recognized: a primary pattern consisting of reproductive condition variables (11KT, E2, GSI), and a secondary pattern including general condition traits (condition factor, hematocrit, fork length). VTG was low in all fish, indicating low estrogenic activity of water at all sites. Wastewater contaminants associated negatively with GSI, 11KT and E2; PCBs associated negatively with GSI and 11KT; and DDTs associated positively with GSI and 11KT. Regression of GSI on sex steroids revealed a novel, nonlinear association between these variables. Inclusion of sex steroids in the GSI regression on contaminants rendered wastewater contaminants nonsignificant in the model and reduced the influence of PCBs and DDTs. Thus, the influence of contaminants on GSI may have been partially driven by organismal modes-of-action that include changes in sex steroid production. The positive association of DDTs with 11KT and GSI suggests that lifetime, sub-lethal exposures to DDTs have effects on male carp opposite of those reported by studies where exposure concentrations were relatively high. Lastly, this study highlighted advantages of multivariate/multiple regression approaches for exploring associations between complex contaminant mixtures and gradients and reproductive condition in wild fishes.

Channel catfish (Ictalurus punctatus) leukocytes express estrogen receptor isoforms ERα and ERß2 and are functionally modulated by estrogens.

Estrogens are recognized as modulators of immune responses in mammals and teleosts. While it is known that the effects of estrogens are mediated via leukocyte-specific estrogen receptors (ERs) in humans and mice, leucocyte-specific estrogen receptor expression and the effects of estrogens on this cell population is less explored and poorly understood in teleosts. Here in, we verify that channel catfish (Ictalurus punctaus) leukocytes express ERα and ERß2. Transcripts of these isoforms were detected in tissue-associated leukocyte populations by PCR, but ERß2 was rarely detected in PBLs. Expression of these receptors was temporally regulated in PBLs following polyclonal activation by concanavalin A, lipopolysaccharide or alloantigen based on evaluation by quantitative and end-point PCR. Examination of long-term leukocyte cell lines demonstrated that these receptors are differentially expressed depending on leukocyte lineage and phenotype. Expression of ERs was also temporally dynamic in some leukocyte lineages and may reflect stage of cell maturity. Estrogens affect the responsiveness of channel catfish peripheral blood leukocytes (PBLs) to mitogens in vitro. Similarly, bactericidal activity and phorbol 12-myristate 13-acetate induced respiratory burst was modulated by 17ß-estradiol. These actions were blocked by the pure ER antagonist ICI 182780 indicating that response is, in part, mediated via ERα. In summary, estrogen receptors are expressed in channel catfish leukocytes and participate in the regulation of the immune response. This is the first time leukocyte lineage expression has been reported in teleost cell lines.

Regulation of gonadal sex ratios and pubertal development by the thyroid endocrine system in zebrafish (Danio rerio).

We examined associations between thyroid condition, gonadal sex and pubertal development in zebrafish. Seventy-two-hour postfertilization larvae were reared in untreated medium or in the presence of goitrogens (sodium perchlorate, 0.82 mM; methimazole, 0.15 and 0.3 mM) or thyroxine (1 and 10 nM) for 30 days. Thyrocyte height, gonadal sex and gonadal development were histologically determined at 45 and 60 days postfertilization (dpf). Thyrocyte hypertrophy, an index of hypothyroidism, was observed at 45 and 60 dpf in perchlorate-treated but only at 45 dpf in methimazole-treated fish. Similarly, gonadal sex ratios were biased toward ovaries relative to control animals at 45 and 60 dpf in perchlorate-treated fish but only at 45 dpf in methimazole-treated fish. Gonadal sex ratios were biased toward testes at 45 and 60 dpf in thyroxine-treated fish. Spermatogenesis was delayed in testes from goitrogen-treated fish at 60 dpf relative to control values, but was unaffected in testes from thyroxine-treated individuals. Oogenesis seemed to be nonspecifically delayed in all treatments relative to control at 60 dpf. This study confirmed the previously reported association between hypothyroid condition and ovarian-skewed ratios, and hyperthyroid condition and testicular-skewed ratios, and also showed that male pubertal development is specifically delayed by experimental hypothyroidism. The simultaneous recovery from the hypothyroid and ovary-inducing effects of methimazole by 60 dpf (27 days post-treatment) suggests that the ovary-skewing effect of goitrogens is reversible when thyroid conditions return to basal levels before developmental commitment of gonadal sex. Conversely, the masculinizing effect of hyperthyroidism seems to be stable and perhaps permanent.

Effects of dietary exposure to brominated flame retardant BDE-47 on thyroid condition, gonadal development and growth of zebrafish.

Little is known about the effects of brominated flame retardants in teleosts and some of the information currently available is inconsistent. This study examined effects of dietary exposure to 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) on thyroid condition, body mass and size, and gonadal development of zebrafish. Pubertal, 49-day-old (posthatch) fish were fed diets without BDE-47 (control) or with 1, 5 or 25 µg/g BDE-47/diet. Treatments were conducted in triplicate 30-L tanks each containing 50 zebrafish, and 15 fish per treatment (5 per tank) were sampled at days 40, 80 and 120 of exposure. Measurements were taken of body mass, standard length, head depth and head length. Sex (at 40-120 days of exposure), germ cell stage (at 40 days) and thyroid condition (at 120 days; follicular cell height, colloid depletion, angiogenesis) were histologically determined. Whole-body BDE-47 levels at study completion were within the high end of levels reported in environmentally exposed (wild) fishes. Analysis of variance was used to determine differences among treatments at each sampling time. No effects were observed on thyroid condition or germ cell stage in either sex. Reduced head length was observed in females exposed to BDE-47 at 80 days but not at 40 or 120 days. In males, no apparent effects of BDE-47 were observed at 40 and 80 days, but fish exposed to 25 µg/g had lower body mass at 120 days compared to control fish. These observations suggest that BDE-47 at environmentally relevant whole-body concentrations does not affect thyroid condition or pubertal development of zebrafish but does affect growth during the juvenile-to-adult transition, especially in males.

Air, land, and water variables associated with the first appearance and current spatial distribution of toxic Prymnesium parvum blooms in reservoirs of the Southern Great Plains, USA.

This study examined the association of air, land, and water variables with the first historical occurrence and current distribution of toxic Prymnesium parvum blooms in reservoirs of the Brazos River and Colorado River, Texas (USA). One impacted and one reference reservoir were selected per basin. Land cover and use variables were estimated for the whole watershed (WW) and a 0.5-km zone on either side of streams (near field, NF). Variables were expressed in annual values. Principal component and trend analyses were used to determine (1) differences in environmental conditions before and after the 2001 onset of toxic blooms in impacted reservoirs (study period, 1992-2017), and (2) traits that uniquely discriminate impacted from reference reservoirs (2001-2017). Of thirty-three variables examined, two positively aligned with the reoccurring appearance of blooms in impacted reservoirs (air CO2 and herbicide Glyphosate) and another two negatively aligned (insecticides Terbufos and Malathion). Glyphosate use was observed throughout the study period but a turning point for an upward trend occurred near the year of first bloom occurrence. While the relevance of the decreased use of insecticides is uncertain, prior experimental studies reported that increasing concentrations of air CO2 and water Glyphosate can enhance P. parvum growth. Consistent with prior findings, impacted reservoirs were of higher salinity than reference reservoirs. In addition, their watersheds had far lower wetland cover at NF and WW scales. The value of wetlands in reducing harmful algal bloom incidence by reducing nutrient inputs has been previously recognized, but wetlands can also capture pesticides. Therefore, a diminished wetland cover could magnify Glyphosate loads flowing into impacted reservoirs. These observations are consistent with a scenario where rising levels of air CO2 and Glyphosate use contributed to the establishment of P. parvum blooms in reservoirs of relatively high salinity and minimal wetland cover over their watersheds.

The hypothalamus-pituitary-thyroid axis in teleosts and amphibians: endocrine disruption and its consequences to natural populations.

Teleosts and pond-breeding amphibians may be exposed to a wide variety of anthropogenic, waterborne contaminants that affect the hypothalamus-pituitary-thyroid (HPT) axis. Because thyroid hormone is required for their normal development and reproduction, the potential impact of HPT-disrupting contaminants on natural teleost and amphibian populations raises special concern. There is laboratory evidence indicating that persistent organic pollutants, heavy metals, pharmaceutical and personal care products, agricultural chemicals, and aerospace products may alter HPT activity, development, and reproduction in teleosts and amphibians. However, at present there is no evidence to clearly link contaminant-induced HPT alterations to impairments in teleost or amphibian population health in the field. Also, with the exception of perchlorate for which laboratory studies have shown a direct link between HPT disruption and adverse impacts on development and reproductive physiology, little is known about if or how other HPT-disrupting contaminants affect organismal performance. Future field studies should focus on establishing temporal associations between the presence of HPT-disrupting chemicals, the occurrence of HPT alterations, and adverse effects on development and reproduction in natural populations; as well as determining how complex mixtures of HPT contaminants affect organismal and population health.

Movement of synthetic organic compounds in the food web after the introduction of invasive quagga mussels (Dreissena bugensis) in Lake Mead, Nevada and Arizona, USA.

Introductions of dreissenid mussels in North America have been a significant concern over the last few decades. This study assessed the distribution of synthetic organic compounds (SOCs) in the food web of Lake Mead, Nevada/Arizona, USA and how this distribution was influenced by the introduction of invasive quagga mussels. A clear spatial gradient of SOC concentrations in water was observed between lake basins downstream of populated areas and more rural areas. Within the food web, trophic magnification factors (TMF) indicated statistically significant biomagnification for nine, and biodilution for two, of 22 SOCs examined. The highest value recorded was for PCB 118 (TMF, 5.14), and biomagnification of methyl triclosan (TMF, 3.85) was also apparent. Biodilution was observed for Tonalide® (0.06) and Galaxolide® (0.38). Total SOC concentration in quagga mussels was higher than in three pelagic fishes. Also, 19 of 20 SOC examined in Largemouth Bass (Micropterus salmoides) had substantially lower concentrations in 2013, when quagga mussels had become well established, than in 2007/08, soon after quagga mussels were introduced. Estimates of SOC concentrations in the water column and quagga mussels suggest that a considerable portion (~10.5%) of the SOC mass in the lake has shifted from the pelagic to the benthic environments due to quagga mussel growth. These observations suggest that benthic species, such as the endangered Razorback Sucker, may be experiencing increased risk of SOC exposure. In addition, stable isotope analysis (carbon and nitrogen) indicated a decrease in the nutritional value of zooplankton to consumers (e.g., Razorback Sucker larvae) since quagga mussels became established. These changes could affect Razorback Sucker larval survival and recruitment. Results from this study strongly suggest that the introduction of quagga mussels has greatly altered the dynamics of SOCs and other processes in the food web of Lake Mead.

Sources and distribution of organic compounds using passive samplers in Lake Mead national recreation area, Nevada and Arizona, and their implications for potential effects on aquatic biota.

The delineation of lateral and vertical gradients of organic contaminants in lakes is hampered by low concentrationsand nondetection of many organic compounds in water. Passive samplers (semipermeable membrane devices [SPMDs] and polar organic chemical integrative samplers [POCIS]) are well suited for assessing gradients because they can detect synthetic organic compounds (SOCs) at pg L(-1) concentrations. Semi-permeable membrane devices and POCIS were deployed in Lake Mead, at two sites in Las Vegas Wash, at four sites across Lake Mead, and in the Colorado River downstream from Hoover Dam. Concentrations of hydrophobic SOCs were highest in Las Vegas Wash downstream from waste water and urban inputs and at 8 m depth in Las Vegas Bay (LVB) where Las Vegas Wash enters Lake Mead. The distribution of hydrophobic SOCs showed a lateral distribution across 10 km of Lake Mead from LVB to Boulder Basin. To assess possible vertical gradient SOCs, SPMDs were deployed at 4-m intervals in 18 m of water in LVB. Fragrances and legacy SOCs were found at the greatest concentrations at the deepest depth. The vertical gradient of SOCs indicated that contaminants were generally confined to within 6 m of the lake bottom during the deployment interval. The high SOC concentrations, warmer water temperatures, and higher total dissolved solids concentrations at depth are indicative of a plume of Las Vegas Wash water moving along the lake bottom. The lateral and vertical distribution of SOCs is discussed in the context of other studies that have shown impaired health of fish exposed to SOCs.

Growth response of the ichthyotoxic haptophyte, Prymnesium parvum Carter, to changes in sulfate and fluoride concentrations.

Golden alga Prymnesium parvum Carter is a euryhaline, ichthyotoxic haptophyte (Chromista). Because of its presumed coastal/marine origin where SO42- levels are high, the relatively high SO42- concentration of its brackish inland habitats, and the sensitivity of marine chromists to sulfur deficiency, this study examined whether golden alga growth is sensitive to SO42- concentration. Fluoride is a ubiquitous ion that has been reported at higher levels in golden alga habitat; thus, the influence of F- on growth also was examined. In low-salinity (5 psu) artificial seawater medium, overall growth was SO42-dependent up to 1000 mg l-1 using MgSO4 or Na2SO4 as source; the influence on growth rate, however, was more evident with MgSO4. Transfer from 5 to 30 psu inhibited growth when salinity was raised with NaCl but in the presence of seawater levels of SO42-, these effects were fully reversed with MgSO4 as source and only partially reversed with Na2SO4. Growth inhibition was not observed after acute transfer to 30 psu in a commercial sea salt mixture. In 5-psu medium, F- inhibited growth at all concentrations tested. These observations support the hypothesis that spatial differences in SO42- -but not F--concentration help drive the inland distribution and growth of golden alga and also provide physiological relevance to reports of relatively high Mg2+ concentrations in golden alga habitat. At high salinity, however, the ability of sulfate to maintain growth under osmotic stress was weak and overshadowed by the importance of Mg2+. A mechanistic understanding of growth responses of golden alga to SO42-, Mg2+ and other ions at environmentally relevant levels and under different salinity scenarios will be necessary to clarify their ecophysiological and evolutionary relevance.

Environmental regulation of sex determination in fishes: Insights from Atheriniformes.

Sex determination is the first step toward the establishment of phenotypic sex in most vertebrates. Aquatic poikilotherms such as teleost fishes exhibit a high diversity of sex-determination mechanisms and gonadal phenotypes that are remarkably plastic and responsive to a variety of environmental factors (e.g., water temperature, pH, salinity, photoperiod, population density). This chapter reviews current knowledge of genotypic and environmental sex determination systems in fishes with special reference to Atheriniformes-one of the best-characterized taxa in this field-and offers perspectives to guide and stimulate further research.

Brain aromatase (Cyp19A2) and estrogen receptors, in larvae and adult pejerrey fish Odontesthes bonariensis: Neuroanatomical and functional relations.

Although estrogens exert many functions on vertebrate brains, there is little information on the relationship between brain aromatase and estrogen receptors. Here, we report the cloning and characterization of two estrogen receptors, alpha and beta, in pejerrey. Both receptors' mRNAs largely overlap and were predominantly expressed in the brain, pituitary, liver, and gonads. Also brain aromatase and estrogen receptors were up-regulated in the brain of estradiol-treated males. In situ hybridization was performed to study in more detail, the distribution of the two receptors in comparison with brain aromatase mRNA in the brain of adult pejerrey. The estrogen receptors' mRNAs exhibited distinct but partially overlapping patterns of expression in the preoptic area and the mediobasal hypothalamus, as well as in the pituitary gland. Moreover, the estrogen receptor alpha, but not beta, were found to be expressed in cells lining the preoptic recess, similarly as observed for brain aromatase. Finally, it was shown that the onset expression of brain aromatase and both estrogen receptors in the head of larvae preceded the morphological differentiation of the gonads. Because pejerrey sex differentiation is strongly influenced by temperature, brain aromatase expression was measured during the temperature-sensitive window and was found to be significantly higher at male-promoting temperature. Taken together these results suggest close neuroanatomical and functional relationships between brain aromatase and estrogen receptors, probably involved in the sexual differentiation of the brain and raising interesting questions on the origin (central or peripheral) of the brain aromatase substrate.

Effects of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in zebrafish: general and reproductive toxicity.

Mixed-sex populations of young adult zebrafish (~2-month-old) were exposed to measured RDX concentrations of 0, 1 or 9.6 ppm for up to 12 weeks followed by a 15-day rearing period in untreated water. RDX caused high mortality at 9.6 ppm, with most deaths occurring within the first 8 weeks of exposure. RDX at 9.6 ppm caused lower body weights at 4 and 8 weeks of exposure; and at 1 ppm, lower body weight was observed only at 4 weeks. Fish length was not affected by treatment at any time during the exposure period. The bioconcentration factor for RDX seemed to be influenced by time of exposure but not by water RDX concentration; its overall values were 1.01+/-0.13, 0.91+/-0.06 and 2.23+/-0.04 at 4, 8 weeks and 12 weeks, respectively. RDX was not detected in fish collected after the 15-day recovery period. In a separate experiment, adult females and males were separately exposed to RDX at measured concentrations of 0, 0.5 and 3.2 ppm for a period of 6 weeks. Reproductive performance was evaluated by biweekly breeding of the fish and measuring packed-egg volume (PEV) as index of fecundity. At 0.5 ppm, RDX caused elevated PEV levels relative to the control value at 2 weeks but not at 4 or 6 weeks, whereas no significant effects were noted at 3.2 ppm. Egg fertilization and embryo hatching rates were not affected by RDX at any of the concentrations tested. In conclusion, RDX at sublethal concentrations causes short-term negative effects on growth and, at 0.5 ppm, positive effects on fecundity.

Low-dose stimulation of growth of the harmful alga, Prymnesium parvum, by glyphosate and glyphosate-based herbicides.

Glyphosate-based herbicides (GBH) are widely used around the globe. While generally toxic to phototrophs, organic phosphorus in glyphosate can become available to glyphosate-resistant phytoplankton and contribute to algal bloom development. Few studies have examined the effects of GBH on growth of eukaryotic microalgae and information for the toxic bloom-forming haptophyte, Prymnesium parvum, is limited. Using a batch-culture system, this study examined the effects on P. parvum growth of a single application of Roundup Weed and Grass Killer Super Concentrate Plus® (Roundup SC), Roundup Weed and Grass Killer Ready-to-Use III® (Roundup RtU), and technical-grade glyphosate at low concentrations [0-1000 µg glyphosate acid equivalent (ae) l-1]. Roundup formulations differ in the percent of glyphosate as active ingredient (Roundup SC, ∼50%; Roundup RtU, 2%), allowing indirect evaluation of the influence of inactive ingredients. Roundup SC enhanced exponential growth rate at 10-1000 µg glyphosate ae l-1, and a positive monotonic association was noted between Roundup SC concentration and early (pre-exponential growth) but not maximum cell density. Glyphosate and both Roundup formulations enhanced growth rate at 100 µg glyphosate l-1, but only Roundup SC and glyphosate significantly stimulated early and maximum density. This observation suggests the higher concentration of inactive ingredients and other compounds in Roundup RtU partially counteracts glyphosate-dependent growth stimulation. When phosphate concentration was varied while maintaining other conditions constant, addition of Roundup SC and glyphosate at 100 µg l-1 influenced growth more strongly than equivalent changes in phosphate-associated phosphorus. It appears, therefore, that low doses of glyphosate stimulate growth by mechanisms unrelated to the associated small increases in total phosphorus. In conclusion, glyphosate and GBH stimulate P. parvum growth at low, environmentally relevant concentrations. This finding raises concerns about the potential contribution to P. parvum blooms by glyphosate-contaminated runoff or by direct application of GBH to aquatic environments.

Growth-suppressing and algicidal properties of an extract from Arundo donax, an invasive riparian plant, against Prymnesium parvum, an invasive harmful alga.

This study examined the ability of acidic and neutral/alkaline fractions of a methanolic extract from giant reed (Arundo donax) and of two of its constituents, gramine and skatole, to inhibit growth of the ichthyotoxic golden alga (Prymnesium parvum) in batch culture. For this study, growth suppression was defined as inhibition of maximum cell density, algicidal activity as early occurrence of negative growth, and algistatic activity as lack of net growth. The acidic fraction did not affect algal growth. The neutral/alkaline fraction showed growth-suppressing and algicidal activities but no signs of algistatic activity - namely, cells in cultures surviving a partial-algicidal exposure concentration (causing transient negative growth) were later able to initiate positive growth but at higher concentrations, algicidal activity was full and irreversible. Gramine suppressed growth more effectively than skatole and at the highest concentration tested, gramine also showed partial-algicidal and algistatic activity. While the partial-algicidal activities of the neutral/alkaline fraction and of gramine were short-lived (≤6days) and thus may share similar mechanisms, algistatic activity was unique to gramine and persisted for >3 weeks. Given gramine's reported concentration in the neutral/alkaline fraction, its corresponding level of algicidal activity is much lower than the fraction's suggesting the latter contains additional potent algicides. Inhibition of maximum cell density by all test compounds was associated with reductions in exponential growth rate, and in the case of the neutral/alkaline fraction and gramine also reductions in early (pre-exponential) growth. These results indicate that giant reed is a potential source of natural products to control golden alga blooms. Giant reed is an invasive species in North America, thus also providing incentive for research into strategies to couple management efforts for both species.

Effects of prolonged exposure to perchlorate on thyroid and reproductive function in zebrafish.

The objectives of this study were to determine the effects of prolonged exposure to perchlorate on (1) thyroid status and reproductive performance of adult zebrafish (Danio rerio) and (2) F1 embryo survival and early larval development. Using a static-renewal procedure, mixed sex populations of adult zebrafish were exposed to 0, 10, and 100 mg/l nominal concentrations of waterborne perchlorate for 10 weeks. Thyroid histology was qualitatively assessed, and females and males were separated and further exposed to their respective treatments for six additional weeks. Eight females in each tank replicate (n = 3) were paired weekly with four males from the same respective treatment, and packed-egg (spawn) volume (PEV) was measured each of the last five weeks. At least once during weeks 14-16 of exposure, other end points measured included fertilization rate, fertilized egg diameter, hatching rate, standard length, and craniofacial development of 4-day-postfertilization larvae and thyroid hormone content of 3.5-h embryos and of exposed mothers. At 10 weeks of exposure, perchlorate at both concentrations caused thyroidal hypertrophy and colloid depletion. A marked reduction in PEV was observed toward the end of the 6-week spawning period, but fertilization and embryo hatching rates were unaffected. Fertilized egg diameter and larval length were increased by parental exposure to perchlorate. Larval head depth was unaffected but the forward protrusion of the lower jaw-associated cartilage complexes, Meckel's and ceratohyal, was decreased. Exposure to both concentrations of perchlorate inhibited whole-body thyroxine content in mothers and embryos, but triiodothyronine content was unchanged. In conclusion, prolonged exposure of adult zebrafish to perchlorate not only disrupts their thyroid endocrine system but also impairs reproduction and influences early F1 development.