Contribution of transformation products towards the total herbicide toxicity to tropical marine organisms.

The toxicity of herbicide degradation (transformation) products is rarely taken into account, even though these are commonly detected in the marine environment, sometimes at concentrations higher than the parent compounds. Here we assessed the potential contribution of toxicity by transformation products of five photosystem II herbicides to coral symbionts (Symbiodinium sp.), the green algae Dunaliella sp., and prawn (Penaeus monodon) larvae. Concentration-dependent inhibition of photosynthetic efficiency (∆F/F m ') was observed for all herbicides in both microalgal species. The toxicity of solutions of aged diuron solutions containing transformation products to Symbiodinium sp. and Dunaliella sp. was greater than could be explained by the concentrations of diuron measured, indicating transformation products contributed to the inhibition of ∆F/F m '. However, the toxicity of aged atrazine, simazine, hexazinone, and ametryn solutions could be explained by the concentration of parent herbicide, indicating no contribution by transformation products. Prawn larval metamorphosis was not sensitive to the herbicides, but preliminary results indicated some toxicity of the transformation products of atrazine and diuron. Risk assessments should take into account the contribution of herbicide transformation products; however, further studies are clearly needed to test the toxicity of a far wider range of transformation products to a representative diversity of relevant taxa.

Degradation of Herbicides in the Tropical Marine Environment: Influence of Light and Sediment.

Widespread contamination of nearshore marine systems, including the Great Barrier Reef (GBR) lagoon, with agricultural herbicides has long been recognised. The fate of these contaminants in the marine environment is poorly understood but the detection of photosystem II (PSII) herbicides in the GBR year-round suggests very slow degradation rates. Here, we evaluated the persistence of a range of commonly detected herbicides in marine water under field-relevant concentrations and conditions. Twelve-month degradation experiments were conducted in large open tanks, under different light scenarios and in the presence and absence of natural sediments. All PSII herbicides were persistent under control conditions (dark, no sediments) with half-lives of 300 d for atrazine, 499 d diuron, 1994 d hexazinone, 1766 d tebuthiuron, while the non-PSII herbicides were less persistent at 147 d for metolachlor and 59 d for 2,4-D. The degradation of herbicides was 2-10 fold more rapid in the presence of a diurnal light cycle and coastal sediments; apart from 2,4-D which degraded more slowly in the presence of light. Despite the more rapid degradation observed for most herbicides in the presence of light and sediments, the half-lives remained > 100 d for the PS II herbicides. The effects of light and sediments on herbicide persistence were likely due to their influence on microbial community composition and its ability to utilise the herbicides as a carbon source. These results help explain the year-round presence of PSII herbicides in marine systems, including the GBR, but more research on the transport, degradation and toxicity on a wider range of pesticides and their transformation products is needed to improve their regulation in sensitive environments.

Diuron tolerance and potential degradation by pelagic microbiomes in the Great Barrier Reef lagoon.

Diuron is a herbicide commonly used in agricultural areas where excess application causes it to leach into rivers, reach sensitive marine environments like the Great Barrier Reef (GBR) lagoon and pose risks to marine life. To investigate the impact of diuron on whole prokaryotic communities that underpin the marine food web and are integral to coral reef health, GBR lagoon water was incubated with diuron at environmentally-relevant concentration (8 µg/L), and sequenced at specific time points over the following year. 16S rRNA gene amplicon profiling revealed no significant short- or long-term effect of diuron on microbiome structure. The relative abundance of prokaryotic phototrophs was not significantly altered by diuron, which suggests that they were largely tolerant at this concentration. Assembly of a metagenome derived from waters sampled at a similar location in the GBR lagoon did not reveal the presence of mutations in the cyanobacterial photosystem that could explain diuron tolerance. However, resident phages displayed several variants of this gene and could potentially play a role in tolerance acquisition. Slow biodegradation of diuron was reported in the incubation flasks, but no correlation with the relative abundance of heterotrophs was evident. Analysis of metagenomic reads supports the hypothesis that previously uncharacterized hydrolases carried by low-abundance species may mediate herbicide degradation in the GBR lagoon. Overall, this study offers evidence that pelagic phototrophs of the GBR lagoon may be more tolerant of diuron than other tropical organisms, and that heterotrophs in the microbial seed bank may have the potential to degrade diuron and alleviate local anthropogenic stresses to inshore GBR ecosystems.

Herbicide Persistence in Seawater Simulation Experiments.

Herbicides are detected year-round in marine waters, including those of the World Heritage listed Great Barrier Reef (GBR). The few previous studies that have investigated herbicide persistence in seawater generally reported half-lives in the order of months, and several studies were too short to detect significant degradation. Here we investigated the persistence of eight herbicides commonly detected in the GBR or its catchments in standard OECD simulation flask experiments, but with the aim to mimic natural conditions similar to those found on the GBR (i.e., relatively low herbicide concentrations, typical temperatures, light and microbial communities). Very little degradation was recorded over the standard 60 d period (Experiment 1) so a second experiment was extended to 365 d. Half-lives of PSII herbicides ametryn, atrazine, diuron, hexazinone and tebuthiuron were consistently greater than a year, indicating high persistence. The detection of atrazine and diuron metabolites and longer persistence in mercuric chloride-treated seawater confirmed that biodegradation contributed to the breakdown of herbicides. The shortest half-life recorded was 88 d for growth-regulating herbicide 2,4-D at 31°C in the dark, while the fatty acid-inhibitor metolachlor exhibited a minimum half-life of 281 d. The presence of moderate light and elevated temperatures affected the persistence of most of the herbicides; however, the scale and direction of the differences were not predictable and were likely due to changes in microbial community composition. The persistence estimates here represent some of the first appropriate data for application in risk assessments for herbicide exposure in tropical marine systems. The long persistence of herbicides identified in the present study helps explain detection of herbicides in nearshore waters of the GBR year round. Little degradation of these herbicides would be expected during the wet season with runoff and associated flood plumes transporting a high proportion of the original herbicide from rivers into the GBR lagoon.

Glyphosate persistence in seawater.

Glyphosate is one of the most widely applied herbicides globally but its persistence in seawater has not been reported. Here we quantify the biodegradation of glyphosate using standard "simulation" flask tests with native bacterial populations and coastal seawater from the Great Barrier Reef. The half-life for glyphosate at 25 °C in low-light was 47 days, extending to 267 days in the dark at 25 °C and 315 days in the dark at 31 °C, which is the longest persistence reported for this herbicide. AMPA, the microbial transformation product of glyphosate, was detected under all conditions, confirming that degradation was mediated by the native microbial community. This study demonstrates glyphosate is moderately persistent in the marine water under low light conditions and is highly persistent in the dark. Little degradation would be expected during flood plumes in the tropics, which could potentially deliver dissolved and sediment-bound glyphosate far from shore.

Phytotoxicity of four photosystem II herbicides to tropical seagrasses.

Coastal waters of the Great Barrier Reef (GBR) are contaminated with agricultural pesticides, including the photosystem II (PSII) herbicides which are the most frequently detected at the highest concentrations. Designed to control weeds, these herbicides are equally potent towards non-target marine species, and the close proximity of seagrass meadows to flood plumes has raised concerns that seagrasses may be the species most threatened by herbicides from runoff. While previous work has identified effects of PSII herbicides on the photophysiology, growth and mortality in seagrass, there is little comparative quantitative toxicity data for seagrass. Here we applied standard ecotoxicology protocols to quantify the concentrations of four priority PSII herbicides that inhibit photochemistry by 10, 20 and 50% (IC10, IC20 and IC50) over 72 h in two common seagrass species from the GBR lagoon. The photosystems of seagrasses Zosteramuelleri and Haloduleuninervis were shown to be generally more sensitive to the PSII herbicides Diuron, Atrazine, Hexazinone and Tebuthiuron than corals and tropical microalgae. The herbicides caused rapid inhibition of effective quantum yield (∆F/F m '), indicating reduced photosynthesis and maximum effective yields (Fv/Fm ) corresponding to chronic damage to PSII. The PSII herbicide concentrations which affected photosynthesis have been exceeded in the GBR lagoon and all of the herbicides inhibited photosynthesis at concentrations lower than current marine park guidelines. There is a strong likelihood that the impacts of light limitation from flood plumes and reduced photosynthesis from PSII herbicides exported in the same waters would combine to affect seagrass productivity. Given that PSII herbicides have been demonstrated to affect seagrass at environmental concentrations, we suggest that revision of environmental guidelines and further efforts to reduce PSII herbicide concentrations in floodwaters may both help protect seagrass meadows of the GBR from further decline.

Fatty acid profiles as a potential lipidomic biomarker of exposure to brevetoxin for endangered Florida manatees (Trichechus manatus latirostris).

Fatty acid signature analysis (FASA) is an important tool by which marine mammal scientists gain insight into foraging ecology. Fatty acid profiles (resulting from FASA) represent a potential biomarker to assess exposure to natural and anthropogenic stressors. Florida manatees are well studied, and an excellent necropsy program provides a basis against which to assess this budding tool. Results using samples from 54 manatees assigned to four cause-of-death categories indicated that those animals exposed to or that died due to brevetoxin exposure (red tide, or RT samples) demonstrate a distinctive hepatic fatty acid profile. Discriminant function analysis indicated that hepatic fatty acids could be used to classify RT versus non-RT liver samples with reasonable certainty. A discriminant function was derived based on 8 fatty acids which correctly classified 100% of samples from a training dataset (10 RT and 25 non-RT) and 85% of samples in a cross-validation dataset (5 RT and 13 non-RT). Of the latter dataset, all RT samples were correctly classified, but two of thirteen non-RT samples were incorrectly classified. However, the "incorrect" samples came from manatees that died due to other causes during documented red tide outbreaks; thus although the proximal cause of death was due to watercraft collisions, exposure to brevetoxin may have affected these individuals in ways that increased their vulnerability. This use of FASA could: a) provide an additional forensic tool to help scientists and managers to understand cause of death or debilitation due to exposure to red tide in manatees; b) serve as a model that could be applied to studies to improve assessments of cause of death in other marine mammals; and c) be used, as in humans, to help diagnose metabolic disorders or disease states in manatees and other species.

Testing the ecotoxicology of vegetable versus mineral based lubricating oils: 1. Degradation rates using tropical marine microbes.

Vegetable-derived lubricants (VDL) might be more biodegradable than mineral-derived lubricants (MDL) due to the absence of high molecular weight aromatics, but this remains largely untested in tropical conditions. In this laboratory study, the degradation rates of 2-stroke, 4-stroke and hydraulic VDLs were compared with their MDL counterparts in the presence of mangrove and coral reef microbial communities. While MDLs were comprised largely of unresolved saturated and some aromatic hydrocarbons, their VDL counterparts contained, potentially more degradable, fatty acid methyl esters. Degradation of some VDL was observed by day 7, with the 2-stroke VDL markedly consumed by mangrove microorganisms and the hydraulic VDL degraded by both microorganism communities after this short period. All of the VDL groups were significantly more degraded than the comparable MDLs mineral oil lubricants over 14 days in the presence of either mangrove or coral reef microbial communities. In general the mangrove-sourced microorganisms more efficiently degraded the lubricants than reef-sourced microorganisms.

Testing the ecotoxicology of vegetable versus mineral based lubricating oils: 2. Induction of mixed function oxidase enzymes in barramundi, Lates calcarifer, a tropical fish species.

An increasing number of vegetable-based oils are being developed as environmentally friendly alternatives to petroleum products. However, toxicity towards key tropical marine species has not been investigated. In this study we used laboratory-based biomarker induction experiments to compare the relative stress of a vegetable-based lubricating oil for marine 2-stroke engines with its mineral oil-based counterpart on tropical fish. The sub-lethal stress of 2-stoke outboard lubricating oils towards the fish Lates calcarifer (barramundi) was examined using liver microsomal mixed function oxidase (MFO) induction assays. This study is the first investigation into the use of this key commercial species in tropical North Queensland, Australia in stress assessment of potential hydrocarbon pollution using ethoxyresorufin O-deethylase (EROD) induction. Our results indicated that barramundi provide a wide range of inducible rates of EROD activity in response to relevant organic stressors. The vegetable- and mineral-based lubricants induced significant EROD activity at 1.0 mg kg(-1) and there was no significant difference between the two oil treatments at that concentration. At increasing concentrations of 2 and 3 mg kg(-1), the mineral-based lubricant resulted in slightly higher EROD activity than the vegetable-based lubricant. The EROD activity of control and treated barramundi are found to be within ranges for other species from temperate and tropical environments. These results indicate that vegetable-based lubricants may be less stressful to barramundi than their mineral counterparts at concentrations of lubricant > or =2 mg kg(-1). There is great potential for this species to be used in the biomonitoring of waterways around tropical North Queensland and SE Asia.

The ecotoxicology of vegetable versus mineral based lubricating oils: 3. Coral fertilization and adult corals.

Biodegradable vegetable-derived lubricants (VDL) might be less toxic to marine organisms than mineral-derived oils (MDL) due to the absence of high molecular weight aromatics, but this remains largely untested. In this laboratory study, adult corals and coral gametes were exposed to various concentrations of a two-stroke VDL-1A and a corresponding MDL to determine which lubricant type was more toxic to each life stage. In the fertilization experiment, gametes from the scleractinian coral Acropora microphthalma were exposed to water-accommodated fractions (WAF) of VDL-1A and MDL for four hours. The MDL and VDL-1A WAFs inhibited normal fertilization of the corals at 200 microg l(-1) total hydrocarbon content (THC) and 150 microg l(-1) THC respectively. Disturbance of a stable coral-dinoflagellate symbiosis is regarded as a valid measure of sub-lethal stress in adult corals. The state of the symbiosis in branchlets of adult colonies of Acropora formosa was monitored using indicators such as dinoflagellate expulsion and dark-adapted photosystem II yields of dinoflagellate (using pulse amplitude modulation fluorescence). An effect on symbiosis was measurable following 48 h exposure to the lubricants at concentrations of 190 microg l(-1) and 37 microg l(-1) THC for the MDL and VDL-1A respectively. GC/MS revealed that the main constituent of the VDL-1A WAF was the compound coumarin, added by the manufacturer to improve odour. The fragrance containing coumarin was removed from the lubricant formulation and the toxicity towards adult corals re-examined. The coumarin-free VDL-2 exhibited significantly less toxicity towards the adult corals than all of the other oil types tested, with the only measurable effect being a slight but significant drop in photosynthetic efficiency at 280 microg l(-1).