The study, published in the journal Environmental Science & Technology, focused on the water flea Daphnia magna, considered the lab rat of ecotoxicology because of its sensitivity to contaminants in its environment. The organism is commonly used by regulators to monitor freshwater toxicity, but the tests used typically look at levels of toxicity that will kill the water flea within 24 hours of exposure.
Those tests employ "a 'kill 'em and count 'em' technique that doesn't provide a great deal of insight into the mechanism of action," said Dr. Chris Vulpe, associate professor of nutritional sciences and toxicology and principal investigator of the study.
There also is a chronic toxicity test that assesses the impact of lower levels of exposure on reproduction, but again, exactly how the toxicant is affecting the organism is unclear, the researchers said.
But with toxicogenomics, scientists are hoping to understand toxicants based upon characteristic changes in an organism's gene expression. "By looking at the pattern of genes turned on and off in response to toxicants, we can get an idea of what is causing the toxicity," said Vulpe, who is also a member of the Berkeley Institute for the Environment on campus, which brings together diverse programs and units focused on environmental research. Vulpe worked with Helen Poynton, UC Berkeley graduate student in nutritional sciences and toxicology and lead author of the study.
In an effort to test the viability of gene expression assays in environmental toxicity screening, the researchers exposed the water flea to copper, cadmium and zinc, three metal contaminants that are commonly found in the environment, particularly in parts of California because of the state's history of mining. The metals are also used in industrial parts ranging from brake pads to batteries, and can be found in urban runoff.
For the study, the researchers chose sublethal exposure levels that are comparable to what is found in the environment.
For each metal, the researchers found a decrease in the expression of alpha amylase genes, which are needed to break down starch and as a result interferes with digestion. They also found that exposure to copper decreased the activity of genes that encode glucan binding proteins and lectins, which are possibly involved in the water flea's ability to recognize an infection.
"It's possible that the decrease in expression of these genes is responsible for the immune system suppression seen in other copper-exposed organisms," said Poynton.
Signs of oxidative stress were discovered when the water flea was exposed to cadmium. The researchers saw an increase in activity of genes related to glutathione-S-transferase and peroxiredoxins, both of which protect cells from oxidative damage.
Exposure to zinc led to a significant decrease in chitinase gene activity, the researchers found. They noted that chitinase is needed to break up the exoskeleton of crustaceans during molting, an activity necessary for growth and reproduction. The researchers followed up with a chronic toxicity test and found that exposure to high levels of zinc decreases reproduction rates for the water flea.
"Our study is one of the first proof-of-concepts that aquatic toxicogenomics is possible," said Poynton. "The extra information we get from looking at gene expression could help us make more informed decisions about how harmful a toxicant is, and it could give regulators a new direction that we should be pursuing in monitoring water quality. For instance, we could find that it's necessary to regulate toxicant levels at lower levels, so we can act before toxicants get to the level of actually killing a population. There are sublethal effects of these metal contaminants suggested by our data."
Toxicogenomics could also be used for chemical screening, the researchers said. "For those in industry, chemicals could be screened for potentially ecological consequences while they are still in development," said Poynton. "In pursuing 10 different chemicals for one application, it may be discovered that one is particularly toxic, so it can be ditched right away. At the same time, if screening reveals that there is little or no impact on gene expression from a particular chemical, why not pursue that one for commercial development?"
However, the researchers are also careful to acknowledge the limitations of relying upon gene expression as the sole indicator of ecotoxicity. "It remains to be seen whether a particular gene expression actually leads to adverse outcomes for the organism," said Vulpe. "Does the gene expression lead to actual changes biologically? Also, some changes may be adaptive, helping an organism survive. Just because a gene is changing isn't bad."
Nevertheless, the results of this study suggest that genomics can play a significant role in assessing the toxicity of potential environmental contaminants, the researchers said.
"A 24-hour acute assay won't tell you that you're messing up the feeding mechanisms of the water flea, and chronic tests only look at reproduction," said Vulpe. "What you really want to know is whether there is something that will impact an organism's ability to survive well, including its ability to eat, escape predators or fight infection. That is what genomics could do."
Other co-authors of the study include researchers from the Children's Hospital Oakland Research Institute, the University of New Hampshire, Eon/Terragenomics, and the U.S. Army Corps of Engineers.
The U.S. Environmental Protection Agency, National Science Foundation, National Institutes of Health and the U.S. Army Engineer Research and Development Center provided support for this research.