Pseudoreplication and the usage of biomarkers in ecotoxicological bioassays.

Pseudoreplication is a widely discussed topic in the scientific community. Its principal critique concerns the lack of independence in flawed experimental designs and the use of inferential statistics to test the hypothesis of such experiments. Thirty years after its appearance, it remains misunderstood by many researchers, including ecotoxicologists. In the present study, we try to clarify some of its concepts by filling in what seems to be a gap in the terminology of manipulative experiments. We propose the term "experimental medium" to refer strictly to the relevant spatial scale of the experiment to preserve the specificity of the experimental and observational units and to display pseudoreplication as a kind of misinterpretation and/or misanalysis of inferential statistics. A classification of the types of experimental designs in ecotoxicology is offered, and the problems in using inferential statistics in suboptimal designs are discussed. We hope to shed some light on such a classic topic for ecotoxicologists. Environ Toxicol Chem 2017;36:2868-2874. © 2017 SETAC.

A multibiomarker evaluation of urban, industrial, and agricultural exposure of small characins in a large freshwater basin in southern Brazil.

Iguaçu River is the second most polluted river of Brazil. It receives agrochemicals and contaminants of urban and industrial sources along its course. A multibiomarker approach was employed here to evaluate the health of a small characin (Astyanax spp.) at two sites along the river, sampled during a dry (autumn) and a rainy (spring) season. Biomarkers were condition factor and somatic indices (gonads and liver); genetic damage (comet assay and micronucleus test); enzyme activities such as hepatic catalase (CAT) and glutathione S-transferase (GST), lipoperoxidation (LPO), branchial and renal carbonic anhydrase (CA), acetylcholinesterase (AChE) in the muscle and the brain, histopathology of the liver and gills, and concentrations of polycyclic aromatic hydrocarbons (PAHs) in bile. There were no consistent differences in biomarker responses between the two study sites. Some biomarkers revealed greater potential impact in the rainy season, when increased amounts of contaminants are washed into the river (combined CAT inhibition and LPO increase, CA upregulation). Other biomarkers, however, revealed potential greater impact in the dry season, when contaminants potentially concentrate (GST induction, AChE inhibition, and liver histopathological alterations). Although of a complex nature, field experiments such as this provide rich data for monitoring protocols and assessment of general risk of exposure to pollutants of river systems.