An assessment of Hyalella azteca burrowing activity under laboratory sediment toxicity testing conditions Lorne E. Doig, Karsten Liber * Toxicology Centre, 44 Campus Drive, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5B3 article info Article history: Received 5 January 2010 Received in revised form 28 May 2010 Accepted 31 May 2010 Available online 29 June 2010 Keywords: Hyalella azteca Burrowing Sediment Toxicity test Amphipods abstract Burrowing of the freshwater amphipod Hyalella azteca was evaluated under laboratory conditions similar to those recommended for standard sediment toxicity testing in Canada (EPS 1/RM/33; Environment Can- ada, 1997) and the United States (EPA/600/R-99/064; US EPA, 2000). Sediment type, time of day (light versus dark), size of animal, and the presence or absence of food were varied to assess their effects on burrowing activity. Hyalella azteca were found to burrow rapidly in fine, organic-rich sediments, but were slower to burrow in a sandy sediment. There was no increase in the number of animals occupying the sediment surface of a fine, organic-rich sediment after 4 h of darkness compared to the previous 4 h of light. Over a 9- to 10-d duration, a higher percentage of animals occupied the surface of the sandy sed- iment. The addition of food promoted burrowing in sandy sediment, as did using smaller animals. Overall, longer-duration tests involving older animals and coarse sediments may require formal observation to confirm burrowing and ensure adequate sediment exposure. The addition of food during a test may pro- mote the burrowing of larger animals in coarse sediments, but may not be necessary in field-collected sediments that are not excessively sandy. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction Over the last two decades, Hyalella azteca, a freshwater talitrid amphipod, has become a routinely used organism in sediment tox- icity testing. The rationales for its use as an animal model in toxic- ity testing have been detailed in previous publications (e.g., Nebeker et al., 1984; Borgmann et al., 1989; Environmental Can- ada, 1997; US EPA, 2000; ASTM, 2002), but include that it is a rel- atively common, ubiquitous North American freshwater organism (Bousfield, 1958), it is relatively sensitive to a variety of contami- nants (Borgmann et al., 1989; Borgmann et al., 2005), and it is often naturally associated with sediments (Hargrave, 1970). However, there has been some debate as to the nature of H. azteca’s behavior both in the wild and under laboratory testing conditions (Wang et al., 2004). In general, H. azteca appears to be a habitat generalist, choosing to live not only in or on sediments (Hargrave, 1970), but also on other surfaces such as macrophytes (Humphreys, 1964; France, 1993), rocks (Winnell and Jude, 1987), and large detrital debris (Wienert, 1950). Wang et al. (2004) questioned the evidence supporting burrowing in the field and pointed to contradictory lab- oratory observations (largely anecdotal) where H. azteca have been seen occupying the sediment surfaces rather than burrowing into the sediment. If these animals do not burrow in sediment, their appropriateness as a benthic animal model in laboratory sediment toxicity/bioaccumulation testing would be questionable. However, routine visual observations made during our previous laboratory research (e.g., Doig and Liber, 2006) indicated that H. azteca do in- deed burrow into sediments under standard 10-d test conditions, but this behavior was not quantified. The objective of this research was therefore to quantify the degree to which H. azteca burrow into freshwater sediments under laboratory conditions and to eval- uate how common test variables influence burrowing behavior. The findings of this study are applicable not only to standard sed- iment toxicity testing protocols, but to the use of H. azteca in sed- iment toxicology research in general. 2. Methods and materials 2.1. Sediments and general test methods Using an Ekman grab sampler (15  15 cm), sediment was col- lected on July 9, 2005, from two sites (A and B) in Little Bear Lake (54°17 0 N, 104°41 0 W), an oligotrophic lake in north-central Sas- katchewan, Canada, which was known to be free from metal con- tamination. These sediments were placed in 20-L plastic pails and allowed to settle for 1 h before the surface water was carefully decanted. The sediment was brought back to the laboratory, homogenized, sieved to remove coarse debris (P2 mm), and refrig- 0045-6535/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.chemosphere.2010.05.054 * Corresponding author. Tel.: +1 306 966 7444; fax: +1 306 931 1664. E-mail address: karsten.liber@usask.ca (K. Liber). Chemosphere 81 (2010) 261–265 Contents lists available at ScienceDirect Chemosphere journal homepage: www.elsevier.com/locate/chemosphere