2171 Environmental Toxicology and Chemistry, Vol. 26, No. 10, pp. 2171–2178, 2007  2007 SETAC Printed in the USA 0730-7268/07 $12.00 + .00 USE OF THE AUSTRALIAN CRIMSON-SPOTTED RAINBOWFISH (MELANOTAENIA FLUVIATILIS) AS A MODEL TEST SPECIES FOR INVESTIGATING THE EFFECTS OF ENDOCRINE DISRUPTORS CARMEL A. POLLINO,† EUGENE GEORGIADES,‡ and DOUGLAS A. HOLDWAY*§ †Integrated Catchment Assessment and Management Centre, Fenner School of Environment and Society, Australian National University, Canberra, Australian Capitol Territory 0200, Australia ‡Department of Biotechnology and Environmental Biology, RMIT University, Bundoora Campus, PO Box 71, Bundoora, Victoria 3083, Australia §Faculty of Science, University of Ontario Institute of Technology, Oshawa, Ontario L1H 7K4, Canada ( Received 3 December 2006; Accepted 26 April 2007) Abstract—Few studies have investigated the potential reproductive effects of toxicants on Australian freshwater fish species. The present study uses the Australian rainbowfish (Melanotaenia fluviatilis) as a model for testing the potential effects of 17-estradiol. Groups of reproductively active rainbowfish were exposed to waterborne 17-estradiol (control, carrier control, and 30, 100, 300, and 1,000 ng/L) for 3- and 14-d periods. Biomarkers of both low ecological relevance (plasma estradiol and testosterone, phos- phoprotein, -glutamyltranspeptidase [GTP]) and high ecological relevance (egg counts, hatchability, larval lengths, histopathology) were measured and the relationships between these examined. Relative to the controls, exposed female rainbowfish had a decline in plasma estradiol. An increase in phosphoprotein (vitellogenin) also was observed after exposure to 1,000 ng/L after 3 d, and histological examination of the gonads showed an increase in oocyte atresia at 1,000 ng/L on days 3 and 14. Changes in egg production were observed at 300 and 1,000 ng/L. No changes were detected to egg hatchability and larval lengths of offspring. Although there were no changes in male rainbowfish plasma testosterone or the histological organization of testes, levels of phosphoprotein increased and GTP activities were increased after 3 d of exposure at all test concentrations, but these declined at day 14. The present study demonstrated that, though measurements at the biochemical level were responsive to 17-estradiol exposure, fewer changes were observed in markers of higher ecological relevance at the exposures concentrations and durations tested. Keywords—Estradiol Crimson-spotted rainbowfish Sex steroids Phosphoprotein -Glutamyltranspeptidase INTRODUCTION Endocrine disruptors are exogenous compounds that inter- fere with the biosynthesis, transport, binding, action, or me- tabolism of endogenous hormones [1]. In endocrine disruptor studies, 17-estradiol has been used as a positive control [2,3]; however, some studies have shown that it also can be regarded as an aquatic contaminant [4–6]. Sewage effluent often has a high content of natural estrogens that have the potential to impact aquatic wildlife [4]. In the United Kingdom, measure- ments of estradiol in sewage effluents can range up to 100 ng/L [5], and concentrations up to 141 ng/L have been mea- sured in the Israeli environment [6]. In Australia, sewage ef- fluent exposure altered the reproductive morphology of male mosquitofish in a tributary of the Hawksbury-Nepean River system in New South Wales [7]. The increased use of recycled water for irrigation in Australia also has the potential to result in increased levels of endocrine disruptors in natural water- ways [8]. Recently in Australia, there has been growing interest in the use of recycled sewage for meeting industry and drinking water needs, as well as for meeting environmental flow re- quirements [9]. Such needs are becoming more urgent given the recent severe drought, with deficiencies in rainfalls being experienced in southern, eastern, and western parts of Aus- tralia. Between August and December 2006, many areas ex- * To whom correspondence may be addressed: (douglas.holdway@uoit.ca). perienced their lowest rainfalls on record (http://www.bom. gov.au). Research examining the hormone-like effects of synthetic chemicals upon fish and their interference with the endocrine system has grown considerably; however, little research using fish species native to the Southern Hemisphere has been con- ducted. The present study examined the Australian crimson- spotted rainbowfish (Melanotaenia fluviatilis) as a model for testing endocrine-disrupting substances. Melanotaenia fluvia- tilis are found throughout the inland Murray-Darling system [10]. This system extends across four states of Australia and the Basin predominantly receives rural runoff, but urban cen- tres located throughout the Basin discharge into the local wa- terways [11]. The crimson-spotted rainbowfish is an ideal test species because they are sexually dimorphic and can reproduce at approximately three months of age. This species is ideal for measuring the effects of xenobiotics and has been used pre- viously in ecotoxicological studies [12,13]. In addition, a base- line study of the reproductive cycle of M. fluviatilis has been conducted previously [14]. Contaminants in the ecosystem can elicit a series of bio- logical responses ranging from changes at the biochemical level, to changes at the population and community levels [15]. Although biochemical responses generally are considered to have low ecological relevance, they are short-term measure- ments that can signify preliminary xenobiotic effects before the appearance of gross lesions, reproductive impairment, or