An interlaboratory study on the use of steroid hormones in examining endocrine disruption.

In recent years, there has been an increased use of the measurement of sex steroid hormone levels in the blood of animals exposed to chemicals as an indicator of reproductive impairment or an alteration in endocrine function. Although levels of hormones are often compared among animals and laboratories, there has been no study to examine the between-laboratory variability in actual steroid measurements. Therefore, we initiated a study with white sucker collected from a site receiving pulp mill effluent, previously documented as having reduced steroid levels, to address this issue. Samples of plasma and media from in vitro gonadal incubations were delivered to eight outside laboratories with the ability to measure steroid hormones. These laboratories ranged from well-established fish endocrine laboratories to wildlife toxicology laboratories, which have recently implemented the methods to measure steroid hormones. In this study, we have considered both the absolute measure of steroid content between laboratories as well as the ability to discriminate between reference and exposed populations as important criteria when evaluating the utility of these measures. Of the eight outside laboratories conducting the analyses, six detected identical site differences in circulating levels of testosterone and 17beta-estradiol to those documented by our Burlington laboratory (ON, Canada). However, the absolute value of the steroid hormones measured in the plasma varied significantly (plasma testosterone 0.6-23.1 ng/ml, 17beta-estradiol 77.6-1782.7 pg/ml) with coefficients of variation of 70.4% and 60.3% respectively. Similar results were demonstrated for the measurement of steroid hormones in media following in vitro gonadal incubation. Although there was a fair amount of variability in the absolute measure of steroid hormone levels, we would predict a far greater coherence of interlaboratory results through the sharing of reagents and the use of a common methodology between laboratories. These results are very promising, providing evidence for the inclusion of steroid hormones in monitoring endocrine disruption in wildlife species.

Capture and confinement stress in white sucker exposed to bleached kraft pulp mill effluent.

This study evaluates the effects of handling and confinement stress and a 3-day recovery period on a number of biochemical parameters used to monitor exposure of fish to bleached kraft mill effluent (BKME). Plasma was collected at four times of the day from male and female white sucker subjected to four levels of handling stress during their spawning migration at a BKME and a reference site. Indicators of a general response to stress (plasma cortisol, glucose, lactate, and total protein) and of reproductive fitness (plasma testosterone, 11-ketotestosterone, and 17 beta-estradiol) were measured. With the exception of 17 beta-estradiol in females, all of the parameters measured varied with time of day and stress level. The general stress indicators ranged from being highly to marginally responsive to handling and confinement stress and were inconsistent in terms of indicating a site difference. The reproductive steroids were moderately responsive to stress and gave the most consistent site difference with testosterone and 11-ketotestosterone levels reduced in BKME-exposed fish under low levels of stress. There was limited evidence for recovery from capture, handling, and transport in the spring for plasma cortisol, lactate, and 17 beta-estradiol at the BKME site, plasma glucose at the reference site, and plasma testosterone at both sites. Fish were also more responsive to an additional acute stress on Day 1 and Day 3 of recovery compared to that on Day 0. This study emphasizes the need for standardized methods in field collections and sampling with the least amount of stress possible, and suggests that holding white sucker for 1 or 3 days does not allow them to recover from the stress of capture.