Monitoring sea lice (Lepeophtheirus salmonis) sensitivity to emamectin benzoate in Atlantic salmon (Salmo salar)

The main objectives of thesis are: (1) to describe sea lice management on the east coast of Canada in the absence of standardized methods for monitoring sea lice abundance on Atlantic salmon production sites, (2) to develop and optimize field bioassays to monitor for changes in sea lice sensitivity toward Emamectin Benzoate (EMB), and (3) to optimize and use monooxygenase testing to describe enzyme activity d collected sea lice populations with exposure to EMB.

Reports from Europe of reduced sensitivity and resistance development of sea lice towards several chemotherapeutants necessitate the development of resistance management strategies, including methods to monitor changes in sensitivity to treatments. A bioassay for sea lice sensitivity towards EMB was developed and validated for field use. Bioassay optimization included an evaluation of the inter-rater reliability of sea lice responsiveness to EMB and an evaluation of gender-related differences in susceptibility. The concordance correlation coefficient was used to compare the agreement between the proportion of lice categorized as moribund or dead by two independent raters prior to and following the adoption of a set of bioassay response criteria. The adoption of a set of bioassay response criteria improved the concordance between raters' assessments and it also improved the model estimation of the EC50 values (the "effective concentration" leading to a response of 50% of the lice not prone to natural response) for subsequent bioassays. An evaluation of gender-related differences in EMB susceptibility for 19 bioassays indicated that preadult stage female sea lice from field populations exhibited a significantly higher sensitivity towards EMB compared to preadult males in 12 of 19 bioassays; 7 of 19 bioassays showed no significant difference in gender susceptibility, although all male EC50 values were higher than female values. Preadult stage females from one population of laboratory cultivated sea lice also exhibited a significantly higher EMB sensitivity compared to preadult males. The EMB bioassay was implemented for field use and a total of 38 field bioassays were completed from 2002 to 2005 using preadult stage sea lice collected from Atlantic salmon farms in four regions in the Bay of Fundy salmon farming area. There was no significant overall effect of region or year on EC50 values, and although the range of sensitivities obtained in this three-year study did not appear sufficient to affect current clinical success in the control of sea lice, the results suggest that there is a seasonal or temperature associated variation in sensitivity of preadult sea lice to EMB with sensitivity decreasing in the fall and winter months.

As a potential means of EMB detoxification by sea lice, oxidase enzymes are of particular interest as an alternative to the use of bioassays for resistance detection. The results of this study indicated significant effects of region and a seasonal trend on median haem peroxidase values for 2001 to 2004 field sea lice samples. There were no significant effects in the 2005 field data. There was a positive, but not statistically significant, correlation between EMB sensitivity (bioassay EC50 values) and median haem peroxidase activity in field sea lice samples for the years 2001 to 2005.

This research has demonstrated the widespread use of EMB for sea lice control in the Bay of Fundy and has resulted in the development of an EMB bioassay which could be used to verify clinical resistance in the field. The developed bioassay does however lack rapidity and simplicity for use as a routine test for a field monitoring program. The measurement of total oxidase activity in individual sea lice is difficult to confirm as a diagnostic tool for resistance detection due to the absence of sea lice populations known to be resistant to EMB, and the inability of the assay to indicate the specific cytochrome P450 enzymes responsible for resistance development. However, the ability of the haem peroxidase assay to identify individuals displaying elevated general oxidative activity is encouraging, as field populations are likely to consist of a mixture of susceptible and less susceptible individuals. This study also highlights the importance of obtaining data on the parameters influencing oxidase levels in EMB-susceptible sea lice populations in order to standardize the haem peroxidase assay methodology before implementation in a resistance monitoring program. The haem peroxidase assay should be used in combination with bioassays or other tests (i.e. molecular) when resistance is suspected and the mechanism has been identified. The finding of a seasonal effect on the bioassay and oxidase values requires further investigation. (Abstract shortened by UMI.).