**ABSTRACT NOT FOR CITATION WITHOUT AUTHOR PERMISSION. The title, authors, and abstract for this completion report are provided below. For a copy of the full completion report, please contact the author via e-mail at mwilkie@wlu.ca or via telephone at 519 884-0710 extension 3313. Questions? Contact the GLFC via email at frp@glfc.org or via telephone at 734-662-3209.**




1Michael P. Wilkie, 2Roger Bergstedt, 3D. Gordon McDonald


1Department of Biology, Wilfrid Laurier University, Waterloo, Ontario, N2L 3C5.

2Hammond Bay Biological Station, Millersburg, Michigan, 49759 (retired)

3Department of Integrative Biology, University of Guelph, Guelph, Ontario, N1G 2W1 (retired)


July 2014





The lampricide, 3-trifluoromethyl-4-nitrophenol (TFM) has been effectively used to control invasive sea lamprey (Petromyzon marinus) populations in the Great Lakes for over 50 years. It is now clearly established that TFM interferes with mitochondrial ATP production in both sea lamprey and in the rainbow trout (Oncorhynchus mykiss), leading to a mismatch between ATP supply and demand that can eventually result in the depletion of energy reserves and death. The specificity of TFM is related to the relative inability of larval sea lampreys to detoxify and eliminate TFM from their bodies compared to most non-target fishes. However, in some instances non-target fishes may experience adverse effects, toxicity and mortality during routine applications of TFM. Accordingly, the objectives of this project were to: (I) more fully explain why TFM is selective to sea lamprey compared to non-target fishes such as the rainbow trout and lake sturgeon (Acipenser fulvescens), (II) determine if routine application of TFM resulted in significant adverse physiological disturbances in non-target fishes and to determine if such effects were reversible, (III) ascertain howchanges in season and nutritional status affected the TFM sensitivity of lampreys.

To better understand how TFM is detoxified and eliminated, sea lampreys, rainbow trout and lake sturgeon were exposed to either the TFM LC100 (MLC; median lethal concentration) or the TFM LC50 of larval sea lampreys (lake sturgeon only) for up to 12 h, and the concentrations of TFM and its detoxification product, TFM-glucuronide, quantified in the carcasses of the animals using solid phase extraction and HPLC. Not surprisingly, TFM was quickly taken-up by all 3 groups, but sea lamprey did not detoxify TFM through the formation of TFM-glucuronide. As a result, TFM was eliminated slowly over several hours by sea lamprey during depuration in TFM-free water. In contrast, rainbow trout and lake sturgeon readily converted TFM to TFM-glucuronide, which was rapidly eliminated within 3 h, along with the parent compound, during recovery. Accompanying experiments using radio-labelled TFM (14C-TFM) indicated that TFM was more rapidly taken-up by larval sea lampreys in more alkaline waters (pH 8.5) compared to circumneutral pH (pH 7.8) and acidic water (pH 6.5), due a greater proportion of TFM being its un-ionized, more lipophilic phenolic form at higher pH. These findings strongly suggest TFM is taken-up in its un-ionized form by diffusion down water-blood TFM gradients. Despite relatively slow rates of TFM elimination, sea lamprey were still able to recover from short bouts of TFM exposure (4 or 6 h), as demonstrated by the restoration of brain and liver glycogen reserves, phosphocreatine concentrations and the elimination of lactate within 4 h of recovery. Collectively, these findings confirm that while sea lampreys have a lower capacity to detoxify TFM, they can recover from short-term TFM exposure, which could lead to treatment residuals if the animals are able to temporarily avoid full TFM doses or if TFM treatments are interrupted.

TFM had relatively few adverse effects in rainbow and lake sturgeon. Reductions in liver glycogen were observed in rainbow trout, and were likely related to increased glycogen requirements due to the need to make up for a shortfall in ATP supply, which was likely compromised, even at the sub-lethal concentrations of TFM to which the fish were exposed. Similar reductions in TFM were not observed in lake sturgeon because they were exposed to lower TFM concentrations. The reductions in liver glycogen in rainbow trout may partially explain why aerobic swim performance was impaired following TFM exposure, as demonstrated by a 28 % reduction in the critical swimming velocity (Ucrit).

In the field, however, it is likely that liver glycogen stores would be rapidly replenished with the resumption of feeding. Surprisingly, anaerobic, burst swim performance was not compromised, and actually improved slightly 4 h following TFM exposure. Light microscopy revealed that exposure to TFM caused minimal damage to the gills. Subsequent measurements of Na+/K+-ATPase and H+-ATPase (V-ATPase) activity and quantity using western blotting, and measurements of unidirectional Na+ movements across the gills indicated that TFM had minimal effects on gillmediated ion regulation in trout and sturgeon in hard water.

To determine how changes in season and physiological condition affected the TFM sensitivity of larval lampreys to TFM, animals were captured in May, June, August and September of 2011 and subjected to acute toxicity tests at the Hammond Bay Biological Station within 5-10 d of capture. Subsequent determination of the 12 h LC50s, indicated that the animals captured in the spring at temperatures of approximately 12C were most sensitive to TFM, with the 12 h LC50 increasing by almost 3-fold by August, when water temperatures were warmest (~ 22C). The greater sensitivity of the larval sea lamprey to TFM in the spring appeared to be related to 75 % lower glycogen stores in the muscle, and 40 % lower glycogen stores in the brain, along with greatly reduced lipid reserves, compared to the animals captured later in the year. These findings suggest that there is justification for altering the timing of applications to take advantage of the greater sensitivity of larval lampreys in the spring, particularly in rivers containing large water volumes and flows, which could reduce overall TFM use. Extra vigilance, however, may be required in the summer to guard against treatment residuals, when the larval lampreys are most robust, larger and tolerant to TFM. Indeed, accompanying experiments exploring how body size impacted TFM sensitivity demonstrated that there was a significant positive correlation between TFM tolerance, body mass and length, but not condition factor.

In conclusion, the present project demonstrates that the effectiveness of TFM treatments is affected by a variety of abiotic and biotic factors. Consideration of these factors and incorporating this knowledge into TFM treatment regimens will allow sea lamprey control supervisors to conserve TFM use and minimize the risk of treatment residuals.

Although non-target rainbow trout and lake sturgeon have efficient means to detoxify TFM, and suffered few adverse effects during TFM exposure, further studies are required to more accurately determine how life stage, environmental factors and nutritional status affect their capacity to deal with this lampricide..