Trace metal concentrations and the physiological role of zinc in the West indian manatee (Trichechus manatus)

Due to the potential impact of contaminants in marine mammals, many studies have examined metal levels in cetaceans and pinnipeds. However, only two studies have been reported for the endangered West Indian manatee (Trichechus manatus). Thus, the objectives of this study are: 1) Determine baseline levels of metals in the Florida manatee in wild and captive West Indian manatees, 2) Evaluate metal loads in various tissues of the Florida manatee, 3) Asses trace metals found in the surrounding environment of Kings Bay, Crystal River, Florida, 4) Examine carbonic anhydrase in the Florida manatee, 5) Evaluate MT as a biomarker for monitoring metal exposure and induction of MT expression in manatees, and 6) Compare metal concentrations and MT expression in other Paenungulata species.;Zinc (11.2+/-4.5 ppm) was of main interest, as levels were higher in manatees than in other related species, such as the Elephas maximus, and other marine mammals. Metal distribution in manatee carcasses was examined and accumulation of Zn was primarily present in the liver with a range of (32.39 - 288.55 ppm, ww). To evaluate potential sources of metal exposure, the surrounding environment and vegetation samples from Kings Bay, Crystal River were evaluated for Zn, Cu, As, and Se. Our study determined that plant material is likely to be a significant source of metal exposure. Diet may be contributing to Zn accumulation in manatees with levels of Zn higher in favorable vegetation species, such as Vallisneria americana and Najas guadalupensis. Moreover, metal levels were higher in the northern areas of Kings Bay with possibly anthropogenic influences.;Due to the fact that zinc levels were high without apparent adverse effect, we explored possible physiological factors underlying the high zinc levels. Carbonic anhydrase is a zinc metalloenzyme which catalyzes the hydration/dehydration of CO2 and functions in respiration, acid-base balance, and fluid formation. Two isozymes of carbonic anhydrase from manatees were confirmed by isoelectrofocusing with 2D SDS-PAGE and LC-MS/MS and confirmation of CA I and CA II was unique compared to other marine mammal species which only have one isozyme.;In addition to CA, metallothionein (MT) was evaluated in the Florida manatee. Metallothionein is a metal-binding protein involved in divalent metal homeostasis that has not been examined in manatees and elephants. We successfully cloned and sequenced MT 1 (381 bp) from isolated peripheral blood mononuclear cells of the Florida manatee that was 83% identical to rhesus macque ( Macaca mulatta) MT 1E and 84% identical to chimpanzee (Pan troglodytes) MT 1x. MT expression was induced in cultured manatee leukocytes exposed to 2x10-9 M CdCl2 and 2x10 -5 M ZnCl2, demonstrating a 7-fold and 13-fold increase following 24 hr exposure, respectively. Maximal induction of MT expression occurred at lower doses of CdCl2 of 2x10-11 M than 2x10-8 M and higher doses of ZnCl2 of 2x10 -4 M compared to 2x10-6 M. Moreover, a partial sequence for MT 4 (182 bp) was obtained and evaluated as a potential biomarker for metal exposure. However, MT 4 expression in manatee skin did not correlate with metal levels in blood or skin. This is the first study to evaluate carbonic anhydrase kinetics and metallothionein expression in Order Sirenia.;Manatees belong to clade Paenungulata, with elephants as the closest relative next to the dugong. Asian elephant MT (376 bp) was cloned and sequenced with phylogenetic analysis resulting in a maximum likelihood of 74% to manatee MT 1. Sirenians are unique amongst the marine mammals, as the only herbivorous species, and are sentinel species to the health of our waterways. The results of this study will aid environmental agencies and management officials to monitor trace metal accumulation in the aquatic environment for manatee health and conservation. (Abstract shortened by UMI.).