Stage 37 38 Stage 39 40 Figure. Stages in development of anurans based on Gosner (1960). These four stages were selected in Lithobates chiricahuensis and L. pipiens for 48 h exposure to static concentrations of CFT Legumine (5% rotenone). Table 1. Toxicity of commercial rotenone products to amphibians. Organism Concentration Threshold Compound Reference Southern leopard frog (Rana sphenocephala) unknown larval stage 0.5 mg/L 96 h LC₅₀ Noxfish Chandler and Marking (1982) African clawed frog (Xenopus laevis) Adults/unknown stages of tadpoles 5-6 mg/L 100% mortality (tadpoles) Prenfish McCoid and Bettoli (1996) Chiricahua leopard frog (R. chiricahuensis) Gosner stage 25 0.79 mg/L 96 h LC₅₀ Prenfish Little and Calfee (2008) unpublished data Chiricahua leopard frog (R. chiricahuensis) Gosner stage 28-31 1.2 mg/L 96 h LC₅₀ Prenfish Little and Calfee (2008) unpublished data Columbia spotted frog (R. luteiventris) Gosner stage 40-45 1.0 mg/L 24 h field exposure (100% mortality) CFT Legumine 5% Rotenone Billman et al. (2011) Columbia spotted frog (R. luteiventris) adults 1.0 mg/L 24 h field exposure (no observed mortality) CFT Legumine 5% Rotenone Billman et al. (2011) Boreal toad (Anaxyrus boreas) tadpoles 1.0 mg/L 24 h field exposure (100% mortality) CFT Legumine 5% Rotenone Billman et al. (2011) Leopard frog (R. pipiens) Unknown stage of juveniles 7.3 mg/L 24 h LC₅₀ Dri-Noxfish Farringer (1972) (from Bradbury 1986)
Leopard frog (R. pipiens) Unknown stage of juveniles 7.9 mg/L 24 h LC₅₀ Dri-Noxfish Farringer (1972) (from Bradbury 1986) Leopard frog (R. pipiens) Unknown stage of tadpoles 4.6 mg/L 96 h LC₅₀ Dri-Noxfish Farringer (1972) (from Bradbury 1986) Leopard frog (R. pipiens) Unknown stage of tadpoles 3.2 mg/L 96 h LC₅₀ Dri-Noxfish Farringer (1972) (from Bradbury 1986) Leopard frog (R. pipiens) Unknown of tadpoles 0.1 mg/L 8-24 h (100% mortality) 5% Rotenone Hamilton (1941) (from Bradbury 1986) Tiger salamander (Ambystoma tigrinum) Unknown gilled stage 0.02 mg/L 8-24 h (toxic but not fatal) 5% Rotenone Hamilton (1941) (from Bradbury 1986) Tiger salamander tigrinum) Unknown stage of tadpoles 0.1 mg/L 8-24 h (100% mortality) 5% Rotenone Hamilton (1941) (from Bradbury 1986) Table 2. Clutch identification of Lithobates chiricahuensis obtained from captive refugia on the Ladder Ranch, Sierra County, New Mexico. Clutch ID Date obtained Estimated Number of Eggs in Clutch Hatch Date Estimated Hatch Rate Estimated Number of Tadpoles Number of Tadpoles Used in Tests LC-01-13 (3 May) 900 10 May 85% ~750 700 LC-02-13 (2 Jun) 900 6 Jun 75% ~650 600 LC-03-13 (14 Aug) 700 20 Aug 80% ~500 445 LC-04-13 (25 Sep) 800 29 Sep 85% ~680 * *Heating malfunction in the NMSU greenhouse resulted in the majority of the tadpoles perishing.
Table 4. Average (± standard error, sample size; 95% confidence intervals are reported in parenthesis when the tests were not replicated) lethal concentrations of 50% of the test populations (LC50) at 48 h using CFT Legumine (Rotenone 5%) on Chiricahua leopard frog (Lithobates chiricahuensis) and northern leopard frog (L. pipiens) at targeted Gosner stages (GS) in greenhouse conditions. Nominal concentrations of rotenone were obtained from CFT Legumine formulation assuming the active ingredient (rotenone) is 5%. Species Gosner CFT Legumine (5%) (mg/L) Nominal Rotenone (ug/L) L. chiricahuensis GS 21-25 Early: Pre-limb bud formation 0.42 mg/L (± 0.125, n=3) 21.0 µg/L L. chiricahuensis GS 26-30 Mid: Limb bud formed 1.03 mg/L (± 0.073, n=3) 51.5 µg/L L. chiricahuensis GS 31-36 Late: Toe differentiation hind limb complete 1.30 mg/L (± 0.037, n=2) 65.0 µg/L L. chiricahuensis GS 37-40 Early froglet: Prior to front limb development 3.24 mg/L (± 0.045, n=2) 162.0 µg/L L. chiricahuensis** (Sub-lethal Exposure) GS 37-40 froglet: Prior to front limb development 3.40 mg/L (3.13-3.70) 170.0 µg/L L. pipiens GS 21-25 Pre-limb bud formation 1.30 mg/L (1.21-1.41) 65.0 µg/L L. pipiens GS 26-30 Mid: Limb bud formed 1.26 mg/L (1.16-1.36) 63.0 µg/L
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