Open literature Review Summary 

Chemical Name: Atrazine 

CAS No: 1912-24-9

ECOTOX Reference Number and Citation: 161314 

Paetow LJ; McLaughlin JD; Pauli BD; Marcogliese DJ. 2013. Mortality of American Bullfrog 

Tadpoles Lithobates catesbeianus Infected by Gyrodactylus jennyae and Experimentally Exposed 

to Batrachochytrium dendrobatidis. J. Aquat. Anim. Health 25(1): 15-26. 

Purpose of Review: Registration Review

Date of Assessment: 4/6/2015 

Brief Summary of Study Findings: This study investigated the effects of atrazine on larval 

American bullfrogs (Lithobates catesbeianus) experimentally exposed to Bd. After the start of 

the experiment, the sample population was found to be infected with gyrodactylus jennyae

leading to mild to severe skin erosions in the experimental population. Increased mortality was 

observed between control groups and Bd and Bd+ atrazine groups, but no difference was found 

between Bd and Bd + atrazine groups. The relationship between the co-infections and the 

severity of skin lesions was correlated with their influence on survival. 

Methods: 102 bullfrog tadpoles ((Gosner stages 26–37) were obtained from an Environment 

Canada laboratory located in Moncton, New Brunswick (originally purchased from a commercial 

source in the United States). A virulent strain of Bd was used (JEL423) provided by J.E. Longcore 

(University of Maine, Orono). Tadpoles were randomly selected for each treatment group. 

Four experimental treatment and control groups were established: (1) a group of tadpoles 

acutely exposed to two doses of Bd zoospores (2) a group exposed for 14 d to atrazine, followed 

by two acute doses of zoospores (3) a group exposed for 14 d to glyphosate, followed by two 

acute doses of zoospores and (4) a group that received no experimental exposures. There were 

four replicates for each group, except for the control group, which had five; each replicate 

consisted of six tadpoles. Nominal chemical exposure concentrations were 21 ʅg of atrazine, 

and 1.5 mg acid equivalents (a.e.) of glyphosate, per liter of water. In the herbicide exposed 

animals, the two acute exposures to Bd occurred at 14 and 16 d after the start of the pesticide 

exposures, and each Bd exposure lasted 24 h. 

Each replicate group was maintained in a 33-L aquarium containing 8 L of aerated, dechlorinated 

water. Three-quarters of the water in each tank was changed every 3 d. During the pesticide 

exposure phase, the herbicides were replenished with each water change. Water volume was 

maintained at 1.3 L per tadpole and reduced by 1 L with each tadpole’s death, to a minimum of 

4 L. At day 45, all surviving tadpoles were killed by immersion in a 0.8% solution of MS-222 

(tricaine methanesulfonate). Tadpoles that died or were killed were immediately fixed in Bouin’s 

solution, in a 70% solution of ethanol, or in a 10% solution of buffered formalin (two tadpoles in 

each solution per aquarium). 
At day 0, a stock solution of each herbicide was prepared. During the pesticide exposure phase 

of the study, each liter of water that was siphoned from the exposure tanks every third day was 

replaced with 1 mL of pesticide stock solution. At two time points (at 12 h after herbicide 

addition and at day 14), water samples were drawn directly from two randomly selected 

pesticide-containing aquariums (one containing atrazine and one containing glyphosate) 

to compare the actual and nominal concentrations. 

The concentrations of glyphosate measured at 12 h and at 14 d post addition of the pesticides to 

the aquariums were both 2.1 mg/L a.e., which was 1.4 times higher than the nominal 

concentration of 1.5 mg/L a.e. The concentrations of atrazine measured on the same days were 

140 ʅg/L and 77 ʅg/L, respectively, which were 6.7 times and 3.7 times higher than the 

nominal concentration of 21 ʅg/L. 

Results: By day 30 of the experiment, a consistent pattern of dermal depigmentation was 

observed around the eyes and at the base of the tail of tadpoles in every group, including the 

control group. At days 30 and 31, two tadpoles that displayed severe depigmentation (each 

from the group exposed to both glyphosate and Bd) were removed from the experiment for 

examination. Depigmentation was found to be the result of skin erosion caused by the tadpoles’ 

infection by the monogenean ectoparasite G. jennyae. Further examination revealed that G. 

jennyae was present in each aquarium. 

Increased mortality was observed between control groups and Bd and Bd+ atrazine groups, but 

no difference was found between Bd and Bd + atrazine groups. Increased susceptibility to Bd

due to atrazine exposure was not demonstrated in the study. 

Description of Use in Document: Qualitative. This study is of sufficient quality for inclusion in 

risk assessment, but with limited utility. 

Rationale for Use: The study provides information on effects of Bd on survival and the effects of 

co-infection of tadpoles with monogenean ectoparasite G. jennyae. However, effects due to 

atrazine alone could not be discerned from those due to Bd alone. 

Limitations of the Study: The presence of the co-infection in all experimental animals with the 

monogenean ectoparasite G. jennyae confounded interpretation of results. 

There is uncertainty in the concentrations of atrazine based on the measured concentrations (14 

ug/L and 77 ug/L) as compared to the nominal concentration of 21 ug/L. 

Water quality parameters not reported in study. 

Reviewer: Colleen M. Rossmeisl, DVM; ERB3, Environmental Fate and Effects Division COLLEEN 

ROSSMEISL

Digitally signed by COLLEEN ROSSMEISL 

DN: c=US, o=U.S. Government, ou=USEPA, 

ou=Staff, cn=COLLEEN ROSSMEISL, 

dnQualifier=0000069649 

Date: 2016.04.07 17:20:44 -04'00'