Description of Use in Document

EPA Primary Reviewer: Amy Blankinship Date: __5/15___________

EPA Secondary Reviewer: Elizabeth Donovan Date: __5/26/16___________

Data Evaluation Record

ECOTOX CODE: 160043
STUDY TYPE: Non-guideline
DP BARCODE: Not provided
PC CODES: 057701 (Malathion); 035001 (Dimethoate)
CAS NO.: 121-75-5 (Malathion); 60-51-5 (Dimethoate)
MRID NO.: Not available
TEST MATERIAL (% purity): Malathion and Dimethoate (purity not provided)
CITATION: El-Gawad, E.A.A., Kandiel, M.M.M., Abbass, A.A., and Shaheen, A.A. (2011). Impact of Some Organophosphorus Insecticides on Growth Performance, Fecundity and Semen Characteristics in Nile Tilapia (Oreochromis niloticus). Lucrări Ştiinţifice Med. Vet. 54(1): 150-160.
SPONSOR: Not applicable (open literature review)
EXECUTIVE SUMMARY: The purpose of this study is to determine endocrine disrupting effects of the organophosphorus compounds, malathion and dimethoate, in Nile Tilapia (Oreochromis niloticus). The 1.6 mg/kg dimethoate and 0.17 mg/kg malathion test concentrations were prepared in the dry fish food. The treatment effects investigated included reproductive (GSI, semen quality, and plasma sex steroids testosterone and 17β-estradiol), growth performance (condition K factor), and gonadal morphology. Malathion and dimethoate are EDSP List 1 chemicals.
Methods: The Nile Tilapia (Oreochromis niloticus) used in the study were obtained from a private fish hatchery in Kafer El-Sheikh, Egypt. The fish mean body weights were 29.93 g for females and 33.56 g for males, and the fish mean body lengths were 12.8 cm for females and 13.33 cm for males. The fish were acclimated to laboratory conditions at 27 ± 1°C for 7 days in fiberglass tanks (110x90x40 cm). The control and treated fish were fed diet daily at a rate of 3% of body weight. About 50% of the tank water was exchanged daily. The malathion and dimethoate (purities not reported) were purchased from Sigma Aldrich Chemical (USA). The test chemicals were dissolved in cod liver oil to form stock test solutions. The 1.6 mg/kg dimethoate and 0.17 mg/kg malathion test concentrations were prepared by mixing one mL of test solutions into 3600 g of dry food. The treated feed was packed in clean plastic jars and stored refrigerated at 4°C until use. The 120-day test included an untreated control, 1.6 mg/kg dimethoate, and 0.17 mg/kg malathion treatment groups. Each treatment group contained one replicate tank with 100 fish.
At test termination, blood samples were collected from 6 male and 6 female fish. The total length and weight of fish were measured at test termination. Absolute fecundity, the total number of ripened eggs in the ovaries per female, was determined by counting immediately after dissection by the gravimetric method. Relative fecundity was calculated based on the number of eggs per length (cm) or body weight (g).
Semen quality was determined from testes samples collected at test termination from 6 male fish per treatment group. Semen was collected from testes and transferred to a clean Eppendorf tube for analysis. The semen evaluations included pH, motility, viability, sperm cell concentration, and sperm cell morphology. The pH was determined using pH indicator papers. The sperm were counted as motile if they exhibited progressive movement or spontaneous flagellar beating, and the percentage motility was calculated by grading the percentage motile cells. Based on 5% eosin and 10% nigrosin staining, the percentages of live spermatozoa were counted by differential coloration using a microscope (100x). Sperm counts (cells/mL) were determined using a hemocytometer. The percentage of abnormal spermatozoa was counted, and morphological abnormalities were reported as a percentage of the total number of all counted spermatozoa.
For gonadosomatic indexes (GSI) determination, the gonads from male and female euthanized fish were excised and weighed. The growth performance was calculated based on the condition (K) factor.
For the sex hormonal assay, blood samples collected from the heart were assayed for estradiol 17β (E2) and total testosterone (T) using radioimmunoassay (RIA) kits.
The gonad tissue sections were collected from treated fish at test termination. The gonad samples were fixed in 10% buffered formalin, dehydrated in alcohol, cleared by xylene, embedded in paraffin, sectioned, stained with Hematoxylin and eosin, and analyzed under a light microscope.
For statistical analyses, the data results were expressed as mean ± SEM. The data were analyzed by one-way analysis of variance (ANOVA) and Duncan’s multiple range tests to determined significant differences between groups. The analyses were conducted using Statistical Package for the Social Sciences (SPSS) software (version 16.0). Statistical significance was determine at p<0.05.
Results (EDSP List 1 chemicals): The 1.6 mg/kg dimethoate and 0.17 mg/kg malathion treatments significantly decreased ovarian activity and lowered the reproductivity of female O. niloticus compared to the control (Table 1, copied without alteration from El-Gawad et al. 2011). Absolute fecundity was significantly decreased in the 1.6 mg/kg dimethoate and 0.17 mg/kg malathion treatment groups. The relative fecundity in relation to total length and bodyweight were significantly decreased in the 1.6 mg/kg dimethoate and 0.17 mg/kg malathion treatment groups.

Significant differences (p<0.05) in semen quality were observed in response to the 1.6 mg/kg dimethoate and 0.17 mg/kg malathion treatments (Table 2, copied without alteration from El-Gawad et al. 2011). The sperm cell concentrations were lower in the dimethoate (6.16 x 10⁹ cells/mL) and malathion (5.66 x 10⁹ cells/mL) treatment groups compared to the control (15.84 x 10⁹ cells/mL). The sperm progressive motility was decreased in the dimethoate (24.17%) and malathion (21.67%) treatment groups compared to the control (60.83%). The live sperm percentage was reduced in the dimethoate (50.07%) and malathion (43.32%) treatment groups compared to the control (73.48%). Based on sperm morphological observations, increased tail deformity was observed in the dimethoate (57.93%) and malathion (48.6%) treatment groups compared to the control (28.81%). No effects on semen pH or percentage of head deformities were observed for either treatment group.

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