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ENSR Cowering end Engineering <br />4.5 Biological Monitoring <br />Observations of mortality and reproduction in each test chamber will be made daily. <br />4.6 Test Duration <br />The test will be terminated when 60 percent of the surviving control organisms have produced <br />three broods (maximum test duration of 8 days). <br />4.7 Calculations <br />Fisher's exact test will be used to test for a significant difference in the survival of the various <br />effluent concentrations and the control. For the purpose of determining the NOEC, <br />reproduction data will be compared from only those effluent concentrations where the <br />mortality was not significantly different from the control. Normality and homogeneity <br />assumptions for reproduction data will be evaluated by the appropriate statistical tests <br />(p 50.01). If the data meet the assumptions, Dunnett's procedure (or Bonferroni's procedure <br />for unequal replicates) will be used to make the comparison (p =0.05). If the data do not <br />meet the assumptions, Steel's many -one rank test (or Wilcoxon's test for unequal replicates) <br />will be used to make the comparison (p = 0.05). Reproduction will be determined by the <br />number of young per original female. if the presence of males is visually confirmed at test <br />termination, these organisms will not be used in the reproduction comparison. The no <br />observable effect concentration (NOEC), lowest observable effect concentration (LOEC), and <br />ChV (the geometric mean of the NOEC and LOEC) will be calculated on the basis of survival <br />and /or reproduction. The lethality lC (25 percent reduction in organism performance based <br />on organism survival) and chronic IC (25 percent reduction in organism performance based <br />on reproduction) will be calculated by linear interpolation. <br />4.8 Quality Criteria <br />ENSR Project No.: 8503 -096 -193 <br />Protocol No.: CD4CO.193.001 <br />Effective: 08/96 <br />Page: 4 of 5 <br />The test will not be considered valid if control mortality exceeds 20 percent or if 60% of the <br />surviving adults do not produce at least three broods with an average of 15 or more young <br />per surviving female. <br />5.0 TEST REPORT <br />The report will be a typed document describing the results of the test and will be signed by <br />the Study Director and Quality Assurance Unit. The report will include, but not be limited to, <br />the following: <br />• 'A Colorado Department of Health summary sheet. <br />• A copy of all raw data. <br />• Name of test, Study Director, and laboratory. <br />• A description of the experimental design and the test chambers, the number of <br />test organisms, replicates per treatment, and the lighting. <br />• Test organism scientific name, age, and diet. <br />