584c 



WOELKE ON PACIFIC OYSTER EMBRYO BIOASSAY 1 1 3 



and clam populations arc present. Bioassays with bivalve larva of simu- 

 lated wastes prenared in the laboratory have been reported by a number 

 of investigators [3-]0]. Dimick and Brcose [I J] propose the bay mussel 

 as a standard bivalve for marine water bioassays because these molluscs 

 are found in nearly all estuarine areas in the world. 



In this paper I shall give a brief description of a rapid, inexpensive, 

 and dependable bioassay technique I developed while working for the 

 Shellfish Research Unit of the Washington Department of Fisheries. 



For the remainder of my presentation I shall deal with my subject 

 in four steps. First, state the assumptions and justifications; second, 

 describe the method itself; third, present data obtained in a series of 

 toxicity bioassays conducted on 29 samples of pulp mill and oil refinery 

 wastes; and fourth, describe how this technique has been extended to 

 actual estuarine pollution problems. 



In developing this technique I assumed a general acceptance of the 

 need for and merit of biological assays in the field of water quality re- 

 search. I further assumed an acceptance of the concept that development 

 of a bioassay procedure with a commercially valuable marine organism, 

 which could be applied at any time of the year with the same ease and 

 reproducibility currently attributed to the biological oxygen demand and 

 coliform mean probable number tests, would be a useful tool for evaluat- 

 ing water quality. It is my contention that the first 48 hr in the develop- 

 ment of fertilized eggs of the Pacific oyster, Crassosirea gigas, provides 

 a biological system whose response can be utilized to satisfy these as- 

 sumptions. During this 48-hr period the fertilized eggs normally develop 

 into free-swimming, fully shelled veliger larvae. My final assumption is 

 that failure to develop to fully shelled (normal) larvae in 48 hr will 

 break the life cycle of the Pacific oyster. I consider failure of the eggs 

 tu develop, or the proportion (per cent) of larvae developing in an ab- 

 normal manner to constitute a measure of the biological response to a 

 particular stimulus. 



Method 



The basic steps I have followed in carrying out bioassays with fertil- 

 ized Pacific oyster eggs are relatively simple and straight-forward. Adult 

 oysters (spawners) are thermally conditioned at 20 C in flowing sea- 

 water until they can be readily spawned. This usually requires about 

 four to six weeks. To insure the availability of spawners during all 

 months of the year, several groups of oysters at various stages of sex al 

 maturity are kept on hand at all times. Several hours before a bioassay 

 is to be conducted, 10 to 20 mature oysters are placed into Pyrex dishes 

 filled with filtered ultraviolet-light-treated water. These dishes are placed 

 in a water bath and the temperature is raised to 28 to 30 C. About 30 

 min before the time the spawning is desired, a sperm suspension from 



