34 V. S. BUREAU OF FISHERIES. 



manipulations required for this experiment, consumed much time. 

 Consequently the end of the breedino^ season for oysters came before 

 there was o]3portunity to work out the relations of all these factors 

 in an entirely quantitative manner. 



Results liiixy be summarized as follows : 



AVith regard to the condition of the eggs, it was observed that 

 embryos from eggs of which 50 per cent or more developed quickly 

 to a free swimming stage withstood deprivation of oxygen better 

 than those from eggs of which only a small proportion developed. 



With respect to age, it appeared that younger larvse endured com- 

 parative oxygen deprivation slightly longer than older ones, but the 

 experiments were not conclusive. 



It was found that embryos show some effect of oxygen lack in 

 water containing 25 per cent of saturation of oxygen at 24° C. and a 

 markedly increased mortality in 10 per cent of saturation. Killing 

 time varied from 12 to 30 hours according to extent of oxygen de- 

 pletion. At 28 to 30° C. embryos die much more rapidly from oxygen 

 starvation than at 22 to 24° C. 



Carbon dioxide seems to have a toxic effect other than that due to 

 hydrogen ion concentration. Embi-yos died earlier in water con- 

 taining sufficient oxygen but excessive COo than in water of the same 

 oxygen content and with the same Ph obtained by adding lactic acid. 



Investigation of the effect of pollution on satisfactory development 

 of eggs in gonads was also conducted. Results showed that from 

 oysters, kept 7 to 10 days in water polluted so as to markedly lower 

 its ox^'gen content, only an abnormally small proportion of eggs 

 could be made to develop after artificial fertilization, conducted ac- 

 cording to standardized technique, and that the embryos so obtained 

 did not develop as far as controls. 



An incidental observation proved of considerable interest. Oysters 

 which had been in cold storage 24 days for the purpose of studying 

 greening were found to have very full gonads. Eggs fertilized with 

 sperm from some of the same lot of oysters developed satisfactorily, 

 and many of the embryos reached the primitive shell stage. Some 

 of them, indeed, were used in oxygen-requirement tests. 



Observations on the cause of copper greening confirmed previous 

 conclusions by showing that not only the amount of copper obtain- 

 able by the oysters, but also conditions of habitat determine whether 

 or not visible green spots occur. Marked appearance of greenness 

 after one or more weeks in cold storage Avas observed. It has been 

 twice noted that Avith gonads filled a smaller proportion of oysters 

 become green during storage than in otlier conditions. Two-year- 

 old spent oysters, set and reared in Wareham River, did not become 

 green during four Aveeks in cold storage. This is in contrast to 

 oysters which at some time during their previous history have been 

 in Avaters known to produce greenness (near Bridgeport and New 

 HaAen). 



Experiments both in the laboratory and in the field indicated a 

 positive influence of diminished oxygen supply in causing the ap- 

 pearance of green spots. Repeated ol)servations of marked greening 

 of oysters in polluted tanks of sea water were also made, thus con- 

 firming the similar results obtained during the previous summer in 

 the Seekonk River. 



