containing 10% Formalin^ in seawater. The sam- 

 ple size was variable — all exceeded 100 eggs, usu- 

 ally several hundred. The vials were then shipped 

 to the laboratory for examination with the aid of a 

 dissecting microscope. The epibiotic organisms 

 were clearly visible using transmitted light for 

 illumination (Fisher et al. 1975). Closer examina- 

 tion of the egg cases was carried out with a phase 

 microscope to aid in the characterization of the 

 fouling organisms. Portions of the samples were 

 categorized as to the comparative developmental 

 state of the eggs, extent of epibiotic fouling, and 

 egg mortality by the following methods: 



1. The following observations of the eyespots 

 which develop as the embryos develop were used to 

 give a comparative estimate of the time the eggs 

 had been carried externally on the female: 



Dl. No visible eyespot. 

 D2. Emerging eyespot. 

 D3. Full eyespot. 



Any samples which showed evidence of hatch- 

 ing were not used. Occasionally, there was varia- 

 tion in the degree of development of the eggs from 

 a single sample, in which case the eggs that had 

 developed furthest were used for observation. 



2. The extent of epibiotic fouling was deter- 

 mined by the following observations of the exter- 

 nal egg membrane: 



Fl. None — no evidence of epibionts at 100 x 

 (Figure 2A). 



F2. Light — occasional short filaments. 



F3. Moderate — the majority of the surface 

 covered with short filaments and occa- 

 sional long filaments (Figure 2B). 



F4. Heavy — the surface extensively covered 

 with short and long filaments (Figure 

 2C). 



F5. Very heavy — the surface extensively 

 covered with short filaments, long fila- 

 ments, and detrital material. 



3. The number of empty egg cases was used as 

 an estimate of mortality. 



Ml. <10% mortality. 

 M2. 10-25% mortality. 



^Reference to trade names does not imply endorsement by the 

 National Marine Fisheries Service, NOAA. 



M3. 26-50% mortality. 

 M4. 51-75% mortality. 

 M5. 76-100% mortality. 



Only empty egg cases (Figure 3) were consid- 

 ered mortalities. Other abnormal conditions, 

 such as discolored eggs which might have eventu- 

 ally led to mortalities, were observed but not used 

 in the estimates. All developmental stage D3 

 samples were checked for emerging embryos to 

 ensure that the empty egg cases were not due to 

 hatching. 



In addition to the field samples, seven ovigerous 

 females from the Point Reyes area were examined 

 before being placed into flow-through seawater 

 tanks at the laboratory. After 25 days the eggs 

 were reexamined to determine the progress of the 

 infestation. In addition, one complete egg mass 

 from an ovigerous female was examined to deter- 

 mine the homogeneity of the fouling condition 

 throughout the egg mass. 



Results 



Observation of eyespot development placed 

 10.5% of the samples into category Dl, 35.2% into 

 D2, and 54.3% into D3. Fouling was observed in all 

 developmental categories, but mortalities were 

 generally higher in the more developed eggs. The 

 histograms presented in Figure 4 show the percent 

 of samples from each region placed in each mortal- 

 ity category (M1-M5) and fouling category (F1-F5) 

 after combining the developmental categories. 



The eggs of the seven females held in the labo- 

 ratory for 25 days showed an average increase 

 in their development, fouling and mortality of 

 one level in each category. The greatest observed 

 change was on an egg mass in developmental 

 stage 2 which originally showed light fouling (cat- 

 egory 2) and were in mortality category M2. 

 After 25 days it was in developmental stage 3 

 and showed very heavy fouling (category 5) and 

 had advanced to mortality category, M5. Another 

 showed no increase in fouling as it matured 

 from developmental stages 1 to 3, but the egg 

 mortality category advanced from Ml to M3. 



Examination of the entire egg mass of one 

 specimen showed that the extent of the fouling 

 was variable and concentrated mostly on the 

 periphery of the mass and on the inner eggs near 

 the fold of the abdomen. This raises the possibility 

 of sampling error; however, it would probably be 

 insignificant since the field samples came primar- 

 ily from the exterior of the egg masses. 



202 



