Ganias et al.: Degeneration of postovulartory follicles in Sordino pilchardus 



133 



XSA=axD>> , 

 where a and b are parameters. 



(1) 



In case of isometry, (i.e., 6 = 2), shrinkage is 

 supposed to occur evenly in all dimensions, and 

 the shape of the regressing POFs does not un- 

 dergo significant changes. In any other case 

 (fe>2, or 6<2), the shape is altered along resorp- 

 tion, and thus its intermediate phases may be 

 used in the staging of POFs. 



Existing histological criteria for the stag- 

 ing of sardine POFs were refined and based on 

 two hauls from the 2005 survey that contained 

 more than 90 histological specimens each. Giv- 

 en that individuals in each sample were caught 

 at the same time and temperature, POFs in 

 each daily cohort should have been at the same 

 stage and have had the same age, maximizing 

 the morphological contrast among daily classes. 

 Furthermore, one of these hauls was performed 

 in the evening, just before the average daily 

 spawning hour for the Iberian sardine (20:00; 

 Zwolinski et al.. 2001; ICES, 2004) and thus 

 provided information on the final histological 

 state of each daily class of POFs. 



After refining the staging criteria and clas- 

 sifying the POFs from all surveys into daily 

 classes, we used the time of capture and daily 

 spawning hour to estimate the exact age of 

 POFs, i.e., the time in hours elapsed between 

 spawning and sampling. The effect of POF age 

 and other factors on POF cross-sectional area 

 were tested with a generalized linear model 

 (GLM) with an overdispersed Poisson distribu- 

 tion and a logarithmic link function. Apart from POF 

 age, the effect of temperature, sampling year, and the 

 one-way interaction of year with age were considered in 

 the model. The significance of the relationship between 

 POF age and size indicated whether our staging and 

 aging criteria were accurate. Furthermore, because the 

 laboratory processing of the ovaries differed from year 

 to year, the year effect and its interaction with age was 

 used to test the effect of preservation medium (AFA, 

 formalin) and embedding material (paraffin, resin) on 

 the size of POFs. Residual inspection plots revealed the 

 adequacy of the fitted model. 



Results 



A total of 249 ovaries with POFs were detected and 

 used in our analysis (1997: 65, 1999: 104, 2005: 80). 

 The two hauls from the 2005 survey that were used for 

 refining POF staging criteria contained many females 

 with POFs at various stages and sizes, facilitating the 

 distinction between successive daily classes of spawners. 

 The frequency distribution of POF cross-sectional areas 

 in the two hauls displayed three size modes, which were 

 considered to correspond to different age classes (Fig. 2). 



0.005 



0.009 



0.013 



0.017 



0.021 



0.005 



0.009 



0.013 



0.017 



0.021 



POF area (mm^) 



Figure 2 



Frequency distribution of postovulatory follicle (POF) cross-sec- 

 tional areas from two samples of the Iberian sardine Sardina 

 pilchardus from the 2005 survey. (A) Sample collected at 12:00, 

 /! = 48; (Bl sample collected at 17:30. n=32. Arrows indicate POF 

 size modes in each sample. 



The most advanced size mode in the sample hauled at 

 12:00 (Fig. 2A) consisted of larger POFs compared to 

 the POFs in the sample hauled at 17:30 (Fig. 2B). Re- 

 examination and comparison of POF size modes in each 

 sample showed that, apart from size, they differed in 

 shape and in the fine histological characteristics of the 

 follicular layers (Fig. 3). In particular, large POFs had 

 an irregular shape and contained a large, convoluted, 

 and thick granulosa layer (Fig. 3A). In the intermediate 

 size mode, the shape of the follicle changed to semirect- 

 angular and the granulosa layer tended to lose its con- 

 voluted appearance and to form a single layer (Fig. 3B). 

 Finally, in the smaller size mode, all POFs displayed a 

 triangular shape (Fig. 3, C-E). However, more detailed 

 examination showed that these triangular POFs could 

 be further separated into 1) a group of slightly larger 

 POFs with a thin layer of the granulosa (Fig. 3C), and 2) 

 a group of very small POFs that contained only granu- 

 losa remnants in the form of residual vacuoles (Fig. 3, 

 D and E). 



POF diameter increased significantly with POF cross- 

 sectional area (Fig. 4) and the relationship was not 

 significantly different between the paraffin samples 

 from 1997 and 2005 (P>0.05). The allometric coefficient 

 b of Equation 1 differed significantly from the square 



