Top ⇑The Proposed Standard Methods: Elements of Both Former Approaches

Clutch Size

A clutch is a group of eggs, which is formed from synchronously developing oocytes and is oviposited within a relatively short interval of time. One of the goals in studying fecundity should be to determine the sizes and frequency of clutches produced during a spawning season. Results from a number of studies have shown that all mature or ripening oocytes in an ovary are ovulated as a group to produce a clutch of ripe eggs which is then spawned (oviposited) over a relatively short period of time whether all at once (e.g. Gasterosteus aculeatus) or in successive bouts as in many darters and minnows.

Because females begin spawning soon after ovulation, we generally expect the ovaries of RE females to contain fewer eggs than the number they typically spawn from a complete clutch. Data show that RE females yield spurious estimates of clutch size and also will result in erroneous statistics for clutch size body size relationship.

One therefore should use only MA or MR females to estimate clutch size. At times, one to a few ripening oocytes from a previous clutch will be present in the follicles of MA females. One should not count them. Ripening oocytes that are not ovulated with the rest of their clutch appear to be resorbed; they do not appear to be ovulated with the next clutch.

Ovum Size: Diameter and Mass

Ripe eggs provide the best estimates of egg size mass or diameter because they represent the final stage of development. Ova removed from females as well as spawned ova may be used. One should be careful to identify the stage of the ova (ovulated from female vs. spawned) as diameter measurements will be affected. In general, spawned ova should have greater diameters ripe ova removed from females due to further hydration of the eggs after oviposition.

Ripening oocytes provide a good estimate of ovum mass as yolk loading has ended by this stage of ovum formation. One should be sure that all follicular material has been removed from the oocytes, as ripening oocytes can be difficult to clean. Diameters of ripening oocytes will underestimate the diameters of ripe eggs in females because they are not yet fully hydrated.

Both the mass and the diameter of ripe eggs in females will be underestimated by using mature oocytes. Mature oocytes may, however, serve as useful indices of the size of ripe eggs for purposes of comparisons, although this has not yet been demonstrated conclusively. At present, data seem to suggest that in some cases one might not detect differences in ripe eggs by using mature oocytes due to greater variability in mature oocytes. In all cases, the use of oocyte/ovum dry mass is preferable to diameter as a measure of nutritive material afforded the young. Correlations between oocyte/ovum diameter and mass decrease sequentially between mature oocytes, ripening oocytes, and ripe eggs due at least in part to the hydration of the latter two stages. Thus, diameter is not as good a predictor of mass in ripening oocytes and ripe eggs as in mature oocytes.

Relative Clutch Mass (RCM)

One approach to estimating relative clutch mass would be to use the mean and variance for the mass of individual ripe eggs from RE females and the clutch sizes of MA and MR females to estimate clutch mass just before spawning. The MA and MR females would, therefore, be dried and weighed to use the body weights to estimate RCM. Clutch and body weights from MA females may provide indices of RCM in females just prior to spawning but there are no data addressing this question. MR females should provide the most straightforward estimates of RCM, but they are often rare in collections.

Drying Temperatures

The temperature at which oocytes, ova, and carcasses are dried will affect the masses of the tissues being weighed. In general, there is a sequential loss of lipids at higher temperatures. Thus, one should dry tissues at the lowest possible stable oven temperature. Many ovens cannot maintain a stable temperature below about 40°C and others are limited below 30°C. Thus, 30-40°C is the recommended temperature range. The oven should be able to maintain a very narrow range of temperature variation. A suggested maximum range of variation is ±1°C.

Weighing Tissues

Transfer tissues from the oven to a dessicator with fresh dessicant as rapidly as possible to avoid allowing the tissue to take up moisture. The balance chamber should also have a dessicant in it. The transfer of tissue from the dessicator to the balance should also be as rapid as possible.

     © Copyright 1995 by David C. Heins, All rights reserved.

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