Metabolic Rates of Rainbow Trout Eggs in Reconstructed Salmonid Egg Pockets

The Kruskal–Wallis test indicated that the fish stage significantly affected the DO demand (H(2) = 31, p ₄⁺ excretion (H(2) = 30, p Figure 3). Pairwise comparisons using Dunn’s test showed that the DO demand (mg O₂ egg h⁻¹) in the eyed eggs (294–310 dd) was 0.002 ± 0.0004 mg O₂ egg⁻¹ h⁻¹, which is significantly lower (p = 0.001) than that of the nearly hatched alevins (350–358 dd), respiring 0.016 ± 0.003 mg O₂ egg⁻¹ h⁻¹. In turn, the stage with completely absorbed yolk sacks (454–462 dd) had a DO demand of 0.07 ± 0.007 mg O₂ egg⁻¹h⁻¹, which is significantly higher (p = 0.03) than that of the previous stage.

This marked increment in metabolic activity from the embryonic to larval stage is not surprising since it was observed earlier by Wicket [25] for pink (O. gorbuscha) and coho (O. kisutch) salmons reared at 8 and 5 °C, respectively. Wicket [25] reported metabolic rates for eyed eggs ranging from 0.0006 to 0.0002 mg O₂ egg⁻¹ h⁻¹, sharply increasing by one order of magnitude (0.009–0.01 mg O₂ egg⁻¹ h⁻¹) for newly hatched alevins. Further evidence comes from Alderdice [19], who found that freshly fertilised chum (O. keta) salmon eggs at 10 °C were respiring 0.00093 mg O₂ egg⁻¹ h⁻¹, while hatching larva respiration peaked at 0.0052 mg O₂ egg⁻¹ h⁻¹. Rombough [26] found that pooled rainbow trout embryo and alevin metabolic rates ranged from 0.028 to 0.078 mg O₂ egg⁻¹ h⁻¹ when reared at 6 and 15 °C, respectively. Also, triploid stages of the same species presented a similar trend, spanning from 0.001 (eyed eggs), to 0.004 (hatched alevins), and to 0.03 mg O₂ egg⁻¹h⁻¹ (yolk sack absorbed) [27]. Finally, for Atlantic salmon, the DO demand ranged from 0.0067 [28] to 0.0048 mg O₂ egg⁻¹ h⁻¹ [29] for the hatching larvae under temperatures of 17 and 10 °C, respectively. The variability in the DO flux is thus related to different ontogenetic phases, reflecting different mass and metabolic activities, which, if not clearly pointed out, prevent the making of meaningful comparisons among different works [26,30]. Metabolic rates should be compared when eggs belong to the same stage and when they are exposed to the same level of stress, which are not guaranteed given the different scopes of studies. Indeed, it is well known that activity and stress, though not easily quantifiable or controllable, can greatly affect baseline metabolic rates, attained under ideal conditions [31,32]. For this reason, we believe that the higher variability in the alevin DO respiration rates, especially those of the last batch, can be attributed to the larger size and the increased mobility of the larvae in the restrained closed respirometry chambers with respect to the embryos. Similar problems arise when burrowing macrofauna or when larvae are incubated in the absence of sediments, where they feel uncomfortable and try to dig across the glass walls of the incubation chambers leading to unrealistic, overestimated metabolic activity, largely exceeding the baseline respiration measured when they are within sediments [33]. Another source of variability can be attributed to the usage in some works [19,26] of a mass unit instead of an egg unit, leading to an underestimation of metabolic rates if the inactive mass of the yolk sack is considered in the calculations. This is especially true for the early stages, when the inactive mass of the yolk sack can represent a major fraction of the embryo mass. However, for those works that report metabolic activity in relation to the whole embryo, inter-genera differences in egg size [6,34,35] can explain the variability among respiration rates. Finally, different incubation temperatures inevitably affect metabolic rates and thus the DO demands; for example, Atlantic salmon eggs at the hatching stage incubated at 17 and 5 °C nearly halved their metabolic activity, shifting from 0.0067 to 0.0039 mg O₂ egg⁻¹ h⁻¹ [28].

Ammonium excretion at the eyed stage averaged 0.23 ± 0.07 μg N-NH₄⁺ egg h⁻¹, which is substantially lower (p = 0.001) than the 0.69 ± 0.20 μg N-NH₄⁺ egg⁻¹ h⁻¹ of the alevins with yolk sack, which, in turn, differed significantly from the excretion of the stage with a completely absorbed yolk sack (3.60 ± 2.0 μg N-NH₄⁺ egg⁻¹ h⁻¹, p = 0.04). The increments in metabolic rates, following the ongoing developmental process and the increase in the biomass, explains the increased excretion in the late stages [36]. We speculate that the variability among rates can be justified by the same reasons discussed earlier for DO fluxes, related to the experimental conditions, the size and some stress increasing along with the different stages. Smith [30] was a pioneer in evaluating NH₄⁺ excretion rates from rainbow trout eggs, ranging from 0.1 for early stages to 0.7 μg N-NH₄⁺ egg⁻¹ h⁻¹ for alevins. Such a range of values includes the excretion rate reported for the pre-hatching stages of the same species by Noronha et al. [37] (0.16 μg N-NH₄⁺ egg⁻¹ h⁻¹). Teles [27], incubating diploid and triploid rainbow trout eggs, measured an increment in the NH₄⁺ excretion rates from 0.08 (early eyed) to 0.25 (hatching phase), to a peak of 1.5 μg N-NH₄⁺ egg⁻¹ h⁻¹ (eggs with an exhausted yolk sack). Besides the aforesaid factors dictating diversity in DO respiration activity, for NH₄⁺, an additional variable can be represented through pH changes. Indeed, the NH₄⁺ excretion is primarily driven by the diffusion of the unionised ammonia form (NH₃), owing to a partial pressure gradient, while the ionised ammonium form (NH₄⁺) cannot diffuse across the almost impermeable chorion [37]. Due to its pK of 9.5, the NH₃ will more easily diffuse into acidic compartments [38], and this can double the excretion rates from eggs incubated at pH 10 when they are incubated at lower pH values (e.g., 6 to 8) [37].