Resistance of the eggs to aquatic moulds
Many researchers have tried to breed Alytes larvae from eggs placed in water, but they failed as all eggs died from Oomycete infections (De L'Isle du Dreneuf, 1873; Fischer-Sigwart, 1885; Boulenger, 1912; Nijs, 1985). Midwife toads in nature breed on land and are normally not exposed to the water dwelling moulds. Moreover, soon after an egg is laid, the exterior gelatinous layer toughens and forms a cover that protects the egg from fungi. So, under natural conditions, they are not exposed to the fungi that attack amphibian eggs in water. Alytes has no recent co-evolutionary history with aquatic fungi such as Saprolegnia and, as a consequence, no resistance against them. Kammerer admits that mortality of Alytes-eggs placed in water is high: “Ultimately, the eggs stay behind in the water, where, indeed, most of them perish (in the first breeding attempt of that kind), but a few of them then still continue their development.” (Kammerer, 1919:326). He claims that mortality decreases with subsequent breeding attempts of the same animals: “Later, the results improve considerably; in later generations of animals with completed instinct variation, the mortality of water eggs is hardly higher than that of other frog eggs that are normally laid in water.” (Kammerer, 1919:356). The only possible mechanism for such a rapid decrease in egg mortality would be that eggs became resistant to Oomycetes. There is substantial evidence that resistance of amphibian eggs to Oomycetes has a genetic basis and that maternal effects do not represent a major contribution to variation in infection of eggs and embryos (Sagvik et al., 2008a, b; Ault et al., 2012; Urban et al., 2015). We fail to recognize any plausible mechanism for how subsequent clutches of the same mother could become increasingly resistant. Kammerer’s (1909) experiments were started with a moderate number of adults, the F1 generation suffered massive mortality, resulting in a genetic bottleneck. Thus, selection in his experiment cannot have resulted in massive resistance against a community of Oomycetes species within one or two generations. It is even more difficult to understand how the epigenetic activation or the inactivation of one or more genes could confer resistance of eggs against pathogens to which they have not been exposed before. Hence, it is simply hard to believe that Kammerer succeeded in breeding the large numbers of water-breeding midwife toads he mentions in 1906 and in his crossing experiments (Kammerer, 1911).