Assessment of double ovulation to halve Xenopus laevis use for eggs

The African clawed-toed frog Xenopus laevis is a widely-used research animal owing to its large egg and embryo size and reliable production of large numbers of eggs. The standard Xenopus ovulation protocol involves hormone injection to induce females to lay eggs, with reuse following three month's rest. Animals used this way flourish and lay useful egg numbers for many years. It has been reported that after an ovulation, a second ovulation just one week later yields an unchanged number of eggs and that successive weekly ovulations show only gradual decline in numbers. This is consistent with reports that Xenopus in the wild mate multiple times in a season and suggests that Xenopus have significant egg-laying capacity above what is currently used.

We have piloted a Double-Ovulation protocol, re-ovulating frogs 7-10 days after a previous ovulation, followed by the standard 3-month rest. Preliminary observations show no signs of reduction in egg yield or health of the animals, despite doubling egg yield in a given calendar period. The proposed project will determine rigorously whether indeed doubling egg yield is possible while maintaining quality and without increasing stress to the animals, either acutely or over multiple cycles. We will directly compare single- and double-ovulation protocols, counting total eggs laid and measuring fertilisation rates. In addition to monitoring animal weight and general health, we will also, innovatively, determine stress levels in the animals by measuring secreted corticosterone levels using a newly established assay. We will assess both acute stress (during ovulation) and chronic stress (testing for build-up versus return-to-baseline between cycles). The results of this study will show whether it is possible to reduce the size of Xenopus colonies by half worldwide, potentially reducing the number of animals by half (up to 25,000 animals) while maintaining animal welfare and research productivity.