Edward Bles, Part 1. Glasgow Winters and Xenopus Frogs

During the latter half of the 20th century African Clawed Frogs (Xenopus) became a major organism for research in developmental biology. They became widely used because for decades they were used for human pregnancy tests. Their use and their spread through hospital laboratories over the world depended on their reliability in responding to the injection of gonadotrophic hormones by laying eggs. The urine of pregnant women contains human chorionic gonadotrophin and the Xenopus pregnancy test became the best biological test before immunological tests were developed. Because they were so widespread and freely available, eggs and embryos for research could be obtained throughout the year by a simple injection in both females and males.

Before any of this was known or put into practice Xenopus and their embryos were studied after natural breeding of Xenopus in captivity. In the wild in South Africa, their breeding behaviour had been described in 1890. Live Xenopus laevis were coming into Britain from South Africa and were being kept successfully, often living for many years, by herpetologists in zoos and in private collection. For example, when Hans Gadow wrote his book in 1901 George Albert Boulenger of the Natural History Museum had kept some in his room in London for at least 11 years. Getting them to spawn though was a different matter. Boulenger’s were seen in amplexus but eggs never appeared.

The first recorded breeding of Xenopus in captivity appears to have been at London Zoo in 1893. The events and development of the resulting tadpoles were described by the Zoological Society’s prosector, Frank Evers Beddard (1858-1925). Eggs were laid in the evening of Saturday 27 May 1893; by Monday 30th they had hatched. It would seem that some were reared to metamorphosis since 5 were admitted to the Zoo’s books on 30 December.

Xenopus tadpole (from Beddard's paper)

Twenty-one specimens of Xenopus had arrived in London Zoo in 1890-93 from several collectors. Beddard states in his paper that those which bred were collected by Frank Finn (1868-1932) in Zanzibar. I will return to this point.

The person associated with first devising a system for breeding Xenopus and in describing their development is Edward J. Bles. In Cambridge Bles was keeping Xenopus but only when a pair was moved into the tropical tank at Cambridge Botanic Gardens did spawning occur. That was in February 1899.

It was while Bles was working in Glasgow that he developed an apparently reliable method for breeding Xenopus although it seems that the female which bred was the same one as in Cambridge. By mid-1903 he was not short of material for embryology. Between April and July 1903 around 15,000 eggs were obtained.

Bles considered it essential that the frogs should be allowed to hibernate. It is easy to forget just how difficult it was in buildings without central heating or electricity to keep tropical aquatic animals warm or to arrange for seasonal changes. In Bles’s time at the University of Glasgow, the zoology department occupied the dark basements and cellars below the Hunterian Museum. To house his frogs, Bles used an arrangement of an inverted bell jar held over a galvanized iron water bath heated from below by a gas micro-burner made by Zeiss. It had been devised by his friend in Cambridge, John Samuel Budgett (1872-1904), a fellow amphibian expert, for keeping tropical fish as well as aquatic frogs. Bles kept the summer temperature at around 25°C but in winter the water in the bell jar was kept at 15-16°C by day, falling to 5-8° at night.

Bles's aquarium for Xenopus

Bles made the point that with a number of amphibians, hibernation appeared essential for the induction of breeding. However, he also arranged for water to be sprayed back into the inverted bell jar aquarium after cooling so as to simulate falling rain. Both these ‘tricks’ are still used to induce frogs and toads to breed.

Bles wrote a long and detailed account of the development of Xenopus, beautifully illustrated by A. Kirkpatrick Maxwell (1884-1975). Bles had been introduced to Maxwell by a friend in Glasgow. It was a lucky break for Maxwell; he was apprenticed to a lithographer and attending classes at the Glasgow School of Art. After his work for Bles  he went on to have an highly successful career as a medical illustrator for the army in the First World War, for example; his drawings were a major improvement for Gray’s Anatomy and he contributed to many other textbooks and journals. His fame extended into the age of molecular biology. Max Perutz commissioned him to draw the model of the haemoglobin molecule.

One of Kirkpatrick Maxwell's plates for Bles
There was an overlay of tracing paper with the lettering

At this stage I must point out that the breeding of Xenopus at London Zoo in 1893 and by Bles in Cambridge and Glasgow between 1899 and 1903 could not have been the same species. All were described at the time as belonging to the species, Xenopus laevis, the Smooth Clawed Frog, and nobody I have found has questioned that assignation. However, the Xenopus aficianados who are reading this article will realise that X. laevis does not occur in Zanzibar. Therefore, and assuming that Beddard had correctly identified the pair collected by Frank Finn, London Zoo bred Müller’s Clawed Frog, Xenopus muelleri (first described by Peters in 1844) which does occur in Zanzibar. Bles states that his animals, two males and two females, came from dealer who had had them for two years. Their previous history was unknown. However, Bles did note that in his frogs the surviving basal portion of the tentacle was very short—a characteristic of X. laevis. By contrast, the tentacle of X. muelleri is as long as the diameter of the eye. Therefore, I think it is safe to assume that Beddard described the breeding and development of X. muelleri while Bles worked on X. laevis. The point is important since the methods described by Bles for X. laevis might not—indeed probably would not—apply to X. muelleri. It is difficult to imagine frogs from Zanzibar, where temperatures range from 20 to 33°C, tolerating a period of hibernation.

Dr Frederic Lionel Vanderkamp (1914-1993) who later became well-known for his research on tsetse flies appears to have been employed in some capacity at Bristol Zoo in the mid-1930s. He wrote an article in The Aquarist magazine questioning whether Bles had failed to discover the real reason for his successful breeding of X. laevis:

…although numerous other zoologists carried out un every detail his methods…they were unsuccessful. The reason, no doubt, was that Bles carried out most of his work at Glasgow where the water is naturally soft (pH about 6.7), whilst other zoologists repeated his methods in their own districts where the pH value of the water was above 7.

It has been discovered, during course of experiment, that X. laevis will spawn, providing they are in fit condition, any time when the pH of their water is lowered under 7. In April 1934, when this species was first bred at Bristol Zoo, they spawned 12 hours after the pH of the water had been lowered, although previously they had been kept seven years together without attempting to mate. Again in December 1934, X. laevis mated about 12 hours after they had been placed in water in which the pH was under 7, similarly again during March this year. Try as we may, we have, so far, failed to get them to mate by any other method except lowering the pH of their surrounding water. It is probable that the Clawed Frog is unable to spawn in alkaline water.

It is difficult to square Vanderplank’s last statement with Bles’s first spawning of X. laevis in Cambridge where the water is hard and alkaline unless for some reason the water in the water lily tank in which they bred had been treated. Similarly, Dr Edward Elkan in his extensive work on the Xenopus pregnancy test in Britain had X. laevis breed in outdoor tanks in London, again not known for having soft, acidic water. In the wild X. laevis is known to breed in acid and alkaline, soft and hard water. I suspect that both Bles and Vanderplank had found that a sudden change in the environment, like water simulating rain, or a shift in pH, also like adding rain water, can be the trigger for reproduction in otherwise quiescent amphibians in captivity. The nature of the change often does not matter, just a change will do.

The observations and descriptions Bles made from fertilization to after metamorphosis have stood the test of time. For example, he showed that the tadpoles within two hours of starting to feed rise to the surface in order to fill their lungs (see my previous post on this topic here). He reasoned that air in the lungs fulfils not only a hydrostatic function but also a respiratory one since in warm water (which holds less oxygen) the tadpoles came to the surface for air more frequently. A similar increase in frequency was seen when the tadpoles were kept in a smaller volume of water, which would lead to hypoxic conditions. He also established that Xenopus tadpoles are suspension feeders, eating microorganisms and other small particles in the water; unlike the tadpoles of the typical frogs they do not scrape surfaces for food.

Bles’s major paper on Xenopus was regarded as a particularly fine piece of work, both by his contemporaries and by those reading it over a hundred years after its publication.

Kirkpatrick Maxwell's depiction of Xenopus tadpoles for Bles

Beddard FE. 1894. Notes upon the tadpole of Xenopus laevis (Dactylethra capensis). Proceedings of the Zoological Society of London 1894,101-107. 

Bles EJ. 1901. On the breeding habits of Xenopus laevis Daud. Proceedings of the Cambbridge Philosophical Society 11, 220-222.

Bles EJ. 1905. The life-history of Xenopus laevis Daud. Transactions of the Royal Society of Edinburgh 41, 789-821. 

Elkan ER. 1938. The Xenopus pregnancy test. British Medical Journal 2, 1253-1256. 

Vanderplanck FL. 1935. The effects of pH on breeding. Aquarist (May-June 1935), 135-136.

For further reading on the history of Xenopus frogs in research, this article contains all the key references:

Gurdon JB, Hopwood N. 2000. The introduction of Xenopus laevis into developmental biology: of empire, pregnancy testing and ribosomal genes. International Journal of Developmental Biology 44, 43-50.