This paper covers experiments which were made with the object of developing suitable media for use in raising anopheline larvae. Two types of media were sought: “routine,” for the purpose of obtaining adults; and “standard,” to serve as a basis for special experiments with larvae. The chief requirements of the routine medium would be simplicity and reliability, and these seemed to be met best by a combination of mud, rain water, and bread crumbs. This medium proved itself well adapted to a wide variety of species and circumstances. A “standard” medium should be made up in such a way that it can be reproduced in any laboratory, so that larval experiments may be duplicated and compared; it would necessarily be based on distilled water. No entirely satisfactory medium was found, but “Medium S,” made up of calcium sulphate, sodium chloride, and magnesium sulphate, in the proportions of 0.5, 0.5, and 1.0 part per thousand respectively, made the best medium for early growth of Anopheles atroparvus, when dried bread crumbs were added as food. Of the substances tried, bread crumbs seemed to be the most satisfactory organic food both for routine and standard purposes.
It was found that the technique of keeping larvae for three days at a constant temperature of 27°, under circumstances as nearly standard as possible, could be adapted to making an index of the suitability of natural waters for anopheline breeding. Results with a few water samples are given. In the only case carefully studied, the unsuitability of the water seemed to be due to a high nitrate content. The addition of even very small amounts of nitrate to otherwise favorable media had an adverse effect on the growth of A. atroparvus.
From studies of the effect of soil (mud and sand), it was concluded that the mud used in the routine breeding method favored larval growth by furnishing necessary minerals in solution, by serving as a source of organic food, and by acting as an adsorbing agent for unfavorable materials.
All experiments were first made with A. atroparvus, since this species makes a convenient standard for European workers. Comparison experiments with other species showed that the routine method could be used for all of the species tried, and that the standard method was about equally favorable for atroparvus, labranchiae, maculipennis, and superpictus, but that it was inadequate for sacharovi. Superpictus was found to be more tolerant of nitrates than the other species, but more exacting in its mineral requirements.