"To appreciate, therefore, according to this method, the pollution, and in general the degree of corruption of a water, the examination ought to be begun immediately on taking the sample. In such an appreciation we ought also to take account of the increase of microbia during the flow of the waters, to the end that an extraordinary number of microbia may be attributed either to a natural increase or to an incidental pollution. With respect to the five microbia per cubic centimeter contained in the Maugfall water at the moment of its arrival in Munich (which from its source to its arrival at Munich takes about twenty-four hours), it has been observed that in this case the figure is not augmented during the course of the water. It has been observed, in fact, that the Maugfall water arrives at Munich under a pressure of five to six atmospheres. It is thence admitted, with much probability, that the vitality of the microbia is abated under this pressure. Dr. Karl Lehmann has experimentally demonstrated that such an influence is exerted upon many of the lower organisms by a strong pressure of oxygen. Prof. Maggi, of the University of Pavia, has found that the water of Lake Maggiore at certain depths no longer contains bacteria.

"As a rapid alteration of the hygienic conditions of a water results from the rapid increase of microbia, it seemed to me of no trifling interest to examine the behavior of carbonic waters which are ordinarilv drunk in a period more or less long from their preparation. For these researches there were prepared ordinary bottles of carbonic water (water saturated with carbonic acid under pressure), and at the same time there were taken as a check samples of the potable water which served for their preparation. Care was taken to use sterilized bottles and stoppers. As for the apparatus for the carbonic water, the water-receiver was always kept filled during the preparation of our samples. Portions both of the carbonic water and of that unprepared were submitted to cultivation, to fix the initial conditions of the experiment. From these cultivations it resulted that the carbonic water contained 186 microbia per cubic centimeter, and the original water only 115. Upon each of the two waters were made comparative examinations at intervals of five days for a period of fifteen days.

"In these researches it was found that while in the non-carbonic water the number of microbia rose in five, ten and fifteen days from hundreds to thousands, in the carbonic water the number of microbia not only did not increase, but it diminished. In five days the number of organisms had fallen from 186 to 87; in ten days, to 30; and in fifteen days, to 20.

"This absence of increase in the carbonic waters may be due to one of the following causes: 1, action of carbonic acid; 2, action of pressure; 3, joint action of carbonic acid and pressure; 4, deficiency of oxygen. We may set pressure aside. I admit, indeed, that it may be sufficient to hinder the development of microbia, but in our case it is not necessary. In examining three qualities of carbonic mineral waters, Giessel, Selters, and Apollinaris, which were under very slight pressures, I have always found a scattered quantity of organisms which went on decreasing. But the decisive proof for excluding the necessity of pressure is in the researches made on carbonated water prepared at an ordinary pressure.

"Into Maugfall water contained in sterilized bottles I caused to bubble for half an hour, with occasional stirring, a current of carbonic acid developed by the action of hydrochloric acid upon calcium carbonate. The carbonic acid before passing into the water under examination was made to pass into two bottles containing solutions of sodium carbonate, to remove the traces of hydrochloric acid which may have been mechanically carried along by the current.

"The carbonic water being thus prepared, the bottles were closed with ground glass stoppers secured with a layer of paraffin. The water being left in this condition, it resulted from researches made in the period of fifteen days that in this case also the quantity of microbia did not increase, but diminished. Pressure being thus excluded, there remained only as the cause hindering the increase of the microbia either the action of the carbonic acid or the want of oxygen. But it has been possible, also, to exclude oxygen. Into the same Maugfall water contained in sterilized bottles there was passed for an hour a current of hydrogen, taking care to stir. The hydrogen, generated by the action of dilute sulphuric acid upon zinc, was washed by a passage through a solution of caustic potash. The bottles thus prepared were hermetically closed, and the water examined from day to day. But the organisms in this water, which, as regards oxygen, would be in the same condition as the carbonic water prepared at the ordinary pressure, increased rapidly and similarly to the microbia in water which was in free contact with the atmosphere.

"These results place it beyond doubt that atmospheric oyxgen is not an element necessary for the increase of microbia in drinking waters, and that the carbonic acid is the sole agent which interferes with the life of these organisms in carbonic waters".