THE CULTURE OF THE SULPHUR BACTERIA 55 



3. The coloured sulphur bacteria require more sulphuretted 



hydrogen than the colourless forms. 



4. Only a trace of organic matter is tolerated. Relatively 



large amounts are toxic, e.g. O-l per cent, peptone 

 was found to be injurious. 



5. The oxygen requirement is very small. 



6. Exposure to light is necessary for the growth of the 



purple sulphur bacteria. 



These conclusions are of special interest because they 

 have formed the starting-point of later researches. Although 

 Winogradsky states that only small quantities of organic 

 matter are tolerated it is noteworthy that in all the experi- 

 ments given above, with the exception of {c), the fluid in which 

 the bacteria multiplied must have contained soluble organic 

 matter in quantity. 



The next important investigation is that of Molisch (3), 

 who cultivated both fresh- water and marine forms. For fresh- 

 water forms his general procedure was as follows. The material 

 was covered with water from the river Moldau, at Prague, 

 and included such substances as hay, boiled egg, bones, earth- 

 worms, snails, and peptone water. It was left to decompose 

 and examined periodically for its bacterial content. For 

 marine species such substances as Zostera (sea-wrack), a 

 bit of crab, or lish, or some other remains, were placed in sea- 

 water which had been brought from Trieste. In some of his 

 cultures a layer of oil was spread over the surface to limit the 

 supply of oxygen. Molisch obtained an abundant supply of 

 both colourless and coloured sulphur bacteria, as well as purple 

 coloured sulphurless bacteria. He was successful in obtaining 

 pure cultures of the last named,* but not with the sulphur 

 organisms. Contrary to Winogradsky, he considered that a 

 larger amount of organic matter was necessary for the growth 

 of these organisms. He confirmed, however, Winogradsky's 

 conclusions that oxygen can be utilized only at diminished 

 pressure, and that exposure to light is necessary for the coloured 



* The cultivation of the sulphurless purple bacteria was first ac- 

 complished by Esmarch when he isolated Spirillum rubvum. 



