84 West and Griffiths . — The Lime-Sulphur Bacteria 
growth owing to the length of time (twenty-four to forty-eight hours) occu- 
pied by each cell-fission. 
It has been found that a good fixative is 40 per cent, commercial 
formalin, which not only fixes the protoplast moderately well, but also 
removes the large globules of calcium carbonate, leaving only the proto- 
plasmic reticulum with the numerous smaller grains of sulphur embedded in 
the threads. Formalin of this strength has, however, a tendency to affect 
adversely the protoplasmic reticulum, since it has to act for some hours in 
order to remove the larger globules. On the other hand, dilute acetic acid 
removes the larger globules at once with slight effervescence, leaving the 
sulphur grains unaffected, but again there is a slight injury to the protoplasmic 
reticulum. 
The best fixative was one recommended by Professor J. B. Farmer, 
and consisted of a mixture of three parts of absolute alcohol and one part 
of glacial acetic acid. This not only removed the globules very rapidly, but 
also the reddish sulphur grains, and at the same time fixed the protoplasmic 
reticulum exceedingly well. By this method there was no appreciable 
shrinkage. 
Since the original note, two distinct species of Hillhousia have been 
recognized. In addition to the large one — H. mirabilis — there is a smaller 
species not more than half the size. This small species, which we propose 
to call H. palustris , is distinguished not merely by its size, but also by the 
more rounded segments during the process of cell-division. Its protoplasmic 
reticulum is very little smaller than that of H. mirabilis , and therefore the 
cell contains comparatively few meshes. 
Both species are widely distributed and appear to have rather different 
habitats. Doubtless they will be found to exist in many parts of the 
world. 
The Protoplast. After the removal of the globules of calcium 
carbonate by means of formalin, the protoplast is seen to consist of a more 
or less uniform and rather coarse reticulum. There is no trace of a nucleus, 
and the reticulum is evenly distributed throughout the cell, as originally 
figured. 1 The whole reticulum is very finely granular and appears to con- 
tain rather larger granules in the corners of the meshes. 
The sulphur grains are rather small (1-2 ^ in diameter), of a reddish 
colour, and are situated within the threads of the protoplasmic reticulum 
between the much larger globules of calcium carbonate. 
Dry-staining is carried out with difficulty owing to the highly resistant 
cell-wall. Specimens were fixed on cover-slips with 40 per cent, commercial 
formalin and allowed to dry in the air, the small amount of mucus on the 
outside of the organisms causing them to adhere. Fairly good stained pre- 
parations were obtained with safranin, iodine green, Ziehl’s carbol-fuchsin, 
1 West and Griffiths, 1. c., Fig. 19. 
