CULTURAL REACTIONS 817 



in 2 to 3 days, deepening to a cafe-au-lait or chocolate tint in the course of a fort- 

 night. Individual strains vary in the depth of colour they produce, some giving 

 a darker brown than others. The pigment is not confined to the layer of growth ; 

 it spreads throughout the potato. It will be recalled that a brown growth on potato 

 is also given by Pf. mallei, Pf. whifmori, V. cholera, and Ps. pyocyanea, and certain 

 other organisms. A similar but less intense brownish colour is sometimes notice- 

 able in old agar slope cultures, particularly of Br. melitensis (Kristensen 1931) ; it 

 is not sufficiently constant, however, to be of differential value. 



Huddleson, Hasley, and Torrey (1927) and Huddleson and Winter (1927) have observed 

 the development of crystals of ammonium magnesium phosphate in cvdtures of Brucella 

 on liver agar incubated aerobicaUy, but not in 5-10 per cent. COj. They regard their 

 formation as being due to the production of ammonia by the growing organisms, and 

 to its combination with the magnesium phosphate in the medium. In their experience 

 crystals are formed much more rapidly by melitensis and paramelitensis than by abortus 

 strains. Om* own experience (Wilson 1933) bears out their observations to some extent, 

 but does not suggest that the differences between different types are sufficient to be of 

 value in the identification of individual strains. Incidentally American suis strains 

 appear to be most active m the production of ammonia. 



Attention has been drawTi by de Santis (1933) to the different appearances presented 

 on PetragnanVs egg medium. Strains of Br. melitensis are said to grow on this medium, 

 and usually to change the colour from Ught yellowish-green to dark green. Br. abortus 

 strains, on the other hand, generally fail to develop. The behaviour of Br. suis strains 

 is doubtful, but in our Hmited experience growth is not infrequent. The reliabhity of 

 this test for differential purposes is stUl under discussion (see Menzani 1934, Tosatti 1934, 

 Messieri 1935, Vittone 1935, Pagnini 1935, Foresti 1935). The general opinion seems 

 to be that it is less satisfactory than the HjS and dye tests (see later). According to 

 Schwarzmaier (1936), Br. melitensis grows on Petragnani's medium both. with and without 

 malachite green, Br. suis grows only without malachite green, and Br. abortus fails to 

 grow under either condition. The mode of preparation of the medium appears to be 

 of importance. If it is heated to a temperature above 80° C. for too long the normal 

 inhibitory action of the egg albumin on the bovine and porcine strams is destroyed, thus 

 allowing some development of these organisms to occur (de Santis 1935). According 

 to Martini (1935), the inhibitory effect of the egg albumin on abortus strains can be removed 

 by the addition of sufficient HCl to lower the reaction of the finished medium from pH 8-4 

 to pH 6-8. If this is true, it suggests that the effect may be related to the greater necessity 

 of abortus strains for COg. 



Growth is rather slow, and unless a fairly heavy inoculum is made, colonies 

 are not usually visible for 2 days or even longer. In broth the maximum turbidity 

 is not reached for a week or more. On the whole the American porcine strains 

 probably g've the best, and the Danish porcine strains the poorest growth, the 

 melitensis and abortus strains occup5dng an intermediate position. The behaviour, 

 however, of different strains of the same type is subject to so much variation 

 that no reliance can be placed on this character for differential purposes, 



Br. abortus, Br. melitensis and Br. suis, when inoculated on to the chorio- 

 allantoic membrane of the developing chick embryo, are able to multiply and to 

 bring about death of the embryo in a few days with lesions in the spleen and 

 liver. All three organisms grow intracellularly — Br. melitensis in the ectodermal 

 epithelium, Br. abortus and Br. suis in cells of mesodermal origin and in the vascular 

 endothelium. Rough strains are non-invasive (Goodpasture and Anderson 1937, 

 Buddingh and Womack 1941, de Ropp 1944). 



