Biochemical Testing 22 appreciation of the techniques of molecular biology in its contribution to science and the mitigation of human suffering, while being reasonable in its use; and an unceasing advancement toward the development and implementation of new methodologies produced through scientific, technical, and social advancement. Phenotypic identification The phenotypic identification of mycobacteria is based on the characteristics of the culture and biochemical features, and comprises a

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... ge variety of tests, some of which are conducted in order to locate the microorganisms into two large groups: www.intechopen.com Biochemical Isolation and Identification of Mycobacteria 23 operons and the efficiency of their transcription. One or two rrn operons are found in mycobacteria, regulated by P1 and P2 promoters. The rrnA operon, located in the murA gene, is found in all species, including M. tuberculosis and M. leprae; but, in some species, like those of rapid growth, an additional rrnB operon is found, located in the tyrS gene (Arnvig et al. 2005 , Verma et al 1999 . Colony morphology Mycobacteria in culture generally appear as two types of colonies: rough or smooth, and with a shiny or opaque aspect (Tasso et al 2003). Pigment Some species of mycobacteria, classified as photochromogens, produce carotenoid pigment in the presence of light; the group designated as scotochromogenic develops yellow colonies independent of light stimulus; the non-chromogenic group does not produce pigment; their colonies appear with shades of pale yellow or cream, in the presence of light or in darkness, and the color does not intensify when exposed to light (Bernardelli et al 2007, Belén et al 2007). Growth in presence of P-nitrobenzoic acid This test contributes to the differentiation of species of the M. tuberculosis complex. Pnitrobenzoic acid inhibits the growth of: M. tuberculosis, M. bovis, M. africanum, and M. microti (Palomino et al 2007). Growth in the presence of hydrazide of thiophene-thiophene-2-carboxilic acid Permits the differentiation of M. tuberculosis that grows in the presence of the compound as opposed to other species such as M. bovis; it is sensitive to the compound when added to the Lowenstein Jensen (LJ) medium. Some species of NMT are positive for this test. Growth in McConkey agar without violet crystal Mycobacteria such as M. fortuitum, M. chelonae, and M. abscessus have the ability to grow in McConkey agar without violet crystal, as opposed to other mycobacteria such as M. tuberculosis and M. bovis that cannot grow in this medium. Growth in 5% sodium chloride Species of mycobacteria such as M. fortuitum show the capacity to grow in a LJ culture medium to which has been added 5% sodium chloride. Growth in the presence of hydroxylamine This test plays a very important role in determining the difference in capacity of growth of species such as M. bovis in LJ media supplemented with a concentration of 250 mg of hydroxylamine. NTM does not have this capability.