Screening for antibiotics active against intracellular bacteria.

A large number of useful antibiotics have been discovered since the isolation of tyrothricin in 1939, yet as time passes and as the number of known antibiotics increases, their rediscovery becomes more and more fre(uent. Improvements in screening techniques are therefore indicated to eliminate undesired antibiotics with a minimum of labor. New techniques should also attempt to recognize biological activities that previously used screening methods were unable to detect. One of these activities concerns effectiveness against intracellular bacteria. Suter (1952) has shown that tubercle bacilli enclosed in phagocytes are as susceptible to isonicotinic acid hydrazide as those suspended in Tween-albumin medium. In contrast, these phagocytized cells of Mycobacterium tuberculosis are about 100fold more resistant to streptomycin than their extracellular counterpart. Rtesults like these suggest that direct screening against intracellular bacteria may have the following advantages: (a) Only substances capable of entering mammalian cells without inactivation would be detected. (b) Substances toxic to mammalian cells would be rejected. In addition, the possibility exists that certain substances may exhibit an antimicrobial action only intracellularly. The development of a technique for screening within the mammalian cell could furnish the necessary tool for the detection of such intracellular factors. Because of their very slow growth, pathogenic mycobacteria are not ideal laboratory tools. Members of the genus Brucella have the advantage of being pathogens with intracellular habits and they grow relatively fast, readily permitting quantitative enumeration. The use of brucellae was warranted, since the control of brucellosis is still a problem (Spink, 1956). However, brucellae were used primarily as a test system which might yield substances of value in many other fields, including the control of virus diseases. The method described by Pomales-Lebron and Stinebring (1957) for the quantitative study of intracellular brucellae was thus the foundation on which the present screening method was built. The efficiency of this method was tested by screening a series of filtrates from actinomycetes isolated at random from various soils.