Folia Microbiol. 38 (1), 25-28 (1993) Antimicrobial Activity Determined in Strains of Bacillus circulans Cluster C. PEREZ a, C. SUAREZ b and G.R. CASTRO c* acdtedra de Farmacologia and bCdtedra de Quimica Biol6gica, Facultad de Odontologia, Universidad de Buenos Aires, 1122 Buenos Aires CCdtedra de Microbiologia Superior, Facultad de Bioquirnica~ Quimica y Farmacia, Universidad Nacional de Tucumdn- PRO1MIMIRCEN, 4000 Tucumdn, Argentina Received October 12, 1992 ABSTRACT. Wild-type strains of the genus Bacillus were screened for antimicrobial activity. Two strains exhibited antimicro- bial activity against Micrococcus luteus and were identified as Bacillus polymyxa MIR-23 and Bacillus circulans MIR-13. Bacillus polymyxa MIR-23 was active against Escherichia coli, Pseudomonas aeruginosa and Aspergillus niger, whereas Bacillus circulans MIR-13 did not show activity against these microorganisms. Bacillus subtilis ATCC 6633 and B. po/ymyxa ATCC 10401 were used as standard antibiotic-producer strain and showed different antimicrobial profiles and yields from B. pobyrnyxa MIR-23. The different antimicrobial profile of the two selected strains could possibly be used as a taxonomic marker. During the last ten years many multiple antibiotic/resistant microorganisms have been clini- cally isolated, and they are very difficult to treat with most of the single antibiotics. For this reason, search for wild-type microorganisms able to produce antibiotics and/or having broad antimicrobial activities is very important. It is also essential to determine to which antibiotics a pathogen is suscep- tible. The genus Bacillus produces several extracellular peptide antibiotics that are synthesized through different mechanisms (Nakano and Zuber 1990). For example, as a result of research in this field, industrial production of bacitracin and polymyxins has become possible. Most of the antibiotics produced by strains of Bacillus are active against Gram-positive bacteria or fungi. However, some microorganisms belonging to the circulans cluster produce several peptide antibiotics such as polymyxins and related compounds, to which Gram-negative bacteria are sensitive (Katz and Demain 1977). In particular, Gram-negative microorganisms, such as Escherichia coli, are associated with enteric infections and neonatal diarrheas, and Pseudomonas aeruginosa is an opportunistic pathogen resistant to multiple antibiotics and related to the hospital-acquired infections. The taxonomy of the genus Bacillus is very complex and genera can be divided into several clusters according to modern systematic studies that use numerical taxonomy. The Bacillus circulans cluster is a wide and heterogeneous group, in which Bacillus circulans, B. rnacerans, B. polymyxa and related species are included (Priest et al. 1988). In this paper, we describe the selection of two wild-type strains belonging to the Bacillus cir- culans cluster that show antimicrobial activity toward some microorganisms with clinical importance. We also describe the microbial characterization and the relationship between the time of antimicrobial production and bacterial growth at different temperatures. MATERIALS AND METHODS Isolation and strain characterization. Soil samples (1 g) from the field were suspended in 10 mL' of sterile physiological solution (NaCI, 15 mmol/L) and heated for 10 rain at 80 ~ Aliquots of the samples were added to Erlenmeyer flasks (50 mL) containing Mueller-Hinton medium (5 mL), and were incubated for 2 d at 37, 45 and 55 ~ Each culture was also spread on agar plates containing the same medium, and then incubated for 1-3d. Isolated colonies were purified by streaking on Mueller-Hinton agar three times. Colony morphology, microscopy and Gram staining were used as criteria to select strains of the genus Bacillus. The selected strains were characterized according to international criteria, with the Jacquard coefficient (Castro 1991). *Corresponding author.