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Micropropagation is one of the main applications of tissue culture of plants. It allows rejuvenation and, consequently, improves explant rooting of woody species. However, one of its main problems is the risk of microbial manifestation. Here, we aimed to evaluate the survival and the reduction in microbial manifestations on in vitro cultures of Eucalyptus microcorys with the addition of chemical agents (sodium hypochlorite or streptomycin) to WPM culture medium. We used in vitro cultivated explants from adventitious shoots from two E. microcorys mother plants, over at 44 years-old. We drew from the multiplication phase explants with clumps of 8 to 12 shoots that displayed bacterial manifestation and submitted them to microbe control, adding different chemical agents at different concentrations to the culture medium: sodium hypochlorite (NaClO) and the antibiotic streptomycin. The results indicated that the best concentrations to reduce the manifestation of microorganisms on in vitro cultivation were 100 mg L− 1 of streptomycin and sodium hypochlorite with 0.003% of active chlorine, since these concentrations maintained 100% of the explants alive and reduced microbial manifestation more efficiently.