The capacity of Pseudomonas sp. to degrade different pesticides has been the object of numerous studies; due to this, its dynamics was evaluated and the effect of the application of glyphosate (Roundup®) in soils of Tolima, taxonomically classified as Entic haplustolls, Typic ustipsamments, and Inceptic haplustalfs with coverage of secondary forest, stubble and sorghum crop, respectively, through the in vitro 14C-glyphosate mineralization. These soils were subjected to presence/absence of heat, and were denominated with heat treatment (HT) and without heat treatment (WHT). In both cases a solution of glyphosate Roundup SL® and radioactive was applied. The statistical design was used at random and the response parameter was the quantification of Pseudomonas sp. viable cells (cfu) during the mineralization process, for the three soils up to 120 days. It was found that adding glyphosate stimulated the increase of Pseudomonas sp., correlated significantly with the production of 14CO2. The rates of mineralization and the bacterial populations in WHT soils were significantly (P≤0.05) greater with respect to HT soils. Finally, the higher averages for mineralization and bacterial dynamics were in the soils under use of secondary forest (63% and 26.8·107).