Pesticides, which are widely used in tropical areas for controlling disease vectors, require the services of urban pest control operators (fumigators). Pesticide use and handling policies and regulations mostly relate to agricultural uses, not to urban pest control purposes. Our objectives included investigating use patterns, pesticide management practices, knowledge on related policies in urban pesticide applicators of southeastern Mexico, and comparing butyrylcholinesterase (BuChE) activity in this group with individuals non-occupationally exposed to pesticides. We conducted a cross-sectional study where all participants were males. The exposed group was integrated by 27 urban pest control operators and the non-exposed group by 26 non-sprayer subjects. A structured questionnaire was applied to participants and BuChE activity in serum was determined. Urban pest control operators had been working with pesticides for 1-17 years, mostly during summer; 67 % did not use personal protection equipment. Eighteen active ingredients were employed as pesticides; chemical groups included pyrethroids (PYR), organophosphates (OP), coumarins, phenylpyrazoles, carbamates (CAR), chloronicotinyls, and amides. Policy knowledge and application were reported in only 30 % of the fumigators. BuChE activity (U/L) was similar between groups: 9837.95 ± 1844.46 U/L in urban pest control vs. 10 414.81 ± 1558.42 U/L for the non-sprayer group. We suggest the inclusion of additional biomarkers in the evaluation of occupational exposure to pesticides, because currently only OPs and CARs are considered, while fumigators are exposed to many other pesticides. Urban pesticide applicators lacked knowledge regarding pesticide management and application policies, hence the need for educational courses for workers and updating regulations.