An estimated 69% of the population of Bhutan is engaged in agriculture. Farmers are exposed to a wide variety of pesticides during the preparation, transport, storage, mixing and application of pesticides posing a significant health risk. A controlled cross-sectional study of farmers in selected sites of Bhutan was conducted to characterize the level of exposure to pesticides and assess their knowledge attitude and practice on the safe handling of pesticides. A total of 399 participants were enrolled in the study comprising of 295 exposed farmers and 104 healthy and unexposed controls. A structured investigator administered questionaries was used to assess their Knowledge, Attitude and practice, and their blood samples were taken for measuring Acetyl Cholinesterase enzyme activity level. There was a significant difference between the Acetyl Cholinesterase enzyme inhibition of exposed and non-exposed control groups observed in the study (P< 0.001). Of the total of 295 farmers, 62 (21.01%) had severe enzyme inhibition of >30% as compared to the unexposed group. Safety practices of handling pesticides were low. The most common symptoms self-reported were headache (OR 1.08, 0.60–1.93) and neurological problems like forgetfulness, lack of concentration (OR 1.12, 0.50–2.48) and increased tiredness (OR 1.075, 0.52–2.19) that were significantly associated with the enzyme inhibition. In addition, we record a very low level of knowledge (17.0%), a fair attitude (63.0%) and poor practice (35.0%) on the safe handling and management of pesticides. This pilot study provides indication of exposure to pesticides in the selected sites of the country. Furthermore, it provides evidence for public health interventions by identifying the exposure patterns and pathways of individuals most at risk in the farming communities of the country. Surveillance and bio-monitoring programs are deemed necessary.