In developing countries, urban areas may be at greater risk of pesticide exposure compared to semi-urban agricultural regions. To investigate this, concentrations of selected pesticides were measured in 81 human milk samples collected in urban Kolkata and semi-urban Nadia in West Bengal, India. Three classes of pesticides were investigated – legacy organochlorines and emerging pyrethroids and neonicotinoids. The average concentration of the majority of the chemicals (DDT, its metabolites, HCH isomers, bifenthrin, endosulfan), showed a clear urban > semi-urban trend. Compared with previous measurements in other Indian cities and developing nations, current HCH and DDT concentrations in urban Kolkata were high. These chemicals were detected in 100% of the samples in both the urban and the semi-urban region. Also in both regions, the Estimated Daily Intake of DDTs, HCHs, aldrin, dieldrin and the pyrethroid bifenthrin for breastfed infants exceeded the Tolerable Daily Intake in a number of samples. Three pyrethroids were detected in human milk samples in India for the first time. This indicates a shift in the usage pattern of pesticides in India from organochlorines to pyrethroids. These findings may be used to drive targeted regulation of pesticides in developing countries with similar histories of pesticide use.