Waterhemp is one of the most troublesome weeds in the US and is spreading into Ontario. In 2014, a waterhemp population was not controlled with glyphosate in a field in Lambton County, ON. This population was the first confirmed glyphosate-resistant (GR) waterhemp in Canada. In 2015, waterhemp seeds were collected from 48 fields in Lambton (32), Chatham-Kent (2), and Essex (14) counties to determine the occurrence and distribution of GR waterhemp in Ontario. Waterhemp plants were grown in a greenhouse and sprayed when 10 cm in height. In addition to glyphosate (a group 9 herbicide), collected populations were screened for resistance to imazethapyr and atrazine, representing herbicide groups 2 and 5, respectively. Visual control estimates for biomass reduction were completed at 1, 3, and 5 wk after application. Glyphosate-resistant waterhemp was confirmed in 40 fields, representing 82% of all sampled fields from the three Ontario counties. Of the 49 populations collected, all were resistant to imazethapyr (group 2) and 76% were resistant to atrazine (group 5). Of all the populations tested, 61% of all samples were found to be resistant to all three herbicide groups. This study is the first to confirm GR waterhemp in Ontario.