Background: Yukon, a territory in northern Canada, has one of the highest reported sexually transmitted chlamydia infection rates in the country. Objective: We examined screening practices among physicians and community nurses to elucidate factors that may be contributing to the high rates. Design: Cross-sectional survey. Methods: A questionnaire was distributed to all physicians in Yukon and all community nurses in Yukon's communities. We surveyed sexual health assessment frequency, chlamydia testing frequency and barriers to screening. Comparison of physician testing practices was performed to another Canadian jurisdiction, which previously undertook a similar survey. Survey results were compared to the available laboratory data in Yukon. Results: Eligible physicians and nurses, 79% and 77%, respectively, participated in the survey. Physicians tested 15 to 24-year-old females more frequently than 15 to 24-year-old males for chlamydia (p=0.007). Physicians who asked sexual health assessment questions were more likely to test for chlamydia in both females (p<0.001) and males (p=0.032). More physicians screened females based on risk factors compared to males. General practice physicians in Yukon were more likely to test females for chlamydia than general practice physicians in Toronto, Canada (p<0.001). Community nurses had different testing patterns than physicians, with a lower overall frequency of testing, equal frequency of testing males and females, and in applying risk factor-based screening to both males and females. Barriers to screening included testing causing patient discomfort, patients reluctant to discuss screening, health provider uncomfortable conducting sexually transmitted infection tests and sexual health assessments, among others. Laboratory data in Yukon appear to confirm provider screening patterns. Conclusions: This survey provides valuable information on health provider screening patterns. We have some evidence which suggests that chlamydia testing rates may be higher among patients seen by physicians in Yukon in comparison to another Canadian jurisdiction. However, more consistent application of optimal screening methods with support to “start the conversation” around sexual health may assist in overcoming barriers to screening and in addressing Yukon's high rate of chlamydia.