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Introduction: Noise is one of the most extensive environmental factors affecting the general population. The present study is focused on the association between discomfort caused by noise and the incidence of certain diseases (ischaemic heart disease, stroke and hypertension). Materials and Methods: This cross-sectional questionnaire study, conducted in 10 cities in the Czech Republic, comprises two stages with 3592 obtained questionnaires in the first phase and 762 in the second phase. Twelve variables describe subjective responses to noise from different sources at different times of day. The intensity of the associations between variables was measured by correlation coefficient. Logistic regression was used for fitting models of morbidity, and confounders such as age and socio-economic status were included. The hypotheses from the first phase were independently validated using data from the second phase. Results: The general rates of noise annoyance/sleep disturbance had greater correlation with traffic noise variables than with neighbourhood noise variables. Factors significantly associated with diseases are: for hypertension − annoyance by traffic noise (the elderly, odds ratio (OR) 1.4) and sleep disturbance by traffic and neighbourhood noise (the elderly, OR 1.6); for ischaemic heart disease − the general rate of noise annoyance (all respondents, OR 1.5 and the adults 30–60 years, OR 1.8) and the general rate of annoyance and sleep disturbance (all respondents, OR 1.3); for stroke − annoyance and sleep disturbance by traffic and neighbourhood noise (all respondents, OR 1.8). Conclusion: Factors that include multiple sources of noise or non-specific noise are associated with the studied diseases more frequently than the source-specific factors.