The purpose was to measure the prevalence and related risk factors of low back pain (LBP) among health care workers (HCWs) at different levels of health care in southwestern Saudi Arabia. A cross-sectional study using a self-administered questionnaire was conducted among HCWs providing primary, secondary and tertiary health care services in the Aseer region, southwestern Saudi Arabia. The questionnaire collected data regarding having LBP in the past 12 months, socio-demographics, work conditions and history of chronic diseases, regular physical exercise and overexertional back trauma. Univariate and multivariable logistic regression analyses were performed. Out of 740 participants, the overall prevalence of LBP in the past 12 months amounted to73.9% (95% CI: 70.7–77.0). The prevalence of LBP with neurological symptoms reached 50.0%. The prevalence of LBP necessitating medications and or physiotherapy was 40.5%, while the prevalence of LBP requiring medical consultation was 20%. Using multivariable logistic regression, the following risk factors were identified: working in secondary and tertiary hospitals (aOR = 1.32, 95% CI:1.01–1.76), increased BMI (aOR = 1.10, 95% CI:1.01–3.65), and positive history of overexertional back trauma (aOR = 11.52, 95% CI:4.14–32.08). On the other hand, practising regular physical exercise was a significant protective factor (aOR = 0.61, 95% CI: 0.42–0.89). LBP is a common problem among HCWs. Many preventable risk factors have been identified, including exertional back trauma, increased BMI and lack of regular physical exercise. Occupational health and safety programmes to build ergonomically safe working conditions and encourage regular physical exercise are needed.