Background: Microorganisms are mostly transmitted through the hands of healthcare workers in any healthcare setting. This transmission can be from healthcare workers to the patients and from their surrounding environment. This ultimately leads to increased healthcare-associated infections and to the resistance against the antimicrobials and the cost of healthcare-associated infections increases accordingly. Objectives: To observe the practice of hand hygiene and to calculate the hand hygiene compliance rate among healthcare workers of Ministry of Health Hospitals in the Kingdom of Saudi Arabia. Methods: The hand hygiene practice of healthcare workers in 318 Ministry of Health hospitals was observed by direct observation during the year 2018. This was done as a part of the National Hand Hygiene Program during the year 2018. The data was collected using the World Health Organization Hand Hygiene observation form. The compliance rate was calculated among different healthcare professionals, hospital departments and work shifts. Results: A total of 511,079 opportunities were observed over a period of one year. The overall hand hygiene compliance rate was found to be 70%. The compliance was found to be highest after the blood/body fluids exposure (85%), followed by after touching the patients (76%), followed by clean aseptic procedures (75%), after touching the patientâs surroundings (66%), and before touching the patients (64%). The overall hand hygiene compliance was found to be higher in nurses (73%) as compared to the doctors which was 67%. Conclusion: Our findings confirm that hand hygiene compliance is higher in the nurses as compared to the physicians and the rest of the health care workers in MOH health care settings. Initiatives such as improved monitoring and training healthcare facilities are needed to improve hand hygiene compliance in future.