Introduction: To avoid the spread of communicable diseases, infection control and preventive measures are essential in all health care settings. Every health care professional, especially nurses, plays an important role in reducing the risk of health care-associated infections. Our goal is to examine the knowledge and attitudes regarding infection control and prevention measures. Methods: A descriptive study was carried out on a convenience sample at King Khalid Hospital and New Najran General Hospital in Najran city, using a self-administered questionnaire to examine the knowledge and attitude of nurses regarding infection control and prevention measures. Results: According to the study, 76.7% of the individuals had good knowledge, while 23.3% had poor knowledge. The majority of participants (78.8%) had a positive attitude toward infection control and prevention measures, whereas 21.2% had a negative attitude. There were statistically significant variations based on total years of experience, gender, education level, and the occurrence of needle stick during years of employment. However, due to the years of experience in the surgical wards, there were statistically significant variances. Conclusion: More training programs and seminars should be held to raise awareness and improve attitudes toward infection control practices and preventative measures.