Background Italy was the first country in Europe to face the coronavirus pandemic. Healthcare workers (HCWs) were at higher risk of contracting COVID-19, because of their closer contact with patients. The present study aimed to analyze HCWs’ knowledge, practices, and attitude towards COVID-19.MethodsWe set up a cross-sectional survey through SurveyMonkey® and circulated the link in Facebook and Whatsapp closed groups. It was conducted in Italy from the end of February to the first week of May 2020. The study participants were general practitioners, pediatricians and other health professionals. Data were collected using a well-structured questionnaire including demographic, scope of knowledge, awareness and practice assessment section.ResultsA total of 958 participants were included. Of these, 320 (33.4%) were general practitioners, 248 (25.9%) were pediatricians and 390 (40.7%) were other health professionals. The highest response rate was from Northern Italy (48.1%), followed by the Center (29.9%) and Southern Italy (22.0%). The reported risk of having contact with a patient affected by Covid-19 was higher in the lockdown period than the pre-lockdown period (19.4% vs 10.2%, p=0.002). Respondents reported to have changed their clinical practice, more during lockdown period (81.1%) than in the pre-lockdown (46.3%). Particularly, they increased the use of masks (87.1%, p<0.001), disinfection and sanitization of doctors’ offices (75.8%, p<0.001), the use of protective glasses (71.2%, p<0.001), alcoholic hand solution (71.2%, p<0.001), and hand washing (31.8%, p=0.028).ConclusionsThe majority of HCWs felt prepared to face the pandemic, and increased significantly their knowledge on infection prevention and control measures less known in the routine clinical practice (such as use of protective glasses and alcoholic hand solution), compared to other well-known measures (such as hand washing). HCWs are at high risk of infection and need extensive knowledge and awareness of the disease to take adequate precautionary measures and they are crucial to disseminate good practices.