As the 2022 human monkeypox (HMPX) multi-country outbreak is spreading, the response of healthcare workers (HCWs) is central to mitigation efforts. The current study aimed to evaluate HMPX knowledge and confidence in diagnosis and management among HCWs in Kuwait. We used a self-administered questionnaire distributed in July-August 2022 through a snowball sampling approach. The survey items evaluated HMPX knowledge, confidence in diagnosis and management of the disease, and the belief in conspiracies regarding emerging virus infections (EVIs). The sample size was 896 HCWs: nurses ( = 485, 54.1%), pharmacists ( = 154, 17.2%), physicians ( = 108, 12.1%), medical technicians/allied health professionals (MT/AHP, = 96, 10.7%), and dentists ( = 53, 5.9%). An overall low level of HMPX knowledge was noticed for items assessing virus transmission and non-cutaneous symptoms of the disease, with higher knowledge among physicians. Approximately one-fifth of the study sample agreed with the false notion that HMPX is exclusive to male homosexuals ( = 183, 20.4%), which was associated with lower knowledge with higher frequency among MT/AHP compared to nurses, physicians, and pharmacists. Confidence levels were low: confidence in diagnosis based on diagnostic tests ( = 449, 50.1%), confidence in the ability to manage the HMPX ( = 426, 47.5%), and confidence in the ability to diagnose HMPX clinically ( = 289, 32.3%). Higher confidence levels were found among nurses and participants with postgraduate degrees. Higher embrace of conspiracy beliefs regarding EVIs was noticed among participants with lower knowledge, and among those who agreed or were neutral/had no opinion regarding the false idea of HMPX exclusive occurrence among male homosexuals, while lower levels of belief in conspiracies were noticed among physicians, dentists, and pharmacists compared to MT/AHP. Variable levels of HMPX knowledge were observed in this study per item, with low level of knowledge regarding virus transmission. Differences in knowledge and confidence levels in diagnosis and management of HMPX should be considered in education and training aiming to prepare for outbreak response. The relatively high prevalence of embracing conspiratorial beliefs regarding EVIs is worrisome and needs proper interventions. The attitude towards male homosexuals' role in monkeypox spread should be evaluated in future studies considering the possibility of stigma and discrimination in this most-at-risk group.