Dengue presents a growing public health concern in the Dar es Salaam region of Tanzania, marked by the recurring incidence of outbreaks. Unfortunately, there is little information available on the region's preparedness in terms of health care workers' knowledge on dengue as well as the availability of reagents and equipment essential for diagnosing and monitoring of dengue infections. To elucidate this, 78 health facilities were visited in Temeke district and structured questionnaires were distributed to 324 health care workers. The aim was to evaluate health care workers' knowledge on dengue and to assess the availability of reagents and equipment essential for diagnosing and monitoring of dengue infections. Content validity of the questionnaire was achieved through extensive literature review and it exhibited high reliability (Cronbach Alpha coefficient = 0.813). Cumulative scores for responses on knowledge questions by health care workers were computed. Characteristics such as level of education, place of work and gender were tested for association with these scores using chi-square tests and logistics regression. Almost all health care workers (99.7%) were aware of dengue disease. However, less than half (46.9%) had knowledge scores of or over 40%. Clinicians had approximately four times higher knowledge scores than other cadres (AOR, 3.637; p-value <= 0.0001), and those who worked in private facilities had twice the knowledge score than those working in government institutions (AOR, 2.071; p-value = 0.007). Only 8.6%, 35.6% and 14.7% of respondents reported the availability of dengue rapid tests, medical guidelines and refresher training respectively, showing a lack of health facilities readiness for the detection of dengue infections. Based on findings from this study, we recommend government authorities to build capacity of health care workers, to improve their understanding of dengue. We also urge the government and stakeholders to work together to ensure availability of diagnostic tests and other tools needed for diagnosis and surveillance of dengue. Dengue is a growing public health concern in Tanzania, particularly in the Dar es Salaam region, where outbreaks are common. However, little is known about the region's preparedness for dengue outbreaks, including the knowledge of health care workers and availability of diagnostic equipment. In a study of 324 health care workers in 78 health facilities in Temeke district, it was found that while almost all workers were aware of dengue, less than half had adequate knowledge of diagnosis, management, and monitoring. Clinicians had higher knowledge scores than other cadres, and those working in private facilities had more knowledge than those in government institutions. Only a small percentage of respondents reported the availability of diagnostic tests, medical guidelines, and refresher training. The study recommends building the capacity of health care workers and ensuring the availability of diagnostic tests and tools to improve readiness for dengue outbreaks. Dengue outbreaks occur every two years in Tanzania, with the last outbreak in 2019 resulting in over 6,800 cases and 13 deaths. As climate change is expected to increase the number of dengue cases, improving health care workers' knowledge and diagnostic capacity is crucial to effectively detect outbreaks.