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Background  Dengue fever is a rapidly emerging infection throughout the tropics and subtropics with extensive public health burden. Adequate training of healthcare providers is crucial to reducing infection incidence through patient education and collaboration with public health authorities. We examined how public sector healthcare providers in a dengue-endemic region of Ecuador view and manage dengue infections, with a focus on the 2009 World Health Organization (WHO) Dengue Guidelines.    Methods  A 37-item questionnaire of dengue knowledge, attitudes, and practices was developed and administered to dengue healthcare providers in Machala, Ecuador. Survey focus areas included: “Demographics,” “Infection and Prevention of Dengue,” “Dengue Diagnosis and the WHO Dengue Guide,” “Laboratory Testing,” “Treatment of Dengue,” and “Opinions Regarding Dengue.”    Results  A total of 76 healthcare providers participated in this study, of which 82 % were medical doctors and 14 % were nurses. Fifty-eight percent of healthcare professionals practiced in ambulatory clinics and 34 % worked in a hospital. Eighty-nine percent of respondents were familiar with the 2009 WHO Dengue Guidelines, and, within that group, 97 % reported that the WHO Dengue Guide was helpful in dengue diagnosis and clinical management. Knowledge gaps identified included  Aedes aegypti  mosquito feeding habits and dengue epidemiology. Individuals with greater dengue-related knowledge were more likely to consider dengue a major health problem. Only 22 % of respondents correctly reported that patients with comorbidities and dengue without warning signs require hospital admission, and 25 % of providers reported never admitting patients with dengue to the hospital. Twenty percent of providers reported rarely (≤25 % of cases) obtaining laboratory confirmation of dengue infection. Providers reported patient presumptive self-medication as an ongoing problem. Thirty-one percent of healthcare providers reported inadequate access to resources needed to diagnose and treat dengue.    Conclusion  Participants demonstrated a high level of knowledge of dengue symptoms and treatment, but additional training regarding prevention, diagnosis, and admission criteria is needed. Interventions should not only focus on increasing knowledge, but also encourage review of the WHO Dengue Guidelines, avoidance of presumptive self-medication, and recognition of dengue as a major health problem. This study provided an assessment tool that effectively captured healthcare providers’ knowledge and identified critical gaps in practice.    Electronic supplementary material  The online version of this article (doi:10.1186/s40794-016-0024-y) contains supplementary material, which is available to authorized users.

Knowledge, attitudes, and practices regarding dengue infection among public sector healthcare providers in Machala, Ecuador