BACKGROUND: Dengue fever, an endemic arboviral disease, represents one of the major public health concerns in Nepal. It is transmitted by bites of infected Aedes aegypti and Aedes albopictus, the former being primary vector. The bacterial community plays a significant role in biology of mosquitoes; however, the bacterial communities of primary vector A. aegypti remain unstudied in Nepal. The study was designed to determine the rate of dengue seropositivity and to explore the bacterial diversity of A. aegypti from dengue endemic districts, Kanchanpur and Parsa of Nepal. METHODS: A cross-sectional study was conducted between June 2013 and November 2013 at two hospitals of Kanchanpur and Parsa. A total of 221 serum samples were collected from patients (inpatients and outpatients) suspected of suffering from dengue fever and attending Mahakali Zonal Hospital, Mahendranagar, Kanchanpur, and Narayani Zonal Hospital, Birgunj, Parsa. Detection of anti-dengue IgM was performed by using human dengue IgM capture ELISA. The larvae and pupae of mosquitoes (A. aegypti) were collected, reared, and emerged. Then, the bacteria were isolated and identified from the gut of identified mosquitoes by using standard methods. RESULTS: Out of total 221 serum samples collected from patients suspected of suffering from dengue fever, 34 (15.38%) were positive for anti-dengue IgM. Gram-negative bacteria were isolated in largest proportion (63%) followed by gram-positive cocci (23.27%) and gram-positive rods (13.73%). The most common cultivable bacteria isolated were Staphylococcus spp., Pseudomonas spp., and Acinetobacter spp. The average bacterial load in the vectors was 3.91 × 104 cfu/ml. CONCLUSIONS: High rate of anti-dengue IgM seropositivity was reported in our study. The environmental bacteria were predominantly isolated and identified in A. aegypti. The paratransgenic approach to control vector might be possible by spreading the genetically modified bacteria in larval habitat or shelter of adult mosquitoes.