Acute respiratory infections are a common and serious public health problem worldwide, especially in resource-limited countries. Although highly multiplexed molecular diagnostic tests allow clinical microbiology laboratories to rapidly detect a wide range of respiratory microorganisms, these commercial tests are expensive. Our aim was to develop affordable in-house multiplex PCR assays and apply them to nasopharyngeal samples from Senegalese patients to assess local epidemiology. Based on available simplex PCR systems, we developed in-house duplexed and triplexed PCR assays targeting 23 respiratory pathogens. Their specificity and sensitivity were evaluated using 965 DNA/RNA extracts from nasopharyngeal swabs, by comparing the results obtained with the same systems in simplex PCR and with a commercial kit (FTD ® Respiratory Pathogens 21) used for routine diagnosis in our French laboratory. In-house multiplexed assays were then applied to swabs from 500 febrile patients from the rural area of Niakhar. In-house multiplex tests had a specificity and sensitivity of 100%, with an estimated cost of which was three to 6.5 times lower than commercial kits and an analysis time of two hours. Streptococcus pneumoniae was the most frequently observed bacterium (n=181; 36.2%). For respiratory viruses, adenovirus was the most prevalent (n=26, 5.2%), followed by RSV (n=15, 3%), SARS-CoV-2 (n=11, 2.2%), rhinovirus (n=10, 2%) and influenza A virus (n=9, 1.8%). Co-infection was detected in 28.2% (n=141) of nasopharyngeal samples. In-house duplex and triplex PCR assays are efficient and affordable. Their use can be modular, targeting the most frequent pathogens according to local epidemiology in order to further limit costs.