Molecular testing for acute respiratory infections (ARIs) has documented value but limited implementation due to questions that typically slow the acceptance of new tests. This study sought to address these questions and achieve implementation. Rhinovirus was added to a nested multiplex PCR (M-PCR), increasing its diagnostic yield. Over one winter, three hospital pediatric departments used the M-PCR to complement their direct fluorescent-antibody assay (DFA) for respiratory syncytial virus (RSV). Clinicians recorded "pretest probability estimates" (using continuous scales for various pathogen groups) for comparison with test results; treatments and test turnaround times were also recorded. Transnasal and throat swabs, with or without nasopharyngeal aspirate (NPA), were M-PCR tested. NPA-containing sample sets found to be RSV positive by DFA were not further tested. Single PCR for human metapneumovirus (hMPV) was performed retrospectively. Of 178 ARI episodes representing 172 patients, NPA was included in 97 sample sets; 54 (56%) were determined to be RSV positive. The other NPA-containing sample sets (n = 43) yielded 27 findings (63%), and the swab-only sets (n = 81) yielded 47 findings (58%); rhinovirus was found most often. Testing for hMPV yielded seven positive results. M-PCR median turnaround times were 4 days in swab-only samples and 5 days with NPA. Antibiotics were prescribed in 50 episodes, at rates similar for RSV and rhinovirus. Pretest probability estimates of a viral cause were lower in episodes caused by rhinovirus than in episodes caused by RSV. The hospitals continued to use M-PCR for NPA-containing samples found to be RSV negative by DFA. Test implementation is more likely with higher diagnostic yield and a protocol that reflects day-to-day clinical and laboratory operations
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