Objectives: To investigate 15 respiratory viruses in children with acute respiratory tract infections (ARTIs) using multiplex reverse-transcriptase polymerase chain reaction (RT-PCR), and to analyze the clinical and epidemiological features of these viruses. Methods: In a cross-sectional study, 135 children, ≤5 years of age who presented with ARTIs in Najran Maternity and Children Hospital, Najran, Saudi Arabia between October 2012 and July 2013 were included. The clinical and sociodemographic data, and the laboratory results were recorded using a standardized questionnaire. Two nasopharyngeal swabs were collected from each child: one for bacteriological examination, and the second for viral detection using multiplex RT-PCR. Results: A single viral pathogen was detected in 76 patients, viral coinfections in 9, and mixed viral and bacterial pathogens in 15. Respiratory syncytial virus was isolated in 33 patients, human rhinovirus (hRV) in 22, adenovirus (AdV) in 19, human metapneumovirus in 13, influenza virus in 10, parainfluenza virus in 7, human corona virus (hCoV) in 4, and human bocavirus in one. Conclusion: Respiratory syncytial virus, hRV, and AdV were the most frequent viruses, accounting for more than two-thirds of the cases. Other viruses, such as MPV, hCoV NL63, and hCoV OC43, may play a role in pediatric ARTIs. Of significance is the potential use of multiplex RT-PCR to provide epidemiological and virological data for early detection of the emergence of novel respiratory viruses in the era of the Middle East respiratory syndrome coronavirus.