Mycoplasma (M.) pneumoniae has been associated with exacerbation of symptoms in asthmatic school children and adults; and an etiological role in asthma has been suggested. The purpose of this study was to investigate whether infection with M. pneumoniae in early childhood has a long-term influence on lung function and bronchial responsiveness. In a retrospective, clinical cohort-study children younger than 5 years-of-age when PCR-tested for M. pneumoniae were enrolled. Sixty-five children with clinical symptoms suggesting infection with M. pneumoniae during an epidemic season completed a clinical follow-up examination including lung function testing (28 PCR-positive and 37 PCR-negative). In addition to the PCR-test for M. pneumoniae all respiratory tract specimens were additionally tested for other atypical bacteria and for viruses by PCR. Lung function was measured as specific airway resistance by whole-body plethysmography and bronchial hyperresponsiveness was assessed by cold, dry air hyperventilation. Neither baseline lung function nor bronchial response to cold dry air hyperventilation differed between M.pneumoniae-positive and -negative children: mean baseline lung function were 1.17 versus 1.21 (kPa sec), P=0.45; and mean change in specific resistance was 13% versus 9%, P=0.42. In conclusion, M. pneumoniae infection in early childhood was not associated with long-term effects on lung function and bronchial hyperresponsiveness 2 years after infection. Pediatr Pulmonol. 2008; 43:567-575. © 2008 Wiley-Liss, Inc.