In recent years, the morbidity of Mycoplasma pneumoniae pneumonia (MPP) has dramatically increased in China. An increasing number of studies indicate that an imbalance in the respiratory microbiota is associated with respiratory infection. We selected 28 hospitalized patients infected with M. pneumoniae and 32 healthy children. Nasopharyngeal (NP) and oropharyngeal (OP) swabs were collected from healthy children, whereas NP, OP and bronchoalveolar lavage (BAL) specimens were collected from patients. Microbiota analysis was performed on all microbial samples using 16 S ribosomal RNA (16 S rRNA) sequencing. The NP microbial samples in healthy children were divided into two groups, which were dominated by either Staphylococcus or mixed microbial components. The respiratory microbiota in pneumonia patients harbored a lower microbial diversity compared to healthy children, and both the NP and OP microbiota of patients differed significantly from that of healthy children. Hospitalized MPP children with a higher abundance of Mycoplasma in the BAL fluid (BALF) microbiota tended to suffer longer hospitalization lengths and higher peak fevers and serum C-reactive protein levels. Concordance analysis explained the succession of imbalanced NP microbiota to the OP and lung in diseased children. However, the association of the abundance of Mycoplasma in BALF microbiota with that in NP or OP microbiota varied among individuals, which suggested the sensitivity of BALF in MPP diagnostics, mirroring MPP severity.