Macrolide-resistant Mycoplasma pneumoniae in adolescents with community-acquired pneumonia

Naoyuki Miyashita 0 Yasuhiro Kawai 1 Hiroto Akaike 1 Kazunobu Ouchi 1 Toshikiyo Hayashi 0 Takeyuki Kurihara 0 Niro Okimoto 0 the Atypical Pathogen Study Group 0 Department of Internal Medicine I, Kawasaki Medical School , 2-1-80 Nakasange, Kita-ku, Okayama 700-8505 , Japan 1 Department of Pediatrics, Kawasaki Medical School , 577 Matsushima, Kurashiki 701-0192 , Japan Background: Although the prevalence of macrolide-resistant Mycoplasma pneumoniae isolates in Japanese pediatric patients has increased rapidly, there have been no reports concerning macrolide-resistant M. pneumoniae infection in adolescents aged 16 to 19 years old. The purpose of this study was to clarify the prevalence and clinical characteristics of macrolide-resistant M. pneumoniae in adolescent patients with community-acquired pneumonia. Methods: A total of 99 cases with M. pneumoniae pneumonia confirmed by polymerase chain reaction (PCR) and culture were analyzed. Forty-five cases were pediatric patients less than 16 years old, 26 cases were 16 to 19-year-old adolescent patients and 28 cases were adult patients. Primers for domain V of 23S rRNA were used and DNA sequences of the PCR products were compared with the sequence of an M. pneumoniae reference strain. Results: Thirty of 45 pediatric patients (66%), 12 of 26 adolescent patients (46%) and seven of 28 adult patients (25%) with M. pneumoniae pneumonia were found to be infected with macrolide-resistant M. pneumoniae (MR patients). Although the prevalence of resistant strains was similar in pediatric patients between 2008 and 2011, an increase in the prevalence of resistant strains was observed in adolescent patients. Among 30 pediatric MR patients, 26 had an A-to-G transition at position 2063 (A2063G) and four had an A-to-G transition at position 2064 (A2064G). In 12 adolescent MR patients, 10 showed an A2063G transition and two showed an A2064G transition, and in seven adult MR patients, six showed an A2063G transition and one showed an A2064G transition. Conclusions: The prevalence of macrolide-resistant M. pneumoniae is high among adolescent patients as well as pediatric patients less than 16-years old. To prevent outbreaks of M. pneumoniae infection, especially macrolide-resistant M. pneumoniae, in closed populations including among families, in schools and in university students, physicians should pay close attention to macrolide-resistant M. pneumoniae. - Background Mycoplasma pneumoniae is a common causative pathogen of respiratory infections in children and adults, accounting for as many as 10-30% of all cases of community-acquired pneumonia (CAP) [1]. M. pneumoniae pneumonia is specified for weekly reporting by specially designated sentinel clinics in accordance with the Japanese Infectious Diseases Control Law. Between 2001 and 2010, the average number of cases of M. pneumoniae pneumonia per sentinel clinic per year was 16.2 [2]. In 2011, especially the second half of the year, an epidemic of M. pneumoniae infection occurred throughout Japan and the incidence was the highest observed during the past decade, with the number of cases per sentinel clinic reaching 36.3 per year [2]. Many outbreaks of M. pneumoniae have occurred in closed populations including among families, in high schools, and in university students. During 2010 and 2011, a similar surge in M. pneumoniae infections was described in other countries especially in northern Europe [3-11]. Although M. pneumoniae pneumonia is usually mild in severity, macrolide-resistant M. pneumoniae with mutations in the 23S rRNA gene has emerged in pediatrics patients with CAP [12,13]. Recently, the prevalence of macrolideresistant M. pneumoniae isolates in Japanese pediatric patients has increased sharply [14]. More than 60% of M. pneumoniae strains in pediatric patients showed high resistance to 14- and 15-membered ring macrolides with MICs greater than or equal to 32 mg/L [14]. In contrast to pediatric patients, the prevalence of macrolide-resistant M. pneumoniae infection in adult patients is low [15]. Previous reports were limited to pediatric patients less than 16 years old or adults (20 years old), and there are no reports investigating high school-aged patients (16 to 18-years old) [12,13,15-18]. Several studies to determine the influence of age in CAP patients indicated that M. pneumoniae pneumonia is significantly more common in younger patients and especially in the 1020-year-old age group [1]. The purpose of this study was to clarify the prevalence and clinical characteristics of macrolide-resistant M. pneumoniae in adolescents, especially high school-aged patients, with CAP. Methods Patients This study was conducted as a part of CAP studies that investigated the prevalence and clinical features of atypical pneumonia and evaluated the usefulness of diagnostic methods for the diagnosis of this condition. All patients with CAP who visited 12 institutions participating in the Atypical Pathogen Study Group from January 2008 to December 2011 were enrolled in this study. The diagnosis was based on clinical signs and symptoms of lower respiratory tract infection (cough, fever, productive sputum, dyspnea, chest pain, or abnormal breath sounds) and the presence of new infiltrates on chest radiographs that were at least segmental and were not caused by preexisting or other known causes. Informed consent was obtained from all patients; the study protocol was approved by the Ethics Committee at Kawasaki Medical School. Microbiological laboratory tests Microbiological tests, such as Gram stain, cultures, realtime polymerase chain reaction (PCR), urinary antigen tests and serological tests, were performed as described previously [19]. Nasopharyngeal swab specimens were obtained from all patients and, if pleural fluid and sputum were available, a Gram stain test and a quantitative culture were obtained. Blood cultures were obtained from all adolescent and adult patients. Sputum data were only evaluated when the Gram stain test revealed numerous leukocytes (>25 in a 100 microscopic field) and few squamous epithelial cells (<10 in a 100 microscopic field). Invasive methods, such as bronchoscopic examination, were employed to obtain specimens in some patients after full explanation of the procedures. A bronchoscopic examination was undertaken for clinical indications. These specimens were also used for culturing and PCR. Cultures for M. pneumoniae and Legionella species were performed on pleuropneumonialike organism broth (Difco, Detroit, MI, USA) and buffered charcoal-yeast extract alpha agar, respectively. Cultures for Chlamydophila pneumoniae and C. psittaci were performed using cycloheximide-treated HEp-2 cells grown in a 24-well cell culture plate. All specimens were examined twice. Culture confirmation was done by fluorescent-antibody staining with C. pneumoniae and C. psittaci species-specific and genusspecific monoclonal antibodies. The target DNA sequences for PCR were a region o (...truncated)