Application of LAMP coupled with NALF for precise detection of mycoplasma pneumoniae.

Mycoplasma pneumoniae (MP),as the most commonly infected respiratory pathogen in community-acquired pneumonia in preschool children,has becoming a prominent factor affecting children's respiratory health.Currently, there is a lack of easy, rapid, and accurate laboratory testing program for MP infection, which causes comparatively difficulty for clinical diagnostic.Here,we utilize loop-mediated isothermal amplification (LAMP) to amplify and characterize the P1 gene of MP, combined with nucleic acid lateral flow (NALF) for fast and visuallized detection of MP.Furthermore, we evaluated and analyzed the sensitivity, specificity and methodological consistency of the method.The results showed that the limit of detection(LoD) of MP-LAMP-NALF assay was down to 100 copys per reaction and there was no cross-reactivity with other pathogens infected the respiratory system. The concordance rate between MP-LAMP-NALF assay with quantitative real-time PCR was 94.3 %,which exhibiting excellent testing performance.We make superior the turnaround time of the MP-LAMP-NALF assay, which takes only about 50 min. In addition, there is no need for precision instruments and no restriction on the laboratory site.Collectively, LAMP-NALF assay targeting the P1 gene for Mycoplasma pneumoniae detection was a easy, precise and visual test which could be widely applied in outpatient and emergency departments or primary hospitals.When further optimized, it could be used as "point-of-care testing" of pathogens or multiple testing for pathogens.

A LAMP-based hydrogen ion selective electrochemical sensor for highly sensitive detection of Mycoplasma pneumoniae.

A concise and rapid detection method for Mycoplasma pneumoniae is urgently required due to its severe impact on human health. To meet such a need, this study proposed and constructed an innovative point-of-care testing (POCT) platform that consists of a hydrogen ion-selective loop-mediated isothermal amplification (H+-LAMP) sensor and an electrochemical detection device. The H+-LAMP sensor successfully integrated the working and reference electrodes and converted the H+ generated during the LAMP process into an electrochemical signal. High sensitivity and stability for pathogen detection were also achieved by treating the working electrode with an electrodeposited polyaniline solid contact layer and by using an ion-selective membrane. As a result, the sensor shows a sensitivity of 68.26 mV per pH, a response time of less than 2 s, and a potential drift of less than 5 mV within one hour, which well meets the urgent need. The results also demonstrated that the detection limit for Mycoplasma pneumoniae was lowered to 1 copy per µL, the nucleic acid extraction and detection process could be completed in 30 minutes, and the impact of interfering ions on the sensor was negligible. Validation with 20 clinical samples yielded satisfactory results. More importantly, the storage lifespan of such an electrochemical sensor is over seven days, which is a great advantage for on-site pathogen detection. Therefore, the hydrogen ion-selective sensor constructed in this investigation is particularly suitable as a core component for instant pathogen detection platforms.

[Metagenomic next-generation sequencing-based retrospective investigation of the drug resistance sites of Mycoplasma pneumoniae in children].

Objective: To analyze the drug-resistant gene loci of Mycoplasma pneumoniae (MP) using metagenomic next-generation sequencing (mNGS). Methods: From November 2022 to October 2023, 697 clinical samples (including sputum, alveolar lavage fluid and blood) of 686 children with Mycoplasma pneumoniae positive detected by mNGS were retrospectively analyzed. Samples were divided into intensive care unit (ICU) group and non-ICU group, Chi-square test was used to compare groups, and Mann-Kendall trend test was used to analyze the change trend of the detection rate of drug resistance gene loci over time. Results: Of the 697 samples, 164 were from the ICU group and 533 were from the non-ICU group. The detection rate of Mycoplasma pneumoniae resistance gene was 44.3% (309/697), and all detected drug-resistant gene loci of MP were A2063G. The detection rate of Mycoplasma pneumoniae in ICU group was 50.0% (82/164), and the detection rates of Mycoplasma pneumoniae resistance gene loci in sputum, alveolus lavage fluid and blood samples were 75.0% (18/24) and 48.4% (62/128), respectively. The detection rate in sputum was higher than alveolus lavage fluid samples (χ2=5.72,P=0.017). The detection rate of Mycoplasma pneumoniae in non-ICU group was 42.6% (227/533), the detection rate of Mycoplasma pneumoniae resistance gene loci in sputum and alveolar lavage fluid was 40.0% (16/40), 44.3% (201/454), and no detection rate in blood samples (0/12). There was no significant difference in the detection rate of alveolar lavage fluid and sputum (χ2=0.27, P=0.602). From November 2022 to October 2023, the detection rate of submitted samples showed an increasing trend month by month (overall: Z=3.99, ICU inspection group: Z=2.93, non-ICU group: Z=3.01, all P<0.01). Among the bacteria commonly detected with Mycoplasma pneumoniae, Streptococcus pneumoniae accounted for the highest proportion, the detection rate was 15.5% (108/697), and Epstein-Barr virus accounted for the highest proportion of 17.6% (123/697). Conclusions: From November 2022 to October 2023, the detection rate of Mycoplasma pneumoniae drug resistance gene loci showed an increasing trend. The detection rate of drug resistance gene loci in sputum samples of ICU group was higher than alveolus lavage fluid. No new drug resistance site were detected.

Novel Variant and Known Mutation in 23S rRNA Gene of Mycoplasma pneumoniae, Northern Vietnam, 2023.

During a 2023 outbreak of Mycoplasma pneumoniae-associated community-acquired pneumonia among children in northern Vietnam, we analyzed M. pneumoniae isolated from nasopharyngeal samples. In almost half (6 of 13) of samples tested, we found known A2063G mutations (macrolide resistance) and a novel C2353T variant on the 23S rRNA gene.

Macrolide resistance of Mycoplasma pneumoniae in several regions of China from 2013 to 2019.

This paper retrospectively analysed the prevalence of macrolide-resistant Mycoplasma pneumoniae (MRMP) in some parts of China. Between January 2013 and December 2019, we collected 4,145 respiratory samples, including pharyngeal swabs and alveolar lavage fluid. The highest PCR-positive rate of M. pneumoniae was 74.5% in Beijing, the highest resistance rate was 100% in Shanghai, and Gansu was the lowest with 20%. The highest PCR-positive rate of M. pneumoniae was 74.5% in 2013, and the highest MRMP was 97.4% in 2019; the PCR-positive rate of M. pneumoniae for adults in Beijing was 17.9% and the MRMP was 10.48%. Among the children diagnosed with community-acquired pneumonia (CAP), the PCR-positive and macrolide-resistant rates of M. pneumoniae were both higher in the severe ones. A2063G in domain V of 23S rRNA was the major macrolide-resistant mutation, accounting for more than 90%. The MIC values of all MRMP to erythromycin and azithromycin were ≥ 64 µg/ml, and the MICs of tetracycline and levofloxacin were ≤ 0.5 µg/ml and ≤ 1 µg/ml, respectively. The macrolide resistance varied in different regions and years. Among inpatients, the macrolide-resistant rate was higher in severe pneumonia. A2063G was the common mutation, and we found no resistance to tetracycline and levofloxacin.

[A multicenter study on respiratory pathogen detection with Mycoplasma pneumoniae pneumonia in children].

Objective: To analyze the status of respiratory pathogen detection and the clinical features in children with Mycoplasma pneumoniae pneumonia (MPP). Methods: A prospective, multicenter study was conducted to collect clinical data, including medical history, laboratory examinations and multiplex PCR tests of children diagnosed with MPP from 4 hospitals in China between November 15th and December 20th, 2023. The multiplex PCR results and clinical characteristics of MPP children in different regions were analyzed. The children were divided into severe and mild groups according to the severity of the disease. Patients in the severe group were further divided into Mycoplasma pneumoniae (MP) alone and Multi-pathogen co-detection groups based on whether other pathogens were detected besides MP, to analyze the influence of respiratory pathogen co-detection rate on the severity of the disease. Mann-Whitney rank sum test and Chi-square test were used to compare data between independent groups. Results: A total of 298 children, 136 males and 162 females, were enrolled in this study, including 204 children in the severe group with an onset age of 7.0 (6.0, 8.0) years, and 94 children in the mild group with an onset age of 6.5 (4.0, 7.8) years. The level of C-reactive protein, D-dimer, lactic dehydrogenase (LDH) were significantly higher (10.0 (5.0, 18.0) vs. 5.0 (5.0, 7.5) mg/L, 0.6 (0.4, 1.1) vs. 0.5 (0.3, 0.6) mg/L, 337 (286, 431) vs. 314 (271, 393) U/L, Z=2.02, 2.50, 3.05, all P<0.05), and the length of hospitalization was significantly longer in the severe group compared with those in mild group (6.0 (6.0, 7.0) vs. 5.0 (4.0, 6.0) d, Z=4.37, P<0.05). The time from onset to admission in severe MPP children was significantly shorter than that in mild MPP children (6.0 (5.0, 9.5) vs. 9.0 (7.0, 13.0) d, Z=2.23, P=0.026). All patients completed the multiplex PCR test, with 142 cases (47.7%) MPP children detected with 21 pathogens including adenovirus 25 cases (8.4%), human coronavirus 23 cases (7.7%), rhinovirus 21 cases (7.0%), Streptococcus pneumoniae 21 cases (7.0%), influenza A virus 18 cases (6.0%). The pathogens with the highest detection rates in Tianjin, Shanghai, Wenzhou and Chengdu were Staphylococcus aureus at 10.7% (8/75), adenovirus at 13.0% (10/77), adenovirus at 15.3% (9/59), and both rhinovirus and Haemophilus influenzae at 11.5% (10/87) each. The multi-pathogen co-detection rate in severe MPP children was significantly higher than that in mild MPP group (52.9% (108/204) vs. 36.2% (34/94), χ²=10.62,P=0.005). Among severe MPP children, there are 89 cases in the multi-pathogen co-detection group and 73 cases in the simple MPP group. The levels of LDH, D-dimer and neutrophil counts in the multi-pathogen co-detection group were significantly higher than those in the simple MPP group (348 (284, 422) vs. 307 (270, 358) U/L, 0.8 (0.5, 1.5) vs. 0.6 (0.4, 1.0) mg/L, 4.99 (3.66, 6.89)×109 vs. 4.06 (2.91, 5.65)×109/L, Z=5.17, 4.99, 6.11, all P<0.05). Conclusions: The co-detection rate of respiratory pathogens, LDH and D-dimer in children with severe MPP were higher than those with mild MPP. Among severe MPP children the stress response of children in co-detection group was more serious than that of children with simple MPP.

[A multicenter prospective study on early identification of refractory Mycoplasma pneumoniae pneumonia in children].

Objective: To explore potential predictors of refractory Mycoplasma pneumoniae pneumonia (RMPP) in early stage. Methods: The prospective multicenter study was conducted in Zhejiang, China from May 1st, 2019 to January 31st, 2020. A total of 1 428 patients with fever >48 hours to <120 hours were studied. Their clinical data and oral pharyngeal swab samples were collected; Mycoplasma pneumoniae DNA in pharyngeal swab specimens was detected. Patients with positive Mycoplasma pneumoniae DNA results underwent a series of tests, including chest X-ray, complete blood count, C-reactive protein, lactate dehydrogenase (LDH), and procalcitonin. According to the occurrence of RMPP, the patients were divided into two groups, RMPP group and general Mycoplasma pneumoniae pneumonia (GMPP) group. Measurement data between the 2 groups were compared using Mann-Whitney U test. Logistic regression analyses were used to examine the associations between clinical data and RMPP. Receiver operating characteristic (ROC) curves were used to analyse the power of the markers for predicting RMPP. Results: A total of 1 428 patients finished the study, with 801 boys and 627 girls, aged 4.3 (2.7, 6.3) years. Mycoplasma pneumoniae DNA was positive in 534 cases (37.4%), of whom 446 cases (83.5%) were diagnosed with Mycoplasma pneumoniae pneumonia, including 251 boys and 195 girls, aged 5.2 (3.3, 6.9) years. Macrolides-resistant variation was positive in 410 cases (91.9%). Fifty-five cases were with RMPP, 391 cases with GMPP. The peak body temperature before the first visit and LDH levels in RMPP patients were higher than that in GMPP patients (39.6 (39.1, 40.0) vs. 39.2 (38.9, 39.7) ℃, 333 (279, 392) vs. 311 (259, 359) U/L, both P<0.05). Logistic regression showed the prediction probability π=exp (-29.7+0.667×Peak body temperature (℃)+0.004×LDH (U/L))/(1+exp (-29.7+0.667×Peak body temperature (℃)+0.004 × LDH (U/L))), the cut-off value to predict RMPP was 0.12, with a consensus of probability forecast of 0.89, sensitivity of 0.89, and specificity of 0.67; and the area under ROC curve was 0.682 (95%CI 0.593-0.771, P<0.01). Conclusion: In MPP patients with fever over 48 to <120 hours, a prediction probability π of RMPP can be calculated based on the peak body temperature and LDH level before the first visit, which can facilitate early identification of RMPP.

Mycoplasma pneumoniae at the rise not only in China: rapid increase of Mycoplasma pneumoniae cases also in Spain.

After the use of facemasks, other isolation measures enacted during the SARS-CoV-2 pandemic were lifted, respiratory pathogens, such as RSV, reappeared, but until the November 2023 WHO alert for China, M. pneumoniae had virtually disappeared. After observing a similar reappearance in our hospital, a retrospective analysis of the number of positive M. pneumoniae tests. Between 2018 and December 2023, 1619 PCR tests were ordered and 43 (2.6%) of them were positive. Two outbreaks, one in 2018 and one in 2023, accounted for the majority of cases. Tests were usually ordered in an outpatient setting (53.54%, n = 23) and most of them were paediatric patients with a mean age (sd) of 10.2 (6.2) years. As for the severity of the cases, in the 2018 outbreak, of 15 children who tested positive, 53.3% (n = 8) were admitted to the ward and 6.7% (n = 1) at the intensive care unit. Whereas in 2023, 2 patients were tested in the ward (10.5%) and one in the intensive care unit (5.2%) from a total of 19 patients. The positive rate in 2023 was significantly higher in comparison with years 2020, 2021 and 2022 and significantly lower in comparison with 2018 (P-value=0.003). The outbreak in late 2023 can be explained by the seasonality of Mycoplasma pneumonia alone, which has shown outbreaks every 3-5 years, and it does not appear to be more severe than the previous one.

Observational retrospective clinical study on clinical features of macrolide-resistant Mycoplasma pneumoniae pneumonia in Chinese pediatric cases.

This study aimed to investigate differences in clinical characteristics and laboratory findings between children infected with Macrolide-Sensitive Mycoplasma pneumoniae (MSMP) and Macrolide-Resistant Mycoplasma pneumoniae (MRMP). Additionally, the research sought to identify laboratory markers for rapidly distinguishing refractory Mycoplasma pneumoniae pneumonia (RMPP) from ordinary Mycoplasma pneumoniae pneumonia (OMPP). In total, 265 Mycoplasma pneumoniae (MP) patients were included, with MRMP identified by specific point mutations in domain V of the 23S rRNA gene. A retrospective analysis compared the clinical courses and laboratory data, revealing that MRMP patients experienced prolonged febrile days (P = 0.004), elevated CRP levels (P < 0.001), and higher MP DNA loads than MSMP patients (P = 0.037). Based on clinical symptoms, MRMP was divided into RMPP (n = 56) and OMPP (n = 70), with RMPP demonstrating significantly increased IL-18, community-acquired respiratory distress syndrome (CARDS) toxins in nasopharyngeal aspirate, and serum CRP levels (P < 0.001; P = 0.006; P < 0.001). In conclusion, timely recognition of RMPP is crucial for enhancing prognosis. The identification of MRMP, coupled with proinflammatory cytokines such as IL-18, CARDS toxins, and CRP, emerges as promising markers with the potential to contribute significantly to diagnostic accuracy and prognosis assessment.

Serum IL-17A and IL-6 in paediatric Mycoplasma pneumoniae pneumonia: implications for different endotypes.

Paediatric Mycoplasma pneumoniae pneumonia (MPP) is a heterogeneous disease with a diverse spectrum of clinical phenotypes. No studies have demonstrated the relationship between underlying endotypes and clinical phenotypes as well as prognosis about this disease. Thus, we conducted a multicentre prospective longitudinal study on children hospitalized for MPP between June 2021 and March 2023, with the end of follow-up in August 2023. Blood samples were collected and processed at multiple time points. Multiplex cytokine assay was performed to characterize serum cytokine profiles and their dynamic changes after admission. Cluster analysis based on different clinical phenotypes was conducted. Among the included 196 patients, the levels of serum IL-17A and IL-6 showed remarkable variabilities. Four cytokine clusters based on the two cytokines and four clinical groups were identified. Significant elevation of IL-17A mainly correlated with diffuse bronchiolitis and lobar lesion by airway mucus hypersecretions, while that of IL-6 was largely associated with lobar lesion which later developed into lung necrosis. Besides, glucocorticoid therapy failed to inhibit IL-17A, and markedly elevated IL-17A and IL-6 levels may correlate with lower airway obliterans. Our study provides critical relationship between molecular signatures (endotypes) and clustered clinical phenotypes in paediatric patients with MPP.

Outbreak of Mycoplasma pneumoniae pneumonia in hospitalized patients: Who is concerned? Nord Franche-Comté Hospital, France, 2023-2024.

We report an outbreak of confirmed Mycoplasma pneumoniae community-acquired pneumonia (CAP) in Nord Franche-Comté Hospital, France, from 14 November 2023 to 31 January 2024. All 13 inpatients (11 adults with a mean age of 45.5 years and 2 children) were diagnosed with positive serology and/or positive reverse transcription polymerase chain reaction (RT-PCR) on respiratory specimens. All patients were immunocompetent and required oxygen support with a mean duration of oxygen support of 6.2 days. Two patients were transferred to the intensive care unit (ICU) but were not mechanically ventilated. Patients were treated with macrolides (n = 12, 92.3%) with recovery in all cases. No significant epidemiological link was reported in these patients.

Molecular epidemiology of Mycoplasma pneumoniae pneumonia in children, Wuhan, 2020-2022.

BACKGROUND: Mycoplasma pneumoniae (M. pneumoniae) is an important pathogen of community-acquired pneumonia in children. The factors contributing to the severity of illness caused by M. pneumoniae infection are still under investigation. We aimed to evaluate the sensitivity of common M. pneumoniae detection methods, as well as to analyze the clinical manifestations, genotypes, macrolide resistance, respiratory microenvironment, and their relationship with the severity of illness in children with M. pneumoniae pneumonia in Wuhan. RESULTS: Among 1,259 clinical samples, 461 samples were positive for M. pneumoniae via quantitative polymerase chain reaction (qPCR). Furthermore, we found that while serological testing is not highly sensitive in detecting M. pneumoniae infection, but it may serve as an indicator for predicting severe cases. We successfully identified the adhesin P1 (P1) genotypes of 127 samples based on metagenomic and Sanger sequencing, with P1-type 1 (113/127, 88.98%) being the dominant genotype. No significant difference in pathogenicity was observed among different genotypes. The macrolide resistance rate of M. pneumoniae isolates was 96% (48/50) and all mutations were A2063G in domain V of 23S rRNA gene. There was no significant difference between the upper respiratory microbiome of patients with mild and severe symptoms. CONCLUSIONS: During the period of this study, the main circulating M. pneumoniae was P1-type 1, with a resistance rate of 96%. Key findings include the efficacy of qPCR in detecting M. pneumoniae, the potential of IgM titers exceeding 1:160 as indicators for illness severity, and the lack of a direct correlation between disease severity and genotypic characteristics or respiratory microenvironment. This study is the first to characterize the epidemic and genomic features of M. pneumoniae in Wuhan after the COVID-19 outbreak in 2020, which provides a scientific data basis for monitoring and infection prevention and control of M. pneumoniae in the post-pandemic era.

Increase of respiratory illnesses among children in Beijing, China, during the autumn and winter of 2023.

In 2023, through an ongoing respiratory pathogen surveillance system, we observed from mid-September onwards, an increase of respiratory illness among children aged ≤ 15 years presenting at hospital outpatient clinics in Beijing, China. Data indicated that illness was caused by multiple pathogens, predominantly Mycoplasma pneumoniae. Seasonality, periodicity and high prevalence of resistance to macrolide (30 of 30 strains sequenced with the A2063G mutation) were important characteristics of the M. pneumoniae epidemic, which resulted in a rise in consultations at specialised paediatric hospitals.

Mycoplasma pneumoniae epidemic in Denmark, October to December, 2023.

We report a surge of patients, especially children and adolescents, with respiratory disease caused by Mycoplasma pneumoniae in Denmark since October 2023. While the surge has reached an epidemic level, no impact on hospital capacity has been observed; only 14% (446/3,195) of cases, primarily adults, required hospitalisation. Macrolide resistance was detected in less than 2% of samples tested. Timely monitoring of hospitalisations linked to M. pneumoniae infections has been established to inform the healthcare system, decisionmakers and the public.

Integrative study of pulmonary microbiome, transcriptome and clinical outcomes in Mycoplasma pneumoniae pneumonia.

BACKGROUND: This study aimed to investigate the interactions among three core elements of respiratory infection-pathogen, lung microbiome, and host response-and their avocation with the severity and outcomes of Mycoplasma pneumoniae pneumonia (MPP) in children. METHODS: We prospectively collected bronchoalveolar lavage fluid from a cohort of 41 children with MPP, including general MPP (GMPP) and complicated MPP (CMPP), followed by microbiome and transcriptomic analyses to characterize the association among pathogen, lung microbiome, and host response and correlate it with the clinical features and outcomes. RESULTS: The lung microbiome of patients with CMPP had an increased relative abundance of Mycoplasma pneumoniae (MP) and reduced alpha diversity, with 76 differentially expressed species. Host gene analysis revealed a key module associated with neutrophil function and several inflammatory response pathways. Patients with a high relative abundance of MP, manifested by a specific lung microbiome and host response type, were more prone to CMPP and had a long imaging recovery time. CONCLUSION: Patients with CMPP have a more disrupted lung microbiome than those with GMPP. MP, lung microbiome, and host response interacts with each other and are closely related to disease severity and outcomes in children with MPP.

Chlamydophila pneumoniae-associated community-acquired pneumonia in paediatric patients of a tertiary care hospital in Mexico: molecular diagnostic and clinical insights.

Chlamydophila pneumoniae is a cause of community-acquired pneumonia (CAP) and responsible for 1-2% of cases in paediatric patients. In Mexico, information on this microorganism is limited. The aim of this study was to detect C. pneumoniae using two genomic targets in a real-time PCR and IgM/IgG serology assays in paediatric patients with CAP at a tertiary care hospital in Mexico City and to describe their clinical characteristics, radiological features, and outcomes. A total of 154 hospitalized patients with diagnosis of CAP were included. Detection of C. pneumoniae was performed by real-time PCR of the pst and arg genes. Complete blood cell count, C-reactive protein measurement and IgM and IgG detection were performed. Clinical-epidemiological and radiological data from the patients were collected. C. pneumoniae was detected in 25 patients (16%), of whom 88% had underlying disease (P = 0.014). Forty-eight percent of the cases occurred in spring, 36% in girls, and 40% in children older than 6 years. All patients had cough, and 88% had fever. Interstitial pattern on chest-X-ray was the most frequent (68%), consolidation was observed in 32% (P = 0.002). IgM was positive in 7% and IgG in 28.6%. Thirty-six percent presented complications. Four percent died. A high proportion showed co-infection with Mycoplasma pneumoniae (64%). This is the first clinical report of C. pneumoniae as a cause of CAP in Mexican paediatric patients, using two genomic target strategy and serology. We found a frequency of 16.2% with predominance in children under 6 years of age. In addition; cough and fever were the most common symptoms. Early detection of this pathogen allows timely initiation of specific antimicrobial therapy to reduce development of complications. This study is one of the few to describe the presence of C. pneumoniae in patients with underlying diseases.

Mycoplasma pneumoniae carriage in children with recurrent respiratory tract infections is associated with a less diverse and altered microbiota.

BACKGROUND: Mycoplasma pneumoniae is a common cause of community-acquired pneumonia in school-aged children and can be preceded by asymptomatic carriage. However, its role in recurrent respiratory tract infections is unclear. We studied the prevalence of M.pneumoniae carriage in children with recurrent respiratory infections and identified associated factors. METHODS: We tested M.pneumoniae carriage by qPCR in children with recurrent infections and their healthy family members in a cross-sectional study. Serum and mucosal total and M.pneumoniae-specific antibody levels were measured by ELISA and nasopharyngeal microbiota composition was characterized by 16S-rRNA sequencing. FINDINGS: Prevalence of M.pneumoniae carriage was higher in children with recurrent infections (68%) than their family members without infections (47% in siblings and 27% in parents). M.pneumoniae carriage among family members appeared to be associated with transmission within the household, likely originating from the affected child. In logistic regression corrected for age and multiple comparisons, IgA (OR 0.16 [0.06-0.37]) and total IgG deficiency (OR 0.15 [0.02-0.74]) were less prevalent in M.pneumoniae carriers (n = 78) compared to non-carriers (n = 36). In multivariable analysis, the nasopharyngeal microbiota of M.pneumoniae carriers had lower alpha diversity (OR 0.27 [0.09-0.67]) and a higher abundance of Haemophilus influenzae (OR 45.01 [2.74-1608.11]) compared to non-carriers. INTERPRETATION: M.pneumoniae carriage is highly prevalent in children with recurrent infections and carriers have a less diverse microbiota with an overrepresentation of disease-associated microbiota members compared to non-carriers. Given the high prevalence of M.pneumoniae carriage and the strong association with H. influenzae, we recommend appropriate antibiotic coverage of M.pneumoniae and H. influenzae in case of suspected pneumonia in children with recurrent respiratory tract infections or their family members. FUNDING: Wilhelmina Children's Hospital Research Fund, 'Christine Bader Stichting Irene KinderZiekenhuis', Sophia Scientific Research Foundation, ESPID Fellowship funded by Seqirus, Hypatia Fellowship funded by Radboudumc and The Netherlands Organisation for Health Research and Development (ZonMW VENI grant to LM Verhagen).

Molecular typing of Mycoplasma pneumoniae and its correlation with macrolide resistance in children in Henan of China.

BACKGROUND/PURPOSE: As a major causative pathogen of community-acquired pneumonia, Mycoplasma pneumoniae (M. pneumoniae) can cause both upper and lower respiratory tract inflammation as well as extrapulmonary syndromes, especially in infants and the elderly. The emergence of macrolide-resistance has significant effects on the treatment of relevant diseases in children. This study aimed to analyze the genotypes and the macrolide resistance-associated mutations in M. pneumoniae sampled from the pediatric patients in Henan, China. METHODS: A segment of gene on the 23S rRNA was amplified and sequenced to detect the mutations related to macrolide resistance. Molecular typing was performed by the method named multiple locus variable-number tandem repeat analysis (MLVA) for macrolide-susceptible and macrolide-resistant specimens. RESULTS: Among the M. pneumoniae-positive samples, 95.7% (111/116) had macrolide-resistant mutation, and all of them consisted of the A2063G mutation. There were only two MLVA types identified in this study, type 4-5-7-2 (51/92, 55.4%) and type 3-5-6-2 (41/92, 44.6%). CONCLUSION: There was no correlation between MLVA types and macrolide resistance (P â€‹> â€‹0.05).

Multicenter evaluation of the Acaruiter Respiratory Panel for diagnosis of respiratory tract infections in Chinese children.

IMPORTANCE: Compared to multiplex PCR assays that are available in the Chinese market, the Acaruiter Respiratory Panel (fluorescent PCR) covers a wider range of pathogens including eight viruses, five bacteria, Mycoplasma pneumoniae and Chlamydia pneumoniae, and has high accuracy and effectiveness in the detection of pathogens. This is the first study to evaluate the performance of the Acaruiter Respiratory Panel. This regent may be a promising assay for comprehensive testing for respiratory pathogens from nasopharyngeal swab specimens in Chinese children.

Mycoplasma pneumoniae: Among the smallest bacterial pathogens with great clinical significance in children.

BACKGROUND: Mycoplasmas are the smallest prokaryotic microorganisms found in nature. Mycoplasma pneumoniae (M. pneumoniae) is the most commonly studied among human mycoplasmas. OBJECTIVES: In this review, we briefly focus on the recent developments that have enhanced our understanding of M. pneumoniae, one of the smallest pathogenic bacteria of great clinical importance in children. CONTENT: M. pneumoniae infections may involve either upper or lower respiratory tract or both of them. Extrapulmonary manifestations have been reported in almost every organ, including the skin and the hematologic, cardiovascular, musculoskeletal, and nervous system due to direct local effects, after dissemination of bacteria or indirect effects. The correct identification of M. pneumoniae infections is vital for prescription of the appropriate therapy.There are scarce specific findings of clinical laboratory results for the diagnosis of M. pneumoniae infection. Detection of M. pneumoniae infections can be achieved using culture, serology, or molecular-based methods. Culture is time-consuming, laborious, and expensive. The major types of serological tests for M. pneumoniae include the microtiter plate enzyme immunoassay (EIA), the membrane EIA, indirect immunofluorescence, and particle agglutination. Nucleic acid amplification tests (NAATs) include traditional PCR, nested PCR, real-time quantitative PCR, nucleic acid sequence-based amplification (NASBA), loop-mediated isothermal amplification (LAMP) technology, and RNA simultaneous amplification and testing (SAT). Macrolides have been the drug of choice for treating M. pneumoniae infection in past years. Clinically significant acquired macrolide-resistant M. pneumoniae (MRMP)has emerged worldwide which may be associated with more extrapulmonary complications, and severe clinical and radiological features. Since molecular-based assays can detect M. pnueumoniae in clinical specimens, there is a need for real point of care testing for fast detection of M. pneumoniae or its DNA and mutations in macrolide resistance gene. It is necessary to develop safe vaccines that provide protective immunity against M.pneumoniae infection.