A clade of Streptococcus pneumoniae clonal complex 320 with increased tolerance to ß-lactam antibiotics in a Chinese metropolitan city.

OBJECTIVES: We characterized the population structure and features of clinical Streptococcus pneumoniae isolates associated with invasive pneumococcal disease (IPD) from 2009 to 2017 in a Chinese metropolitan city using a whole-genome sequencing approach. METHODS: Seventy-nine pneumococcal strains, including 60 serogroup-19 strains from children enduring IPD from a paediatric hospital in Shenzhen, were subjected to whole-genome sequencing. Population structure was characterized through phylogenetic analysis, sequence typing, serotyping, virulence factor, and antimicrobial drug resistance (AMR) gene profiling, combining the publicly available related WGS data. Clinical demography and antibiotic susceptibility profiles were compared among different populations to emphasize the higher-risk populations. Genetic regions associated with AMR gene mobilization were identified through comparative genomics. RESULTS: These IPD strains mainly belonged to clonal complex 320 (CC320) and were composed of serotypes 19A and 19F. In addition to sporadic possible importation-related isolates (ST320), we identified an independent clade, CC320_SZpop (ST271), that predominantly circulated in Shenzhen and possibly expanded its range. Clinical features and antibiotic susceptibility analysis revealed that CC320_SZpop might manifest much higher pathogenicity and tolerance to ß-lactams. Specific virulence factors in Shenzhen isolates of CC320_SZpop were identified. Furthermore, an ca. 40 kb hotspot genomic region enduring frequent recombination was identified, possibly associated with the divergence of S. pneumoniae strains. CONCLUSION: A novel pneumococcal clade, CC320_SZpop, circulating in Shenzhen and other regions in China, possibly under expansion, was found and deserves more study and surveillance. Our study also emphasizes the importance of continuous genomic surveillance of clinical S. pneumoniae isolates, especially IPD isolates.

Molecular characteristics and antimicrobial resistance of invasive pneumococcal isolates from children in the post-13-valent pneumococcal conjugate vaccine era in Shenzhen, China.

OBJECTIVES: This study aimed to evaluate the molecular epidemiology and antimicrobial resistance of invasive pneumococcal isolates from children in Shenzhen, China, in the early stage of the pneumococcal 13-valent conjugated vaccine (PCV-13) era from 2018 to 2020. METHODS: Invasive pneumococcal strains were isolated from hospitalized children with invasive pneumococcal diseases (IPDs) from January 2018 to December 2020. The serotype identification, multilocus sequence typing (MLST), and antibiotic susceptibility tests were performed on all culture-confirmed strains. RESULTS: Sixty-four invasive strains were isolated mainly from blood (70.3%). Prevalent serotypes were 23F (28.1%), 14 (18.8%), 19F (15.6%), 6A/B (14.1%), and 19A (12.5%), with a serotype coverage rate of 96.9% for PCV13. The most common sequence types (STs) were ST876 (17.1%), ST271 (10.9%), and ST320 (7.8%). Half of the strains were grouped in clonal complexes (CCs): CC271 (21.9%), CC876 (20.3%), and CC90 (14.1%). Meningitis isolates showed a higher resistance rate (90.9% and 45.5%) to penicillin and ceftriaxone than the rate (3.8% and 9.4%) of non-meningitis isolates. The resistance rates for penicillin (oral), cefuroxime, and erythromycin were 53.13%, 73.4%, and 96.9%, respectively. The dual ermB and mefA genotype was found in 81.3% of erythromycin-resistant strains. The elevated minimum inhibitory concentration (MIC) of ß-lactam antibiotics and dual-genotype macrolide resistance were related mainly to three major serotype-CC combinations: 19F-CC271, 19A-CC271, and 14-CC876. CONCLUSION: Invasive pneumococcus with elevated MICs of ß-lactams and increased dual ermB and mefA genotype macrolide resistance were alarming. Expanded PCV13 vaccination is expected to reduce the burden of paediatric IPD and to combat antibiotic-resistant pneumococcus in Shenzhen.

LincR-PPP2R5C Promotes Th2 Cell Differentiation Through PPP2R5C/PP2A by Forming an RNA-DNA Triplex in Allergic Asthma.

PURPOSE: The roles and mechanisms of long noncoding RNAs (lncRNAs) in T helper 2 (Th2) differentiation from allergic asthma are poorly understood. We aimed to explore a novel lncRNA, LincR-protein phosphatase 2 regulatory subunit B' gamma (PPP2R5C), in Th2 differentiation in a mouse model of asthma. METHODS: LincR-PPP2R5C from RNA-seq data of CD4+ T cells of asthma-like mice were validated and confirmed by quantitative reverse transcription polymerase chain reaction, northern blotting, nuclear and cytoplasmic separation, and fluorescence in situ hybridization (FISH). Lentiviruses encoding LincR-PPP2R5C or shRNA were used to overexpress or silence LincR-PPP2R5C in CD4+ T cells. The interactions between LincR-PPP2R5C and PPP2R5C were explored with western blotting, chromatin isolation by RNA purification assay, and fluorescence resonance energy transfer. An ovalbumin-induced acute asthma model in knockout (KO) mice (LincR-PPP2R5C KO, CD4 conditional LincR-PPP2R5C KO) was established to explore the roles of LincR-PPP2R5C in Th2 differentiation. RESULTS: LncR-PPP2R5C was significantly higher in CD4+ T cells from asthmatic mice ex vivo and Th2 cells in vitro. The lentivirus encoding LincR-PPP2R5C suppressed Th1 differentiation; in contrast, the short hairpin RNA (shRNA) lentivirus decreased LincR-PPP2R5C and Th2 differentiation. Mechanistically, LincR-PPP2R5C deficiency suppressed the phosphatase activity of the protein phosphatase 2A (PP2A) holocomplex, resulting in a decline in Th2 differentiation. The formation of an RNA-DNA triplex between LincR-PPP2R5C and the PPP2R5C promoter enhanced PPP2R5C expression and activated PP2A. LincR-PPP2R5C KO and CD4 conditional KO decreased Th2 differentiation, airway hyperresponsiveness and inflammatory responses. CONCLUSIONS: LincR-PPP2R5C regulated PPP2R5C expression and PP2A activity by forming an RNA-DNA triplex with the PPP2R5C promoter, leading to Th2 polarization in a mouse model of acute asthma. Our data presented the first definitive evidence of lncRNAs in the regulation of Th2 cells in asthma.

Azalomycin F4a targets peptidoglycan synthesis of Gram-positive bacteria revealed by high-throughput CRISPRi-seq analysis.

Azalomycin F4a is a promising 36-membered polyhydroxy macrolide that shows antibacterial activity against drug-resistant Gram-positive bacteria, but its exact working mechanism remains to be elusive. Here, we isolated the azalomycin F4a product from a Streptomyces solisilvae and demonstrated its antibacterial activity against Gram-positive pathogens including Streptococcus pneumoniae, Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA). We further showed that combination of azalomycin F4a with methicillin has an additive antimicrobial effect on MRSA, where the minimal inhibitory concentrations (MIC) of methicillin to MRSA was decreased by 1000-fold in the presence of sublethal concentration of azalomycin F4a. A CRISPRi-seq based whole genome screen was employed to identify the potential targets of azalomycin F4a, which revealed that peptidoglycan synthesis (PGS) was inhibited by azalomycin F4a. Furthermore, azalomycin F4a treatment could significantly impair S. aureus biofilm formation. Our research highlights that cell wall synthesis is an additional target for novel classes of macrolide besides ribosome.

The usnic acid derivative peziculone targets cell walls of Gram-positive bacteria revealed by high-throughput CRISPRi-seq analysis.

Usnic acid, a representative dibenzofuran metabolite, is known to have antimicrobial properties. However, despite considerable interest as an antimicrobial agent, the mechanism by which usnic acid and its derivatives exert their action is not fully characterized. This article describes the synthesis of peziculone, a 5:1 equilibrium mixture of two inseparable usnic acid derivatives: peziculone A and peziculone B. The antibacterial activity of peziculone against several Gram-positive bacterial pathogens was found to be significantly better compared with usnic acid. Clustered regularly interspaced short palindromic repeats interference sequencing analysis and membrane fluorescent staining were used to demonstrate that peziculone destabilizes the cell walls of Gram-positive bacteria. Additionally, peziculone 2.5 and 3.5 µg/mL impaired cell surface appendages and biofilm formation by Staphylococcus aureus. Taken together, these data demonstrate that peziculone, a derivative compound of usnic acid, has significant antimicrobial activity against Gram-positive bacteria by targeting the cell walls; this provides a platform for development of novel antibacterial drugs.

Etiology and clinical features of children with bronchiectasis in China: A 10-year multicenter retrospective study.

INTRODUCTION: The current study aims to investigate the etiology spectrum and the clinical characteristics of bronchiectasis in Chinese children. METHODS: The study is designed as a multicenter retrospective study. 193 cases were enrolled in 13 centers in China between 2008 and 2017. The inclusive cases must meet the clinical as well as the HRCT criteria. Only if both two radiologists confirmed the diagnosis, the case could be enrolled. The cases that could not provide clinical and imageology data were excluded. The data were entered into the specialized system and then analyzed. RESULTS: One hundred sixty-nine cases (87%) were found to have the underlying etiology. Post-infective (46%), primary immunodeficiency (14%), and PCD (13%) were the common causes. All cases came from 28 provinces in Mainland China. The median age of symptom onset was 5.8 (2.0, 8.9) years. The median age of diagnosis was 8.4 (4.5, 11.6) years. The main symptoms were cough, sputum expectoration, and fever during the exacerbation. Nineteen percent of patients suffered from limited exercise tolerance. Clubbing was found in 17% of cases. Nearly 30% of patients presented growth limitations. On the HRCT findings, 126 cases had diffused bronchiectasis, and bilateral involvement was found in 94 cases. The lower lobes and right middle lobes were most commonly involved. Approximately 30% of cultures of sputum and bronchoalveolar lavage were positive. CONCLUSION: A majority of cases could be found the underlying etiology. Post-infective, primary immunodeficiency, and PCD were the most common causes. Some clinical figures might indicate a specific etiology.

Talaromyces marneffei infection and complicate manifestation of respiratory system in HIV-negative children.

BACKGROUND: Respiratory symptoms are the earliest clinical manifestation of Talaromyces marneffei (TM) infection. In this study, we aimed to improve the early identification of TM infection in human immunodeficiency virus (HIV)-negative children with respiratory symptoms as the first manifestation, analyze the risk factors, and provide evidence for diagnosis and treatment. METHODS: We retrospectively analyzed six cases of HIV-negative children with respiratory system infection symptoms as the first presentation. RESULTS: All subjects (100%) had cough and hepatosplenomegaly, and five subjects (83.3%) had a fever; other symptoms and signs included lymph node enlargement, rash, rales, wheezing, hoarseness, hemoptysis, anemia, and thrush. Additionally, 66.7% of the cases had underlying diseases (three had malnutrition, one had severe combined immune deficiency [SCID]). The most common coinfecting pathogen was Pneumocystis jirovecii, which occurred in two cases (33.3%), followed by one case of Aspergillus sp. (16.6%). Furthermore, the value of ß-D-glucan detection (G test) increased in 50% of the cases, while the proportion of NK decreased in six cases (100%). Five children (83.3%) were confirmed to have the pathogenic genetic mutations. Three children (50%) were treated with amphotericin B, voriconazole, and itraconazole, respectively; three children (50%) were treated with voriconazole and itraconazole. All children were tested for itraconazole and voriconazole plasma concentrations throughout antifungal therapy. Two cases (33.3%) relapsed after drug withdrawal within 1 year, and the average duration of antifungal treatment for all children was 17.7 months. CONCLUSION: The first manifestation of TM infection in children is respiratory symptoms, which are nonspecific and easily misdiagnosed. When the effectiveness of anti-infection treatment is poor for recurrent respiratory tract infections, we must consider the condition with an opportunistic pathogen and attempt to identify the pathogen using various samples and detection methods to confirm the diagnosis. It is recommended the course for anti-TM disease be longer than one year for children with immune deficiency. Monitoring the blood concentration of antifungal drugs is important.

Prevalence and resistance characteristics of multidrug-resistant Streptococcus pneumoniae isolated from the respiratory tracts of hospitalized children in Shenzhen, China.

Background: PCV13 introduction in China has led to a significant reduction of vaccine serotype Streptococcus pneumoniae. However, non-vaccine serotypes with highly resistance and invasiveness were often reported in the post-pneumococcal conjugate vaccine era and there was regional differences. Methods: A total of 669 S. pneumoniae strains were collected from the respiratory tracts of hospitalized children at Shenzhen Children's Hospital in 2021 and 2022. Antimicrobial resistance (AMR) characteristics were assessed through antibiotic susceptibility testing performed with the VITEK 2 compact system. AMR genes and single nucleotide polymorphisms (SNPs) in pbp1a, pbp2b, and pbp2x were identified via analysis of whole genome sequencing data. Statistical examination of the data was conducted employing chi-square and Fisher's exact tests. Results: We found that non-vaccine serotypes strains had accounted for 46.6% of all the pneumococcal isolated strains. The most common non-vaccine serotype is 23A, with a prevalence rate of 8.9%, followed by 15A (6.6%), 6E (5.7%), 34 (3.2%), and 15B (2.9%). The multidrug resistance rates (MDR) of vaccine serotypes were 19F (99.36%), 19A (100%), 23F (98.08%), 6B (100%), and 6C (100%). Meanwhile, the MDR of non-vaccine serotypes were 15B (100.00%), 6E (100%), 15C (100%), 34 (95.24%), and 23A (98.31%). Resistance rates of 6E to more than six antibiotic classes reached 89.47%, which is similar to 19F (83.33%) and 19A (90%). Unique resistance profiles were also identified for non-vaccine serotypes, including significantly higher resistance to chloramphenicol in 6E, 15B, and 15C than in 19F and 19A. Furthermore, through genome sequencing, we revealed strong correlation of cat-TC with chloramphenicol resistance, patA/patB with tetracycline resistance, ermB and pmrA with erythromycin resistance. Conclusion: The introduction of PCV13 into China from 2017 has led to a shift in the dominant composition of pneumococcal strains. There has been a notable rise and spread of multidrug-resistant non-vaccine serotypes among children. Specifically, the non-vaccine serotype 6E, which was not widely reported in China previously, has emerged. To comprehend the resistance mechanisms, it is crucial to further investigate the molecular and genetic characteristics of these non-vaccine serotypes.

KIF2A decreases IL-33 production and attenuates allergic asthmatic inflammation.

BACKGROUND: The microtubule-dependent molecular motor protein Kinesin Family Member 2A (KIF2A) is down-regulated in asthmatic human airway epithelium. However, little is known about the roles of KIF2A as well as the possible underlying mechanisms in asthma. METHODS: House dust mite (HDM) extract was administered to establish a murine model of asthma. The expression of KIF2A, IL-33 and the autophagy pathways were detected. The plasmid pCMV-KIF2A was used to overexpress KIF2A in the airway epithelial cells in vitro and in vivo. IL-4, IL-5, IL-33 and other cytokines in bronchoalveolar lavage fluid (BALF) and lung tissues homogenates were measured. RESULTS: In response to the challenge of house dust mite (HDM) in vitro and in vivo, airway epithelial cells displayed decreased production of KIF2A. Meanwhile, autophagy and IL-33 were increased in HMD-treated epithelial cells. Mechanistically, KIF2A decreased autophagy via suppressing mTORC1 pathway in HDM-treated epithelial cells, which contributed to the reduced production of IL-33. Moreover, in vivo KIF2A transfection reduced IL-33 and autophagy in the lung, leading to the attenuation of allergic asthma. CONCLUSION: KIF2A suppressed mTORC1-mediated autophagy and decreased the production of epithelial-derived cytokine IL-33 in allergic airway inflammation. These data indicate that KIF2A may be a novel target in allergic asthma.

Molecular Characteristics of Streptococcus pyogenes Isolated From Chinese Children With Different Diseases.

Streptococcus pyogenes is a bacterial pathogen that causes a wide spectrum of clinical diseases exclusively in humans. The distribution of emm type, antibiotic resistance and virulence gene expression for S. pyogenes varies temporally and geographically, resulting in distinct disease spectra. In this study, we analyzed antibiotic resistance and resistance gene expression patterns among S. pyogenes isolates from pediatric patients in China and investigated the relationship between virulence gene expression, emm type, and disease categories. Forty-two representative emm1.0 and emm12.0 strains (n = 20 and n = 22, respectively) isolated from patients with scarlet fever or obstructive sleep apnea-hypopnea syndrome were subjected to whole-genome sequencing and phylogenetic analysis. These strains were further analyzed for susceptibility to vancomycin. We found a high rate and degree of resistance to macrolides and tetracycline in these strains, which mainly expressed ermB and tetM. The disease category correlated with emm type but not superantigens. The distribution of vanuG and virulence genes were associated with emm type. Previously reported important prophages, such as φHKU16.vir, φHKU488.vir, Φ5005.1, Φ5005.2, and Φ5005.3 encoding streptococcal toxin, and integrative conjugative elements (ICEs) such as ICE-emm12 and ICE-HKU397 encoding macrolide and tetracycline resistance were found present amongst emm1 or emm12 clones from Shenzhen, China.

[Clinical features of sleep-disordered breathing in children with neuromuscular disease].

OBJECTIVE: To study the clinical features of sleep-disordered breathing (SDB) in children with neuromuscular disease (NMD). METHODS: A retrospective analysis was performed on the medical data of 18 children who were diagnosed with NMD and underwent polysomnography (PSG) (NMD group). Eleven children without NMD who had abnormal sleeping habit and normal sleep structure on PSG were enrolled as the control group. The two groups were compared in terms of the daily and nocturnal symptoms of SDB, incidence rate of obstructive sleep apnea (OSA), pulmonary function, end-tidal partial pressure of carbon dioxide (PetCO2), features of sleep structure, and sleep respiratory events. RESULTS: In the NMD group, 16 children (89%) had related daily and nocturnal symptoms of SDB, and the youngest age was 1 year at the onset of such symptoms. Compared with the control group, the NMD group had significant reductions in total sleep time and sleep efficiency (P < 0.05), a significant reduction in the proportion of rapid eye movement (REM) sleep (P < 0.05), significant increases in obstructive apnea and hypopnea events (P < 0.05) and oxygen reduction events during REM sleep (P < 0.05), and a significant reduction in blood oxygen saturation during REM sleep (P < 0.05). In the NMD group, 17 children (94%) were diagnosed with OSA, and all children had normal lung function and PetCO2. CONCLUSIONS: There is a high proportion of children with SDB among the children with NMD, and SDB can be observed in the early stage of NMD, which results in the damage of sleep structure and the reduction in sleep efficiency. Respiratory events are mainly obstructive events, and oxygen reduction events are mainly observed during REM sleep.

Severe acute respiratory syndrome coronavirus 2-induced multisystem inflammatory syndrome in children.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-induced multisystem inflammatory syndrome in children (MIS-C) is a life-threatening illness that has been reported in the United States and Europe. It affects multiple organ systems and often requires patient admission to an intensive care unit. Although some features of MIS-C overlap with Kawasaki disease, MIS-C is more common among older children and adolescents, more often affects cardiovascular and gastrointestinal systems, and more frequently presents with elevated inflammatory markers. Rapid and complete clinical recovery is possible in nearly all patients following immunomodulation therapy. Thus far, MIS-C pathophysiology and long-term prognosis are not sufficiently clear; further studies are needed.

Divergent Roles of miR-3162-3p in Pulmonary Inflammation in Normal and Asthmatic Mice as well as Antagonism of miR-3162-3p in Asthma Treatment.

MicroRNA (miRNA) mimics or antagomirs hold great promise for asthma treatment compared with glucocorticoids as mainstay therapy for asthma. But the role of miRNA in regulating asthmatic inflammation is largely unclear. We previously reported that miR-3162-3p in the peripheral blood of children with asthma was obviously upregulated compared to that in healthy children. This study aimed to elucidate the role of miR-3162-3p in pulmonary inflammation in normal and asthmatic mice as well as preliminarily explore the potential of miR-3162-3p antagomir in asthma treatment. A noninvasive whole-body plethysmograph measured airway responsiveness. Both qRT-PCR and Western blot were used to detect the expression of miRNA, mRNA, or protein. Cells in bronchoalveolar lavage fluid were counted by platelet counting and Wright's staining. Inflammatory infiltration and mucus secretion were identified by hematoxylin and eosin and periodic acid-Schiff  staining, respectively. Cytokines in the lungs were detected by ELISA. The miR-3162-3p mimic intraperitoneally administered to normal mice decreased ß-catenin levels in the lungs without obviously altering the lung histology and cytokine levels. Antagonizing miR-3162-3p in ovalbumin-induced asthmatic mice effectively alleviated the typical features of asthma, such as airway hyper-responsiveness, airway inflammation, and Th1/Th2 cytokine imbalance, and concomitantly rescued the total and active ß-catenin expression. Collectively, we discovered divergent roles of miR-3162-3p in lung inflammation between normal and asthmatic mice. The anti-inflammatory effects of the miR-3162-3p antagomir were comparable to those of glucocorticoid treatment. Our study helped in understanding the contribution of miRNAs to the pathogenesis of asthma.

[Clinical features of influenza with plastic bronchitis in children].

OBJECTIVE: To study the clinical features of influenza with plastic bronchitis (PB) in children, and to improve the awareness of the diagnosis and treatment of PB caused by influenza virus. METHODS: A retrospective analysis was performed for the clinical data of 70 children with lower respiratory influenza virus infection from October 2018 to October 2019. According to the presence or absence of PB, they were divided into an influenza+PB group with 12 children and a non-PB influenza group with 58 children. Related clinical data were collected for the retrospective analysis, including general information, clinical manifestations, laboratory examination, imaging findings, treatment, and prognosis. RESULTS: In the influenza+PB group, most children experienced disease onset at the age of 1-5 years, with the peak months of January, February, July, and September. Major clinical manifestations in the influenza+PB group included fever, cough, and shortness of breath. The influenza+PB group had significantly higher incidence rates of shortness of breath and allergic diseases such as asthma than the non-PB influenza group (P<0.05). Of the 12 children in the influenza+PB group, 7(58%) had influenza A virus infection and 5 (42%) had influenza B virus infection, among whom 1 had nephrotic syndrome. For the children in the influenza+PB group, major imaging findings included pulmonary consolidation with atelectasis, high-density infiltration, pleural effusion, and mediastinal emphysema. Compared with the non-PB influenza group, the influenza+PB group had a significantly higher proportion of children who were admitted to the pediatric intensive care unit (P<0.05). Bronchoscopic lavage was performed within 1 week after admission, and all children were improved and discharged after anti-infective therapy and symptomatic/supportive treatment. CONCLUSIONS: Influenza with PB tends to have acute onset and rapid progression, and it is important to perform bronchoscopy as early as possible. The possibility of PB should be considered when the presence of shortness of breath, allergic diseases such as asthma or nephrotic syndrome in children with influenza.

The changing phenotypes and genotypes of invasive pneumococcal isolates from children in Shenzhen during 2013-2017.

BACKGROUND: The phenotypes and genotypes of Streptococcus pneumoniae isolated from invasive pneumococcal diseases (IPDs) were changing all the time. To monitor these changes of phenotypes and genotypes of S. pneumoniae isolates from children, we examined antibiotic susceptibility, serotype distribution and sequence types (STs) of S. pneumoniae, which were isolated before the 13-valent pneumococcal conjugate vaccine (PCV13) introduced into China. METHODS: Strains were isolated from children less than 14 years old between January 2013 and May 2017 from Shenzhen Children's Hospital. Serotypes, antibiotic resistance, and genotypes of these isolates were determined using capsular swelling, E-test, and multi-locus sequence typing, respectively. RESULTS: A total of 94 S. pneumoniae strains were isolated, which belonged to 15 serotypes. The five most prevalent serotypes were 19F (25.5%), 19A (19%), 14 (17%), 23F (7.5%), and 6B (9.6%). We found 42 STs for these isolates. The most abundant STs were ST271 (24.4%), ST876 (17%), and ST320 (10.6%), mainly related to 19F, 14, and 19A, respectively. The potential coverage of PCV13 was 87.2%. Among non-meningitis isolates, the resistance rates to penicillin and ceftriaxone were 0% and 2%. However, the meningitis isolates showed high resistance to penicillin (80%) and ceftriaxone (20%). Most of these isolates (95.7%) were resistant to erythromycin, and 66 (70.2%) strains carried the ermB gene and 24 (25.5%) strains carried both the ermB and mefA/E genes. Serotype 19A showed the highest mean minimum inhibitory concentration (MIC) for penicillin (MIC = 1.486) than the other serotypes, but no significant difference in penicillin MIC among the three main STs (ST271, ST320, and ST876). CONCLUSIONS: The phenotypes and genotypes of invasive pneumococcal isolates from Shenzhen Children's Hospital have changed with the passage of time. Compared with PCV7, PCV13 can more effectively protect Chinese children from IPDs. To some extent, these changes are possibly related to the usage of antibiotics and vaccines.

An integrated respiratory microbial gene catalogue to better understand the microbial aetiology of Mycoplasma pneumoniae pneumonia.

BACKGROUND: The imbalanced respiratory microbiota observed in pneumonia causes high morbidity and mortality in childhood. Respiratory metagenomic analysis demands a comprehensive microbial gene catalogue, which will significantly advance our understanding of host-microorganism interactions. RESULTS: We collected 334 respiratory microbial samples from 171 healthy children and 76 children with pneumonia. The respiratory microbial gene catalogue we established comprised 2.25 million non-redundant microbial genes, covering 90.52% of prevalent genes. The major oropharyngeal microbial species found in healthy children were Prevotella and Streptococcus. In children with Mycoplasma pneumoniae pneumonia (MPP), oropharyngeal microbial diversity and associated gene numbers decreased compared with those of healthy children. The concurrence network of oropharyngeal microorganisms in patients predominantly featured Staphylococcus spp. and M. pneumoniae. Functional orthologues, which are associated with the metabolism of various lipids, membrane transport, and signal transduction, accumulated in the oropharyngeal microbiome of children with pneumonia. Several antibiotic resistance genes and virulence factor genes were identified in the genomes of M. pneumoniae and 13 other microorganisms reconstructed via metagenomic data. Although the common macrolide/ß-lactam resistance genes were not identified in the assembled M. pneumoniae genome, a single-nucleotide polymorphism (A2063G) related to macrolide resistance was identified in a 23S ribosomal RNA gene. CONCLUSIONS: The results of this study will facilitate exploration of unknown microbial components and host-microorganism interactions in studies of the respiratory microbiome. They will also yield further insights into the microbial aetiology of MPP.

Bronchoalveolar lavage fluid microbiota dysbiosis in infants with protracted bacterial bronchitis.

BACKGROUND: Protracted bacterial bronchitis (PBB) is a chronic purulent bronchitis which could cause recurrent coughing and wheezing in infants. Based on previous reports, main pathogens which caused PBB were identified in the patients, but their impacts on lung microbiota dysbiosis remain unclear. METHODS: In this study, bronchoalveolar lavage fluid (BALF) was collected from PBB infants and tracheomalacia (TM) infants younger than 3 years old under the instruction of Shenzhen Children's Hospital, and 12 samples were randomly selected for 16S rDNA analysis in each group. Based on the results of bacterial composition, the microbiota diversity and co-occurrence network in PBB and TM group were detected and compared. RESULTS: Microbiota diversity was significantly lower in PBB group than it in TM group (P<0.001 for the comparison of Shannon and Simpson indexes). The PBB group was found to harbor 25 accumulated bacterial agents by comparison with TM group, including Haemophilus (P<0.001) and Bacteroides (P<0.001). Whilst, the populations of Lactococcus (P<0.001) and Lactobacillus (P<0.001) were dramatically smaller in PBB group. The co-occurrence network in PBB group also differed from that of TM group. It contained four core nodes in PBB patients, including Haemophilus, Parabacteroides, Porphyromonas, and Cronobacter. Haemophilus was found to be negatively associated with most counterparts, including Clostridium and Bacillus. CONCLUSIONS: PBB infants contained discrepant lung genera and co-occurrence network when compared with TM infants. This retrospective study may deepen our understanding of PBB pathogenesis, and it also provided a foundation for bacterial adjunctive therapy of infantile PBB in accordance with clinical treatment.