Objective The prevalence of drug-resistant Mycobacterium tuberculosis (Mtb) strains is exacerbating the global burden of tuberculosis (TB), highlighting the urgent need for new treatment strategies for TB. Methods The recombinant adenovirus vaccine expressing cyclic di-adenosine monophosphate (c-di-AMP) phosphodiesterase B (CnpB) (rAd-CnpB), was administered to normal mice via mucosal immunization, either alone or in combination with drug therapy, to treat Mtb respiratory infections in mice.Enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of antibodies in serum and bronchoalveolar lavage fluid (BALF). Real-time quantitative PCR was performed to assess the transcription levels of cytokines interferon γ(IFN-γ) and interleukin 10(IL-10) in mouse lungs. Flow cytometry was used to determine the proportions of CD4⁺ and CD8⁺ T cell subsets in the lungs and spleens. ELISA was employed to measure the levels of cytokines IFN-γ, IL-2, IL-10, inflammatory factors IL-6, and tumor necrosis factor α (TNF-α) secreted by spleen cells following antigen stimulation. The bacteria loads in the lungs and spleens of Mtb-infected mice were enumerated by plate counting methods. Resluts Intranasal immunization with rAd-CnpB induced high titers of IgG in mouse serum and the production of IgG and IgA in BALF, along with alterations in T lymphocyte subsets in the lungs and spleens. Administration of rAd-CnpB, either alone or in combination with drugs, to Mtb-infected mice significantly increased serum IgG levels as well as IgA and IgG levels in BALF. rAd-CnpB immunization promoted the secretion of CnpB-specific cytokines and inflammatory factors by splenocytes in Mtb-infected mice. However, rAd-CnpB immunotherapy, either alone or combined with drugs, did not significantly affect the bacterial loads in the lungs and spleens of mice with Mtb respiratory infections. Conclusion Mucosal immunization with rAd-CnpB induced significant mucosal, humoral and cellular immune responses in mice, and significantly enhanced CnpB-specific cellular immune responses in Mtb-infected mice.
Animals ; Adenoviridae/immunology* ; Mycobacterium tuberculosis/genetics* ; Mice ; Female ; Phosphoric Diester Hydrolases/genetics* ; Tuberculosis Vaccines/administration & dosage* ; Tuberculosis/prevention & control* ; Mice, Inbred BALB C ; Cytokines ; Lung/microbiology* ; Immunization ; Bronchoalveolar Lavage Fluid/immunology* ; Immunity, Mucosal

OBJECTIVES: To study the characteristics of bronchoalveolar lavage fluid (BALF) microbial distribution at different stages of refractory Mycoplasma pneumoniae pneumonia (RMPP) in children and its relationship with immune function. METHODS: A total of 108 children with RMPP were enrolled. The relative abundance, richness, and diversity of BALF microbiota, as well as immune function, were compared between the acute phase (n=61) and recovery phase (n=47). The correlations between the richness and diversity of BALF microbiota and immune function were analyzed. RESULTS: The relative abundance of Propionibacterium, as well as the Simpson index, Shannon index, Chao1 index, and Observed species index of BALF microbiota in the acute phase were significantly lower than those in the recovery phase (P<0.05). The relative abundances of Streptococcus and Prevotella, as well as the levels of complement C3, complement C4, immunoglobulin A (IgA), immunoglobulin G (IgG), and immunoglobulin M (IgM), were significantly higher in the acute phase than in the recovery phase (P<0.05). Simpson, Shannon, Chao1, and Observed species indices were negatively correlated with levels of complement C3, complement C4, IgA, IgM, and IgG (P<0.05). CONCLUSIONS In children with RMPP, the relative abundance of Propionibacterium and the richness and diversity of BALF microbiota in the acute phase are lower than those in the recovery phase, while the relative abundances of Streptococcus and Prevotella are higher in the acute phase. Microbial richness and diversity are closely related to immune function.
Humans ; Male ; Pneumonia, Mycoplasma/microbiology* ; Female ; Bronchoalveolar Lavage Fluid/microbiology* ; Child, Preschool ; Child ; Infant ; Microbiota

OBJECTIVE: To investigate the application value of bronchoalveolar lavage fluid (BALF) metagenomic next-generation sequencing (mNGS) in etiological diagnosis of children with hematological malignancies complicated with pneumonia. METHODS: We retrospectively analyzed the clinical data of children with hematological malignancies who underwent BALF mNGS pathogenic detection due to pneumonia. All patients underwent mNGS detection of bronchoalveolar lavage fluid as well as traditional methods(including sputum culture, bronchoalveolar lavage fluid culture, blood culture, serological detection of pathogens, etc.). By analyzing the results of mNGS and traditional methods, we compared key indicators such as the positive rate, etiological distribution. RESULTS: A total of 26 children with hematological malignancies enrolled in the study, including 12 males and 14 females, with a median age of 4.9 (1.8-14.9) years, underwent bronchoalveolar lavage (BAL) 35 times. A total of 17 pathogenic microorganisms were detected in BALF mNGS, including 9 cases of bacterial infection, 10 cases of viral infection, 3 cases of fungal infection, 2 cases of mycoplasma infection and 8 cases of mixed infection, and the most commonly detected bacteria, viruses and fungi were streptococcus pneumoniae, cytomegalovirus and pneumocystis jirovecii, respectively. The positive rate of mNGS detection (91.43%) was significantly higher than that of traditional methods detection (20%, P <0.001). A total of 25 cases were adjusted according to BALF mNGS results. CONCLUSION The application of BALF mNGS technology can improve the detection rate of the pathogens in children with hematological malignancies complicated with pneumonia, initially revealed the pathogen spectrum of pulmonary infection in this group, and effectively guide clinical medication, improve treatment outcomes.
Humans ; Bronchoalveolar Lavage Fluid/microbiology* ; Hematologic Neoplasms/complications* ; Child ; Child, Preschool ; Infant ; Retrospective Studies ; Male ; Female ; Adolescent ; High-Throughput Nucleotide Sequencing ; Pneumonia/microbiology*

Pneumocystis jirovecii pneumonia (PJP) is an opportunistic pulmonary infection that commonly occurs in immunocompromised children. We report a case of infantile leukemia complicated by PJP and review the relevant literature. A summary and analysis of 10 infantile leukemia patients with PJP infection (9 cases reported in the literature and 1 case from our center) showed that PJP mostly occurred in the early stages of chemotherapy (80%, 8/10). The main clinical manifestations were dyspnea (100%, 10/10) and hypoxemia (50%, 5/10), while pulmonary imaging findings lacked specificity. In most cases (50%, 5/10), diagnosis was established by identifying pathogens in bronchoalveolar lavage fluid under microscopy. In our case, diagnosis was confirmed using targeted next-generation sequencing (tNGS) of bronchoalveolar lavage fluid. Treatment with intravenous sulfamethoxazole complex was administered in 8 patients, all of whom eventually recovered. PJP may occur in the early stages of chemotherapy for infantile leukemia, thus early prevention is necessary. tNGS facilitates early diagnosis of PJP, and sulfamethoxazole complex remains an effective therapeutic option.
Humans ; Infant ; Bronchoalveolar Lavage Fluid/microbiology* ; Immunocompromised Host ; Leukemia/complications* ; Pneumocystis carinii/isolation & purification* ; Pneumonia, Pneumocystis/diagnosis* ; Trimethoprim, Sulfamethoxazole Drug Combination/therapeutic use*

OBJECTIVES: To investigate the protective effect of the probiotic bacterium Streptococcus salivarius K12 (K12) against Mycoplasma pneumoniae (Mp) infection in mice. METHODS: Forty male BALB/c mice were randomized into normal control group, K12 treatment group, Mp infection group, and K12 pretreatment prior to Mp infection group. The probiotic K12 was administered daily by gavage for 14 days before Mp infection induced by intranasal instillation of Mp. Three days after Mp infection, the mice were euthanized for analysis of bronchoalveolar lavage fluid (BALF) cell counts and serum levels of secretory immunoglobulin A (sIgA), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6). RT-qPCR was performed to detect the P1 and community-acquired respiratory distress syndrome ( CARDS ) toxin of Mp in the lung tissues and the mRNA expressions of TNF-α, IL-6, chemokine 1 (CXCL1), matrix metalloproteinase 9 (MMP9), mucin 5ac (MUC5ac), collagen 3a1 (Col3a1), Toll-like receptor 2 (TLR2) and TLR4; the protein expressions of TLR2 and TLR4 in the lung tissue were detected using Western blotting. Pathological changes in the lung tissue and airway remodeling were examined with HE staining and AB/PAS staining. RESULTS: Compared with the Mp-infected mice with PBS treatment, the infected mice with K12 treatment showed significantly lowered mRNA levels of P1 and CARDS in the lung tissue and reduced white blood cell counts in the BALF (P<0.05). In spite of the absence of significant differences in serum levels of inflammatory factors between the two groups, the mRNA expressions of TNF‑α, IL-6, CXCL1, MMP9, MUC5ac and COL3A1 and the mRNA and protein levels of TLR2 and TLR4 in the lung tissues were significantly lower in K12-treated mice, in which AB/PAS staining showed obviously decreased mucus secretion. CONCLUSIONS K12 pretreatment can effectively reduce pulmonary inflammatory responses, improve airway remodeling and alleviate lung injury in Mp-infected mice.
Animals ; Mice ; Pneumonia, Mycoplasma/metabolism* ; Mice, Inbred BALB C ; Toll-Like Receptor 2/metabolism* ; Mycoplasma pneumoniae ; Male ; Tumor Necrosis Factor-alpha/metabolism* ; Interleukin-6/metabolism* ; Lung/microbiology* ; Toll-Like Receptor 4/metabolism* ; Streptococcus salivarius ; Probiotics/administration & dosage* ; Bronchoalveolar Lavage Fluid ; Matrix Metalloproteinase 9/metabolism* ; Mucin 5AC/metabolism* ; Chemokine CXCL1/metabolism* ; Immunoglobulin A, Secretory/metabolism* ; Bacterial Toxins ; Bacterial Proteins

This paper was mainly to discuss the potential role and mechanism of Lianhua Qingwen Capsules(LHQW) in inhibiting pathological inflammation in the model of acute lung injury caused by bacterial infection. For in vitro study, the mRNA expression of MCP-1 in RAW264.7 cells and THP-1 cells, the content of MCP-1 in cell supernatant, as well as the effect of LHQW on chemotaxis of macrophages were detected. For in vivo study, mice were randomly divided into 7 groups, including normal group, model group(LPS 5 mg·kg~(-1)), LHQW 300, 600 and 1 200 mg·kg~(-1)(low, middle and high dose) groups, dexamethasone 5 mg·kg~(-1) group and penicillin-streptomycin group. Then, the anal temperature was detected two hours later. Dry weight and wet weight of lung tissues in mice were determined; TNF-α and MCP-1 levels in alveolar lavage fluid and MCP-1 in serum were detected. In addition, the infiltration of alveolar macrophages was also observed and the infiltration count of alveolar macrophages was measured by CCK-8 method. HE staining was also used to observe the inflammatory infiltration of lung tissues in mice. Both of the in vitro and in vivo data consistently have confirmed that: by down-regulating the expression of MCP-1, LHWQ could efficiently decrease the chemotaxis of monocytes toward the pulmonary infection foci, thus blocking the disease development in ALI animal model.
Acute Lung Injury ; microbiology ; Animals ; Bacterial Infections ; drug therapy ; Bronchoalveolar Lavage Fluid ; Capsules ; Chemokine CCL2 ; metabolism ; Chemotaxis ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Lipopolysaccharides ; Lung ; Macrophages ; drug effects ; Mice ; RAW 264.7 Cells ; Random Allocation ; THP-1 Cells ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVENoncystic fibrosis (non-CF) bronchiectasis remains as a common health problem in Asia. Pathogens' distribution in airways of patients with non-CF bronchiectasis is important for doctors to make right decision.

DATA SOURCESWe performed this systematic review on the English language literatures from 1966 to July 2014, using various search terms included "pathogens" or "bacteria" or "microbiology" and "bronchiectasis" or "non-cystic fibrosis bronchiectasis" or "non-CF bronchiectasis" or "NCFB."

STUDY SELECTIONWe included studies of patients with the confirmed non-CF bronchiectasis for which culture methods were required to sputum or bronchoalveolar lavage fluid (BALF). Weighted mean isolation rates for Haemophilus influenzae, Pseudomonas aeruginosa, Streptococcus pneumoniae, Stapylococcus aureus, Moxarella catarrhails were compared according to different methodology.

RESULTSThe total mean bacterial culture positive rates were 63%. For studies using sputum samples, the mean positive culture rates were 74%. For studies using BALF alone or BALF and sputum, it was 48%. The distributions of main bacterial strains were 29% for H. influenzae, 28% for P. aeruginosa, 11% for S. pneumoniae, 12% for S. aureus, and 8% for M. catarrhails with methodology of sputum. Meanwhile, the bacterial distributions were 37% for H. influenzae, 8% for P. aeruginosa, 14% for S. pneumoniae, 5% for S. aureus, and 10% for M. catarrhails with methodology of BALF alone or BALF and sputum. Analysis of the effect of different methodology on the isolation rates revealed some statistically significant differences.

CONCLUSIONSH. influenzae accounted for the highest percentage in different methodology. Our results suggested that the total positive culture rates and the proportion of P. aeruginosa from sputum and BALF specimens had significant differences, which can be used in further appropriate recommendations for the treatment of non-CF bronchiectasis.

Bronchiectasis ; microbiology ; Bronchoalveolar Lavage Fluid ; microbiology ; Haemophilus influenzae ; pathogenicity ; Humans ; Pseudomonas aeruginosa ; pathogenicity ; Sputum ; microbiology

OBJECTIVETo analyze the clinical characteristics of protracted bacterial bronchitis (PBB) in children.

METHODThe clinical data of patients seen from October, 2010 to March, 2014 in Department of Respiratory Diseases of our hospital were retrospectively analyzed. Inclusion criteria were over 4 weeks cough, receiving fiberoptic bronchoscopy, positive bacterial culture and (or) the increased percentage of neutral granulocytes in bronchoalveolar lavage fluid (BALF).

RESULTTwenty eight patients were involved, 26 were male (93%) and two were female (7%). The median age of patients was 8.5 months. The median duration of cough was four weeks. The average length of hospital stay was (8.3 ± 3.9)days. The main clinical feature was wet cough in 28 cases, wet cough with wheezing was seen in 21 cases. The wet cough phase distribution was irregular in 21 cases. The crackles with wheeze (in 21 cases) was main signs of PBB. The percentage of CD3⁻ CD16⁺ 56⁺ cells increased in peripheral blood. The fiberoptic bronchoscopic manifestations of PBB were luminal mucosal edema. Eleven patients also had airway malacia. The neutrophil median in BALF was 0.2. The positive rate of bacterial culture of BALF was 36%. The main bacteria were Streptococcus pneumoniae (50%) and Haemophilus influenzae (30%). The main treatment for PBB patients included amoxycillin/clavulanate potassium and second-generation cephalosporins. The average duration of treatment was (17.3 ± 3.2)days, the prognosis was good.

CONCLUSIONPBB is common in male infants. Persistent wet cough with wheezing was the main characteristic of PBB. PBB is commonly accompanied by immune dysfunction and airway malacia, and the pathogens were Streptococcus pneumoniae and Haemophilus influenzae.

Bacterial Infections ; drug therapy ; pathology ; Bronchitis ; drug therapy ; microbiology ; pathology ; Bronchoalveolar Lavage Fluid ; Bronchoscopy ; Cough ; Female ; Haemophilus influenzae ; isolation & purification ; Humans ; Infant ; Male ; Respiratory Sounds ; Retrospective Studies ; Streptococcus pneumoniae ; isolation & purification

OBJECTIVETo compare the detection rates of Mycoplasma pneumoniae (MP) from nasopharyngeal aspirates (NPA) and bronchoalveolar lavage fluid (BALF) in children with pneumonia.

METHODSA total of 164 hospitalized children with pneumonia were enrolled. NPA and BALF of these children were collected within 24 hours of admission, and MP-DNA was detected by fluorescence quantitative PCR. Venous blood samples of all these children were collected within 24 hours of admission and on days 7-10 of treatment, and serum MP-IgM was detected using ELISA.

RESULTSThe positive rate of MP-DNA in NAP of the 164 cases was 51.8% , which was lower than 63.4% as the detection rate of MP-IgM in serum (P=0.044), and the two detection rates were moderately consistent with each other (Kappa=0.618, P<0.01). The positive rate of MP in BALF was 71.3%, which was not significantly different with that of MP-IgM in serum (P>0.05), and the detection rates were well consistent (Kappa=0.793, P<0.01). The detection rate of MP in NPA was lower than that in BALF (P<0.01), with moderate consistency between two of them (Kappa=0.529, P<0.01). The median MP copy number in BALF was significantly higher than that in NPA (P<0.01). The MP detection rates in NPA and BALF were significantly different among different courses of disease (P<0.05). As the course of disease extended, the MP detection rates in both NPA and BALF showed a declining trend; children with MP pneumonia of 1-2 weeks' duration and 2-4 weeks' duration had a higher MP-DNA detection rate in BALF than in NPA (P<0.05).

CONCLUSIONSMP-DNA in BALF has a high sensitivity, with a great significance for early diagnosis of MP pneumonia, while NPA MP-DNA tests may lead to a missed diagnosis.

Adolescent ; Bronchoalveolar Lavage Fluid ; microbiology ; Child ; Child, Preschool ; DNA, Bacterial ; analysis ; Female ; Humans ; Infant ; Male ; Nasopharynx ; microbiology ; Pneumonia, Mycoplasma ; diagnosis

OBJECTIVETo analyze the clinical characteristics of Mycoplasma pneumoniae-associated hemophagocytic syndrome (MP-HLH).

METHODA retrospective investigation of the clinical manifestation, laboratory test, imagelogy, clinical course and outcome of 3 cases with MP-HLH seen between June 2013 and July 2013 in Shenzhen Children's Hospital, and review of relevant literature were conducted.

RESULTOf the 3 cases of MP-HLH, 2 were males, one was female, the ages were 1 year, 3 years and 6 years, respectively. They had no underlying disease previously. All the 3 cases had onset of fever, cough as main symptoms. Diagnosis of refractory Mycoplasma pneumoniae pneumonia was made, which was accompanied by decreased neutrophils [(0.08-0.68)×10(9)/L], hemoglobin [(79-103) g/L], platelet [(64-157)×10(9)/L], plasma fibrinogen [(1.3-1.5) g/L], lactate dehydrogenase [(1,170-1,285) U/L] and increased serum ferritin [(936.7-39 789.0) µg/L] in the third week of course. In two cases the T lymphocytes decreased, and the NK cell activity decreased significantly in one. Bone marrow cytology showed prompted bone marrow hyperplasia, and the phenomenon of phagocytosed blood cells. CT scan was performed for all the cases and consolidation with pleural effusion were shown. Two cases were admitted to PICU, and required endotracheal intubation and mechanical ventilation. Flexible bronchoscopy and bronchial lavage were performed and bronchial cast was found in two cases. All of them were treated with macrolide combined with other antibiotics, glucocorticoids and gamma globulin combination therapy, including one case given dexamethasone [10 mg/(m2·d)], cyclosporine[6 mg/(kg·d)], etoposide [150 mg/(m2·d)] chemotherapy. Two cases were cured, and 1 case died. The authors summarized the 18 cases reported in domestic and foreign literature. Foreign children were diagnosed and treated with steroids in 1-2 weeks, and 10 cases were cured, and 2 cases died. They died of massive hemorrhage and meningoencephalitis, and domestic children were diagnosed and treated within two to 4 weeks after onset, 5 cases were cured, one case died of severe pneumonia.

CONCLUSIONMP-HLH is a rare disease in children, and had acute onset, rapid progression and high mortality. Early treatment with steroids was associated with a good prognosis, the key to successful treatment is early diagnosis and treatment, avoiding the immune cascade. Too late a diagnosis or development of serious complications may lead to death.

Anti-Bacterial Agents ; therapeutic use ; Bronchoalveolar Lavage Fluid ; Bronchoscopy ; Child ; Child, Preschool ; Fatal Outcome ; Female ; Fever ; Glucocorticoids ; therapeutic use ; Humans ; Infant ; Lymphohistiocytosis, Hemophagocytic ; diagnosis ; drug therapy ; microbiology ; Macrolides ; therapeutic use ; Male ; Mycoplasma pneumoniae ; isolation & purification ; Pleural Effusion ; Pneumonia, Mycoplasma ; complications ; diagnosis ; drug therapy ; Respiration, Artificial ; Retrospective Studies ; Tomography, X-Ray Computed ; Treatment Outcome