树突状细胞在结核病免疫应答中的作用及机制

作为最有效的专职抗原提呈细胞（DC）,树突状细胞在结核病免疫中的作用日益受到关注。机体感染结核杆菌后,未成熟DC（iDC）捕获抗原并逐渐发育成熟,携带抗原成份从感染组织迁移至外周免疫器官,将抗原成份提呈给T、B淋巴细胞激发免疫应答,起着连接固有免疫和适应性免疫作用,并通过分泌IL-12、IFN-γ等细胞因子参与机体免疫调节。深入研究DC在结核病免疫中的作用机制将为抗结核新型疫苗的开发及免疫治疗方案的设计提供依据。     

A user-friendly, highly sensitive assay to detect the IFN-gamma secretion by T cells

OBJECTIVES: Development of a user-friendly test alternative to ELISA-based assays to detect IFN-gamma by in vitro cultured peripheral blood mononuclear cells (PBMC) stimulated with pathogen-derived antigens. DESIGN AND METHODS: The molecular components of an operational IFN-gamma ELISA-based test were applied in a lateral flow (LF) immuno-sandwich assay using up-converting phosphor (UCP) reporter particles. The analytical sensitivity of the UCP-LF IFN-gamma assay (ULIGA) was determined and the assay was qualitatively validated with a selection of 60 supernatants derived from PBMC cultures stimulated with M. leprae derived antigens, mitogen or medium alone. RESULTS: ULIGA indicated an analytical sensitivity better than 2 pg/mL, and demonstrated four orders of magnitude dynamic range. The assay correlated well with the IFN-gamma ELISA. CONCLUSIONS: ULIGA allows detection well below the cutoff value (100 pg/mL) used to define positive responses in the IFN-gamma ELISA. The test procedure is less demanding in respect to equipment and labor, and is suited for testing single samples.     

Catheter-related bloodstream infections caused by rapidly growing nontuberculous mycobacteria: a case series including rare species

Rapidly growing nontuberculous mycobacteria (RGMs) are responsible for a variety of clinical syndromes in humans including catheter-related blood stream infections (CRBSIs). Recently, we identified a cluster of RGM-associated CRBSI at our institution. We describe the epidemiologic and clinical patterns associated with these infections. We conducted a retrospective single-center review of RGM CRBSI between May 2004 and June 2005. RGMs isolated from blood cultures of 6 patients included Mycobacterium mucogenicum (2), Mycobacterium fortuitum (2), and the rare RGM species, Mycobacterium neoaurum (1) and Mycobacterium septicum (1). All of the patients had a long-term intravascular catheter (mean duration, 6.5 months). Bacteremia was resolved in all patients after catheter removal and appropriate antibiotics. None of the patients suffered a relapse of RGM CRBSI, and all survived to 1 year. RGMs are causative pathogens in both immunosuppressed and immunocompetent individuals with long-term intravascular catheters and blood stream infections. Recent trends at our center suggest that infections with these pathogens are rising.     

Mycobacterial pulmonary infections in patients with idiopathic pulmonary fibrosis

Patients with idiopathic pulmonary fibrosis (IPF) have an increased risk for developing tuberculosis (TB). However, no studies have been reported regarding the development of nontuberculous mycobacterium (NTM) lung disease (NTMLD). We reviewed 795 patients with IPF from five university hospitals who were diagnosed by histological or radio-clinical criteria. In the 795 patients with IPF, pulmonary infections with mycobacterium tuberculosis (MTB) and NTM were found in 35 (4.4%) and 16 patients (2.0%), respectively, which was a higher frequency than that found in the general population. TB was more common in patients treated with immunosuppressants than in those who did not receive immunosuppressants (2.6% vs 1.4%, P = 0.12). Among the IPF patients who had mycobacterial infections,immunosuppressant users developed TB or NTMLD within 1 yr after treatment with immunosuppressants,while those occurred later than 2 yr after diagnosis of IPF in the subjects that did not receive immunosuppressants. Among 51 IPF patients who had mycobacterial infections, 9 (18%) died during follow-up. Of these, three died due to progression of pulmonary tuberculosis. TB and NTMLD is relatively common in patients with IPF in Korea and may be fatal in some groups. Careful evaluation of TB and NTMLD is necessary not only for immunosuppressant users, but also for nonusers with IPF.     

13种抗生素对快生长分枝杆菌的体外抑菌效果评价

目的 评价快生长分枝杆菌(rapidly growing mycobacteria,RGM)临床分离株对13种抗生素的敏感性特征。方法 收集首都医科大学附属北京胸科医院2015年1月至2019年12月从临床样本中分离出的98株RGM,包括25株偶发分枝杆菌、70株脓肿分枝杆菌和3株龟分枝杆菌。使用Sensititre RAPMYCO药敏板测定13种抗生素(阿米卡星、妥布霉素、阿莫西林-克拉维酸、头孢西丁、亚胺培南、环丙沙星、莫西沙星、克拉霉素、利奈唑胺、米诺环素、多西霉素、替加环素、甲氧苄啶-磺胺甲噁唑)对98株RGM临床分离株的体外抑菌活性。结果 在所测定的13种抗生素中,阿米卡星对所检测的3种RGM临床分离株的体外抑菌活性最佳,对偶发分枝杆菌和脓肿分枝杆菌临床分离株的敏感率分别100.0%(25/25)和90.0%(63/70)。与多西环素、米诺环素相比,替加环素对RGM临床分离株的体外抑菌效果更好,对偶发分枝杆菌和脓肿分枝杆菌临床分离株的敏感率分别为100.0%(25/25)和72.9%(51/70)。莫西沙星和环丙沙星对偶发分枝杆菌临床分离株表现出较强的抑菌活性,敏感率分别为100.0%(25/25)和92.0%(23/25),而对脓肿分枝杆菌临床分离株几乎没有体外抑菌活性,耐药率均为95.7%(67/70)。此外,几乎所有检测的RGM临床分离株均对头孢西丁、多西环素、利奈唑胺、亚胺培南和甲氧嘧啶-磺胺甲噁唑耐药。结论 阿米卡星、替加环素和克拉霉素对脓肿分枝杆菌和偶发分枝杆菌临床分离株具有较好的体外抑菌活性,但在不同菌株间依然存在差异,需要对每个菌种或临床分离菌株进行单独的药物敏感性试验。     

Phenotypic characterization of bovine memory cells responding to mycobacteria in IFNγ enzyme linked immunospot assays

Bovine tuberculosis (bTB) remains a globally significant veterinary health problem. Defining correlates of protection can accelerate the development of novel vaccines against TB. As the cultured IFNγ ELISPOT (cELISPOT) assay has been shown to predict protection and duration of immunity in vaccinated cattle, we sought to characterize the phenotype of the responding T-cells. Using expression of CD45RO and CD62L we purified by cytometric cell sorting four distinct CD4⁺ populations: CD45RO⁺CD62L^hi, CD45RO⁺CD62L^lo, CD45RO⁻CD62L^hi and CD45RO⁻CD62L^lo (although due to low and inconsistent cell recovery, this population was not considered further in this study), in BCG vaccinated and Mycobacterium bovis infected cattle. These populations were then tested in the cELISPOT assay. The main populations contributing to production of IFNγ in the cELISPOT were of the CD45RO⁺CD62L^hi and CD45RO⁺CD62L^lo phenotypes. These cell populations have been described in other species as central and effector memory cells, respectively. Following in vitro culture and flow cytometry we observed plasticity within the bovine CD4⁺ T-cell phenotype. Populations switched phenotype, increasing or decreasing expression of CD45RO and CD62L within 24 h of in vitro stimulation. After 14 days all IFNγ producing CD4⁺ T cells expressed CD45RO regardless of the original phenotype of the sorted population. No differences were detected in behavior of cells derived from BCG-vaccinated animals compared to cells derived from naturally infected animals. In conclusion, although multiple populations of CD4⁺ T memory cells from both BCG vaccinated and M. bovis infected animals contributed to cELISPOT responses, the dominant contributing population consists of central-memory-like T cells (CD45RO⁺CD62L^hi).     

16S～23S rDNA内转录间隔区序列分析及其在分支杆菌鉴定中的应用价值

目的：研究16S－23SrDNA内转录间隔区（internal transcribed spacr,ITS)序列在分支杆菌鉴定中的应用价值。方法：应用PCR－直接测序法对22例30株分支杆菌参考菌株和16株分支杆菌临床分离菌株16S－23SrDNA ITS测序。对所测定序列以及GenBank所报道的有关序列用Clustal程序（MegAlign Package[Windows Version4.01]；DNASTAR，Madson,Wis)处理分析，计算种间相似性，用PHYLIP Package构建分支杆菌菌种聚类分析树状谱。结果：除结核分支杆菌复合群（MTC）菌种（结核分支杆菌、牛分支杆菌、非洲分支杆菌和田鼠分支杆菌）16S－23SrDNA ITS序列完全一致外，其它分支杆菌菌种间核苷酸排列顺序及长度变异较大，种间相似性为30．4％－86．5％，聚类分析树状谱能将各分支杆菌菌种相分离，结果表明16S－23SrDNA ITS序列分析能将MTC与非结核分支杆菌（NTM）相鉴别，能将NTM鉴定至种的水平。结论：16S－23SrDNA ITS可做为分支杆菌鉴定的靶基因。