After infection with the varicella-zoster virus (VZV), the virus becomes latent in the sensory ganglia. Immune senescence may lead to its reactivation, resulting in herpes zoster (HZ). The limited immunogenicity of current vaccines in elderly populations remains a significant challenge for prevention and control. This study investigated the immune-enhancing effects of the novel compound adjuvant BC02 on a recombinant VZV glycoprotein E (gE) subunit vaccine in a serum-positive elderly mouse model. A seropositive state was simulated through pre-immunization with the Oka strain of VZV, and the impacts of vaccines with various adjuvant formulations on humoral and cellular immunity in aged mice were systematically compared. Results demonstrated that the number of gE-specific IFN-γ- and IL-2-secreting cells induced by the BC02-adjuvanted vaccine (gE+BC02-1) increased 11.8- and 5.7-fold compared to the single-adjuvant group, significantly enhancing the multifunctionality of CD4+ T cells. The neutralizing antibody titer reached 1:122, comparable to that of the commercial vaccine Shingrix®, while the ratio of memory T/B cells was markedly higher than in the control group. Cross-age group experiments revealed that BC02 could overcome the limitations imposed by immune senescence in elderly models, inducing a balanced Th1/Th2 response and long-term immune memory comparable to that observed in younger groups (antibody titers maintained for ≥8 months). This study confirmed that the BC02 adjuvant synergistically activates innate and adaptive immunity, significantly enhancing the immune efficacy of the gE vaccine in serum-positive elderly individuals, thereby providing an potential strategy for optimizing herpes zoster vaccines for the elderly population.

Objective There are limited data on the use of the creation tuberculin skin test (C-TST) for detecting tuberculosis (TB) infection (TBI) in individuals under 18 years of age. We conducted a study to assess the diagnostic accuracy of C-TST in this population. Methods A double-blind, randomized controlled trial was conducted across 4 tertiary hospitals in China to evaluate the diagnostic accuracy of the C-TST in detecting TBI in individuals under 18 years of age. Participants with suspected pulmonary TB, extrapulmonary TB, or non-TB pulmonary disease were enrolled. The primary outcome was the diagnostic accuracy of the C-TST. Secondary outcomes included the consistency among C-TST, the traditional tuberculin skin test (TST), and T-SPOT.TB assays in different subgroups, as well as the safety of C-TST. Each participant underwent all 3 tests simultaneously: T-SPOT.TB assay, TST, and C-TST. Results C-TST showed a sensitivity of 83.0 % (95 % CI, 68.7 %–91.9 %), while TST and T-SPOT.TB demonstrated sensitivities of 80.9 % (95 % CI, 66.3 %–90.4 %) and 76.6 % (95 % CI, 61.6 %–87.2 %), respectively. The specificities of C-TST, TST, and T-SPOT.TB were 100 % (95 % CI, 91.9 %–100 %), 98.0 % (95 % CI, 87.8 %–99.9 %), and 100 % (95 % CI, 90.9 %–100 %), respectively. The consistency between C-TST and T-SPOT.TB was high (kappa = 0.847). No serious adverse events (AEs) were reported. Conclusions This study demonstrates that C-TST is a reliable and safe diagnostic tool for detecting TBI in children and adolescents. It shows higher sensitivity than both T-SPOT.TB and the traditional TST, with no associated serious AEs. Therefore, C-TST is an effective and safe option for diagnosing TBI in this age group.

This study aimed to evaluate the diagnostic performance of ESAT6-CFP10 (EC) skin test in healthy population and determine the factors influencing the booster effect. We conducted a randomized, double-blind, parallel controlled trial in healthy population. The experiment was divided into two stages. In the first stage, all participants underwent T-SPOT, TB-PPD, and EC tests. In the second stage, to evaluate whether BCG vaccination affected the efficacy of skin tests, the participants with three negative results in the first stage were randomly assigned to the BCG and placebo groups at a ratio of 2:1 and underwent three tests. The positivity rates and concordance of the three tests were calculated in both stages, and a logistic regression model was constructed to determine the factors influencing the booster effect of EC in the second stage. Safety observations were continued until the skin test results were available. In the first stage, 1,564 participants were enrolled in the study. The positivity rates of the T-SPOT, EC, and TB-PPD tests in all the participants were 18.89

Introduction:Newer skin tests, including the ESAT6-CFP10 (EC) skin test, were recommended for diagnosing Mycobacterium tuberculosis (M. tb) infection. However, no data exist assessing the diagnostic performance of the EC skin test among foreign students with different skin tones. Methods:A cohort study at Nanjing Medical University screened incoming foreign freshmen. The EC skin test was used to assess for M. tb infection, and results were read at 24, 48, 72, and 96-hours post-administration. Results:Among 96 participants, M. tb infection rates at 24, 48, 72, and 96-hours post-injection were 3.13%, 7.29%, 13.54%, and 9.38%, respectively. While infection rates were lower among individuals with darker skin tones, the difference was not statistically significant (P=0.186), and variations were consistent across different measurement times. Trajectory analysis revealed 5.3% in the continuous-increasing group, 86.5% in the low-stable group, and 5.2% in the elevated-decreasing group. Notably, participants in the elevated-decreasing group had lighter skin tones, with trajectory patterns consistent across different skin colors. Discussion:The EC skin test is safe, and redness diameter is a more reliable indicator than induration. Results should be collected within 48 to 72 hours, with verification at 72 hours crucial if initial results are negative. Importantly, skin color does not affect EC skin test outcomes.

Novel adjuvants and innovative combinations of adjuvants (Adjuvant Systems) have facilitated the development of enhanced and new vaccines against re-emerging and challenging pathogenic microorganisms. Nonetheless, the efficacy of adjuvants is influenced by various factors, and the same adjuvant may generate entirely different immune responses when paired with different antigens. Herein, we combined the MPXV-B6R antigen with BC02, a novel adjuvant with proprietary technology, to assess its capability to induce both cellular and humoral immunity in mouse models. Mice received two intramuscular injections of B6R-BC02, which resulted in the production of MPXV-specific IgG, IgG1, and IgG2a antibodies. Additionally, it elicited strong MPXV-specific Th1-oriented cellular immunity and persistent effector memory B-cell responses. The advantages of BC02 were further validated, including rapid initiation of the immune response, robust recall memory, and sustained immune response induction. Although the potential of immunized mice to produce serum-neutralizing antibodies against the vaccinia virus requires further improvement, the exceptional performance of BC02 as an adjuvant for the MPXV-B6R antigen has been consistently demonstrated.

In view of the new tuberculosis infection test recommended by WHO in 2022, the MTB antigen-based skin tests（TBST） for the diagnosis of tuberculosis infection, the authors sorted out the name of new varieties of the same kind that have been listed and are under research, discussed the nomenclature of such products with reference to the principles related to nomenclature of biological products in China, expected to provide discussion and reference for experts from research and development, use and regulatory departments.

To achieve maximum efficacy, vaccines, such as subunit, recombinant, and conjugate vaccines, necessitate the incorporation of immunostimulators/adjuvants. Adjuvants play a vital role in bolstering and extending the strength of the immune response while also influencing its type. As antigen and adjuvant formulations become more intricate, it becomes imperative to establish a well-characterized and robust formulation to ensure consistent and reproducible outcomes in preclinical and clinical studies. In the present study, an HPV bivalent vaccine was developed using a BC02 adjuvant in conjunction with HPV 16 and 18 L1 VLP antigens produced from an E. coli expression system. The study involved evaluating the adjuvant formulation and in vivo immunogenicity in mice. Remarkably, a medium-dose of BCG-CpG-DNA combined with a low-dose of aluminum hydroxide substantially enhanced the immunogenicity of HPV16 and 18 VLPs, resulting in improved cellular and humoral immune responses.

Objectives: To verify the diagnostic utility of recombinant fusion protein ESAT6-CPF10 (EC), a novel skin test reagent to detect Mycobacterium tuberculosis infection. Methods: A multi-centered, double-blind, randomized controlled trial was conducted from December 17, 2015, to March 2, 2018. Participants involved in this study included those with active tuberculosis (TB), suspected pulmonary TB, or non-TB pulmonary disease. Each participant received three tests simultane-ously, TB-specific enzyme-linked immunospot assay (T-SPOT.TB), tuberculin skin test (TST), and EC skin test (ECST), and adverse events were reported. Results: Diagnostic accuracy was analyzed using data from 1085 protocol-compliant participants. The sen-sitivities of the ECST, TST, and T-SPOT.TB were 91.2% (95% CI, 89.0-93.2%), 91.4% (95% CI, 89.1-93.3%), and 92.1% (95% CI, 89.9-93.9%), respectively. The specificities of the ECST (69.7%, 95% CI, 64.5-74.5%) and T-SPOT.TB (76.1%, 95% CI, 71.2-80.5%) were significantly higher than the TST (54.4%, 95% CI, 48.9-59.7%). The agreements between ECST and TST (kappa = 0.632) and between ECST and T-SPOT.TB (kappa = 0.780) were substantial. No severe adverse event was reported. Conclusion: The diagnostic performance of the ECST was close to the T-SPOT.TB assay in the detection of TB infection and indicated good potential for clinical application in common scenarios. & COPY; 2022 The Author(s). Published by Elsevier Ltd on behalf of International Society for Infectious Diseases. This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/ )

目的 初步评价卡介苗(Bacillus Calmette-Guerin vaccine,BCG)新菌种单细胞克隆株NIFDC 945 SⅢ的免疫效应及其安全性.方法 将BCG NIFDC 945 SⅢ株、NIFDC 944株、D2 PB302株和Danish 1331株按同一条件制备成菌悬液,经小鼠尾静脉注射,于免疫后24 h及1、3、6、9、12周称量体重,无菌取小鼠脾脏,称脾重,计算脾激活指数;相同时间点进行小鼠脾组织BCG活菌计数,计算脾内BCG存活率.结果 免疫后12周内,BCG NIFDC 945 SⅢ株的脾激活指数总体上低于Danish 1331株,与NIFDC 944株和D2 PB302株相当;BCG NIFDC 945 SⅢ株脾内的BCG存活率总体上低于Danish 1331株和NIFDC 944株,高于D2 PB302株.结论 BCG新菌种单细胞克隆株NIFDC 945 SⅢ具有良好的免疫效应,且安全性较高,有望成为加强型BCG候选株.

Herpes zoster (HZ) is one of two distinct syndromes caused by Varicella-zoster virus (VZV). A primary infection with VZV causes varicella in susceptible young children. After resolution of the primary infection, VZV establishes a lifelong latency within the cranial or dorsal root ganglia. With increasing age, family history of shingles, immunosuppression or other risk factors, there is a decline in the virus-specific T-cell-mediated immune (CMI) response which allows reactivation of latent VZV in the root ganglia resulting in HZ. There are currently two vaccines that have been approved to prevent HZ and postherpetic neuralgia (PHN) but one is a live attenuated vaccine, the protective effect of which is considered to decrease significantly with the age of the recipient. However, a recombinant subunit vaccine may provide targeted VZV-specific cellular and humoral immunity, giving it a more potent and longer-lasting protective effect against HZ. The current study reports the development of a novel adjuvant, BC02 (BCG CpG DNA compound adjuvants system 02), composed of Al(OH)3 inorganic salt adjuvant and BC01 (BCG CpG DNA compound adjuvants system 01), a Toll-like receptor 9 (TLR9) agonist. Immunogenicity and compatibility with recombinant VZV glycoprotein E (gE) in mice were studied. The BC02-adjuvanted gE experimental vaccine was highly effective in eliciting both humoral and cellular immune responses to the recombinant gE glycoprotein and VZV-Oka in a mouse model. The efficient production and long-term persistence of gE and VZV-Oka-specific IFN-γ, IL-2-specific T cells and memory B cells in the early (1W), middle (7W), middle-late (15W), and final (27W) immune stages were established. Results of fluorescent antibody to membrane antigen (FAMA) and serum antibody plaque reduction tests also showed that the BC02 adjuvanted-gE experimental vaccine induced mice to secrete neutralizing antibodies against clinically isolated VZV strains. In combination, the current data suggest that the BC02 compound adjuvant offers a strategy to induce an appropriately strong cellular and humoral immunity against the VZV gE protein subunit to improve vaccine efficacy.

目的 初步探讨新型生物佐剂BC01(BCG CpG DNA compound adjuvant system 01)对机体的免疫激活作用.方法 将C57BL/6小鼠随机分为4组并分别肌肉注射75 μg新型生物佐剂BC01,实验组于免疫后6、12和24 h解剖取脾脏,对照组于注射BC01后0h解剖取脾脏.提取各组小鼠总RNA并进行全基因转录组分析.基因芯片数据经归一化处理后进行基因集富集分析(gene set enrichment analysis,GSEA).同时,采用实时定量PCR法对GSEA中 GO(gene ontology,GO)基因集分组归类中生物过程(biological process,BP)、细胞成分(cellular component,CC)和分子功能(molecular function,MF)相关上调基因进行验证分析.结果 小鼠全基因转录组芯片数据经GSEA分析后发现,BC01佐剂具有活化免疫细胞,激活固有免疫和适应性免疫应答,调节炎症反应、活化免疫细胞迁移与趋化,激活免疫细胞吞噬功能以及促分裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)和细胞外调节蛋白激酶1/2(extracellular-regulated kinase 1/2,ERK1/2)信号级联反应作用.结论 新型生物佐剂BC01经肌肉注射24 h后具有较强的机体免疫激活作用.

The BCG vaccine is prepared from a weakened strain of Mycobacterium bovis (M. bovis), a bacterium closely related to Mycobacterium tuberculosis (MTB), which causes tuberculosis (TB). The vaccine was developed over 13 years, from 1908 to 1921, in the French Institut Pasteur by Léon Charles Albert Calmette and Jean-Marie Camille Guérin, who named the product Bacillus Calmette–Guérin (BCG). BCG, the only licensed vaccine currently available to prevent TB, is given to infants at high risk of TB shortly after birth to protect infants and young children from pulmonary, meningeal, and disseminated TB. The BCG vaccine, one of the safest and most widely used live attenuated vaccines in the world, recently celebrated its 100th anniversary (from 1921 to 2021); its record of use in preventing TB in China is also approaching 100 years. In 2022, a new century of BCG vaccine immunization will begin. In this article, we briefly review the history of BCG vaccine use in China, describe its current status, and offer a preliminary outlook on the future of the vaccine, to provide BCG researchers with a clearer understanding of its use in China.

目的 探讨复合佐剂BC02(BCG CpG DNA combination adjuvant system 02)组成成分对机体固有免疫应答的协同增强作用.方法 将C57BL/6小鼠随机分为4组,分别肌肉注射Al(OH)3佐剂、BC01佐剂和BC02复合佐剂及PBS溶液(对照).免疫后6h,将各组部分小鼠解剖取腹股沟淋巴结进行全基因组转录谱分析.免疫后24、48、72和96 h,将各组剩余小鼠解剖取右后肢内侧注射部位肌肉组织,经组织HE和免疫荧光染色,分析注射部位炎性细胞招募情况.同时,采用qPCR法对信号通路中富集的显著性差异表达基因(significantly differentially expressed genes,SDEG)进行验证分析.结果 SDEG数据经IPA(Ingenuity Pathway Analysis)软件进行GO富集和信号通路网络分析后发现,BC02复合佐剂组成成分具有协同增强免疫细胞活化网络激活、免疫细胞招募信号网络激活、巨噬细胞吞噬功能、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)信号通路网络激活和CCL2相互作用网络激活作用.BC02复合佐剂组成成分也具有协同增强佐剂在注射部位炎性细胞招募的能力.结论 BC02复合佐剂组成成分Al(OH)3无机盐佐剂及BC01生物佐剂具有协同增强机体固有免疫应答的作用.

Both adaptive and innate immunity responses are necessary for the efficient elimination of different pathogens. However, the magnitude, quality and desired type of immune response specific to the co-administered antigen is largely determined by adjuvants. BC02 (BCG CpG DNA compound adjuvants system 02) is a novel compound adjuvant with independent intellectual properties, which is composed of BCG CpG DNA biological adjuvant with Al(OH)3 inorganic salt adjuvant acting as a delivery system. Its safety and strong adjuvant efficacy have been effectively verified in preclinical and clinical trials (Phase Ib, ClinicalTrials.gov Identifier: NCT04239313 and Phase II, ClinicalTrials.gov Identifier: NCT05284812). In this study, we report the level of cell-mediated immunity (CMI) and humoral immune response induced by the BC02 novel adjuvant combined with different doses of varicella-zoster virus (VZV) glycoprotein E (gE) in a mouse model. In addition, we conducted preliminary in vitro experiments to explore the enhancement of RAW264.7 cell immune activity by BC02 adjuvanted-gE experimental vaccine to activate innate immune response. The results showed that the BC02-adjuvanted low, medium or high dose of gE were highly effective in eliciting both CMI and humoral immune responses to the immunized mice, respectively. The production of gE-specific IFN-γ and IL-2-specific T cells was established within 28 days after booster immunization. In particular, the effect of BC02-adjuvanted medium dose of gE has been shown to be more prominent. Meanwhile, fluorescent antibody to membrane antigen (FAMA) and serum antibody plaque reduction tests have also shown that the BC02 adjuvanted-medium dose of gE antigen could induce the secretion of neutralizing antibodies against clinically isolated VZV strains in mice. In addition, our findings have shown that 1/25 dose of gE+BC02 medium dose experimental vaccine can significantly induce the secretion of innate immune cytokines TNF-A, MCP-1, IL-6 and GM-CSF and up-regulate the costimulatory molecules CD40, CD80 and I-A/I-E on RAW264.7 cells; and it has also been activated to form M2 macrophages. At the same time, RAW264.7 cells were stimulated for 12 h, and their phagocytosis was significantly enhanced. Taken together, these results suggest that the BC02 compound adjuvant offers a strategy to induce an appropriate innate and adaptive immunity against the different doses of the VZV gE protein to improve subunit vaccine efficacy, and BC02 may be a promising adjuvant candidate for subunit HZ vaccines.

Abstract Background Diagnostics to identify tuberculosis infection are limited. We aimed to assess the diagnostic accuracy and safety of ESAT6-CFP10 (EC) skin test for tuberculosis infection in Chinese adults. Methods We conducted 2 randomized, parallel-group clinical trials in healthy participants and tuberculosis patients. All participants were tested with the T-SPOT.TB test, then received an EC skin test and tuberculin skin test (TST). The diameter of skin indurations and/or redness at injection sites were measured at different time periods. A bacillus Calmette Guerin (BCG) model was established to assess the diagnosis of tuberculosis infection using an EC skin test. Results In total, 777 healthy participants and 96 tuberculosis patients were allocated to receive EC skin test at 1.0 μg/0.1 mL or 0.5 μg/0.1 mL. The area under the curve was 0.95 (95% confidence interval [CI], .91–.97) for the EC skin test at 1.0 μg/0.1 mL at 24–72 hours. Compared with the T-SPOT.TB test, the EC skin test demonstrated similar sensitivity (87.5, 95% CI, 77.8–97.2 vs 86.5, 95% CI, 79.5–93.4) and specificity (98.9, 95% CI, 96.0–99.9 vs 96.1, 95% CI, 93.5–97.8). Among BCG vaccinated participants, the EC skin test had high consistency with the T-SPOT.TB test (96.3, 95% CI, 92.0–100.0). No serious adverse events related to the EC skin test were observed. Conclusions The EC skin test demonstrated both high specificity and sensitivity at a dose of 1.0 μg/0.1 mL, comparable to the T-SPOT.TB test. The diagnostic accuracy of the EC skin test was not impacted by BCG vaccination. Clinical Trials Registration NCT02389322 and NCT02336542.

Background: Tuberculosis (TB) is a major global health problem in the whole world, especially in the developing countries. We aimed to assess diagnostic accuracy and safety of a novel ESAT6-CFP10 (EC) skin test for TB infection in Chinese adults.Methods: We conducted randomized, double-blinded, parallel-group clinical trials at two sites in healthy adults and TB/non-TB patients, respectively. TB infection-free healthy participants in a ratio of 1:1:1:1 to receive BCG vaccine or placebo, followed by EC skin test (0·5μg/0·1ml), or EC (1μg/0·1ml), along with the TST, T-SPOT.TB test, 12 weeks after the vaccination. TB patients or non-TB patients with pulmonary disease were randomly allocated in a ratio of 1:1 to receive EC skin test at 0·5μg/0·1ml, or 1μg/0·1ml, along with the TST and T-SPOT.TB test. The diameter of the skin indurations and/or redness at the injection sites were measured at 24 hours, 48 hours, and 72 hours after the skin tests. The studies was registered with ClinicalTrials.gov, number NCT02389322 and NCT02336542.Findings: We enrolled 223 TB infection-free healthy participants after screening, and randomly allocated them to receive BCG vaccination or placebo vaccination. Of them, 83 and 86 complete the EC skin test at 0·5μg/0·1ml or 1μg/0·1ml after vaccination. 96 TB patients and 95 non-TB patients with pulmonary diseases were allocated to receive the EC skin test at 0·5μg/0·1ml or 1μg/0·1ml, respectively. EC skin test at 1μg/0·1ml was found to be superior to 0·5μg/0·1ml based on the estimated AUCs, and the maximum AUCs of 0·927 was obtained at 1μg/0·1ml EC skin test at 48 hours using diameters of redness or induration as indicator. The highest diagnostic values achieved with cutoff values between 3·7~6·3 mm, and 5mm was determined as positive cutoff value for the responses of EC skin test. The incidence of adverse reaction following the EC skin tests were 27·8%, and no serious adverse event related to the EC skin tests was observed.Interpretation: The experimental EC skin test has an acceptable safety profile. EC skin test at 1μg/0·1ml can achieve both high specificity and sensitivity, and was selected as optimal dose for future studies.Funding: National Science and Technology Major Project (2012ZX10004710) Zhifei Longcom Biologic Pharmacy Co., China.Declaration of Interest: We declare no competing interests.Ethical Approval: We obtained approvals from the institutional review boards of the Jiangsu Provincial Center of Disease Control and Prevention, and Shanghai Public Health Clinical Center before initiation of the studies. Written informed consent were obtained from all participants. The trials were conducted in accordance with the Declaration of Helsinki and Good Clinical Practice.

目的 以冻干重组结核杆菌融合蛋白(EC)[该制品名称是国家药典委员会确定的药品中文通用名称,“EC”为重组融合蛋白“结核分枝杆菌早期分泌性抗原靶6(ESAT-6)和培养滤液蛋白10(CFP-10)”](以下简称“EC”)原液为原材料,研制EC变态反应原原液效力评价用国家参考品.方法 以电泳法、高效液相色谱法分别测定备选EC原液纯度,纯度合格后经精密称量、分装、冻干制成候选国家参考品,再测定其蛋白质和水分含量.将候选国家参考品稀释成不同稀释度后,皮内注射结核分枝杆菌活菌致敏的豚鼠,根据皮肤试验硬结或红晕反应平均直径大小(简称“反应平均直径大小”)结果探索原液效价测定的适宜稀释度.在此基础上,将候选国家参考品用于EC原液的效价测定并进行方法验证,初步评价其适用性能.研究还初步观察了候选品在-20℃放置24个月的稳定性.结果 EC原液电泳法纯度为100.00％,高效液相色谱法纯度为95.33％,候选国家参考品蛋白质含量为508 μg/瓶,水分含量为1.57％,分装精度控制在±1％以内.稀释度探索结果显示:候选品稀释度为2.5 μg/ml(12.5U/ml)、5μμg/ml(25 U/ml)和10 μg/ml(50 U/ml)时剂量对数反应平均直径大小曲线有良好的线性关系(R2=0.9944),可作为后续研究的适宜稀释度.将候选国家参考品用于EC原液效价测定,两者剂量对数反应平均直径大小曲线趋势基本一致(R2=0.9878,R2=0.9643),且每个稀释度EC原液与相应稀释度候选国家参考品反应平均直径大小的比值均满足1.0±0.2.方法 验证结果显示:候选国家参考品和EC原液的剂量对数反应平均直径大小曲线趋势基本一致(R2=0.9999,R2=0.9815),且两者相应稀释度反应平均直径大小的比值均满足1.0±0.2,方法可行.初步稳定性研究结果显示:-20℃放置24个月的候选国家参考品效价测定性能稳定.结论 候选国家参考品的纯度、蛋白质含量、水分含量及均匀性检测均符合国家参考品质量要求,将其稀释到适宜稀释度后可用于EC原液的效价测定;经方法验证与稳定性观察,显示其性能良好,为国家参考品申报提供了基础.

The potency of synthetic CpG-oligo-deoxynucleotides (CpG-ODNs) adjuvants in modulating the immune cell functions through the TLR pathway has been tested and reported previously. However, the cellular signaling involved in the stimulation of macrophages by natural, CpG motif-containing adjuvant and the effector functions modulated by such stimulation has not been well studied. Here, we used in vitro and ex vivo murine macrophage assay systems, and mouse model of in vivo stimulation to explore the signaling pathway and the effector functions mediated by BC01. Results show that BC01 can induce the production of TNF-α and MCP-1 in macrophages by up-regulating the activation of NF-κB and MAPKs signaling pathway, and elevated the expression of MHC-II, CD40, CD80, and CD86. Upon stimulation with BC01, the peritoneal macrophages isolated from TLR9 −/− mice produced significantly low levels of pro-inflammatory cytokines, attenuated the activation of NF-κB and MAPKs signaling pathways, and showed reduced phagocytosis. Following in vivo stimulation with BC01, the TLR9 −/− mice produced significantly lower levels of pro-inflammatory cytokines in the serum and lymph nodes showed reduced cell proliferation. These results indicate that BC01 is an efficient agonist of TLR9 that can significantly enhance the host-protective immune functions of macrophages. Keywords Adjuvant , innate immunity , macrophage , MAPKs pathway , NF-κB pathway , TLR9

目的 分析国内外不同卡介菌亚株的抗生素敏感特性.方法 对国内外不同卡介菌亚株采用比例法进行一线抗结核药物[异烟肼(isoniazid,INH)、利福平(rifampicin,RFP)、链霉素(streptomycin,SM)及乙胺丁醇(ethambutol,EMB)]、二线抗结核药物[左旋氧氟沙星(levofloxacin,LFX)、丁胺卡那霉素(amikacin,AK)、卷曲霉素(capreomycin,CPM)、乙硫异烟胺(ethionamide,TH)及对氨基水杨酸(p-aminosalicylic acid,PAS)]敏感性分析,采用BACTECMGIT960培养系统进行一线抗结核药物及吡嗪酰胺(pyrazinamide,PZA)抗生素敏感性分析.结果 经比例法分析,不同卡介菌亚株均对一线抗结核药物敏感,除对TH耐药外,对其他二线抗结核药物均敏感;经BACTEC MGIT960系统分析,不同卡介菌亚株对RFP、SM、EMB均敏感,对PZA均耐药;卡介菌亚株BCG Pasteur 1173及BCG Tokyo 172对INH敏感,而BCG Danish 1331菌株及中国卡介菌菌种BCG D2 PB302、BCG NIFDC 945 SⅢ均对INH耐药.结论 中国生产用卡介菌菌种BCG D2 PB302及BCG NIFDC 945 SⅢ在抗生素敏感性方面与BCG Danish1331菌株一致,与其他亚株有差异.