Mycobacterium smegmatis enhances shikonin-induced immunogenic cell death—an efficient in situ tumor vaccine strategy

Tumor vaccines are a promising avenue in cancer immunotherapy.Despite the progress in targeting specific immune epitopes,tumor cells lacking these epitopes can evade the treatment.Here,we aimed to construct an efficient in situ tumor vaccine called Vac-SM,utilizing shikonin(SKN)to induce immunogenic cell death(ICD)and Mycobacterium smegmatis as an immune adjuvant to enhance in situ tumor vaccine efficacy.SKN showed a dose-dependent and time-dependent cytotoxic effect on the tumor cell line and induced ICD in tumor cells as evidenced by the CCK-8 assay and the detection of the expression of relevant indicators,respectively.Compared with the control group,the in situ Vac-SM injection in mouse subcutaneous metastatic tumors significantly inhibited tumor growth and distant tumor metastasis,while also improving survival rates.Mycobacterium smegmatis effectively induced maturation and activation of bone marrow-derived dendritic cells(DCs),and in vivo tumor-draining lymph nodes showed an increased maturation of DCs and a higher proportion of effector memory T-cell subsets with the Vac-SM treatment,based on flow cytometry analysis results.Collectively,the Vac-SM vaccine effectively induces ICD,improves antigen presentation by DCs,activates a specific systemic antitumor T-cell immune response,exhibits a favorable safety profile,and holds the promise for clinical translation for local tumor immunotherapy.

Inferring Mycobacterium Tuberculosis Drug Resistance and Transmission using Whole-genome Sequencing in a High TB-burden Setting in China

Objective China is among the 30 countries with a high burden of tuberculosis(TB)worldwide,and TB remains a public health concern.Kashgar Prefecture in the southern Xinjiang Autonomous Region is considered as one of the highest TB burden regions in China.However,molecular epidemiological studies of Kashgar are lacking.Methods A population-based retrospective study was conducted using whole-genome sequencing(WGS)to determine the characteristics of drug resistance and the transmission patterns.Results A total of 1,668 isolates collected in 2020 were classified into lineages 2(46.0%),3(27.5%),and 4(26.5%).The drug resistance rates revealed by WGS showed that the top three drugs in terms of the resistance rate were isoniazid(7.4%,124/1,668),streptomycin(6.0%,100/1,668),and rifampicin(3.3%,55/1,668).The rate of rifampicin resistance was 1.8%(23/1,290)in the new cases and 9.4%(32/340)in the previously treated cases.Known resistance mutations were detected more frequently in lineage 2 strains than in lineage 3 or 4 strains,respectively:18.6%vs.8.7 or 9%,P<0.001.The estimated proportion of recent transmissions was 25.9%(432/1,668).Multivariate logistic analyses indicated that sex,age,occupation,lineage,and drug resistance were the risk factors for recent transmission.Despite the low rate of drug resistance,drug-resistant strains had a higher risk of recent transmission than the susceptible strains(adjusted odds ratio,1.414;95%CI,1.023–1.954;P=0.036).Among all patients with drug-resistant tuberculosis(DR-TB),78.4%(171/218)were attributed to the transmission of DR-TB strains.Conclusion Our results suggest that drug-resistant strains are more transmissible than susceptible strains and that transmission is the major driving force of the current DR-TB epidemic in Kashgar.

Molecular Identification of Mycobacterium Strains Responsible of Bovine Tuberculosis Cases in Bobo-Dioulasso Slaughterhouse, Burkina Faso

Bovine tuberculosis (bTB) is an endemic zoonosis significantly affects animal health in Burkina Faso. The primary causative agent is Mycobacterium tuberculosis (M. tuberculosis) complex, mainly M. bovis. Cattle are considered as natural reservoir of M. bovis. However, in Burkina Faso, the circulation of these strains remains poorly understood and documented. This study aimed to identify and characterize Mycobacterium strains from suspected carcasses during routine meat inspection at Bobo-Dioulasso refrigerated slaughterhouse. A prospective cross-sectional study was conducted from January 2021 to December 2022 on cases of seizures linked to suspected bovine tuberculosis. Microbiological and molecular analyzes were used for mycobacterial strain isolation and characterization. Out of 50 samples, 24% tested positive by microscopy and 12% by culture. Molecular analysis identified 6 strains of Mycobacteria, exclusively Mycobacterium bovis specifically the subspecies bovis (Mycobacterium bovis subsp bovis). In conclusion, M. bovis subsp bovis is the primary agent responsible for bovine tuberculosis in Bobo-Dioulasso. Continuous monitoring of mycobacterial strains is therefore necessary for the effective control of this pathology in the local cattle population.

Mycobacterium gordoniasis of the cervical lymph nodes:A case report

BACKGROUND Owing to the advancement in bacterial identification techniques,the detection rate of non-tuberculous mycobacterium(NTM)has been on the rise.Different from Mycobacterium tuberculosis,the clinical symptoms of NTM are not easily detected,and the clinical efficacy and prognosis are somewhat heterogeneous.To report a case of Mycobacterium gordoniasis of cervical lymph node diagnosed in Anhui Chest Hospital in July 2022.CASE SUMMARY Upon examination,the patient who weighed 67.5 kg,was human immunodeficiency virus negative,healthy,without hypertension,diabetes,heart disease and other basic diseases microscopic analysis revealed granulomatous inflammation with coagulation necrosis in the lymphocyte,and tuberculosis was not ruled out.Plain computed tomography scans of the neck and chest indicated the presence of a single grayish-yellow and grayish-brown tissue,the dimensions of which was top of form 10.5 cm×3.0 cm×1.5 cm.After pathological consultation in our hospital,the diagnosis was confirmed as NTM infection.CONCLUSION This case report and the clinical epidemiological research on improving NTM have important guiding significance for improving decision-making in clinical treatments.

Overlapping infections of Mycobacterium canariasense and Nocardia farcinica in an immunocompetent patient:A case report

BACKGROUND Infections by non-tuberculous mycobacteria(NTM)have become more common in recent years.Mycobacterium canariasense(M.canariasense)was first reported as an opportunistic pathogen in 2004,but there have been very few case reports since then.Nocardia is a genus of aerobic and Gram-positive bacilli,and these species are also opportunistic pathogens and in the Mycobacteriales order.Conventional methods for diagnosis of NTM are inefficient.Metagenomic next-generation sequencing(mNGS)can rapidly detect many pathogenic microorganisms,even rare species.Most NTM and Nocardia infections occur in immunocompromised patients with atypical clinical symptoms.There are no previous reports of infection by M.canariasense and Nocardia farcinica(N.farcinica),especially in immunocompetent patients.This case report describes an immunocompetent 52-year-old woman who had overlapping infections of M.canariasense,N.farcinica,and Candida parapsilosis(C.parapsilosis)based on mNGS.CASE SUMMARY A 52-year-old woman presented with a productive cough and chest pain for 2 wk,and recurrent episodes of moderate-grade fever for 1 wk.She received antibiotics for 1 wk at a local hospital,and experienced defervescence,but the productive cough and chest pain persisted.We collected samples of a lung lesion and alveolar lavage fluid for mNGS.The lung tissue was positive for M.canariasense,N.farcinica,and C.parapsilosis,and the alveolar lavage fluid was positive for M.canariasense.The diagnosis was pneumonia,and application of appropriate antibiotic therapy cured the patient.CONCLUSION Etiological diagnosis is critical for patients with infectious diseases.mNGS can identify rare and novel pathogens,and does not require a priori knowledge.

Epidemiology and Clinical Features of Mycobacterium avium Complex Pulmonary Disease in a Brazilian Amazon Cohort

The incidence of Mycobacterium avium complex pulmonary disease (MACPD) has been globally increasing. The present study aims to provide a comprehensive analysis of clinical characteristics and treatment response among patients in the Amazon region of Brazil and conduct a retrospective cohort study at a prominent referral hospital in the State of Par, Brazil, from 2012 to 2020. The M. avium group represented 58.9% of cases, followed by M. intracellulare (35.7%), M. colombiense (3.6%), and M. chimaera (1.8%). The majority of patients (73.2%) were female and with an average age of 63 years. Primary clinical manifestations included persistent cough and sputum production. The Charlson comorbidity index (CCI) averaged 2.47, with 36.3% of patients having a CCI score equal to higher than 3. Notably, over 70% of patients had received treatment for pulmonary tuberculosis before initiating treatment for MAC infection. The fibrocavitary radiologic form was the most prevalent (55.4%), frequently exhibiting a bilateral distribution (53.6%). Antimicrobial susceptibility revealed a significant prevalence of MAC resistance to drugs in the therapeutic regimen. Despite this, most of the patients experienced clinical improvement (50%). In conclusion, this study highlights a higher prevalence of MAC infections among middle-aged women, with a history of pulmonary tuberculosis treatment and presenting the fibrocavitary radiological form was predominant.

Tuberculosis in the Carcinal Environment in Chad Due to the Mycobacterium Tuberculosis Circulante Complex

Tuberculosis (TB) is a serious infectious disease and its control is considered a challenge, particularly among vulnerable populations such as prisoners. The prevalence of TB in prisons is an alarming public health problem in many countries. The aim of this study is to describe the epidemiology of tuberculosis and the strategies for controlling this disease in the Chadian prison population. During the course of our study, the prevalence of tuberculosis in prisons was 9.64%. The age group between 55 years and over (43.33%) was the most represented in this study, and all patients were men with a frequency of 62.66%. The cross-tabulation of Culture_7H9G and Culture_7H9P showed that out of our total positive sample, we found a total of 87 positive strains and 63 negative strains. Our study shows that it is essential to know the prevalence of tuberculosis in all the country’s prisons. Indeed, this can serve as an indication of the need for action in prisons to reduce TB rates, in particular by improving the structure of prison environments, diagnosing new cases quickly and accurately, identifying drug-resistant strains and implementing effective, direct treatment observed in people with tuberculosis.

Early Prosthetic Valve Endocarditis with Mycobacterium Tuberculosis after Mitral Valve Replacement: A Case Report

Background: Tuberculous endocarditis is a rare but serious complication of heart valve replacement surgery. We report the case of a 24-year-old patient, who presented with tuberculous endocarditis after mechanical mitral valve replacement, with a favorable clinical course following anti-tuberculosis treatment. Case Presentation: We report a 24-year-old male patient, admitted to the cardiac surgery department of the Fann Hospital (Dakar, Senegal), for the management of severe mixed (rheumatic and endocarditic) mitral insufficiency with associated tricuspid insufficiency. He had a history of recurrent angina and polyarthralgia in childhood, was hospitalized several times for refractory global cardiac decompensation, and for a suspected infective endocarditis a month before his admission. On admission, the clinical examination revealed signs suggestive of mitral and tricuspid insufficiency. Transthoracic echocardiography revealed severe post-endocarditic mitral insufficiency with A3 amputation, highly mobile 15 mm vegetations on the free edge of the large valve, moderate tricuspid insufficiency, and severe pulmonary artery hypertension. Mechanical mitral valve replacement and tricuspid valve annuloplasty using autologous pericardial strip were performed via median sternotomy. After ten days, the patient presented with global cardiac decompensation associated with a clinico-biological infectious syndrome, and tans-oesophageal echography revealed an abscess at the sinotubular junction, communicating with the aorta. A thoraco-abdomino-pelvic CT scan was done, which revealed a bilateral alveolar-interstitial syndrome with mediastinal lymphadenopathy. Anti-tuberculosis treatment with RHZE was initiated for 06 months. The clinical course was favorable. Conclusion: Tuberculous endocarditis in prostheses is a serious complication of heart valve replacement surgery, which may evolve favorably under medical treatment.

Mutation Characteristics of inhA and katG Genes in Isoniazid-Resistant Mycobacterium Tuberculosis Patients in Xinjiang

Objective:To analyze the mutation characteristics of inhA and katG genes in isoniazid-resistant Mycobacterium tuberculosis in Xinjiang.Methods:The katG and inhA in 148 strains of isoniazid-resistant Mycobacterium tuberculosis were amplified through fluorescence quantitative PCR,and the amplified products were sequenced and compared.Results:The inhA gene mutation rate of 148 strains of isoniazid-resistant mycobacterium tuberculosis was 13.51%(20/148),among which the inhA gene mutation rate among patients of Han,Uygur,and Kazakh ethnicity were 15.87%,13.21%,and 17.65%,respectively.There was no significant difference in the inhA mutation rate among nationalities(c^(2)=2.897,P>0.05).The mutation rate of the katG gene was 84.46%(125/148),among which the mutation rates of patients of Han,Uyghur,and Kazak ethnicities were 82.54%,84.91%,and 76.47%,respectively.The Hui and other ethnic groups were all affected by the katG gene mutation.There was no significant difference in the mutation rate of the katG gene among different ethnicities(c^(2)=3.772,P>0.05).The mutation rates of the inhA gene in southern Xinjiang,northern Xinjiang,and other provinces were 18.60%,9.28%,and 37.50%,respectively.The mutation rates of the inhA gene in different regions were statistically different(c^(2)=6.381,P<0.05).There was no significant difference in the inhA mutation rate between patients from southern and northern Xinjiang(c^(2)=2.214,P>0.05)and between southern Xinjiang and other provinces(c^(2)=1.424,P>0.05).However,the mutation rate of the inhA gene in patients from other provinces was higher than that in northern Xinjiang(c^(2)=5.539,P<0.05).The mutation rates of the katG gene in southern Xinjiang,northern Xinjiang,and other provinces were 81.40%,87.63%,and 62.50%,respectively.There was no significant difference in the mutation rates of the katG gene among different regions(c^(2)=3.989,P>0.05).Conclusion:katG gene mutation was predominant in isoniazid-resistant tuberculosis patients in Xinjiang Uygur Autonomous Region,and inhA and katG gene mutation were no different among different ethnic groups.

Characteristics of Mycobacterium tuberculosis serine protease Rv1043c in enzymology and pathogenicity in mice

The serine proteases of Mycobacteria tuberculosis(Mtb)are important contributors to the process of bacterial invasion and its pathogenesis.In the present study,we systematically characterized the role of the Rv1043c protein in Mycobacterium infection by purifying the Rv1043c protein in Escherichia coli and constructing a Mycobacterium smegmatis(Msg)strain overexpressing Rv1043c(Msg_Rv1043c).We found that Rv1043c had serine protease activity and localized to the surface of Mtb.We determined that the optimal pH and temperature for the Rv1043c serine protease were 9.0 and 45°C,respectively.Moreover,the serine protease activity of Rv1043c was enhanced by divalent metal ions of Ca^(2+)and Mg^(2+).Site-directed mutagenesis studies demonstrated that the serine 279 residue in Rv1043c plays a catalytic role.Additionally,mouse model studies confirmed that Rv1043c significantly enhanced the survival of Msg in vivo,induced pulmonary injury and lung cell apoptosis,and promoted the release of pro-inflammatory cytokines interleukin-1βand interleukin-6 in mice.This study presents novel insights into the relationship between mycobacterial serine protease and the pathogenesis of the disease.

Evaluation of Microsphere-based xMAP Test for gyrA Mutation Identification in Mycobacterium Tuberculosis

Drug-resistant(DR) tuberculosis(TB) is a growing threat to public health[1]. Fluoroquinolone(FQ) antibiotics have long been used as anti-tuberculosis drugs, and their widespread application has led to the development of resistance in clinical isolates of Mycobacterium tuberculosis complex(MTBC).

Characterization of Mutations in Genes Related to Rifampicin and Isoniazid Resistance in Multidrug-resistant Mycobacterium tuberculosis Strains from Hangzhou,China

Tuberculosis(TB),caused by the Mycobacterium tuberculosis complex(MTBC),is the 13^(th)leading cause of death and the second leading cause of infectious killer after COVID-19(above HIV and AIDS),according to the World Health Organization's 2022 Global TB Report.In 2021,10.6 million people were diagnosed with a new episode of TB and 1.6 million people died as a result.

Involvement of 2'-5'oligoadenylate synthetase-like proteinin the survivalof Mycobacterium tuberculosis avirulent strain in macrophages

Mycobacterium tuberculosis(M.tuberculosis)can replicate in the macrophage by interfering with many host protein functions.While it is far from known these host proteins for controlling M.tuberculosis infection.Herein,we infected macrophages including THP-1 and Raw264.7 cells with M.tuberculosis and identified the differentially expressed genes(DEGs)in the interferon signaling pathway.Among them,2'-5'oligoadenylate synthetase-like(OASL)underwent the greatest upregulation in M.tuberculosis-infected macrophages.Knockdown of the expression of OASL attenuated M.tuberculosis survival in macrophages.Further,bioinformatics analysis revealed the potential interaction axis of OASL-TAB3-RvO127,which was further validated by the yeast-two-hybrid(Y2H)assay and Co-IP.This interaction axis might regulate the M.tuberculosis survival and proliferation in macrophages.The study reveals a possible role of OASL during M.tuberculosis infection as a target to control its propagation.

Contribution to the Study of Resistance to Anti-Tuberculosis Drugs in the Mycobacterium tuberculosis Complex Isolated at the National Laboratory of Clinical Biology and Public Health in Bangui in the Central African Republic in 2022: Case of Rifampicin

Microscopy-positive and drug-resistant pulmonary tuberculosis (MPT+) is one of the most feared diseases due to the cost of its management and the associated mortality. The GeneXpert, a new molecular test, is in greater demand for the diagnosis of MPT+ resistance cases. The application of GeneXpert to new cases of MPT+ is not effective in the country’s TB screening centres. The objective of this study is to assess the contribution of GeneXpert to the determination of MPT+ resistance cases in Bangui. The study was cross-sectional and covered the period from February to July 2022. The diagnosis of tuberculosis was first performed by microscopy with Ziehl Neelsen hot stain. The GeneXpert was then used to test for resistance in the sputum of all patients with positive microscopy. The collected data was entered into Excel 2013 and analysed with Epi Info 3.3.7. We analysed data from 755 patients, 80 of whom had resistance. The 80 patients ranged in age from 6 to 68 years (mean age = 35 years). The prevalence of resistant TB was 10.60% (80/755). Primary resistance accounted for 73.75% and secondary resistance for 26.25%. The age group 20 - 39 years (57.50%), male (72.50%), 8th district (17.50%), people living in couples (53.75%), farmers (13.75%) were the socio-demographic characteristics most affected by resistance. Treatment failure (13.75%), relapses (13.75%), the notion of contagion (28.75%), a history of smoking (40%) and alcohol (61.25%) were the clinical antecedents reported by the patients. Treatment failure and relapse were the variables associated with the occurrence of resistant PMT+ (p 0.05). A considerable proportion of the overall Mycobacterium tuberculosis resistance to anti-tuberculosis drugs (10.60%) was identified by GeneXpert. Treatment failure and relapse were the factors associated with the risk of resistance.

Research progress on the drug action and resistance mechanism in Mycobacterium tuberculosis

Tuberculosis(TB)is a chronic infectious disease caused by Mycobacterium Tuberculosis(MTB).It is the second largest single cause of death besides novel coronavirus pneumonia.Along with the abuse of antibiotics and extensive use of anti-tuberculosis drugs,multidrug-resistant(MDR)TB,drug-resistant(XDR)TB and totally drug-resistant(TDR)TB became obstacles to the tuberculosis eradication worldwide.According to the World Health Organization(WHO)statistics,China is not only a high burden tuberculosis country in the world,but also a country with a serious epidemic of MDR.Traditional drugs fail to meet the needs of tuberculosis control.Therefore,it is urgent to find new targets of anti-tuberculosis drugs and develop new anti-tuberculosis drugs.Hence,this paper systematically summarizes the mechanism of traditional and newly developed anti-tuberculosis drugs,in which stressing the research progress of drug resistance mechanisms.This work provides us with new insights of new anti-tuberculosis drug developments,and may contribute to a reduction in the harm that tuberculosis brings to society.

Value of ultrasound guided biopsy combined with Xpert Mycobacterium tuberculosis/resistance to rifampin assay in the diagnosis of chest wall tuberculosis

BACKGROUND The thoracic wall lesions,particularly chest wall tuberculosis,and chest wall tumors and other pyogenic wall and actinomycetes infections,almost always present as a diagnostic challenge.AIM To explore the value of ultrasound-guided biopsy combined with the Xpert Mycobacterium tuberculosis/resistance to rifampin(MTB/RIF)assay to diagnose chest wall tuberculosis.METHODS We performed a retrospective study of patients with chest wall lesions from March 2018 to March 2021.All patients received the ultrasound-guided biopsy for pathology examination,acid-fast Bacillus staining,mycobacterial culture,and Xpert MTB/RIF analysis.The sensitivity,specificity,and area under the curve(AUC)were calculated for these diagnostic tests,either individually or combined.Rifampicin resistance results were compared between the mycobacterial culture and the Xpert MTB/RIF assay.RESULTS In 31 patients with the chest wall lesion biopsy,22 patients were diagnosed with chest wall tuberculosis.Of them,3,6,and 21 patients tested positive for mycobacterial culture,acid-fast stain,and Xpert MTB/RIF assay,respectively.The rifampicin resistance results of the 3 culture-positive patients were consistent with their Xpert MTB/RIF assay results.When considering the sensitivity,specificity,and AUC value,the Xpert MTB/RIF assay(95.5%,88.9%,and 0.92,respectively)was a better choice than the acid-fast Bacillus stain(27.3%,100.0%,and 0.64,respectively)and mycobacterial culture(13.6%,100.0%,0.57,respectively).No complications were reported during the procedure.CONCLUSION Ultrasound guided biopsy combined with Xpert MTB/RIF has high value in the diagnosis of chest wall tuberculosis,and can also detect rifampicin resistance.

Mycobacterium Phlei在黄铁矿和方铅矿表面的选择性吸附作用机理

运用吸附量测定、ζ-电位研究、红外光谱和X射线光电子能谱分析,探讨Mycobacterium Phlei在黄铁矿和方铅矿表面的选择性吸附作用机理。结果表明,在其他实验条件固定的情况下,当溶液的初始pH值大于5时,Mycobacterium Phlei在黄铁矿表面的吸附量远远大于方铅矿;静电作用不是致使Mycobacterium Phlei在黄铁矿和方铅矿表面产生选择性吸附的主要作用力;Mycobacterium Phlei细胞表面的C、N和O元素通过矿物表面的Fe,Pb和S元素与矿物表面发生了化学吸附,而且与黄铁矿表面元素的作用程度大于方铅矿。矿物的表面结构及其性质、细胞表面基团与矿物表面金属离子的键合作用大小可能是致使Mycobacterium Phlei产生选择性吸附的主要原因。

Mycobacterium phlei菌对重金属Pb^(2+),Zn^(2+),Ni^(2+),Cu^(2+)的吸附规律

研究了Mycobacteriumphlei菌株分别对水相中重金属离子Pb2+,Zn2+,Ni2+,Cu2+的吸附规律·结果表明,Mycobacteriumphlei菌株对水相中的这四种重金属离子均有一定的吸附作用;吸附过程均在10min内就达到平衡;pH对吸附过程影响较大,在pH=3 0～4 0时,Mycobacteriumphlei菌对这四种金属离子吸附效果最好;温度升高对Pb2+,Ni2+和Cu2+的吸附过程不利,但Mycobacteriumphlei菌对Zn2+的吸附则是一个吸热过程·重金属离子在Mycobacteriumphlei菌上的吸附的选择性大小的顺序为:Pb2+>Zn2+>Cu2+>Ni2+·

Novel Species Including Mycobacterium fukienense sp. Is Found from Tuberculosis Patients in Fujian Province, China, Using Phylogenetic Analysis of Mycobacterium chelonae/abscessus Complex

Objective To identify the novel species ‘Mycobacterium fukienense＇ sp. nov of Mycobacterium chelonoe/abscessus complex from tuberculosis patients in Fujian Province, China. Methods Five of 27 clinical Mycobucterium isolates （CIs） were previously identified as M. chelonoe/obscessus complex by sequencing the hsp65, rpoB, 165-235 rRNA internal transcribed spacer region （its）, recA and sodA house-keeping genes commonly used to describe the molecular characteristics of Mycobocterium. Clinical Mycobecterium isolates were classified according to the gene sequence using a clustering analysis program. Sequence similarity within clusters and diversity between clusters were analyzed. Results The 5 isolates were identified with distinct sequences exhibiting 99.8% homology in the hsp65 gene. However, a complete lack of homology was observed among the sequences of the rpoB, 165-235 rRNA internal tronscribed spacer region （its）, sodA, and recA genes as compared with the M. obscessus. Furthermore, no match for rpoB, sodA, and recA genes was identified among the published sequences. Conclusion The novel species, Mycobacterium fukienense, is identified from tuberculosis patients in Fujian Province, China, which does not belong to any existing subspecies of M. cheloneo/abscessus complex.

Expression,purification and characterization of Mycobacterium tuberculosis RpfE protein

Resuscitation promoting factor E （RpfE） is one of the five Rpf-like proteins in Mycobacterium tuberculos＆ （M. tuberculosis）. These Rpf-like proteins are secretory, which make them candidates for recognition by the host immune system. In this study, the RpfE gene was amplified from M. tuberculosis, cloned into the expression vectors pDE22 and pPRO EXHT, and were expressed in Mycobacterium vaccae （M. vaccae） and Escherichia coli DHSa, respec- tively. Both recombinant RpfE proteins were purified by Ni-Sepharose affinity chromatography, and were given to C57BL/6 mice. The RpfE proteins elicited T cell proliferation, and stimulated the production of gamma interferon （IFN-y）, interleukin-10 （IL-10） and IL-12. Our results indicated that the RpfE protein expressed in M. vaccae could more efficiently stimulate cellular immune response, making it a promising candidate as a subunit vaccine.