Objective Macrolide susceptibility and drug resistance mechanisms of clinical non-tuberculous mycobacteria(NTM) isolates were preliminarily investigated for more accurate diagnosis and treatment of the infection in ...Objective Macrolide susceptibility and drug resistance mechanisms of clinical non-tuberculous mycobacteria(NTM) isolates were preliminarily investigated for more accurate diagnosis and treatment of the infection in China. Methods Four macrolides, including clarithromycin(CLAR), azithromycin(AZM), roxithromycin(ROX), and erythromycin(ERY), were used to test the drug susceptibility of 310 clinical NTM isolates from six provinces of China with the broth microdilution method. Two resistance mechanisms, 23 S r RNA and erm, were analyzed with nucleotide sequence analysis. Results Varied effectiveness of macrolides and species-specific resistance patterns were observed. Most Mycobacterium abscessus subsp. massiliense were susceptible and all M. fortuitum were highly resistant to macrolides. All the drugs, except for erythromycin, exhibited excellent activities against slow-growing mycobacteria, and drug resistance rates were below 22.2%. Only four highly resistant strains harbored 2,058/2,059 substitutions on rrl and none of other mutations were related to macrolide resistance. G2191 A and T2221 C on rrl were specific for the M. abscessus complex(MABC). Seven sites, G2140 A, G2210 C, C2217 G, T2238 C, T2322 C, T2404 C, and A2406 G, were specifically carried by M. avium and M. intracellulare. Three sites, A2192 G, T2358 G, and A2636 G, were observed only in M. fortuitum and one site G2152 A was specific for M. gordonae. The genes erm(39) and erm(41) were detected in M. fortuitum and M. abscessus and inducible resistance was observed in relevant sequevar. Conclusion The susceptibility profile of macrolides against NTM was demonstrated. The well-known macrolide resistance mechanisms, 23 S r RNA and erm, failed to account for all resistant NTM isolates, and further studies are warranted to investigate macrolide resistance mechanisms in various NTM species.展开更多
Background:It is difficult to quickly distinguish non-tuberculous mycobacterial(NTM)infection from tuberculosis(TB)infection in human immunodeficiency virus(HIV)-infected patients because of many similarities between ...Background:It is difficult to quickly distinguish non-tuberculous mycobacterial(NTM)infection from tuberculosis(TB)infection in human immunodeficiency virus(HIV)-infected patients because of many similarities between these diseases.A simple and effective way to determine the differences using routine blood tests is necessary in developing countries.Methods:A retrospective cohort study was conducted to recruit HIV-infected patients with either NTM infection or TB infection diagnosed for the first time according to mycobacterial culture and microscopic identification from May 2010 to March 2016.These data included the analysis of blood cells,liver function,renal function,C-reactive protein(CRP),and erythrocyte sedimentation rate(ESR),and were compared between the HIV/TB and HIV/NTM groups.Results:A total of 240 patients were enrolled.The number of HIV/TB and HIV/NTM patients was 113 and 127,respectively.There were no significant differences in the CD4 T-cell count,age,sex,percentage of patients initiating antiretroviral therapy(ART)before the explicit diagnosis of TB or NTM infection.NTM infection was more likely to be restricted in the pulmonary while TB infection also involves extra-pulmonary sites.Both the leukocyte count(5.60×109/L)and the proportion of neutrophils in the leukocyte count(76.70%)in the HIV/TB group were significantly higher than those in the HIV/NTM group(4.40×10^(9)/L[P=0.0014]and 69.30%[P<0.001].The analysis of liver function markers indicated that the concentration of albumin but not ALT and AST was significantly lower in the HIV/TB group than in the HIV/NTM group(P<0.001).The creatinine and urea levels were not significantly different between the two groups.The ESR(84.00 mm/h)and the concentration of CRP(59.60 mg/L)were significantly higher in the HIV/TB group than in the HIV/NTM group(52.00 mm/h and 19.60 mg/L,respectively)(P<0.001).To distinguish TB infection from NTM infection,the best cut-off value was 69.5 mm/h for ESR,with a positive predictive value(PPV)of 0.740 and negative predictive value(NPV)of 0.721,and 48.8 mg/L for CRP,with a PPV of 0.676 and NPV of 0.697.Conclusion:The dissemination character as well as stronger immune response characterized by higher inflammation markers(e.g.WBC,ESR,CRP)can help distinguish TB from NTM infection in HIV-infected patients who need empirical therapy or diagnostic therapy immediately in low-income areas.展开更多
Background Non-tuberculous mycobacteria (NTM) have emerged as important opportunistic pathogens of the human being in recent years. Patients with pre-existing bronchiectasis are susceptible to NTM. However, informat...Background Non-tuberculous mycobacteria (NTM) have emerged as important opportunistic pathogens of the human being in recent years. Patients with pre-existing bronchiectasis are susceptible to NTM. However, information about its occurrence among bronchiectatic patients in Shenzhen, China is lacking and its impact on the course of bronchiectasis following surgical intervention is unknown. This preliminary study aimed to investigate the prevalence of NTM in bronchiectasis that required surgery in our center, evaluate the role of intraoperative routine screening for NTM, and summarize our initial experience in thoracoscopic management for bronchiectatic patients with NTM. Methods A retrospective analysis of clinical, microbiological data of our bronchiectatic patients with NTM over 5 years was made and 40 patients with bronchiectasis were studied to determine the role of intraoperative routine screening for NTM. Results The prevalence of NTM in this population of patients with bronchiectasis in our center was 6.7% (7/105). The diagnostic yield of the 40 intraoperative specimens was 7.5% (3/40). Of the 7 patients with bronchiectasis and NTM, 3 patients developed postoperative wound infections. All were cured with chemotherapy for 8-12 months along with vigorous surgical debridement. Another patient had a slow growth of mycobacteria involving double lungs and the right thoracic cavity and recovered after chemotherapy for nearly 14 months and tube drainage. The affected tissue was completely resected in the remaining 3 patients with no operative mortality and postoperative morbidity, and routine intraoperative screening for NTM was initiated in these patients. Conclusions NTM is not uncommon in bronchiectatic patients which deserves surgeons' utmost attention. Routine intraoperative screening for NTM identified otherwise unsuspected patients has shown favorable outcomes. Thoracoscopic management for bronchiectasis with NTM is technically feasible although its role remains to be defined.展开更多
<b><span style="font-family:Verdana;">Objective: </span></b><span style="font-family:;" "=""><span style="font-family:Verdana;">To under...<b><span style="font-family:Verdana;">Objective: </span></b><span style="font-family:;" "=""><span style="font-family:Verdana;">To understand the distribution of drug susceptibility test results of opportunistic infections of tuberculosis and non-tuberculous bacilli in AIDS patients. </span><b><span style="font-family:Verdana;">Methods: </span></b><span style="font-family:Verdana;">The AIDS patients who were hospitalized in our hospital from January 2016 to June 2019 were collected as the research objects, and patients with opportunistic tuberculosis and non-tuberculous bacilli from AIDS patients were screened for drug susceptibility tests, and the distribution characteristics of drug susceptibility were analyzed. </span><b><span style="font-family:Verdana;">Results: </span></b><span><span style="font-family:Verdana;">179 strains of tuberculosis and non-tuberculous mycobacteria were isolated from the specimens of AIDS patients, including 135 cases of tuberculosis mycobacteria and 44 cases of non-tuberculous mycobacteria. In the results of the drug susceptibility test, most strains of </span><i></i></span><i><i><span style="font-family:Verdana;">Mycobacterium tuberculosis</span></i><span></span></i><span style="font-family:Verdana;"> showed sensitivity to commonly used drugs, and a few strains showed resistance;most strains </span></span><span style="font-family:Verdana;">of non-tuberculous mycobacteria showed resistance, and a few strains showed</span><span style="font-family:Verdana;"> sensitivity. </span><b><span style="font-family:Verdana;">Conclusion: </span></b><span><span style="font-family:Verdana;">AIDS opportunistic infection of </span><i></i></span><i><i><span style="font-family:Verdana;">Mycobacterium tuberculosis</span></i><span></span></i><span style="font-family:Verdana;"> and non-tuberculous mycobacteria have significant differences in drug sensitivity test results. Timely detection and analysis are of great significance to the diagnosis and treatment of the disease.</span>展开更多
Objective:To determine the accuracy of TB diagnosis of TB in Zambia in the era of increasing HIV prevalence.Methods:Sputum of the clinically diagnosed TB cases was additionally subjected to liquid culture and molecula...Objective:To determine the accuracy of TB diagnosis of TB in Zambia in the era of increasing HIV prevalence.Methods:Sputum of the clinically diagnosed TB cases was additionally subjected to liquid culture and molecular identification.This study distinguished between TB cases confirmed by positive Mycobacterium tuberculosis(M.tuberculosis) cultures and mycobacterial disease caused by non-tuberculous mycobacteria(NTM).Results:Only 49% of the 173 presumptively diagnosed TB cases was M.tuberculosis cultured,while in 13% (22) cases,a combination of M.tuberculosis and NTM was found.In 18% of the patients only NTM were cultured.In 28% ,no mycobacteria was cultivable.HIV positive status was correlated with the isolation of NTM(P【0.05).Conclusions:The diagnosis of tuberculosis based on symptoms, sputum smear and/or chest X-ray leads to significant numbers of false-positive TB cases in Zambia,most likely due to the increased prevalence of HIV.The role of NTM in tuberculosislike disease also seems relevant to the false diagnosis of TB in Zambia.展开更多
We report two cases of pulmonary sequestration infected with nontuberculous mycobacteria(NTM): Mycobacterium avium and Mycobacterium abscessus.Chest computed tomography showed pneumonic consolidation in the right lowe...We report two cases of pulmonary sequestration infected with nontuberculous mycobacteria(NTM): Mycobacterium avium and Mycobacterium abscessus.Chest computed tomography showed pneumonic consolidation in the right lower lobe,which received a systemic blood supply from the descending aorta in both patients.Video-assisted thoracoscopic surgeries were successfully performed and pathologica]examinations revealed multiple caseating granulomas.A review of the literature revealed only seven previous case reports of pulmonary sequestration infected with NTM,and no case with Mycobacterium abscessus has been reported.展开更多
By employing the pUC19 as a backbone,the regulatory and signal sequences which encode kanamycin resistance, and mycobacterial plasmid origin of replication (oriM) were cloned into the pUC19. The recombinant E. Coli-my...By employing the pUC19 as a backbone,the regulatory and signal sequences which encode kanamycin resistance, and mycobacterial plasmid origin of replication (oriM) were cloned into the pUC19. The recombinant E. Coli-mycobacteria shuttle expression plasmid PBCG-8000 was constructed. The PBCG-8000 was able to replicate in both E. Coli and mycobacteria (including BCG) systems, and to confer stable kanamycin resistance upon transformants. The study should facilitate the development of BCG and other mycobacteria into multivalent vaccine vectors.展开更多
Background: The incidence of mycobacterial infection, in particular M. tuberculosis complex (MTC), is increasing in some Western countries, while nontuberculous mycobacteria (NTM) may be recognized more frequently in ...Background: The incidence of mycobacterial infection, in particular M. tuberculosis complex (MTC), is increasing in some Western countries, while nontuberculous mycobacteria (NTM) may be recognized more frequently in clinical specimens worldwide. The clinical scenario and available histopathology alone are often insufficient to separate these two categories of mycobacterial disease, whose behavior and treatment differ. In particular, NTM may be clinically unsuspected in pathological specimens and the opportunity for culturing missed. Methods: We developed two multiplex PCR assays, which distinguish MTC from NTM by detecting the IS6110 insert in the first tube and discriminating up to 14 NTM reference strains in the second by targeting the 16S-23S rRNA internal transcribed spacer. Test material included 594 routine clinical specimens with diverse pathology;many were granulomas unrelated to mycobacterial infection. About 75% were formalin-fixed paraffin blocks, the remainder mainly cytologic imprints or aspirates on FTA cards submitted on suspicion of mycobacterial infection either to avoid frozen sectioning (with the attendant risk of aerosolisation) or at the time of fine needle aspiration. Results: The paraffinized material yielded 53 MTC positives and the cytological 21 positives. A subset consisting of 337 specimens was also analyzed for NTM and yielded 51 positives. The frequency of simultaneous NTM infection in tuberculous patients was about 17%. Mycobacterium avium complex represented the dominant NTM species overall, showed a predilection for lung and lymph node, and together with M. haemophilum were the second most frequent NTM just behind M. ulcerans/M. marinum in skin and soft tissue, the category displaying the largest NTM diversity. Conclusions: Cytological and deparaffinized tissue analyzed in a new two-tube multiplex PCR allows for specific discrimination of causative agents in mycobacterial infection. MTC is readily distinguished from NTM for appropriate therapy, and NTM presumptively diagnosed at the species level allows appropriate choices of antimicrobials.展开更多
Objective Mycobacterium tuberculosis(Mtb),the causative agent of tuberculosis(TB),causes an estimated 1.6 million human deaths annually,but the pathogenesis of TB remains unclear.Immunity plays a critical role in the ...Objective Mycobacterium tuberculosis(Mtb),the causative agent of tuberculosis(TB),causes an estimated 1.6 million human deaths annually,but the pathogenesis of TB remains unclear.Immunity plays a critical role in the onset and outcome of TB.This study aimed to uncover the roles of innate and adaptive immunity in TB.Methods The gene expression profiles generated by RNA sequencing from human peripheral blood mononuclear cells(PBMCs)stimulated with or without Mtb strain H37Rv antigens were analyzed.A total of 973 differentially expressed mRNAs were identified.Results The differentially expressed genes were enriched in innate immunity signaling functions.The mesenchymal-epithelial transition factor(MET)gene was significantly upregulated in CD14^(+)monocytes.A MET inhibitor improved the uptake of the BCG strain by monocytes and macrophages as well as inhibited the expression of indoleamine 2,3-dioxygenase(IDO).The expression of IDO was increased in PBMCs stimulated with Mtb antigens,and the IDO inhibitor promoted the expression of CD40,CD83,and CD86.Conclusion Our results might provide clues regarding the immunomodulatory mechanisms used by Mtb to evade the host defense system.展开更多
Over the past few decades, there has been a significant increase in the number of mycobacterial species described. Currently, the genus?Mycobacterium?consists of 170 species. Most species are called nontuberculous myc...Over the past few decades, there has been a significant increase in the number of mycobacterial species described. Currently, the genus?Mycobacterium?consists of 170 species. Most species are called nontuberculous mycobacteria (NTM) and are potentially or rarely pathogenic and ubiquitous. One of the main challenges in mycobacteriology is the rapid and precise identification of these microorganisms. In this work, we compared two protein extraction protocols for the identification of 38 reference strains and clinical isolates, representing 27 species, by mass spectrometry (MALDI-TOF MS) to subsequently use the best method for identifying environmental mycobacteria. The results obtained with reference strains and clinical isolates showed that protocol A was effective in identifying 92.1% of mycobacterial specimens at the species level and protocol B, 50%. Therefore, protocol A was evaluated for the rapid identification of 27 environmental mycobacterial isolates. These isolates were subjected to PCR-restriction enzyme analysis (PRA-hsp65). Two isolates were misidentified by PRA-hsp65, whereas MALDI-TOF MS was able to identify them correctly. The results were confirmed by?hsp65 and 16S rRNA gene sequencing. Mass spectrometry has the advantage of being a simpler and faster technique than PRA-hsp65, and our results showed that MALDI-TOF MS is a valuable tool for the identification of environmental mycobacterial isolates.展开更多
基金supported by the Key Project of the State Key Laboratory for Infectious Disease Prevention and Control[2014SKLID104]the National Key Programs of Mega Infectious Diseases[2013ZX10003002-001]
文摘Objective Macrolide susceptibility and drug resistance mechanisms of clinical non-tuberculous mycobacteria(NTM) isolates were preliminarily investigated for more accurate diagnosis and treatment of the infection in China. Methods Four macrolides, including clarithromycin(CLAR), azithromycin(AZM), roxithromycin(ROX), and erythromycin(ERY), were used to test the drug susceptibility of 310 clinical NTM isolates from six provinces of China with the broth microdilution method. Two resistance mechanisms, 23 S r RNA and erm, were analyzed with nucleotide sequence analysis. Results Varied effectiveness of macrolides and species-specific resistance patterns were observed. Most Mycobacterium abscessus subsp. massiliense were susceptible and all M. fortuitum were highly resistant to macrolides. All the drugs, except for erythromycin, exhibited excellent activities against slow-growing mycobacteria, and drug resistance rates were below 22.2%. Only four highly resistant strains harbored 2,058/2,059 substitutions on rrl and none of other mutations were related to macrolide resistance. G2191 A and T2221 C on rrl were specific for the M. abscessus complex(MABC). Seven sites, G2140 A, G2210 C, C2217 G, T2238 C, T2322 C, T2404 C, and A2406 G, were specifically carried by M. avium and M. intracellulare. Three sites, A2192 G, T2358 G, and A2636 G, were observed only in M. fortuitum and one site G2152 A was specific for M. gordonae. The genes erm(39) and erm(41) were detected in M. fortuitum and M. abscessus and inducible resistance was observed in relevant sequevar. Conclusion The susceptibility profile of macrolides against NTM was demonstrated. The well-known macrolide resistance mechanisms, 23 S r RNA and erm, failed to account for all resistant NTM isolates, and further studies are warranted to investigate macrolide resistance mechanisms in various NTM species.
基金This study was supported by grants from the fourth round public health 3-year action plan key disciplines construction project of infectious diseases and health microbiology(Grant No.15GWZK0103).
文摘Background:It is difficult to quickly distinguish non-tuberculous mycobacterial(NTM)infection from tuberculosis(TB)infection in human immunodeficiency virus(HIV)-infected patients because of many similarities between these diseases.A simple and effective way to determine the differences using routine blood tests is necessary in developing countries.Methods:A retrospective cohort study was conducted to recruit HIV-infected patients with either NTM infection or TB infection diagnosed for the first time according to mycobacterial culture and microscopic identification from May 2010 to March 2016.These data included the analysis of blood cells,liver function,renal function,C-reactive protein(CRP),and erythrocyte sedimentation rate(ESR),and were compared between the HIV/TB and HIV/NTM groups.Results:A total of 240 patients were enrolled.The number of HIV/TB and HIV/NTM patients was 113 and 127,respectively.There were no significant differences in the CD4 T-cell count,age,sex,percentage of patients initiating antiretroviral therapy(ART)before the explicit diagnosis of TB or NTM infection.NTM infection was more likely to be restricted in the pulmonary while TB infection also involves extra-pulmonary sites.Both the leukocyte count(5.60×109/L)and the proportion of neutrophils in the leukocyte count(76.70%)in the HIV/TB group were significantly higher than those in the HIV/NTM group(4.40×10^(9)/L[P=0.0014]and 69.30%[P<0.001].The analysis of liver function markers indicated that the concentration of albumin but not ALT and AST was significantly lower in the HIV/TB group than in the HIV/NTM group(P<0.001).The creatinine and urea levels were not significantly different between the two groups.The ESR(84.00 mm/h)and the concentration of CRP(59.60 mg/L)were significantly higher in the HIV/TB group than in the HIV/NTM group(52.00 mm/h and 19.60 mg/L,respectively)(P<0.001).To distinguish TB infection from NTM infection,the best cut-off value was 69.5 mm/h for ESR,with a positive predictive value(PPV)of 0.740 and negative predictive value(NPV)of 0.721,and 48.8 mg/L for CRP,with a PPV of 0.676 and NPV of 0.697.Conclusion:The dissemination character as well as stronger immune response characterized by higher inflammation markers(e.g.WBC,ESR,CRP)can help distinguish TB from NTM infection in HIV-infected patients who need empirical therapy or diagnostic therapy immediately in low-income areas.
文摘Background Non-tuberculous mycobacteria (NTM) have emerged as important opportunistic pathogens of the human being in recent years. Patients with pre-existing bronchiectasis are susceptible to NTM. However, information about its occurrence among bronchiectatic patients in Shenzhen, China is lacking and its impact on the course of bronchiectasis following surgical intervention is unknown. This preliminary study aimed to investigate the prevalence of NTM in bronchiectasis that required surgery in our center, evaluate the role of intraoperative routine screening for NTM, and summarize our initial experience in thoracoscopic management for bronchiectatic patients with NTM. Methods A retrospective analysis of clinical, microbiological data of our bronchiectatic patients with NTM over 5 years was made and 40 patients with bronchiectasis were studied to determine the role of intraoperative routine screening for NTM. Results The prevalence of NTM in this population of patients with bronchiectasis in our center was 6.7% (7/105). The diagnostic yield of the 40 intraoperative specimens was 7.5% (3/40). Of the 7 patients with bronchiectasis and NTM, 3 patients developed postoperative wound infections. All were cured with chemotherapy for 8-12 months along with vigorous surgical debridement. Another patient had a slow growth of mycobacteria involving double lungs and the right thoracic cavity and recovered after chemotherapy for nearly 14 months and tube drainage. The affected tissue was completely resected in the remaining 3 patients with no operative mortality and postoperative morbidity, and routine intraoperative screening for NTM was initiated in these patients. Conclusions NTM is not uncommon in bronchiectatic patients which deserves surgeons' utmost attention. Routine intraoperative screening for NTM identified otherwise unsuspected patients has shown favorable outcomes. Thoracoscopic management for bronchiectasis with NTM is technically feasible although its role remains to be defined.
文摘<b><span style="font-family:Verdana;">Objective: </span></b><span style="font-family:;" "=""><span style="font-family:Verdana;">To understand the distribution of drug susceptibility test results of opportunistic infections of tuberculosis and non-tuberculous bacilli in AIDS patients. </span><b><span style="font-family:Verdana;">Methods: </span></b><span style="font-family:Verdana;">The AIDS patients who were hospitalized in our hospital from January 2016 to June 2019 were collected as the research objects, and patients with opportunistic tuberculosis and non-tuberculous bacilli from AIDS patients were screened for drug susceptibility tests, and the distribution characteristics of drug susceptibility were analyzed. </span><b><span style="font-family:Verdana;">Results: </span></b><span><span style="font-family:Verdana;">179 strains of tuberculosis and non-tuberculous mycobacteria were isolated from the specimens of AIDS patients, including 135 cases of tuberculosis mycobacteria and 44 cases of non-tuberculous mycobacteria. In the results of the drug susceptibility test, most strains of </span><i></i></span><i><i><span style="font-family:Verdana;">Mycobacterium tuberculosis</span></i><span></span></i><span style="font-family:Verdana;"> showed sensitivity to commonly used drugs, and a few strains showed resistance;most strains </span></span><span style="font-family:Verdana;">of non-tuberculous mycobacteria showed resistance, and a few strains showed</span><span style="font-family:Verdana;"> sensitivity. </span><b><span style="font-family:Verdana;">Conclusion: </span></b><span><span style="font-family:Verdana;">AIDS opportunistic infection of </span><i></i></span><i><i><span style="font-family:Verdana;">Mycobacterium tuberculosis</span></i><span></span></i><span style="font-family:Verdana;"> and non-tuberculous mycobacteria have significant differences in drug sensitivity test results. Timely detection and analysis are of great significance to the diagnosis and treatment of the disease.</span>
文摘Objective:To determine the accuracy of TB diagnosis of TB in Zambia in the era of increasing HIV prevalence.Methods:Sputum of the clinically diagnosed TB cases was additionally subjected to liquid culture and molecular identification.This study distinguished between TB cases confirmed by positive Mycobacterium tuberculosis(M.tuberculosis) cultures and mycobacterial disease caused by non-tuberculous mycobacteria(NTM).Results:Only 49% of the 173 presumptively diagnosed TB cases was M.tuberculosis cultured,while in 13% (22) cases,a combination of M.tuberculosis and NTM was found.In 18% of the patients only NTM were cultured.In 28% ,no mycobacteria was cultivable.HIV positive status was correlated with the isolation of NTM(P【0.05).Conclusions:The diagnosis of tuberculosis based on symptoms, sputum smear and/or chest X-ray leads to significant numbers of false-positive TB cases in Zambia,most likely due to the increased prevalence of HIV.The role of NTM in tuberculosislike disease also seems relevant to the false diagnosis of TB in Zambia.
基金supported by Mid-career Researcher Program through NRF grant funded by the MEST(2011-0015546)
文摘We report two cases of pulmonary sequestration infected with nontuberculous mycobacteria(NTM): Mycobacterium avium and Mycobacterium abscessus.Chest computed tomography showed pneumonic consolidation in the right lower lobe,which received a systemic blood supply from the descending aorta in both patients.Video-assisted thoracoscopic surgeries were successfully performed and pathologica]examinations revealed multiple caseating granulomas.A review of the literature revealed only seven previous case reports of pulmonary sequestration infected with NTM,and no case with Mycobacterium abscessus has been reported.
文摘By employing the pUC19 as a backbone,the regulatory and signal sequences which encode kanamycin resistance, and mycobacterial plasmid origin of replication (oriM) were cloned into the pUC19. The recombinant E. Coli-mycobacteria shuttle expression plasmid PBCG-8000 was constructed. The PBCG-8000 was able to replicate in both E. Coli and mycobacteria (including BCG) systems, and to confer stable kanamycin resistance upon transformants. The study should facilitate the development of BCG and other mycobacteria into multivalent vaccine vectors.
文摘Background: The incidence of mycobacterial infection, in particular M. tuberculosis complex (MTC), is increasing in some Western countries, while nontuberculous mycobacteria (NTM) may be recognized more frequently in clinical specimens worldwide. The clinical scenario and available histopathology alone are often insufficient to separate these two categories of mycobacterial disease, whose behavior and treatment differ. In particular, NTM may be clinically unsuspected in pathological specimens and the opportunity for culturing missed. Methods: We developed two multiplex PCR assays, which distinguish MTC from NTM by detecting the IS6110 insert in the first tube and discriminating up to 14 NTM reference strains in the second by targeting the 16S-23S rRNA internal transcribed spacer. Test material included 594 routine clinical specimens with diverse pathology;many were granulomas unrelated to mycobacterial infection. About 75% were formalin-fixed paraffin blocks, the remainder mainly cytologic imprints or aspirates on FTA cards submitted on suspicion of mycobacterial infection either to avoid frozen sectioning (with the attendant risk of aerosolisation) or at the time of fine needle aspiration. Results: The paraffinized material yielded 53 MTC positives and the cytological 21 positives. A subset consisting of 337 specimens was also analyzed for NTM and yielded 51 positives. The frequency of simultaneous NTM infection in tuberculous patients was about 17%. Mycobacterium avium complex represented the dominant NTM species overall, showed a predilection for lung and lymph node, and together with M. haemophilum were the second most frequent NTM just behind M. ulcerans/M. marinum in skin and soft tissue, the category displaying the largest NTM diversity. Conclusions: Cytological and deparaffinized tissue analyzed in a new two-tube multiplex PCR allows for specific discrimination of causative agents in mycobacterial infection. MTC is readily distinguished from NTM for appropriate therapy, and NTM presumptively diagnosed at the species level allows appropriate choices of antimicrobials.
基金This study was supported by the Thirteen-Fifth Mega-Scientific Project on“Prevention and Treatment of AIDS,Viral Hepatitis and Other Infectious Diseases”(No.2017ZX10201301-007-002)the National Natural Science Foundation of China(No.81571961 and No.82072233)the 309th Hospital(No.2017ZD-007).
文摘Objective Mycobacterium tuberculosis(Mtb),the causative agent of tuberculosis(TB),causes an estimated 1.6 million human deaths annually,but the pathogenesis of TB remains unclear.Immunity plays a critical role in the onset and outcome of TB.This study aimed to uncover the roles of innate and adaptive immunity in TB.Methods The gene expression profiles generated by RNA sequencing from human peripheral blood mononuclear cells(PBMCs)stimulated with or without Mtb strain H37Rv antigens were analyzed.A total of 973 differentially expressed mRNAs were identified.Results The differentially expressed genes were enriched in innate immunity signaling functions.The mesenchymal-epithelial transition factor(MET)gene was significantly upregulated in CD14^(+)monocytes.A MET inhibitor improved the uptake of the BCG strain by monocytes and macrophages as well as inhibited the expression of indoleamine 2,3-dioxygenase(IDO).The expression of IDO was increased in PBMCs stimulated with Mtb antigens,and the IDO inhibitor promoted the expression of CD40,CD83,and CD86.Conclusion Our results might provide clues regarding the immunomodulatory mechanisms used by Mtb to evade the host defense system.
文摘Over the past few decades, there has been a significant increase in the number of mycobacterial species described. Currently, the genus?Mycobacterium?consists of 170 species. Most species are called nontuberculous mycobacteria (NTM) and are potentially or rarely pathogenic and ubiquitous. One of the main challenges in mycobacteriology is the rapid and precise identification of these microorganisms. In this work, we compared two protein extraction protocols for the identification of 38 reference strains and clinical isolates, representing 27 species, by mass spectrometry (MALDI-TOF MS) to subsequently use the best method for identifying environmental mycobacteria. The results obtained with reference strains and clinical isolates showed that protocol A was effective in identifying 92.1% of mycobacterial specimens at the species level and protocol B, 50%. Therefore, protocol A was evaluated for the rapid identification of 27 environmental mycobacterial isolates. These isolates were subjected to PCR-restriction enzyme analysis (PRA-hsp65). Two isolates were misidentified by PRA-hsp65, whereas MALDI-TOF MS was able to identify them correctly. The results were confirmed by?hsp65 and 16S rRNA gene sequencing. Mass spectrometry has the advantage of being a simpler and faster technique than PRA-hsp65, and our results showed that MALDI-TOF MS is a valuable tool for the identification of environmental mycobacterial isolates.