Differential in vivo expression of mycobacterial antigens in Mycobacterium tuberculosis infected lungs and lymph node tissues

BMC Infectious Diseases

Harald Gotten Wiker²  Lisbet Sviland¹  Nils Anders Leversen⁴  Tehmina Mustafa³ 

[1] Department of Pathology, Haukeland University Hospital, Bergen, Norway;Department of Microbiology, Haukeland University Hospital, Bergen, Norway;Department of Thoracic Medicine, Haukeland University Hospital, Bergen, Norway;Department of Clinical Science, University of Bergen, Bergen, Norway

关键词: MPT57;    Hsp65;    Mce1A;    MPT64;    MPT63;    MTP59;    MPT53;    MPT51;    MPT46;    MPT44;    MPT32;    Granuloma;    Pathology;    Intracellular infection;    Mycobacterium tuberculosis;   

Others  :  1125455
DOI  :  10.1186/1471-2334-14-535

 received in 2014-05-08, accepted in 2014-09-22,  发布年份 2014

【 摘 要 】

Background

The clinical course of tuberculosis (TB) infection, bacterial load and the morphology of lesions vary between pulmonary and extrapulmonary TB. Antigens expressed in abundance during infection could represent relevant antigens in the development of diagnostic tools, but little is known about the in vivo expression of various M. tuberculosis antigens in different clinical manifestations. The aim of this study was to study the differences in the presence of major secreted as well as somatic mycobacterial antigens in host tissues during advanced rapidly progressing and fatal pulmonary disease with mainly pneumonic infiltrates and high bacterial load, and to compare this to the presence of the same antigens in TB lymphadenitis cases, which is mainly chronic and self-limiting disease with organised granulomas and lower bacterial load.

Methods

Human pulmonary (n = 3) and lymph node (n = 17) TB biopsies, and non-TB controls (n = 12) were studied. Ziehl-Neelsen stain, nested PCR 1S6110 and immunohistochemistry were performed. Major secreted (MPT32, MPT44, MPT46, MPT51, MPT53, MPT59, MPT63, and MPT64) and somatic mycobacterial antigens (Mce1A, Hsp65, and MPT57) were detected by using rabbit polyclonal antibodies.

Results

Plenty of bacilli were detectable with Ziehl-Neelsen stain in the lung biopsies while no bacilli were detected in the lymph node biopsies. All the cases were shown to be positive by PCR. Both secretory and somatic antigens were expressed in abundance in pulmonary infiltrates, while primarily somatic antigens were detected in the lymphadenitis cases. Of the secreted antigens, only MPT64 was consistently detected in both cases, indicating a preferential accumulation of this antigen within the inflammatory cells, even if the cells of the granuloma can efficiently restrict bacterial growth and clear away the secreted antigens.

Conclusions

This study shows that major secreted mycobacterial antigens were found in high amounts in advanced pulmonary lesions without proper granuloma formation, while their level of staining was very low, or absent, in the lymph node TB lesions with organised granulomas and very low bacillary load, with one exception of MPT64, suggesting its role in the persistence of chronic infection. These findings have implication for development of new diagnostic tools.

【 授权许可】

   
2014 Mustafa et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]World Health Organization: Global tuberculosis report. Geneva, Switzerland: WHO Press; 2012.
• [2]Small PM, Schecter GF, Goodman PC, Sande MA, Chaisson RE, Hopewell PC: Treatment of tuberculosis in patients with advanced human immunodeficiency virus infection. N Engl J Med 1991, 324:289-294.
• [3]Clark RA, Blakley SL, Greer D, Smith MH, Brandon W, Wisniewski TL: Hematogenous dissemination of Mycobacterium tuberculosis in patients with AIDS. Rev Infect Dis 1991, 13:1089-1092.
• [4]Seibert AF, Haynes J Jr, Middleton R, Bass JB Jr: Tuberculous pleural effusion. Twenty-year experience. Chest 1991, 99:883-886.
• [5]Marais BJ, Pai M: Recent advances in the diagnosis of childhood tuberculosis. Arch Dis Child 2007, 92:446-452.
• [6]Peto HM, Pratt RH, Harrington TA, LoBue PA, Armstrong LR: Epidemiology of extrapulmonary tuberculosis in the United States, 1993–2006. Clin Infect Dis 2009, 49:1350-1357.
• [7]Wilsher ML, Hagan C, Prestidge R, Wells AU, Murison G: Human in vitro immune responses to Mycobacterium tuberculosis. Tuber Lung Dis 1999, 79:371-377.
• [8]Hussain R, Toossi Z, Hasan R, Jamil B, Dawood G, Ellner JJ: Immune response profile in patients with active tuberculosis in a BCG vaccinated area. Southeast Asian J Trop Med Public Health 1997, 28:764-773.
• [9]Emile JF, Patey N, Altare F, Lamhamedi S, Jouanguy E, Boman F, Quillard J, Lecomte-Houcke M, Verola O, Mousnier JF, Dijoud F, Blanche S, Fischer A, Brousse N, Casanova JL: Correlation of granuloma structure with clinical outcome defines two types of idiopathic disseminated BCG infection. J Pathol 1997, 181:25-30.
• [10]Barbolini G, Bisetti A, Colizzi V, Damiani G, Migaldi M, Vismara D: Immunohistologic analysis of mycobacterial antigens by monoclonal-antibodies in tuberculosis and mycobacteriosis. Hum Pathol 1989, 20:1078-1083.
• [11]Fenhalls G, Stevens L, Moses L, Bezuidenhout J, Betts JC, Helden Pv P, Lukey PT, Duncan K: In situ detection of Mycobacterium tuberculosis transcripts in human lung granulomas reveals differential gene expression in necrotic lesions. Infect Immun 2002, 70:6330-6338.
• [12]Fenhalls G, Stevens-Muller L, Warren R, Carroll N, Bezuidenhout J, van Helden P, Bardin P: Localisation of mycobacterial DNA and mRNA in human tuberculous granulomas. J Microbiol Methods 2002, 51:197-208.
• [13]Goel MM, Budhwar P: Immunohistochemical localization of mycobacterium tuberculosis complex antigen with antibody to 38 kDa antigen versus Ziehl Neelsen staining in tissue granulomas of extrapulmonary tuberculosis. Indian J Tuberc 2007, 54:24-29.
• [14]Andersen P: Host responses and antigens involved in protective immunity to Mycobacterium tuberculosis. Scand J Immunol 1997, 45:115-131.
• [15]Wiker HG, Harboe M, Lea TE: Purification and characterization of two protein antigens from the heterogeneous BCG85 complex in Mycobacterium bovis BCG. Int Arch Allergy Appl Immunol 1986, 81:298-306.
• [16]Målen H, Berven FS, Fladmark KE, Wiker HG: Comprehensive analysis of exported proteins from Mycobacterium tuberculosis. Proteomics 2007, 10:1702-1718.
• [17]Nagai S, Wiker HG, Harboe M, Kinomoto M: Isolation and partial characterization of major protein antigens in the culture fluid of Mycobacterium tuberculosis. Infect Immun 1991, 59:372-382.
• [18]de Souza GA, Malen H, Softeland T, Saelensminde G, Prasad S, Jonassen I, Wiker HG: High accuracy mass spectrometry analysis as a tool to verify and improve gene annotation using Mycobacterium tuberculosis as an example. BMC Genomics 2008, 9:316. BioMed Central Full Text
• [19]Harboe M, Malin AS, Dockrell HS, Wiker HG, Ulvund G, Holm A, Jørgensen MC, Andersen P: B-cell epitopes and quantification of the ESAT-6 protein of Mycobacterium tuberculosis. Infect Immun 1998, 66:717-723.
• [20]Wiker HG, Harboe M, Nagai S: A localization index for distinction between extracellular and intracellular antigens of M tuberculosis. J Gen Microbiol 1991, 137:875-884.
• [21]Baba K, Dyrhol-Riise AM, Sviland L, Langeland N, Hoosen AA, Wiker HG, Mustafa T: Rapid and specific diagnosis of tuberculous pleuritis with immunohistochemistry by detecting Mycobacterium tuberculosis complex specific antigen MPT64 in patients from a HIV endemic area. Appl Immunohistochem Mol Morphol 2008, 16:554-561.
• [22]Mustafa T, Wiker HG, Mfinanga SG, Morkve O, Sviland L: Immunohistochemistry using a Mycobacterium tuberculosis complex specific antibody for improved diagnosis of tuberculous lymphadenitis. Mod Pathol 2006, 19:1606-1614.
• [23]Purohit MR, Mustafa T, Wiker HG, Morkve O, Sviland L: Immunohistochemical diagnosis of abdominal and lymph node tuberculosis by detecting Mycobacterium tuberculosis complex specific antigen MPT64. Diagn Pathol 2007, 2:36. BioMed Central Full Text
• [24]Purohit MR, Mustafa T, Wiker HG, Sviland L: Rapid diagnosis of tuberculosis in aspirate, effusions, and cerebrospinal fluid by immunocytochemical detection of Mycobacterium tuberculosis complex specific antigen MPT64. Diagn Cytopathol 2012, 40:782-791.
• [25]Harboe M, Closs O, Deverill J: Production of monospecific antisera against antigenic components of M bovis (BCG). Scand J Immunol 1976, 5:861-866.
• [26]Nyka W: Studies on the effect of starvation on mycobacteria. Infect Immun 1974, 9:843-850.
• [27]Deb C, Lee CM, Dubey VS, Daniel J, Abomoelak B, Sirakova TD, Pawar S, Rogers L, Kolattukudy PE: A novel in vitro multiple-stress dormancy model for Mycobacterium tuberculosis generates a lipid-loaded, drug-tolerant, dormant pathogen. PLoS One 2009, 4:e6077.
• [28]Velayati AA, Farnia P, Masjedi MR, Zhavnerko GK, Merza MA, Ghanavi J, Tabarsi P, Farnia P, Poleschuyk NN, Ignatyev G: Sequential adaptation in latent tuberculosis bacilli: observation by atomic force microscopy (AFM). Int J Clin Exp Med 2011, 4:193-199.
• [29]Schnappinger D, Ehrt S, Voskuil MI, Liu Y, Mangan JA, Monahan IM, Dolganov G, Efron B, Butcher PD, Nathan C, Schoolnik GK: Transcriptional adaptation of mycobacterium tuberculosis within macrophages: insights into the phagosomal environment. J Exp Med 2003, 198:693-704.
• [30]Rohde KH, Abramovitch RB, Russell DG: Mycobacterium tuberculosis invasion of macrophages: linking bacterial gene expression to environmental cues. Cell Host Microbe 2007, 2:352-364.
• [31]Aagaard C, Hoang T, Dietrich J, Cardona PJ, Izzo A, Dolganov G, Schoolnik GK, Cassidy JP, Billeskov R, Andersen P: A multistage tuberculosis vaccine that confers efficient protection before and after exposure. Nat Med 2011, 17:189-194.
• [32]Alifano M, De Pascalis R, Sofia M, Faraone S, Del Pezzo M, Covelli I: Detection of IgG and IgA against the mycobacterial antigen A60 in patients with extrapulmonary tuberculosis. Thorax 1998, 53:377-380.
• [33]Abebe F, Holm-Hansen C, Wiker HG, Bjune G: Progress in serodiagnosis of Mycobacterium tuberculosis infection. Scand J Immunol 2007, 66:176-191.
• [34]Johnson S, Brusasca P, Lyashchenko K, Spencer JS, Wiker HG, Bifani P, Shashkina E, Kreiswirth B, Harboe M, Schluger N, Gomez M, Gennaro ML: Characterization of the secreted MPT53 antigen of Mycobacterium tuberculosis. Infect Immun 2001, 69:5936-5939.
• [35]Wang Z, Potter BM, Gray AM, Sacksteder KA, Geisbrecht BV, Laity JH: The solution structure of antigen MPT64 from Mycobacterium tuberculosis defines a new family of beta-grasp proteins. J Mol Biol 2007, 366:375-381.
• [36]Chu TP, Yuann JM: Expression, purification, and characterization of protective MPT64 antigen protein and identification of its multimers isolated from nontoxic Mycobacterium tuberculosis H37Ra. Biotechnol Appl Biochem 2011, 58:185-189.
• [37]Lima P, Sidders B, Morici L, Reader R, Senaratne R, Casali N, Riley LW: Enhanced mortality despite control of lung infection in mice aerogenically infected with a Mycobacterium tuberculosis mce1 operon mutant. Microbes Infect 2007, 9:1285-1290.
• [38]Shimono N, Morici L, Casali N, Cantrell S, Sidders B, Ehrt S, Riley LW: Hypervirulent mutant of Mycobacterium tuberculosis resulting from disruption of the mce1 operon. Proc Natl Acad Sci U S A 2003, 100:15918-15923.
• [39]Uchida Y, Casali N, White A, Morici L, Kendall LV, Riley LW: Accelerated immunopathological response of mice infected with Mycobacterium tuberculosis disrupted in the mce1 operon negative transcriptional regulator. Cell Microbiol 2007, 9:1275-1283.