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Background    Mycobacterium tuberculosis  ( Mtb ) infection may cause overt disease or remain latent. Interferon gamma release assays (IGRAs) detect  Mtb  infection, both latent infection and infection manifesting as overt disease, by measuring whole-blood interferon gamma (IFN-γ) responses to  Mtb  antigens such as early secreted antigenic target-6 (ESAT-6), culture filtrate protein 10 (CFP-10), and TB7.7. Due to a lack of adequate diagnostic standards for confirming latent  Mtb  infection, IGRA sensitivity for detecting  Mtb  infection has been estimated using patients with culture-confirmed tuberculosis (CCTB) for whom recovery of  Mtb  confirms the infection. In this study, cytokines in addition to IFN-γ were assessed for potential to provide robust measures of  Mtb  infection.    Methods  Cytokine responses to ESAT-6, CFP-10, TB7.7, or combinations of these  Mtb  antigens, for patients with CCTB were compared with responses for subjects at low risk for  Mtb  infection (controls). Three different multiplexed immunoassays were used to measure concentrations of 9 to 20 different cytokines. Responses were calculated by subtracting background cytokine concentrations from cytokine concentrations in plasma from blood stimulated with  Mtb  antigens.    Results  Two assays demonstrated that ESAT-6, CFP-10, ESAT-6+CFP-10, and ESAT-6+CFP-10+TB7.7 stimulated the release of significantly greater amounts of IFN-γ, IL-2, IL-8, MCP-1 and MIP-1β for CCTB patients than for controls. Responses to combination antigens were, or tended to be, greater than responses to individual antigens. A third assay, using whole blood stimulation with ESAT-6+CFP-10+TB7.7, revealed significantly greater IFN-γ, IL-2, IL-6, IL-8, IP-10, MCP-1, MIP-1β, and TNF-α responses among patients compared with controls. One CCTB patient with a falsely negative IFN-γ response had elevated responses with other cytokines.    Conclusions  Multiple cytokines are released when whole blood from patients with CCTB is stimulated with  Mtb  antigens. Measurement of multiple cytokine responses may improve diagnostic sensitivity for  Mtb  infection compared with assessment of IFN-γ alone.

Multiple Cytokines Are Released When Blood from Patients with Tuberculosis Is Stimulated with Mycobacterium tuberculosis Antigens