The cytokine mRNA profiles of primary (arising from inhaled bacilli) and secondary (arising from hematogenous reseeding of the lung) granulomas from the lung lobes of BCG-vaccinated and unimmunized guinea pigs challenged with virulent M.tuberculosis by the pulmonary route were assessed in situ using laser capture microdissection (LCM) at six weeks post-infection. The challenge dose chosen was so low that some lung lobes did not to receive an implant from the airway. In unimmunized guinea pigs, some lobes contained either large, necrotic primary lesions or small, non-necrotic secondary lesions, or both. The lobes of BCG-vaccinated animals contained only non-necrotic primary tubercles, and no secondary lesions were visible. Real-time PCR analysis of the acquired RNA clearly demonstrated that primary tubercles from BCG-vaccinated guinea pigs were overwhelmed with mRNA from the anti-inflammatory cytokine, TGF, with some IFN and IL-12p40 mRNA. In contrast, primary lesions from unimmunized animals were dominated by pro-inflammatory TNF mRNA. The cytokine mRNA profile of secondary lesions from unimmunized animals was strikingly similar to the profile of primary lesions from BCG-vaccinated guinea pigs (i.e., a predominance of TGF mRNA with some IL-12p40 and IFN mRNA), indicating that the lung lobes from which these lesions were retrieved had been naturally "vaccinated" by the time the blood-borne bacilli returned to the lung at 3-4 weeks post-infection.