Cytokine expression in lymph nodes from cattle inoculated intranasally with Mycobacterium bovis was compared to that of non-infected animals using real-time polymerase chain reaction. The effect of M. bovis infection, 4 months post-challenge, was to suppress the expression of anti-inflammatory cytokines interleukin (IL)-4 and IL-10 as well as the pro-inflammatory cytokines tumour necrosis factor (TNF) and IL-6. Expression of interferon (IFN)-γ and IL-12 was maintained. Animals vaccinated with bacille Calmette–Guérin responded differently to challenge with M. bovis. In particular, no decrease in expression of IL-4 or IL-6 was observed following challenge of vaccinated animals and decreased IFN-γ was detected. Also, vaccinated animals had higher levels of IL-4 and IL-10 transcripts compared to unvaccinated animals following challenge. These changes in cytokine expression levels led to a significant shift in the IFN-γ/IL-4 or IFN-γ/IL-10 ratio within the lymph node following challenge. Challenged animals generally showed a strong Th1 bias that was not seen in animals vaccinated prior to challenge. An inverse correlation between the level of pathology and bacterial load within the lymph node and the expression of IL-4, IL-10 and TNF was also observed. These results suggest that in the lymph nodes of cattle with established tuberculosis and a persisting bacterial infection, maintenance of the pro-inflammatory response in combination with a suppressed anti-inflammatory response may control the infection but contribute to host-induced tissue damage. Vaccination, which reduces the bacterial load and consequently the IFN-γ response, may result in less suppression of anti-inflammatory cytokines.