Tuberculosis control relies on the identification and preventive treatment of individuals who are latently infected with Mycobacterium tuberculosis. However, direct identification of latent tuberculosis infection is not possible. The diagnostic tests used to identify individuals latently infected with M. tuberculosis, the in vivo tuberculin skin test and the ex vivo interferon-c release assays (IGRAs), are designed to identify an adaptive immune response against, but not necessarily a latent infection with, M. tuberculosis. The proportion of individuals who truly remain infected with M. tuberculosis after tuberculin skin test or IGRA conversion is unknown. It is also uncertain how long adaptive immune responses towards mycobacterial antigens persist in the absence of live mycobacteria. Clinical management and public healthcare policies for preventive chemotherapy against tuberculosis could be improved, if we were to gain a better understanding on M. tuberculosis latency and reactivation. This statement by the TBNET summarises knowledge and limitations of the currently available tests used in adults and children for the diagnosis of latent tuberculosis infection.In summary, the main issue regarding testing is to restrict it to those who are known to be at higher risk of developing tuberculosis and who are willing to accept preventive chemotherapy.

T-cell based IFN-γ release assays do not permit distinction of active tuberculosis (TB) from successfully treated disease or latent M. tuberculosis infection. We postulated that IFN-γ and IL-2 cytokine profiles of antigen-specific T cells measured by flow-cytometry ex vivo might correlate with TB disease activity in vivo. Tuberculin (PPD), ESAT-6 and CFP-10 were used as stimuli to determine antigen-specific cytokine profiles in CD4 T cells from 24 patients with active TB and 28 patients with successfully treated TB using flow-cytometry. Moreover, 25 individuals with immunity consistent with latent M. tuberculosis infection and BCG-vaccination, respectively, were recruited. Although the frequency of cytokine secreting PPD reactive CD4 T cells was higher in patients with active TB compared to patients with treated TB (median 0.81% vs. 0.39% of CD4 T cells, p = 0.02), the overlap in frequencies precluded distinction between the groups on an individual basis. When assessing cytokine profiles, PPD specific CD4 T cells secreting both IFN-γ and IL-2 predominated in treated TB, latent infection and BCG-vaccination, whilst in active TB the cytokine profile was shifted towards cells secreting IFN-γ only (p<0.0001). Cytokine profiles of ESAT-6 or CFP-10 reactive CD4 T cells did not differ between the groups. Receiver operator characteristics (ROC) analysis revealed that frequencies of PPD specific IFN-γ/IL-2 dual-positive T cells below 56% were an accurate marker for active TB (specificity 100%, sensitivity 70%) enabling effective discrimination from non-active states. In conclusion, a frequency lower than 56% IFN-γ/IL-2 dual positive PPD-specific circulating CD4 T-cells is strongly indicative of active TB.

Unlike the skin test, measurement of PPD reactivity by in vitro quantitation of PPD-specific T cells was unaffected by uremia-associated immunosuppression. This whole-blood assay may thus be a valuable alternative to skin testing, and detection of ESAT-6-specific T cells could moreover allow distinction of latent M. tuberculosis infection from BCG-induced reactivity to PPD. The assay is well suited for clinical use and may facilitate targeting of preventative therapy in high-risk individuals.

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The biological relevance of four iron-containing fractions previously detected in rat intestinal mucosal cells has been studied. The distribution of iron in these fractions obtained by chromatography on Sepharose 6B has been examined after in vivo and in vitro incubation of mucosal cells with 59Ce. In addition, the effects of phenobarbitone, cycloheximide, iron-deficiency and iron-loading on the uptake and distrubution of iron within the four mucosal cell fractions was studied. The iron in fraction I was mostly bound to intracellular membrane particles. Fraction II was shown to be ferritin. Fraction III contained some transferrin and also a protein of molecular weight similar to transferrin but which was not precipitable by antitransferrin antiserum. Quantited with the results of 'chaser' experiments suggested that, in addition to ferritin, at least two of the fractions (I and III) were involved in the process of iron absorption by the mucosal cell.

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