Immunology and Cell Biology (2002) 80, 544–549 Research Article Characterization of immune responses during infection with Mycobacterium avium strains 100, 101 and the recently sequenced 104 BERNADETTE M SAUNDERS, 1 , 2 ALISON DANE, 1 HELEN BRISCOE 1,2 and WARWICK J BRITTON 1,2 1 Centenary Institute of Cancer Medicine and Cell Biology, Newtown and 2 Department of Medicine, University of Sydney, New South Wales, Australia Summary Mycobacterium avium strain 104 was chosen as the M. avium isolate to sequence, as it is virulent to humans, stable and readily transfectable. As this strain has not been widely studied we sought to investigate the pattern of 104 infection in mice. Bacterial growth and the immune response generated were compared with infection with the low virulence M. avium strain 100, and the high virulence common laboratory strain, 101. Mycobacterium avium strains 104 and 101 grew progressively within mice, while strain 100 was gradually cleared. Strains 104 and 101 induced strong T cell activation and spleen cell cultures produced similar levels of IFN- γ. In mice infected with strain 100 no significant T cell activation or IFN- γ production was measured. Further, mice infected with strain 104 or 101 also displayed comparable inflammatory responses and similar granuloma formation, while only minimal inflammation was seen in mice infected with strain 100. Strains 101 and 104 also grew in a similar fashion in bone- marrow-derived macrophages and induced significant levels of TNF and nitric oxide. Thus infection with M. avium strain 104 induced an immunological response comparable to M. avium strain 101 and, with the availability of its sequence, should be a useful tool for designing new vaccines or drugs therapies to treat the increasing incidence of M. avium infection in humans. Key words: granuloma formation, Mycobacterium avium strains, T cell activation. Introduction Infection with Mycobacterium avium is a significant health problem among the HIV-infected population, 1 those with genetic deficiencies in IFN-γ and IL-12 production or function 2 and also among subsets of elderly woman, though no underlying immune deficiency has been identified in this population. 3,4 Development of a vaccine or more effective therapies for the treatment of M. avium infection is an ongoing concern. This research is hindered by the high degree of variability seen within the M. avium family. To date 28 M. avium serovars have been identified 5 with continent, country and regional variations in infective strains commonly reported. In Australia and the USA the major serovars isolated from AIDS patients are four and eight, but in Europe six, four and eight are more common. 6–9 Along with the multiple serovars found, variations in strain morphology and morphotype instability have also been problems. For instance M. avium strain 2–151 exhibits variable virulence in mice, with virulence correlating to morphological appearance on agar. Smooth transparent colonies have low virulence in mice, smooth opaque colonies are highly virulent in mice, while colonies with a rough appearance display intermediate virulence. Conversion from one morphotype, and associated virulence, to another is common, although the mechanism behind this is not well understood. 10–12 Investigations into the virulence mechanisms of M. avium will be aided by the sequencing of the bacterial genome. Due to the high number of serotypes identified and the degree of morphotype variation seen in M. avium, selecting an appro- priate strain to sequence was a complex problem. Mycobacte- rium avium strain 104 was chosen because it fulfilled key criteria: (i) it is pathogenic for humans, M. avium 104 (serovar 1) was originally isolated from an AIDS patient, (ii) the isolate is stable, unlike the commonly used laboratory 2–151 strains, and (iii) the bacteria are easily transfectable, unlike other common laboratory strains such as M. avium 101 (D. Russell personal communication). The full sequence of M. avium strain 104 should be available soon (http://www.tigr.org). However, studies detail- ing the virulence of this strain in mice and the immune responses initiated following infection are limited. Since mice are usually the first line of testing for potential new drugs and vaccines we undertook a characterization of the virulence of and immune responses to M. avium strain 104 in C57BL/6 mice. This response was compared with two other M. avium strains, 101, a common laboratory strain of M. avium, 13–15 and a low virulence strain M. avium 100. In all of the parameters tested, infection with M. avium strain 104 induced an essen- tially equivalent response to infection with M. avium strain 101. Both strains multiply in vivo, induce strong T cell responses, inflammatory cell recruitment and granuloma for- mation, and neither infection completely resolved during the Correspondence: Dr Bernadette M Saunders Centenary Institute of Cancer Medicine and Cell Biology, Locked Bag no. 6, Newtown, New South Wales 2042, Australia. Email: b.saunders@centenary.usyd.edu.au Received 30 April 2002; accepted 28 June 2002.