M. avium subsp. paratuberculosis is a very slow growing mycobacterial species, requiring 4-6 weeks of culture before colonies can be counted visually. This has seriously hampered experimental work with M. ptb. We have previously reported on a luminescent M. tuberculosis H37Rv isolate that was transformed with pSMT1 encoding the luxAB genes of Vibrio harveyi (1). With this luminescent isolate, fastidious and expensive enumeration of colony forming units (CFU) by plating on Middlebrook agar can be replaced by easy and inexpensive luminometry. Here we have used pSMT1 to transform two isolates of M. ptb i.e. ATCC 19698 and S23. C57BL/6 and mutant B6^bg/bg mice were infected intravenously with the luminescent M. ptb isolates and bacterial replication in the spleen was monitored over a period of 5 months by CFU plating and Relative Light Unit (RLU) detection. Our results show that luminometry can be used as an alternative for CFU counting. Luminometry was used to compare the protective efficacy of a gamma- irradiated M.ptb ATCC19698 vaccine in adjuvant and of a live attenuated M. bovis BCG vaccine. Luminescent M. ptb may be a valuable tool for the easy and rapid screening of experimental paratuberculosis vaccines in vivo.

Sponsorship

This work was partially supported by grants from the FWO-Vlaanderen (G.0266.00) and from Brussels Capital region. Va.Ro. holds a grant from DG6- (Former Federal Ministry of Agriculture, now Federal Public Service for Health, Food Chain Security and Environment). Vi.Ro. is a FRIA bursary.