Mutation in mce operons attenuates Mycobacterium tuberculosis virulence.

ANDREA GIOFFRE; EDUARDO INFANTE; DIANA AGUILAR; MARIA DE LA PAZ SANTANGELO; LAURA KLEPP; ARIEL AMADIO; VIRGINIA MEIKLE; IGNACIO ETCHECHOURY; MARIA ISABEL ROMANO; CATALDI ANGEL; ROGELIO PANDO HERNANDEZ; FABIANA BIGI

MICROBES AND INFECTION

ELSEVIER SCIENCE BV

Lugar: Amsterdam; Año: 2005 vol. 7 p. 325 - 334

1286-4579

On the Mycobacterium tuberculosis genome there are four mce operons, all of which are similar in sequence and organization, and code for putatively exported proteins. To investigate whether Mce proteins are essential for virulence, we generated knock-out mutants in mce1, mce2 and mce3 operons of M. tuberculosis and evaluated their ability to multiply in a mammalian host. The allelic replacement was confirmed in each mutant strain by Southern blotting. RT-PCR experiments demonstrated the lack of in vitro expression of mutated genes in Dmce1 and Dmce2 mutants. On the other hand, no expression of mce3 was detected in either the wild-type or mutant strains. Similar doubling time and growth characteristics in in vitro culture were observed for mutants and parental strains. The intratracheal route was used to infect BALB/c mice with the Dmce3, Dmce2 and Dmce1 mutants. Ten weeks after infection, all mice infected with the Dmce mutants survived, while those infected with the wild-type strain died. This long survival correlated with very low counts of colony-forming units (CFU) in the lungs. Dmce1-infected mice developed very few and small granulomas, while animals infected with Dmce3 or Dmce2 mutants showed delayed granuloma formation. Mice infected with Dmce1 did not develop pneumonia, while animals infected with Dmce3 and Dmce2 mutants showed small pneumonic patches. In spleens, bacterial counts of mutant strains were less reduced than in lungs, compared with those of wild-type. In contrast, no such attenuation was observed when the intraperitoneal route was used for infection. Moreover, Dmce1 mutants appear to be more virulent in lungs after intraperitoneal inoculation. In conclusion, mce operons seem to affect the virulence of M. tuberculosis in mice, depend- ing on the route of infection. Hypotheses are discussed to explain this last issue. Thus, mutants in these genes seem to be good candidates for vaccine testing.