Impact of Methoxymycolic Acid Production by Mycobacterium bovis BCG Vaccines

Abstract

ABSTRACT BCG vaccines are a family of closely related daughter strains of an attenuated isolate of Mycobacterium bovis derived by in vitro passage from 1908 to 1921. During subsequent laboratory propagation of the vaccine strain until its lyophilization in 1961, BCG Pasteur underwent at least seven further genomic mutations. The impact of these mutations on the properties of the vaccine is currently unknown. One mutation, a glycine-to-aspartic acid substitution in the mmaA3 gene, occurred between 1927 and 1931 and impairs methoxymycolic acid synthesis in BCG strains obtained from the Pasteur Institute after this period. Mycolic acids of the cell wall are classified into three functional groups (alpha-, methoxy-, and ketomycolic acids), and together these lipids form a highly specialized permeability barrier around the bacterium. To explore the impact of methoxymycolic acid production by BCG strains, we complemented the functional gene of mmaA3 into BCG Denmark and tested a number of in vitro and in vivo phenotypes. Surprisingly, restoration of methoxymycolic acids alone had no effect on cell wall permeability, resistance to antibiotics, or growth in cultured macrophages and C57BL/6 mice. Our results demonstrate that the loss of methoxymycolic acid production did not apparently affect the virulence of BCG strains.