Impact of lgt mutation on lipoprotein biosynthesis and in vitro phenotypes of Streptococcus agalactiae

Abstract

Although Streptococcus agalactiae, the group B Streptococcus, is a leading cause of invasive neonatal disease worldwide the molecular basis of its virulence is still poorly understood. To investigate the role of lipoproteins in the physiology and interaction of this pathogen with host cells, we generated a mutant S. agalactiae strain (A909DLgt) deficient in the Lgt enzyme and thus unable to lipidate lipoprotein precursors (pro-lipoproteins). The loss of pro-lipoprotein lipidation did not affect the viability of S. agalactiae or its growth in several different media, including cationdepleted media. The processing of two well-characterized lipoproteins, but not a non-lipoprotein, was clearly shown to be aberrant in A909DLgt. The mutant strain was shown to be more sensitive to oxidative stress in vitro although the molecular basis of this increased sensitivity was not apparent. The inactivation of Lgt also resulted in changes to the bacterial cell envelope, as demonstrated by reduced retention of both the group B carbohydrate and the polysaccharide capsule and a statistically significant reduction (P50.0079) in A909DLgt adherence to human endothelial cells of fetal origin. These data confirm that failure to process lipoproteins correctly has pleiotropic effects that may be of significance to S. agalactiae colonization and pathogenesis.

The prolipoprotein diacylglyceryl transferase (Lgt) of Enterococcus faecalis contributes to virulence

“…However, an additional cross-reactive band of 100 kDa was observed in the Slgt mutant. Bray et al (2009) observed the same phenomenon and suggested that the band corresponding to the higher molecular mass represented aggregated forms of the prolipoproteins.…”

“…Indeed, the growth of the Slgt mutant was less affected by H 2 O 2 than that of the wild-type strain. Involvement of Lgt in oxidative stress has been reported for S. agalactiae (Bray et al, 2009), although in that study, the mutant appeared more sensitive to oxidative stress. Using RT-qPCR, we determined the expression of genes encoding particular lipoproteins [ef0063, ef0273, ef0577, ef0685, ef1234, ef1362, ef1534, ef2076 (efaA) and ef3082 (fatB)] during growth in GM17 supplemented with H 2 O 2 .…”

“…Loss of Lgt or Lsp leads to attenuation of virulence in Streptococcus pneumoniae (Petit et al, 2001), to a decrease in immune activation for group B Streptococcus, Staphylococcus aureus and Listeria monocytogenes (Henneke et al, 2008;Stoll et al, 2005;Machata et al, 2008), and to a reduction in adherence of Streptococcus agalactiae (Bray et al, 2009) and in intracellular growth of L. monocytogenes (Baumgärtner et al, 2007). While S. agalactiae and Staphylococcus aureus lgt mutants show increased sensitivity to oxidative stress (Bray et al, 2009) and growth attenuation in whole human blood (Stoll et al, 2005), respectively, both display a hypervirulent phenotype (Henneke et al, 2008;Bubeck Wardenburg et al, 2006). In E. faecalis, transport of lipoproteins across the membrane uses the Sec system, as no Tat mechanism is present and none of the 90 lipoproteins predicted in E. faecalis V583 harbours a Tat signal peptide (Reffuveille et al, 2011).…”

Lipoproteins of Gram-Positive Bacteria: Key Players in the Immune Response and Virulence

“…However, in many low-GC Gram-positive pathogens, mutations in either lgt or lsp also led to impaired pathogenicity. This was the case in various pathogenic streptococcal species such as Streptococcus pneumoniae (21,22), Streptococcus agalactiae (23,24), Streptococcus pyogenes, and Streptococcus equi (25,26). In Listeria monocytogenes, the lgt mutant was impaired in invasion and intracellular survival, was markedly attenuated in a mouse infection model, was unable to induce TLR2-dependent activation of NF-B, and exhibited increased susceptibility to cationic peptides (27,28).…”

Role of Pilus Proteins in Adherence and Invasion of Streptococcus agalactiae to the Lung and Cervical Epithelial Cells

“…Pilus protein gbs80-deficient, san1518-deficient, and gbs67-deficient mutants of S. agalactiae were generated by allelic replacement (24). The mutant for gbs80 was constructed in type Ia, III, V, and VII; san1518 was constructed in type Ia, III, and VII; and gbs67 was constructed in type V and VII.…”