Roles for wbtC, wbtI, and kdtA Genes in Lipopolysaccharide Biosynthesis, Protein Glycosylation, Virulence, and Immunogenicity in Francisella tularensis Strain SCHU S4

Abstract

Using a strategy of gene deletion mutagenesis, we have examined the roles of genes putatively involved in lipopolysaccharide biosynthesis in the virulent facultative intracellular bacterial pathogen, Francisella tularensis subspecies tularensis, strain SCHU S4 in LPS biosynthesis, protein glycosylation, virulence and immunogenicity. One mutant, ΔwbtI, did not elaborate a long chain O-polysaccharide (OPS), was completely avirulent for mice, and failed to induce a protective immune response against challenge with wild type bacteria. Another mutant, ΔwbtC, produced a long chain OPS with altered chemical and electrophoretic characteristics. This mutant showed markedly reduced glycosylation of several known glycoproteins. Additionally this mutant was highly attenuated, and elicited a protective immune response against systemic, but not respiratory challenge with wild type SCHU S4. A third mutant, ΔkdtA, produced an unconjugated long chain OPS, lacking a detectable core structure, and which was not obviously expressed at the surface. It was avirulent and elicited partial protection against systemic challenge only.

From the Outside-In: The Francisella tularensis Envelope and Virulence

“…Follow-up studies have confirmed that Schu S4 wbtC and wbtI also play roles in LPS O-antigen synthesis, as both mutants had altered or missing LPS O-antigen, were attenuated in mice infected i.d., and did not protect against low dose Schu S4 aerosol challenge. In addition to its role in LPS O-antigen synthesis, wbtC also appears to play a role in protein glycosylation, as wbtC mutants displayed fewer glycosylated isoforms of both DsbA (see Outer Membrane Proteins Section) and PilA (see Type IV Pili Section), which previously were reported to be glycosylated in F. tularensis (Twine et al, 2012 ).…”

“…In agreement with other studies demonstrating the importance of LPS in virulence and immunogenicity, Schu S4 kdtA mutants were avirulent in mice via an i.d. infection route but were not protective against Schu S4 aerosol challenge (Twine et al, 2012 ).…”

FmvB: A Francisella tularensis Magnesium-Responsive Outer Membrane Protein that Plays a Role in Virulence

“…For the purposes of this study, we focused our attention on transcriptional changes linked to the bacterial envelope, including changes in inner membrane proteins, outer membrane proteins, LPS/capsule synthesis, stress responses, and genes involved in metal uptake/sensing ( Table 2 ). Interestingly, kdtA (required for core oligosaccharide synthesis;[ 63 ]) and wbtA (required for F . tularensis capsule and LPS production; [ 64 ]) were found to be upregulated 1.56- and 1.47-fold ( Table 2 ), respectively, in Δ fmvB , possibly explaining the 50% LPS increase in Δ fmvB ( Fig 5 ).…”

Mass Spectrometric Analysis of Natural and Antigen-Modified Strains of Tularemia Agent

“…вместе с тем в литературе мы встретили единственную работу по массспектрометрическому изучению аттенуированных мутантов штамма F. tularensis Schu4, дефектных по различным генам синтеза лпс. как было показано, такие штаммы имели либо идентичный с родительским белковый профиль, либо незначительное снижение высоты основных пиков в зависимости от локализации мутаций [13]. в нашей работе мы провели сравнительный анализ масс-спектров коллекции из 8 изогенных лпс-дефектных (с различными повреждениями структуры лпс) и родительских вирулентных штаммов трех основных подвидов.…”