A Comparative Study of Outer Membrane Proteome between Paired Colistin-Susceptible and Extremely Colistin-Resistant Klebsiella pneumoniae Strains

In
the present report we characterized the outer membrane proteome, genomic,
and lipid A remodelling changes following the evolution of a colistin-susceptible K. pneumoniae ATCC 13883 strain into an extremely
colistin-resistant strain. Lipid A profiling revealed the outer membrane
of the colistin-susceptible strain is decorated primarily by hexa- and hepta-acylated lipid A species
and a minor tetra-acylated species. In the lipid
A profile of the extremely colistin-resistant strain, in addition
to the aforementioned lipid A species, the obligatory 4-amino-4-deoxy-l-arabinose modification of the hexa-acylated
lipid A was detected. Comparative genomic analysis revealed that the mgrB gene of the colistin-resistant strain is inactivated
by a single nucleotide insertion which produces a frame-shift, resulting
in premature termination. We also detected two synonymous mutations
in the two-component system genes phoP and phoQ. Comparative profiling of the outer membrane proteome
of each strain revealed that outer membrane proteins from bacterial
stress response, glutamine degradation, pyruvate, aspartate, and asparagine
metabolic pathways were over-represented in the extremely colistin-resistant K. pneumoniae ATCC 13883 strain. In comparison, in
the sensitive strain, outer membrane proteins from carbohydrate metabolism,
H+-ATPase, cell division, and peptidoglycan biosynthesis
were over-represented. Notably, there were no discernible differences
between the OmpK35 and OmpK36 major outer membrane porins between
the polymyxin-susceptible and -resistant strains suggesting porin
deficiency is not involved in the colistin resistance in the ATCC
13883 strain. These findings shed new light on the outer membrane
remodelling events accompanying the development of extremely high
levels of colistin resistance in K. pneumoniae.