Klebsiella pneumoniae is responsible for nosocomial infections causing significant morbidity and mortality. Treatment of newly emerging multi-drug resistant strains is hampered due to severely limited antibiotic choices. Passive immunization targeting LPS O-antigens has been proposed as an alternative therapeutic option, given the limited variability of Klebsiella O-antigens. Here we report that the O3 serogroup, previously considered to have uniform O-antigen built of mannan, represents three different subtypes differing in the number of mannose residues within the O-antigen repeating units. Genetic analysis of the genes encoding mannose polymerization revealed differences that underline the observed structural alterations. The O3 variants represent antigenically different types based on the different reactivity pattern of murine monoclonal antibodies raised against a K. pneumoniae O3 strain. Typing of a collection of K. pneumoniae O3 clinical isolates showed that strains expressing the novel O3b antigen, the tri-mannose form, were more prevalent than those having the penta-mannose form, traditionally called O3, while the tetra-mannose variant, termed here O3a, seems to be rare. A monoclonal antibody cross-reacting with all three O3 sub-serogroups was also selected and shown to bind to the surface of various K. pneumoniae strains expressing different O3 subtypes and capsular antigens.

O-antigen, monoclonal antibodies, vaccines, Klebsiella pneumoniae, K.pneumoniae O3

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
2,2,2,3	aDManp	102.9	70.4	79.2	66.9	74.2	61.8
2,2,2	aDManp	102.9	70.4	78.8	66.9	74.2	61.9
2,2	aDManp	101.4	79.3	70.8	67.8	74.1	61.8
2	aDManp	101.4	79.5	70.8	67.8	74.1	61.8
	aDManp	101.4	79.5	70.8	67.7	74.1	61.8


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
2,2,2,3	aDManp	5.11	4.21	3.98	3.75	3.81	3.73-3.90
2,2,2	aDManp	5.03	4.21	3.93	3.75	3.79	3.73-3.88
2,2	aDManp	5.29	4.10	3.94	3.69	3.74	3.74-3.84
2	aDManp	5.29	4.08	3.94	3.69	3.73	3.74-3.84
	aDManp	5.37	4.07	3.98	3.68	3.78	3.74-3.89


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
2,2,2,3	aDManp	102.9/5.11	70.4/4.21	79.2/3.98	66.9/3.75	74.2/3.81	61.8/3.73-3.90
2,2,2	aDManp	102.9/5.03	70.4/4.21	78.8/3.93	66.9/3.75	74.2/3.79	61.9/3.73-3.88
2,2	aDManp	101.4/5.29	79.3/4.10	70.8/3.94	67.8/3.69	74.1/3.74	61.8/3.74-3.84
2	aDManp	101.4/5.29	79.5/4.08	70.8/3.94	67.8/3.69	74.1/3.73	61.8/3.74-3.84
	aDManp	101.4/5.37	79.5/4.07	70.8/3.98	67.7/3.68	74.1/3.78	61.8/3.74-3.89

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

[*]O[C@H]1[C@H](O)[C@@H](CO)O[C@H](O[C@@H]2[C@H](O)[C@@H](O[C@H]3[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]3O[C@H]3[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]3O[C@H]3[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]3[*])O[C@H](CO)[C@H]2O)[C@H]1O

810.705 g/mol (C₃₀H₅₀R₂O₂₅, R = next and previous repeats, not counted in mol weight)