Bacteriophages and phage polysaccharide-degrading enzymes (depolymerases) are garnering attention as possible alternatives to antibiotics. Here, we describe the antimicrobial properties of bacteriophage KpV74 and phage depolymerase Dep_kpv74 specific to the hypervirulent Klebsiella pneumoniae of the K2 capsular type. The depolymerase Dep_kpv74 was identified as a specific glucosidase that cleaved the K2 type capsular polysaccharides of the K. pneumoniae by a hydrolytic mechanism. This depolymerase was effective against thigh soft tissue K. pneumoniae infection in mice without inducing adverse behavioral effects or toxicity. The depolymerase efficiency was similar to or greater than the bacteriophage efficiency. The phage KpV74 had a therapeutic effect only for treating the infection caused by the phage-propagating K. pneumoniae strain and was completely inactive against the infection caused by the K. pneumoniae strain that did not support phage multiplication. The depolymerase was effective in both cases. A mutant resistant to phage and depolymerase was isolated during the treatment of mice with bacteriophage. A confirmed one-base deletion in the flippase-coding wzx gene of this mutant is assumed to affect the polysaccharide capsule, abolishing the KpV74 phage adsorption and reducing the K. pneumoniae virulence.

Klebsiella pneumoniae, bacteriophage, polysaccharide depolymerase, glucosidase, phage resistance, phage therapy

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
4,4	bDGlcp	103.7	73.5	84.0	71.6-71.7	77.6	62.2
4,3	aDGlcpA	102.8	72.8	74.7	72.8	73.0	?
4	bDManp	101.1	72.2	82.3	73.5	76.7	61.2
	aDGlcp	100.4	72.7	72.8	79.9	71.6-71.7	61.2


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
4,4	bDGlcp	4.52	3.37	3.62	3.52	3.52	3.72-3.96
4,3	aDGlcpA	5.18	3.54	3.79	3.55	4.25	-
4	bDManp	4.77	4.20	3.86	4.06	3.57	3.83-4.00
	aDGlcp	5.28	3.59	3.88	3.68	4.05	3.74-3.78


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
4,4	bDGlcp	103.7/4.52	73.5/3.37	84.0/3.62	71.6-71.7/3.52	77.6/3.52	62.2/3.72-3.96
4,3	aDGlcpA	102.8/5.18	72.8/3.54	74.7/3.79	72.8/3.55	73.0/4.25	
4	bDManp	101.1/4.77	72.2/4.20	82.3/3.86	73.5/4.06	76.7/3.57	61.2/3.83-4.00
	aDGlcp	100.4/5.28	72.7/3.59	72.8/3.88	79.9/3.68	71.6-71.7/4.05	61.2/3.74-3.78

The spectrum also has 1 signal at unknown position (not plotted).

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

662.547 g/mol (C₂₄H₃₈R₂O₂₁, R = next and previous repeats, not counted in mol weight)

Bacteriophages and phage polysaccharide-degrading enzymes (depolymerases) are garnering attention as possible alternatives to antibiotics. Here, we describe the antimicrobial properties of bacteriophage KpV74 and phage depolymerase Dep_kpv74 specific to the hypervirulent Klebsiella pneumoniae of the K2 capsular type. The depolymerase Dep_kpv74 was identified as a specific glucosidase that cleaved the K2 type capsular polysaccharides of the K. pneumoniae by a hydrolytic mechanism. This depolymerase was effective against thigh soft tissue K. pneumoniae infection in mice without inducing adverse behavioral effects or toxicity. The depolymerase efficiency was similar to or greater than the bacteriophage efficiency. The phage KpV74 had a therapeutic effect only for treating the infection caused by the phage-propagating K. pneumoniae strain and was completely inactive against the infection caused by the K. pneumoniae strain that did not support phage multiplication. The depolymerase was effective in both cases. A mutant resistant to phage and depolymerase was isolated during the treatment of mice with bacteriophage. A confirmed one-base deletion in the flippase-coding wzx gene of this mutant is assumed to affect the polysaccharide capsule, abolishing the KpV74 phage adsorption and reducing the K. pneumoniae virulence.

Klebsiella pneumoniae, bacteriophage, polysaccharide depolymerase, glucosidase, phage resistance, phage therapy

13C NMR data:
Linkage	Residue	C1	C2	C3	C4	C5	C6
3,4,3	aDGlcpA	101.3	72.9	74.0	73.1	73.5	?
3,4	bDManp	102.0	72.0	82.3	67.3	77.5	62.3
3	aDGlcp	100.2	72.9	72.9	80.0	71.6	61.1
	bDGlcp	97.3	74.2	83.7	71.4	77.0	62.0


1H NMR data:
Linkage	Residue	H1	H2	H3	H4	H5	H6
3,4,3	aDGlcpA	5.25	3.60	3.80	3.54	4.16	-
3,4	bDManp	4.77	4.18	3.76	3.77	3.46	3.75-3.93
3	aDGlcp	5.33	3.61	3.89	3.68	4.09	3.76-3.82
	bDGlcp	4.65	3.32	3.62	3.62	3.47	3.71-3.88


1H/13C HSQC data:
Linkage	Residue	C1/H1	C2/H2	C3/H3	C4/H4	C5/H5	C6/H6
3,4,3	aDGlcpA	101.3/5.25	72.9/3.60	74.0/3.80	73.1/3.54	73.5/4.16	
3,4	bDManp	102.0/4.77	72.0/4.18	82.3/3.76	67.3/3.77	77.5/3.46	62.3/3.75-3.93
3	aDGlcp	100.2/5.33	72.9/3.61	72.9/3.89	80.0/3.68	71.6/4.09	61.1/3.76-3.82
	bDGlcp	97.3/4.65	74.2/3.32	83.7/3.62	71.4/3.62	77.0/3.47	62.0/3.71-3.88

The spectrum also has 1 signal at unknown position (not plotted).

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

O=C(O)[C@H]1O[C@H](O[C@@H]2[C@H](O)[C@H](O[C@@H]3[C@@H](CO)O[C@H](O[C@@H]4[C@@H](O)[C@H](O)O[C@H](CO)[C@H]4O)[C@H](O)[C@H]3O)O[C@H](CO)[C@H]2O)[C@H](O)[C@@H](O)[C@@H]1O

680.562 g/mol (C₂₄H₄₀O₂₂)

Bacteriophages and phage polysaccharide-degrading enzymes (depolymerases) are garnering attention as possible alternatives to antibiotics. Here, we describe the antimicrobial properties of bacteriophage KpV74 and phage depolymerase Dep_kpv74 specific to the hypervirulent Klebsiella pneumoniae of the K2 capsular type. The depolymerase Dep_kpv74 was identified as a specific glucosidase that cleaved the K2 type capsular polysaccharides of the K. pneumoniae by a hydrolytic mechanism. This depolymerase was effective against thigh soft tissue K. pneumoniae infection in mice without inducing adverse behavioral effects or toxicity. The depolymerase efficiency was similar to or greater than the bacteriophage efficiency. The phage KpV74 had a therapeutic effect only for treating the infection caused by the phage-propagating K. pneumoniae strain and was completely inactive against the infection caused by the K. pneumoniae strain that did not support phage multiplication. The depolymerase was effective in both cases. A mutant resistant to phage and depolymerase was isolated during the treatment of mice with bacteriophage. A confirmed one-base deletion in the flippase-coding wzx gene of this mutant is assumed to affect the polysaccharide capsule, abolishing the KpV74 phage adsorption and reducing the K. pneumoniae virulence.

Klebsiella pneumoniae, bacteriophage, polysaccharide depolymerase, glucosidase, phage resistance, phage therapy

There is only one chemically distinct structure:

SVGMWSMILES3D molIonize

 

O=C(O)[C@H]1O[C@H](O[C@@H]2[C@H](O)[C@H](O[C@@H]3[C@@H](CO)O[C@H](O[C@@H]4[C@@H](O)[C@H](O[C@@H]5[C@@H](CO)O[C@@H](O[C@@H]6[C@@H](CO)O[C@H](O[C@@H]7[C@@H](O)C(O)O[C@H](CO)[C@H]7O)[C@H](O)[C@H]6O)[C@@H](O)[C@H]5O[C@H]5O[C@H](C(=O)O)[C@@H](O)[C@H](O)[C@H]5O)O[C@H](CO)[C@H]4O)[C@H](O)[C@H]3O)O[C@H](CO)[C@H]2O)[C@H](O)[C@@H](O)[C@@H]1O

1343.109 g/mol (C₄₈H₇₈O₄₃)