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Development of stable Vibrio cholerae O1 Hikojima type vaccine strains co-expressing the Inaba and Ogawa lipopolysaccharide antigens.

Karlsson SL, Ax E, Nygren E, Källgård S, Blomquist M, Ekman A, Benktander J, Holmgren J, Lebens M - PLoS ONE (2014)

Bottom Line: The strains express approximately equal amounts of Inaba- and Ogawa-LPS antigens which are preserved after formalin-inactivation of the bacteria.Oral immunizations of both inbred and outbred mice with formalin-inactivated whole-cell vaccine preparations of these strains elicited strong intestinal IgA anti-LPS as well as serum vibriocidal antibody responses against both Inaba and Ogawa that were fully comparable to the responses induced by the licensed Dukoral vaccine.This study illustrates the power of using genetic manipulation to improve the properties of bacteria strains for use in killed whole-cell vaccines.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology at Institute of Biomedicine, The Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden.

ABSTRACT
We describe here the development of stable classical and El Tor V. cholerae O1 strains of the Hikojima serotype that co-express the Inaba and Ogawa antigens of O1 lipopolysaccharide (LPS). Mutation of the wbeT gene reduced LPS perosamine methylation and thereby gave only partial transformation into Ogawa LPS on the cell surface. The strains express approximately equal amounts of Inaba- and Ogawa-LPS antigens which are preserved after formalin-inactivation of the bacteria. Oral immunizations of both inbred and outbred mice with formalin-inactivated whole-cell vaccine preparations of these strains elicited strong intestinal IgA anti-LPS as well as serum vibriocidal antibody responses against both Inaba and Ogawa that were fully comparable to the responses induced by the licensed Dukoral vaccine. Passive protection studies in infant mice showed that immune sera raised against either of the novel Hikojima vaccine strains protected baby mice against infection with virulent strains of both serotypes. This study illustrates the power of using genetic manipulation to improve the properties of bacteria strains for use in killed whole-cell vaccines.

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Determinations of the relative amounts of Ogawa antigen in LPS preparations of the generated Hikojima strains MS1568 and MS1580 based on amounts of methylated and un–methylated perosamine analyzed with mass spectrometry in relation to similar analyses of LPS preparations from reference Inaba Phil6973 and Ogawa Cairo 50 strains.The area under the curve for the methylated part (m/z ratio 756.6) divided by the area under the curve for the non–methylated part (m/z ratio 742.6) subtracted with the background (this ratio from the Inaba strain Phil6973) was used for the two Hikojima strain (MS1568 and MS1580) LPS preparations to calculate their percentage of Ogawa LPS (compared to the 100% Ogawa in the LPS from the Cairo50 reference strain). Diagrams are from one of 3 such experiments showing closely similar patterns and with the calculated mean ± SEM percentages Ogawa antigen indicated for MS1568 and MS1580.
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pone-0108521-g003: Determinations of the relative amounts of Ogawa antigen in LPS preparations of the generated Hikojima strains MS1568 and MS1580 based on amounts of methylated and un–methylated perosamine analyzed with mass spectrometry in relation to similar analyses of LPS preparations from reference Inaba Phil6973 and Ogawa Cairo 50 strains.The area under the curve for the methylated part (m/z ratio 756.6) divided by the area under the curve for the non–methylated part (m/z ratio 742.6) subtracted with the background (this ratio from the Inaba strain Phil6973) was used for the two Hikojima strain (MS1568 and MS1580) LPS preparations to calculate their percentage of Ogawa LPS (compared to the 100% Ogawa in the LPS from the Cairo50 reference strain). Diagrams are from one of 3 such experiments showing closely similar patterns and with the calculated mean ± SEM percentages Ogawa antigen indicated for MS1568 and MS1580.

Mentions: In order to quantify the relative amount of Ogawa antigen using a non–immunological method, LPS preparations from different Hikojima strains and from reference Inaba and Ogawa strains were analyzed using mass spectrometry. It is known that each Ogawa LPS molecule consists of a chain of perosamine residues in which the terminal perosamine is methylated. In contrast, Inaba LPS lacks any methylated perosamine [11]. By measuring the amount of specifically methylated and non–methylated perosamine in the LPS preparations we were able to calculate the relative amount of Ogawa LPS in MS1568 and MS1580. All samples were run in triplicate and gave 50±4.9% and 31.5±1.8% for each strain respectively (presented as mean ± SEM), see Figure 3.


Development of stable Vibrio cholerae O1 Hikojima type vaccine strains co-expressing the Inaba and Ogawa lipopolysaccharide antigens.

Karlsson SL, Ax E, Nygren E, Källgård S, Blomquist M, Ekman A, Benktander J, Holmgren J, Lebens M - PLoS ONE (2014)

Determinations of the relative amounts of Ogawa antigen in LPS preparations of the generated Hikojima strains MS1568 and MS1580 based on amounts of methylated and un–methylated perosamine analyzed with mass spectrometry in relation to similar analyses of LPS preparations from reference Inaba Phil6973 and Ogawa Cairo 50 strains.The area under the curve for the methylated part (m/z ratio 756.6) divided by the area under the curve for the non–methylated part (m/z ratio 742.6) subtracted with the background (this ratio from the Inaba strain Phil6973) was used for the two Hikojima strain (MS1568 and MS1580) LPS preparations to calculate their percentage of Ogawa LPS (compared to the 100% Ogawa in the LPS from the Cairo50 reference strain). Diagrams are from one of 3 such experiments showing closely similar patterns and with the calculated mean ± SEM percentages Ogawa antigen indicated for MS1568 and MS1580.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4232259&req=5

pone-0108521-g003: Determinations of the relative amounts of Ogawa antigen in LPS preparations of the generated Hikojima strains MS1568 and MS1580 based on amounts of methylated and un–methylated perosamine analyzed with mass spectrometry in relation to similar analyses of LPS preparations from reference Inaba Phil6973 and Ogawa Cairo 50 strains.The area under the curve for the methylated part (m/z ratio 756.6) divided by the area under the curve for the non–methylated part (m/z ratio 742.6) subtracted with the background (this ratio from the Inaba strain Phil6973) was used for the two Hikojima strain (MS1568 and MS1580) LPS preparations to calculate their percentage of Ogawa LPS (compared to the 100% Ogawa in the LPS from the Cairo50 reference strain). Diagrams are from one of 3 such experiments showing closely similar patterns and with the calculated mean ± SEM percentages Ogawa antigen indicated for MS1568 and MS1580.
Mentions: In order to quantify the relative amount of Ogawa antigen using a non–immunological method, LPS preparations from different Hikojima strains and from reference Inaba and Ogawa strains were analyzed using mass spectrometry. It is known that each Ogawa LPS molecule consists of a chain of perosamine residues in which the terminal perosamine is methylated. In contrast, Inaba LPS lacks any methylated perosamine [11]. By measuring the amount of specifically methylated and non–methylated perosamine in the LPS preparations we were able to calculate the relative amount of Ogawa LPS in MS1568 and MS1580. All samples were run in triplicate and gave 50±4.9% and 31.5±1.8% for each strain respectively (presented as mean ± SEM), see Figure 3.

Bottom Line: The strains express approximately equal amounts of Inaba- and Ogawa-LPS antigens which are preserved after formalin-inactivation of the bacteria.Oral immunizations of both inbred and outbred mice with formalin-inactivated whole-cell vaccine preparations of these strains elicited strong intestinal IgA anti-LPS as well as serum vibriocidal antibody responses against both Inaba and Ogawa that were fully comparable to the responses induced by the licensed Dukoral vaccine.This study illustrates the power of using genetic manipulation to improve the properties of bacteria strains for use in killed whole-cell vaccines.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology at Institute of Biomedicine, The Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden.

ABSTRACT
We describe here the development of stable classical and El Tor V. cholerae O1 strains of the Hikojima serotype that co-express the Inaba and Ogawa antigens of O1 lipopolysaccharide (LPS). Mutation of the wbeT gene reduced LPS perosamine methylation and thereby gave only partial transformation into Ogawa LPS on the cell surface. The strains express approximately equal amounts of Inaba- and Ogawa-LPS antigens which are preserved after formalin-inactivation of the bacteria. Oral immunizations of both inbred and outbred mice with formalin-inactivated whole-cell vaccine preparations of these strains elicited strong intestinal IgA anti-LPS as well as serum vibriocidal antibody responses against both Inaba and Ogawa that were fully comparable to the responses induced by the licensed Dukoral vaccine. Passive protection studies in infant mice showed that immune sera raised against either of the novel Hikojima vaccine strains protected baby mice against infection with virulent strains of both serotypes. This study illustrates the power of using genetic manipulation to improve the properties of bacteria strains for use in killed whole-cell vaccines.

Show MeSH
Related in: MedlinePlus