Giuseppe
Emeritus
Regulated programmed lysis of recombinant Salmonella in host tissues to release protective antigens and confer biological containment
Wei Kong*,, Soo-Young Wanda*,, Xin Zhang,, Wendy Bollen,, Steven A. Tinge?, Kenneth L. Roland*, and Roy Curtiss, III*,,||
*Center for Infectious Diseases and Vaccinology, The Biodesign Institute and School of Life Sciences, Arizona State University, Tempe, AZ 85287-5401; Department of Biology, Washington University, St. Louis, MO 63130; and ?Megan Health, Inc., 3655 Vista Avenue, St. Louis, MO 63110
Contributed by Roy Curtiss III, April 21, 2008 (received for review March 6, 2008)
We have devised and constructed a biological containment system designed to cause programmed bacterial cell lysis with no survivors.
We have validated this system, using Salmonella enterica serovar Typhimurium vaccines for antigen delivery after colonization of host lymphoid tissues.
The system is composed of two parts.
The first component is Salmonella typhimurium strain 8937, with deletions of asdA and arabinose-regulated expression of murA, two genes required for peptidoglycan synthesis and additional mutations to enhance complete lysis and antigen delivery.
The second component is plasmid pYA3681, which encodes arabinose-regulated murA and asdA expression and C2-regulated synthesis of antisense asdA and murA mRNA transcribed from the P22 PR promoter.
An arabinose-regulated c2 gene is present in the chromosome. 8937(pYA3681) exhibits arabinose-dependent growth.
Upon invasion of host tissues, an arabinose-free environment, transcription of asdA, murA, and c2 ceases, and concentrations of their gene products decrease because of cell division.
The drop in C2 concentration results in activation of PR, driving synthesis of antisense mRNA to block translation of any residual asdA and murA mRNA.
A highly antigenic -helical domain of Streptococcus pneumoniae Rx1 PspA was cloned into pYA3681, resulting in pYA3685 to test antigen delivery.
Mice orally immunized with 8937(pYA3685) developed antibody responses to PspA and Salmonella outer membrane proteins.
No viable vaccine strain cells were detected in host tissues after 21 days.
This system has potential applications with other Gram-negative bacteria in which biological containment would be desirable.
programmed cell lysis | rPspA | Rx1
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Author contributions: W.K. and R.C. designed research; W.K., S.-Y.W., X.Z., W.B., and S.A.T. performed research; W.K. analyzed data; and W.K., K.L.R., and R.C. wrote the paper.
Present address: Department of Pathology and Immunology, Washington University, St. Louis, MO 63110.
Present address: Agnesian Healthcare, 430 East Division Street, Fond du Lac, WI 54935.
The authors declare no conflict of interest.
This article contains supporting information online at www.pnas.org/cgi/content/full/0803801105/DCSupplemental.
To whom correspondence should be addressed. E-mail: rcurtiss@asu.edu
? 2008 by The National Academy of Sciences of the USA
-
http://hwmaint.pnas.org/cgi/content/short/105/27/9361?rss=1
------
Wei Kong*,, Soo-Young Wanda*,, Xin Zhang,, Wendy Bollen,, Steven A. Tinge?, Kenneth L. Roland*, and Roy Curtiss, III*,,||
*Center for Infectious Diseases and Vaccinology, The Biodesign Institute and School of Life Sciences, Arizona State University, Tempe, AZ 85287-5401; Department of Biology, Washington University, St. Louis, MO 63130; and ?Megan Health, Inc., 3655 Vista Avenue, St. Louis, MO 63110
Contributed by Roy Curtiss III, April 21, 2008 (received for review March 6, 2008)
We have devised and constructed a biological containment system designed to cause programmed bacterial cell lysis with no survivors.
We have validated this system, using Salmonella enterica serovar Typhimurium vaccines for antigen delivery after colonization of host lymphoid tissues.
The system is composed of two parts.
The first component is Salmonella typhimurium strain 8937, with deletions of asdA and arabinose-regulated expression of murA, two genes required for peptidoglycan synthesis and additional mutations to enhance complete lysis and antigen delivery.
The second component is plasmid pYA3681, which encodes arabinose-regulated murA and asdA expression and C2-regulated synthesis of antisense asdA and murA mRNA transcribed from the P22 PR promoter.
An arabinose-regulated c2 gene is present in the chromosome. 8937(pYA3681) exhibits arabinose-dependent growth.
Upon invasion of host tissues, an arabinose-free environment, transcription of asdA, murA, and c2 ceases, and concentrations of their gene products decrease because of cell division.
The drop in C2 concentration results in activation of PR, driving synthesis of antisense mRNA to block translation of any residual asdA and murA mRNA.
A highly antigenic -helical domain of Streptococcus pneumoniae Rx1 PspA was cloned into pYA3681, resulting in pYA3685 to test antigen delivery.
Mice orally immunized with 8937(pYA3685) developed antibody responses to PspA and Salmonella outer membrane proteins.
No viable vaccine strain cells were detected in host tissues after 21 days.
This system has potential applications with other Gram-negative bacteria in which biological containment would be desirable.
programmed cell lysis | rPspA | Rx1
-
Author contributions: W.K. and R.C. designed research; W.K., S.-Y.W., X.Z., W.B., and S.A.T. performed research; W.K. analyzed data; and W.K., K.L.R., and R.C. wrote the paper.
Present address: Department of Pathology and Immunology, Washington University, St. Louis, MO 63110.
Present address: Agnesian Healthcare, 430 East Division Street, Fond du Lac, WI 54935.
The authors declare no conflict of interest.
This article contains supporting information online at www.pnas.org/cgi/content/full/0803801105/DCSupplemental.
To whom correspondence should be addressed. E-mail: rcurtiss@asu.edu
? 2008 by The National Academy of Sciences of the USA
-
http://hwmaint.pnas.org/cgi/content/short/105/27/9361?rss=1
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