scholarly journals Use of Lambda Phage S and R Gene Products in an Inducible Lysis System for Vibrio cholerae- and Salmonella enterica Serovar Typhimurium-Based DNA Vaccine Delivery Systems

2000 ◽  
Vol 68 (2) ◽  
pp. 986-989 ◽  
Author(s):  
Vivek Jain ◽  
John J. Mekalanos
Keyword(s):  
Vibrio Cholerae ◽  
Dna Vaccine ◽  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Dna Delivery ◽  
Lambda Phage ◽  
Regulated Expression ◽  
Mutant Strains ◽  
Serovar Typhimurium ◽  
Integrity Of Dna

ABSTRACT Novel methods for adapting DNA vaccine technology to the prevention of mucosal diseases are greatly needed. Here we show that regulated expression of phage lambda lysis genes S and R causes dramatic lysis of both Vibrio cholerae and Salmonella entericaserovar Typhimurium cells with concomitant release of plasmid DNA into the surrounding media. We also used single and double DNase mutant strains to show that secreted V. cholerae DNases can adversely affect the integrity of DNA molecules released upon lysis. These results suggest that incorporation of lambda SR-mediated lysis constructs and DNA stabilizing mutations into candidate live attenuated bacterial vaccines offers a promising approach for the development of effective mucosal DNA delivery vectors for humans.

scholarly journals Aspartic Peptide Hydrolases in Salmonella enterica Serovar Typhimurium

2001 ◽  
Vol 183 (10) ◽  
pp. 3089-3097 ◽  
Author(s):  
Rachel A. Larsen ◽  
Tina M. Knox ◽  
Charles G. Miller
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Gene Encoding ◽  
E Coli ◽  
Mutant Strains ◽  
Serovar Typhimurium ◽  
Measurable Rate ◽  
Mature Enzyme ◽  
Aminopeptidase A ◽  
Peptide Hydrolases

ABSTRACT Two well-characterized enzymes in Salmonella entericaserovar Typhimurium and Escherichia coli are able to hydrolyze N-terminal aspartyl (Asp) dipeptides: peptidase B, a broad-specificity aminopeptidase, and peptidase E, an Asp-specific dipeptidase. A serovar Typhimurium strain lacking both of these enzymes, however, can still utilize most N-terminal Asp dipeptides as sources of amino acids, and extracts of such a strain contain additional enzymatic activities able to hydrolyze Asp dipeptides. Here we report two such activities from extracts of pepB pepEmutant strains of serovar Typhimurium identified by their ability to hydrolyze Asp-Leu. Although each of these activities hydrolyzes Asp-Leu at a measurable rate, the preferred substrates for both are N-terminal isoAsp peptides. One of the activities is a previously characterized isoAsp dipeptidase from E. coli, the product of theiadA gene. The other is the product of the serovar Typhimurium homolog of E. coli ybiK, a gene of previously unknown function. This gene product is a member of the N-terminal nucleophile structural family of amidohydrolases. Like most other members of this family, the mature enzyme is generated from a precursor protein by proteolytic cleavage and the active enzyme is a heterotetramer. Based on its ability to hydrolyze an N-terminal isoAsp tripeptide as well as isoAsp dipeptides, the enzyme appears to be an isoAsp aminopeptidase, and we propose that the gene encoding it be designated iaaA (isoAsp aminopeptidase). A strain lacking both IadA and IaaA in addition to peptidase B and peptidase E has been constructed. This strain utilizes Asp-Leu as a leucine source, and extracts of this strain contain at least one additional, as-yet-uncharacterized, peptidase able to cleave Asp dipeptides.

scholarly journals Altering the Length of the Lipopolysaccharide O Antigen Has an Impact on the Interaction of Salmonella enterica Serovar Typhimurium with Macrophages and Complement

2006 ◽  
Vol 188 (7) ◽  
pp. 2735-2739 ◽  
Author(s):  
Gerald L. Murray ◽  
Stephen R. Attridge ◽  
Renato Morona
Keyword(s):  
Vibrio Cholerae ◽  
Salmonella Enterica ◽  
Double Mutant ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Shigella Flexneri ◽  
O Antigen ◽  
Repeat Units ◽  
Complement Resistance ◽  
Serovar Typhimurium ◽  
First Time

ABSTRACT A panel of isogenic Salmonella enterica serovar Typhimurium strains that vary only in the length of the O antigen was constructed through complementation of a wzz double mutant (displaying unregulated O-antigen length) with one of two homologous (wzz ST and wzz fepE) or three heterologous (wzz O139 of Vibrio cholerae and wzz SF and wzz pHS-2 of Shigella flexneri) wzz genes. Each gene was functional in the S. enterica serovar Typhimurium host and specified production of O-antigen polymers with lengths typical of those synthesized by the donor bacteria (ranging from 2 to >100 O-antigen repeat units). By use of this panel of strains, it was found that O-antigen length influences invasion/uptake by macrophage cells; this is the first time this has been shown with Salmonella. O-antigen length was confirmed to be related to complement resistance, with a minimum protective length of >4 and <15 repeat units. O antigen of 16 to 35 repeat units was found to activate complement more efficiently than other lengths, but this was unrelated to complement resistance. No evidence was found to suggest that modifying the length of the O-antigen polymer affected expression of the O1, O4, or O5 antigenic factors.

Immunological responses induced by asd and wzy/asd mutant strains of Salmonella enterica serovar Typhimurium in BALB/c mice

2010 ◽  
Vol 48 (4) ◽  
pp. 486-495 ◽  
Author(s):  
Hong Hua Piao ◽  
Vo Thi Minh Tam ◽  
Hee Sam Na ◽  
Hyun Ju Kim ◽  
Phil Youl Ryu ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Immunological Responses ◽  
Mutant Strains ◽  
Serovar Typhimurium

sopB of Salmonella enterica serovar Typhimurium is a potential DNA vaccine candidate in conjugation with live attenuated bacteria

Vaccine ◽  
2009 ◽  
Vol 27 (21) ◽  
pp. 2804-2811 ◽  
Author(s):  
Arvindhan G. Nagarajan ◽  
Sudhagar V. Balasundaram ◽  
Jessin Janice ◽  
Guruswamy Karnam ◽  
Sandeepa M. Eswarappa ◽  
...  
Keyword(s):  
Dna Vaccine ◽  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Vaccine Candidate ◽  
Serovar Typhimurium

scholarly journals Contribution of the PhoP/Q regulon to survival and replication of Salmonella enterica serovar Typhimurium in macrophages

Microbiology ◽  
2011 ◽  
Vol 157 (7) ◽  
pp. 2084-2093 ◽  
Author(s):  
Jessica A. Thompson ◽  
Mei Liu ◽  
Sophie Helaine ◽  
David W. Holden
Keyword(s):  
Mutant Strain ◽  
Salmonella Enterica ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Systemic Disease ◽  
Hydrolytic Enzymes ◽  
Regulatory System ◽  
Murine Macrophages ◽  
Mutant Strains ◽  
Serovar Typhimurium

The ability of serovars of Salmonella enterica to cause systemic disease is dependent upon their survival and replication within macrophages. To do this, bacteria must withstand or surmount bacteriostatic and bactericidal responses by the host cell, including the delivery of hydrolytic enzymes from lysosomes to the phagosome. The bacterial two-component regulatory system PhoP/Q has been implicated in avoidance of phagolysosomal fusion by S. enterica serovar Typhimurium (S. Typhimurium) in murine macrophages. In this study, the involvement of PhoP/Q-activated genes in avoidance of phagolysosomal fusion was analysed: of all the S. Typhimurium mutant strains tested, only an mgtC mutant strain partially reproduced the phenotype of the phoP mutant strain. As this gene is required for bacterial growth in magnesium-depleted conditions in vitro, the contributions of PhoP/Q to intramacrophage replication and survival were reappraised. Although PhoP/Q was required for both replication and survival of S. Typhimurium within murine macrophages, subsequent analysis of the kinetics of phagolysosomal fusion, taking account of differences in the replication rates of wild-type and phoP mutant strains, provided no evidence for a PhoP/Q-dependent role in this process. PhoP/Q appeared to act subsequent to the process of phagolysosomal avoidance and to promote replication of those bacteria that had already escaped a phagolysosomal fate. Therefore, we conclude that the PhoP/Q regulon enables S. Typhimurium to adapt to intramacrophage stresses other than phagolysosomal fusion.

scholarly journals The stm4066 Gene Product of Salmonella enterica Serovar Typhimurium Has Aminoimidazole Riboside (AIRs) Kinase Activity and Allows AIRs To Satisfy the Thiamine Requirement of pur Mutant Strains

2003 ◽  
Vol 185 (1) ◽  
pp. 332-339 ◽  
Author(s):  
Michael Dougherty ◽  
Diana M. Downs
Keyword(s):  
Gene Product ◽  
Salmonella Enterica ◽  
Kinase Activity ◽  
Salmonella Enterica Serovar Typhimurium ◽  
Mutant Strains ◽  
Serovar Typhimurium ◽  
Pyrimidine Moiety

ABSTRACT In bacteria the biosynthetic pathways for purine mononucleotides and the hydroxymethyl pyrimidine moiety of thiamine share five reactions that result in the formation of aminoimidazole ribotide, the last metabolite common to both pathways. Here we describe the characterization of a Salmonella enterica mutant strain that has gained the ability to efficiently use exogenous aminoimidazole riboside (AIRs) as a source of thiamine. The lesion responsible for this phenotype is a null mutation in a transcriptional regulator of the GntR family (encoded by stm4068). Lack of this protein derepressed transcription of an associated operon (stm4065-4067) that encoded a predicted kinase. The stm4066 gene product was purified and shown to have AIRs kinase activity in vitro. This activity was consistent with the model presented to explain the phenotype caused by the original mutation. This mutation provides a genetic means to isolate the synthesis of the hydroxymethyl pyrimidine moiety of thiamine from the pathway for purine mononucleotide biosynthesis and thus facilitate in vivo analyses.

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