Differentially regulated promoters for antigen expression in Salmonella vaccine strains

Stojanov, Miloš; Besançon, Hervé; Snäkä, Tiia; Nardelli-Haefliger, Denise; Curtiss, Roy; Baud, David

doi:10.1016/j.vaccine.2020.04.047

Cited by 2 publications

(3 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…We therefore tested infection of RAW246.7, and particularly human-monocytederived macrophages (hMDMs) as relevant target cells for Ty21a BLS strains. In contrast to the infection experiments with P asr -mRFP reporter fusions performed by others [43], in which Ty21a was unable to induce active infection of THP-1 derived macrophages, the strains constructed here with a similar reporter fusion infected and in some cases even multiplied in macrophages as was measured with a dual reporter system and live cell imaging (see Figure 6 and Supplementary Movie S1) [34]. As expected, bacterial fluorescence decreased during the time course of live cell imaging however (Supplementary Figure S9), which indicates that the majority of internalized Ty21a is degraded by the hMDM cells.…”

Section: Discussionmentioning

confidence: 82%

“…For an additional control strain, one FRT site flanking the KanR was deleted from this plasmid via In-Fusion to prevent KanR excision, resulting in the BLS-ActxB_B0 KanR variant after stabilization. For intracellular expression analysis, previously described in vivo inducible promoters P asr (acid induced [43]) and P pagC (low Mg 2+ [44]) were cloned upstream of mRFP using In-Fusion cloning (Takara Biotech) according to the manufacturer's protocol (see Figure 1). Promoter sequences were amplified from the Ty21a genome and ribosome binding sides were optimized with according primers using the UTR designer tool (https://sbi.postech.ac.kr/utr_designer/reverse/ accessed on 9 November 2022 [45]).…”

Section: Design and Construction Of The Psalvac Antigen Delivery Plas...mentioning

confidence: 99%

See 1 more Smart Citation

The JMU-SalVac-System: A Novel, Versatile Approach to Oral Live Vaccine Development

Iwanowitsch,

Diessner,

Bergmann

et al. 2024

Vaccines

View full text Add to dashboard Cite

Salmonella enterica Serovar Typhi Ty21a (Ty21a) is the only licensed oral vaccine against typhoid fever. Due to its excellent safety profile, it has been used as a promising vector strain for the expression of heterologous antigens for mucosal immunization. As the efficacy of any bacterial live vector vaccine correlates with its ability to express and present sufficient antigen, the genes for antigen expression are traditionally located on plasmids with antibiotic resistance genes for stabilization. However, for use in humans, antibiotic selection of plasmids is not applicable, leading to segregational loss of the antigen-producing plasmid. Therefore, we developed an oral Ty21a-based vaccine platform technology, the JMU-SalVac-system (Julius-Maximilians-Universität Würzburg) in which the antigen delivery plasmids (pSalVac-plasmid-series) are stabilized by a ΔtyrS/tyrS+-based balanced-lethal system (BLS). The system is made up of the chromosomal knockout of the essential tyrosyl-tRNA-synthetase gene (tyrS) and the in trans complementation of tyrS on the pSalVac-plasmid. Further novel functional features of the pSalVac-plasmids are the presence of two different expression cassettes for the expression of protein antigens. In this study, we present the construction of vaccine strains with BLS plasmids for antigen expression. The expression of cytosolic and secreted mRFP and cholera toxin subunit B (CTB) proteins as model antigens is used to demonstrate the versatility of the approach. As proof of concept, we show the induction of previously described in vivo inducible promoters cloned into pSalVac-plasmids during infection of primary macrophages and demonstrate the expression of model vaccine antigens in these relevant human target cells. Therefore, antigen delivery strains developed with the JMU-SalVac technology are promising, safe and stable vaccine strains to be used against mucosal infections in humans.

show abstract

Section: Discussionmentioning

confidence: 82%

Section: Design and Construction Of The Psalvac Antigen Delivery Plas...mentioning

confidence: 99%

The JMU-SalVac-System: A Novel, Versatile Approach to Oral Live Vaccine Development

Iwanowitsch,

Diessner,

Bergmann

et al. 2024

Vaccines

View full text Add to dashboard Cite

show abstract

“…Compared with the other fluorescence reporter L. monocytogenes strains in the previous studies (Fortineau et al 2000;Freitag and Jacobs 1999), the fluorescence reporter L. monocytogenes strain based on P 18 exhibited better functionality in vivo. During vaccine development based on microbial carrier, selecting an appropriate promoter is the key to optimize the expressions of foreign antigens (Chapman and Rybicki 2019;Galvin et al 2000;Stojanov et al 2020). In this study, the selected constitutive promoters P 18 , P 7 , P 12 , P 9 , and P 24 with different strengths exhibited high activities during the production of the specific antigen UreB.…”

Section: Discussionmentioning

confidence: 96%

Identification and evaluation of a panel of strong constitutive promoters in Listeria monocytogenes for improving the expression of foreign antigens

Wang

et al. 2021

Appl Microbiol Biotechnol

View full text Add to dashboard Cite

Attenuated Listeria monocytogenes could be a potential vaccine vector for the immunotherapy of tumors or pathogens. However, the lack of reliable promoters has limited its ability to express foreign antigens. In the present study, 21 promoters were identified from Listeria monocytogenes through RNA-seq analysis under two pH conditions of pH 7.4 and pH 5.5. Based on the constructed fluorescence report system, 7 constitutive promoters exhibited higher strength than P help (1.8-fold to 5.4-fold), a previously reported strong promoter. Furthermore, the selected 5 constitutive promoters exhibited higher UreB production activity than P help (1.1-fold to 8.3-fold). Notably, a well-characterized constitutive promoter P 18 was found with the highest activity of fluorescence intensity and UreB production. In summary, the study provides a panel of strong constitutive promoters for Listeria monocytogenes and offers a theoretical basis for mining constitutive promoters in other organisms. Key points• Twenty-one promoters were identified from L. monocytogenes through RNA-seq.• Fluorescent tracer of L. monocytogenes (P 18 ) was performed in vitro and in vivo. • A well-characterized constitutive promoter P 18 could improve the expression level of a foreign antigen UreB in L. monocytogenes Keywords L. monocytogenes . Constitutive promoter . RNA-seq . Acid stress * Qing Liu

show abstract

Differentially regulated promoters for antigen expression in Salmonella vaccine strains

Cited by 2 publications

References 34 publications

The JMU-SalVac-System: A Novel, Versatile Approach to Oral Live Vaccine Development

The JMU-SalVac-System: A Novel, Versatile Approach to Oral Live Vaccine Development

Identification and evaluation of a panel of strong constitutive promoters in Listeria monocytogenes for improving the expression of foreign antigens

Contact Info

Product

Resources

About