group leader: Prof. Guzman

Recent Submissions

  • Large-scale production of megakaryocytes in microcarrier-supported stirred suspension bioreactors.

    Eicke, Dorothee; Baigger, Anja; Schulze, Kai; Latham, Sharissa L; Halloin, Caroline; Zweigerdt, Robert; Guzman, Carlos A; Blasczyk, Rainer; Figueiredo, Constança; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2018-07-05)
    Megakaryocytes (MKs) are the precursors of platelets (PLTs) and may be used for PLT production in vivo or in vitro, as well as a source for PLT-derived growth factors. Induced pluripotent stem cells represent an unlimited cell source for the in vitro production of MKs. This study aimed at developing an effective, xeno-free and scalable system to produce high numbers of MKs. In particular, microcarrier beads-assisted stirred bioreactors were evaluated as a means of improving MK yields. This method resulted in the production of 18.7 × 10
  • Induction of CD4(+) and CD8(+) anti-tumor effector T cell responses by bacteria mediated tumor therapy.

    Stern, Christian; Kasnitz, Nadine; Kocijancic, Dino; Trittel, Stephanie; Riese, Peggy; Guzman, Carlos A; Leschner, Sara; Weiss, Siegfried; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2015-10-15)
    Facultative anaerobic bacteria like E. coli can colonize solid tumors often resulting in tumor growth retardation or even clearance. Little mechanistic knowledge is available for this phenomenon which is however crucial for optimization and further implementation in the clinic. Here, we show that intravenous injections with E. coli TOP10 can induce clearance of CT26 tumors in BALB/c mice. Importantly, re-challenging mice which had cleared tumors showed that clearance was due to a specific immune reaction. Accordingly, lymphopenic mice never showed tumor clearance after infection. Depletion experiments revealed that during induction phase, CD8(+) T cells are the sole effectors responsible for tumor clearance while in the memory phase CD8(+) and CD4(+) T cells were involved. This was confirmed by adoptive transfer. CD4(+) and CD8(+) T cells could reject newly set tumors while CD8(+) T cells could even reject established tumors. Detailed analysis of adoptively transferred CD4(+) T cells during tumor challenge revealed expression of granzyme B, FasL, TNF-α and IFN-γ in such T cells that might be involved in the anti-tumor activity. Our findings should pave the way for further optimization steps of this promising therapy.
  • Self-Amplifying Replicon RNA Delivery to Dendritic Cells by Cationic Lipids

    Englezou, Pavlos C.; Sapet, Cedric; Démoulins, Thomas; Milona, Panagiota; Ebensen, Thomas; Schulze, Kai; Guzman, Carlos-Alberto; Poulhes, Florent; Zelphati, Olivier; Ruggli, Nicolas; McCullough, Kenneth C.; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany.
  • Engineered trivalent immunogen adjuvanted with a STING agonist confers protection against Trypanosoma cruzi infection.

    Sanchez Alberti, Andrés; Bivona, Augusto E; Cerny, Natacha; Schulze, Kai; Weißmann, Sebastian; Ebensen, Thomas; Morales, Celina; Padilla, Angel M; Cazorla, Silvia I; Tarleton, Rick L; Guzmán, Carlos A; Malchiodi, Emilio L; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017)
    The parasite Trypanosoma cruzi is the causative agent of Chagas disease, a potentially life-threatening infection that represents a major health problem in Latin America. Several characteristics of this protozoan contribute to the lack of an effective vaccine, among them: its silent invasion mechanism, T. cruzi antigen redundancy and immunodominance without protection. Taking into account these issues, we engineered Traspain, a chimeric antigen tailored to present a multivalent display of domains from key parasitic molecules, combined with stimulation of the STING pathway by c-di-AMP as a novel prophylactic strategy. This formulation proved to be effective for the priming of functional humoral responses and pathogen-specific CD8+ and CD4+ T cells, compatible with a Th1/Th17 bias. Interestingly, vaccine effectiveness assessed across the course of infection, showed a reduction in parasite load and chronic inflammation in different proof of concept assays. In conclusion, this approach represents a promising tool against parasitic chronic infections.
  • Targeted antigen delivery to dendritic cells elicits robust antiviral T cell-mediated immunity in the liver.

    Volckmar, Julia; Gereke, Marcus; Ebensen, Thomas; Riese, Peggy; Philipsen, Lars; Lienenklaus, Stefan; Wohlleber, Dirk; Klopfleisch, Robert; Stegemann-Koniszewski, Sabine; Müller, Andreas J; Gruber, Achim D; Knolle, Percy; Guzman, Carlos A; Bruder, Dunja; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr.7, 38124 Braunschweig, Germany. (2017-03-07)
    Hepatotropic viruses such as hepatitis C virus cause life-threatening chronic liver infections in millions of people worldwide. Targeted in vivo antigen-delivery to cross-presenting dendritic cells (DCs) has proven to be extraordinarily efficient in stimulating antigen-specific T cell responses. To determine whether this approach would as well be suitable to induce local antiviral effector T cells in the liver we compared different vaccine formulations based on either the targeting of DEC-205 or TLR2/6 on cross-presenting DCs or formulations not involving in vivo DC targeting. As read-outs we used in vivo hepatotropic adenovirus challenge, histology and automated multidimensional fluorescence microscopy (MELC). We show that targeted in vivo antigen delivery to cross-presenting DCs is highly effective in inducing antiviral CTLs capable of eliminating virus-infected hepatocytes, while control vaccine formulation not involving DC targeting failed to induce immunity against hepatotropic virus. Moreover, we observed distinct patterns of CD8+ T cell interaction with virus-infected and apoptotic hepatocytes in the two DC-targeting groups suggesting that the different vaccine formulations may stimulate distinct types of effector functions. Our findings represent an important step toward the future development of vaccines against hepatotropic viruses and the treatment of patients with hepatic virus infection after liver transplantation to avoid reinfection.
  • Superior immunogenicity of HCV envelope glycoproteins when adjuvanted with cyclic-di-AMP, a STING activator or archaeosomes.

    Landi, A; Law, J; Hockman, D; Logan, M; Crawford, K; Chen, C; Kundu, J; Ebensen, T; Guzman, C A; Deschatelets, L; Krishnan, L; Tyrrell, D L J; Houghton, M; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr.7, 38124 Braunschweig, Germany. (2017-12-15)
    Three decades after the discovery, hepatitis C virus (HCV) is still the leading cause of liver transplantation and poses a major threat to global health. In spite of recent advances in the development of direct acting antivirals, there is still a need for a prophylactic vaccine to limit the virus spread and protect at-risk populations, especially in developing countries, where the cost of the new treatments may severely limit access. The use of recombinant HCV glycoproteins E1E2 (rE1E2) in combination with the MF59, an oil-in-water emulsion-based adjuvant, has previously been shown to reduce the rate of chronicity in chimpanzees and to induce production of cross-neutralizing antibodies and cellular immune responses in human volunteers. To further improve neutralizing antibody responses in recipients along with robust T cell responses, we have explored the immunogenicity of different adjuvants when formulated with the HCV rE1E2 vaccine in mice. Our data show that cyclic di-adenosine monophosphate (c-di-AMP) and archaeosomes elicit strong neutralizing antibodies similar to those elicited using aluminum hydroxide/monophosphoryl lipid A (Alum/monophos. /MPLA) and MF59. However, both c-di-AMP and archaeosomes induced a more robust cellular immune response, which was confirmed by the detection of vaccine-specific poly-functional CD4+ T cells. We conclude that these adjuvants may substantially boost the immunogenicity of our E1E2 vaccine. In addition, our data also indicates that use of a partial or exclusive intranasal immunization regimen may also be feasible using c-di-AMP as adjuvant.
  • Mucosal Administration of Cycle-Di-Nucleotide-Adjuvanted Virosomes Efficiently Induces Protection against Influenza H5N1 in Mice.

    Ebensen, Thomas; Debarry, Jennifer; Pedersen, Gabriel K; Blazejewska, Paulina; Weissmann, Sebastian; Schulze, Kai; McCullough, Kenneth C; Cox, Rebecca J; Guzmán, Carlos A; Helmholtz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017)
    The need for more effective influenza vaccines is highlighted by the emergence of novel influenza strains, which can lead to new pandemics. There is a growing population of susceptible subjects at risk for severe complications of influenza, such as the elderly who are only in part protected by current licensed seasonal vaccines. One strategy for improving seasonal and pandemic vaccines takes advantage of adjuvants to boost and modulate evoked immune responses. In this study, we examined the capacity of the recently described adjuvant cyclic di-adenosine monophosphate (c-di-AMP) to serve as an adjuvant for improved mucosal influenza vaccines, and induce effective protection against influenza H5N1. In detail, c-di-AMP promoted (i) effective local and systemic humoral immune responses, including protective hemagglutination inhibition titers, (ii) effective cellular responses, including multifunctional T cell activity, (iii) induction of long-lasting immunity, and (iv) protection against viral challenge. Furthermore, we demonstrated the dose-sparing capacity of the adjuvant as well as the ability to evoke cross-clade protective immune responses. Overall, our results suggest that c-di-AMP contributes to the generation of a protective cell-mediated immune response required for efficacious vaccination against influenza, which supports the further development of c-di-AMP as an adjuvant for seasonal and pandemic influenza mucosal vaccines.
  • Roads to advanced vaccines: influenza case study.

    Riese, Peggy; Guzmán, Carlos A; Helmholz Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-09)
    Vaccines represent a cornerstone to ensure healthy lives and promote well-being for all at all ages. However, there are many diseases for which vaccines are not available, are relatively ineffective or need to be adapted periodically. Advances in microbial biotechnology will contribute to overcoming these roadblocks by laying the groundwork for improving and creating new approaches for developing better vaccines, as illustrated here in the case of influenza.
  • Bivalent mucosal peptide vaccines administered using the LCP carrier system stimulate protective immune responses against Streptococcus pyogenes infection.

    Schulze, Kai; Ebensen, Thomas; Chandrudu, Saranya; Skwarczynski, Mariusz; Toth, Istvan; Olive, Colleen; Guzman, Carlos A; Helmholtz Centre for infection researchGmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-09-05)
    Despite the broad knowledge about the pathogenicity of Streptococcus pyogenes there is still a controversy about the correlate of protection in GAS infections. We aimed in further improving the immune responses stimulated against GAS comparing different vaccine formulations including bis-(3',5')-cyclic dimeric adenosine monophosphate (c-di-AMP) and BPPCysMPEG, a derivative of the macrophage-activating lipopeptide (MALP-2), as adjuvants, respectively, to be administered with and without the universal T helper cell epitope P25 along with the optimized B cell epitope J14 of the M protein and B and T cell epitopes of SfbI. Lipopeptide based nano carrier systems (LCP) were used for efficient antigen delivery across the mucosal barrier. The stimulated immune responses were efficient in protecting mice against a respiratory challenge with a lethal dose of a heterologous S. pyogenes strain. Moreover, combination of the LCP based peptide vaccine with c-di-AMP allowed reduction of antigen dose at the same time maintaining vaccine efficacy.
  • Intranasal vaccination with an adjuvanted polyphosphazenes nanoparticle-based vaccine formulation stimulates protective immune responses in mice.

    Schulze, Kai; Ebensen, Thomas; Babiuk, Lorne A; Gerdts, Volker; Guzman, Carlos A.; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr.7, 38124 Braunschweig, Germany. (2017-06-01)
    The most promising strategy to sustainably prevent infectious diseases is vaccination. However, emerging as well as re-emerging diseases still constitute a considerable threat. Furthermore, lack of compliance and logistic constrains often result in the failure of vaccination campaigns. To overcome these hurdles, novel vaccination strategies need to be developed, which fulfill maximal safety requirements, show maximal efficiency and are easy to administer. Mucosal vaccines constitute promising non-invasive approaches able to match these demands. Here we demonstrate that nanoparticle (polyphosphazenes)-based vaccine formulations including c-di-AMP as adjuvant, cationic innate defense regulator peptides (IDR) and ovalbumin (OVA) as model antigen were able to stimulate strong humoral and cellular immune responses, which conferred protection against the OVA expressing influenza strain A/WSN/OVAI (H1N1). The presented results confirm the potency of nanoparticle-based vaccine formulations to deliver antigens across the mucosal barrier, but also demonstrate the necessity to include adjuvants to stimulate efficient antigen-specific immune responses.
  • Cell aggregation enhances bone formation by human mesenchymal stromal cells.

    Chatterjea, A; LaPointe, V L; Barradas, A; Garritsen, H; Yuan, H; Renard, A; van Blitterswijk, C A; de Beor, J; Hemholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Vraunschweig, Germany. (2017-02-15)
    The amount of bone generated using current tissue engineering approaches is insufficient for many clinical applications. Previous in vitro studies suggest that culturing cells as 3D aggregates can enhance their osteogenic potential, but the effect on bone formation in vivo is unknown. Here, we use agarose wells to generate uniformly sized mesenchymal stromal cell (MSC) aggregates. When combined with calcium phosphate ceramic particles and a gel prepared from human platelet-rich plasma, we generated a tissue engineered construct which significantly improved in vivo bone forming capacity as compared to the conventional system of using single cells seeded directly on the ceramic surface. Histology demonstrated the reproducibility of this system, which was tested using cells from four different donors. In vitro studies established that MSC aggregation results in an up-regulation of osteogenic transcripts. And finally, the in vivo performance of the constructs was significantly diminished when unaggregated cells were used, indicating that cell aggregation is a potent trigger of in vivo bone formation by MSCs. Cell aggregation could thus be used to improve bone tissue engineering strategies.
  • Type I IFN and not TNF, is Essential for Cyclic Di-nucleotide-elicited CTL by a Cytosolic Cross-presentation Pathway.

    Lirussi, Darío; Ebensen, Thomas; Schulze, Kai; Trittel, Stephanie; Duran, Veronica; Liebich, Ines; Kalinke, Ulrich; Guzmán, Carlos Alberto; Helmholtz Centre for infection research, Inhoffenstr.7, 38124 Braunschweig, Germany. (2017-07-19)
    Cyclic di-nucleotides (CDN) are potent stimulators of innate and adaptive immune responses. Cyclic di-AMP (CDA) is a promising adjuvant that generates humoral and cellular immunity. The strong STING-dependent stimulation of type I IFN represents a key feature of CDA. However, recent studies suggested that this is dispensable for adjuvanticity. Here we demonstrate that stimulation of IFN-γ-secreting CD8(+) cytotoxic T lymphocytes (CTL) is significantly decreased after vaccination in the absence of type I IFN signaling. The biological significance of this CTL response was confirmed by the stimulation of MHC class I-restricted protection against influenza virus challenge. We show here that type I IFN (and not TNF-α) is essential for CDA-mediated cross-presentation by a cathepsin independent, TAP and proteosome dependent cytosolic antigen processing pathway, which promotes effective cross-priming and further CTL induction. Our data clearly demonstrate that type I IFN signaling is critical for CDN-mediated cross-presentation.
  • Anti-nuclear autoantibodies in the general German population: prevalence and lack of association with selected cardiovascular and metabolic disorders-findings of a multicenter population-based study.

    Akmatov, Manas K; Röber, Nadja; Ahrens, Wolfgang; Flesch-Janys, Dieter; Fricke, Julia; Greiser, Halina; Günther, Kathrin; Kaaks, Rudolf; Kemmling, Yvonne; Krone, Bastian; Linseisen, Jakob; Meisinger, Christa; Moebus, Susanne; Obi, Nadia; Guzman, Carlos A; Conrad, Karsten; Pessler, Frank; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-06-06)
    We determined the prevalence of anti-nuclear autoantibodies (ANAs) in the German adult population and examined the association between ANAs and cardiovascular and metabolic disorders.
  • Gene Expression Driven by a Strong Viral Promoter in MVA Increases Vaccination Efficiency by Enhancing Antibody Responses and Unmasking CD8⁺ T Cell Epitopes.

    Becker, Pablo D; Nörder, Miriam; Weissmann, Sebastian; Ljapoci, Ronny; Erfle, Volker; Drexler, Ingo; Guzmán, Carlos Alberto; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2014-07-22)
    Viral vectors are promising tools for vaccination strategies and immunotherapies. However, CD8⁺ T cell responses against pathogen-derived epitopes are usually limited to dominant epitopes and antibody responses to recombinant encoded antigens (Ags) are mostly weak. We have previously demonstrated that the timing of viral Ag expression in infected professional Ag-presenting cells strongly shapes the epitope immunodominance hierarchy. T cells recognizing determinants derived from late viral proteins have a clear disadvantage to proliferate during secondary responses. In this work we evaluate the effect of overexpressing the recombinant Ag using the modified vaccinia virus early/late promoter H5 (mPH5). Although the Ag-expression from the natural promoter 7.5 (P7.5) and the mPH5 seemed similar, detailed analysis showed that mPH5 not only induces higher expression levels than P7.5 during early phase of infection, but also Ag turnover is enhanced. The strong overexpression during the early phase leads to broader CD8 T cell responses, while preserving the priming efficiency of stable Ags. Moreover, the increase in Ag-secretion favors the induction of strong antibody responses. Our findings provide the rationale to develop new strategies for fine-tuning the responses elicited by recombinant modified vaccinia virus Ankara by using selected promoters to improve the performance of this viral vector.
  • Blockade of Neutrophil's Chemokine Receptors CXCR1/2 Abrogate Liver Damage in Acute-on-Chronic Liver Failure.

    Khanam, Arshi; Trehanpati, Nirupma; Riese, Peggy; Rastogi, Archana; Guzman, Carlos A.; Sarin, Shiv Kumar; Helmholtz-Zentrum für Infektionsforschung GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017)
    Neutrophils serve as critical players in the pathogenesis of liver diseases. Chemokine receptors CXCR1 and CXCR2 are required for neutrophil chemotaxis to the site of inflammation/injury and are crucial in hepatic inflammatory response. However, key mechanism of neutrophil-mediated liver injury in acute-on-chronic liver failure (ACLF) remains highly elusive; which could be targeted for the development of new therapeutic interventions.
  • Immunization with Tc52 or its amino terminal domain adjuvanted with c-di-AMP induces Th17+Th1 specific immune responses and confers protection against Trypanosoma cruzi.

    Matos, Marina N; Cazorla, Silvia I; Schulze, Kai; Ebensen, Thomas; Guzmán, Carlos Alberto; Malchiodi, Emilio L; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-02)
    The development of new adjuvants enables fine modulation of the elicited immune responses. Ideally, the use of one or more adjuvants should result in the induction of a protective immune response against the specific pathogen. We have evaluated the immune response and protection against Trypanosoma cruzi infection in mice vaccinated with recombinant Tc52 or its N- and C-terminal domains (NTc52 and CTc52) adjuvanted either with the STING (Stimulator of Interferon Genes) agonist cyclic di-AMP (c-di-AMP), a pegylated derivative of α-galactosylceramide (αGC-PEG), or oligodeoxynucleotides containing unmethylated CpG motifs (ODN-CpG). All groups immunized with the recombinant proteins plus adjuvant: Tc52+c-di-AMP, NTc52+c-di-AMP, CTc52+c-di-AMP, NTc52+c-di-AMP+αGC-PEG, NTc52+CpG, developed significantly higher anti-Tc52 IgG titers than controls. Groups immunized with c-di-AMP and Tc52, NTc52 or CTc52 showed the highest Tc52-specific IgA titers in nasal lavages. All groups immunized with the recombinant proteins plus adjuvant developed a strong specific cellular immune response in splenocytes and lymph node cells with significant differences for groups immunized with c-di-AMP and Tc52, NTc52 or CTc52. These groups also showed high levels of Tc52-specific IL-17 and IFN-γ producing cells, while NTc52+CpG group only showed significant difference with control in IFN-γ producing cells. Groups immunized with c-di-AMP and Tc52, NTc52 or CTc52 developed predominantly a Th17 and Th1immune response, whereas for NTc52+CpG it was a dominant Th1 response. It was previously described that αGC-PEG inhibits Th17 differentiation by activating NKT cells. Thus, in this work we have also included a group immunized with both adjuvants (NTc52+c-di-AMP+αGC-PEG) with the aim to modulate the Th17 response induced by c-di-AMP. This group showed a significant reduction in the number of Tc52-specific IL-17 producing splenocytes, as compared to the group NTc52+c-di-AMP, which has in turn correlated with a reduction in protection against infection. These results suggest that the Th17 immune response developed after immunizing with NTc52+c-di-AMP could have a protective role against T. cruzi infection. Groups NTc52+c-di-AMP, Tc52+c-di-AMP and NTc52PB, were the ones that showed better protection against infection with lower parasitemia and weight loss, and higher survival.
  • Whole-Genome Enrichment Provides Deep Insights into Vibrio cholerae Metagenome from an African River.

    Vezzulli, L; Grande, C; Tassistro, G; Brettar, I; Höfle, M G; Pereira, R P A; Mushi, D; Pallavicini, A; Vassallo, P; Pruzzo, C; Helmholtz Centre for infection research, Ihoffenstr. 7, 38124 Braunschweig, Germany. (2016-11-25)
    The detection and typing of Vibrio cholerae in natural aquatic environments encounter major methodological challenges related to the fact that the bacterium is often present in environmental matrices at very low abundance in nonculturable state. This study applied, for the first time to our knowledge, a whole-genome enrichment (WGE) and next-generation sequencing (NGS) approach for direct genotyping and metagenomic analysis of low abundant V. cholerae DNA (<50 genome unit/L) from natural water collected in the Morogoro river (Tanzania). The protocol is based on the use of biotinylated RNA baits for target enrichment of V. cholerae metagenomic DNA via hybridization. An enriched V. cholerae metagenome library was generated and sequenced on an Illumina MiSeq platform. Up to 1.8 × 10(7) bp (4.5× mean read depth) were found to map against V. cholerae reference genome sequences representing an increase of about 2500 times in target DNA coverage compared to theoretical calculations of performance for shotgun metagenomics. Analysis of metagenomic data revealed the presence of several V. cholerae virulence and virulence associated genes in river water including major virulence regions (e.g. CTX prophage and Vibrio pathogenicity island-1) and genetic markers of epidemic strains (e.g. O1-antigen biosynthesis gene cluster) that were not detectable by standard culture and molecular techniques. Overall, besides providing a powerful tool for direct genotyping of V. cholerae in complex environmental matrices, this study provides a 'proof of concept' on the methodological gap that might currently preclude a more comprehensive understanding of toxigenic V. cholerae emergence from natural aquatic environments.
  • Immune modulator adamantylamide dipeptide stimulates efficient major histocompatibility complex class I-restricted responses in mice.

    Becker, Pablo D; Nörder, Miriam; Guzmán, Carlos Alberto; Grinstein, Saul; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2007-05)
    Adamantylamide L-alanyl-D-isoglutamine (AdDP) is a synthetic adjuvant which belongs to the family of the desmuramyl peptides. AdDP exerts its adjuvant properties when it is administered either by the parenteral or by the mucosal route, leading to the elicitation of strong humoral responses at both the systemic and the mucosal levels. However, very little is known about the effect of AdDP on cellular immunity. Here we demonstrate that AdDP is able to stimulate cellular responses, which are characterized by the release of gamma interferon by CD8+ T cells when they are restimulated with a major histocompatibility complex class I-restricted peptide and strong in vivo lymphocyte-mediated cytotoxic activity. The capacity of AdDP to stimulate the elicitation of both cellular and humoral adaptive responses makes this adjuvant a promising tool for the development of mucosal vaccine formulations.
  • Exploitation of prokaryotic expression systems based on the salicylate-dependent control circuit encompassing nahR/P(sal)::xylS2 for biotechnological applications.

    Becker, Pablo D; Royo, Jose L; Guzman, Carlos A; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2017-02-15)
    Expression vectors appear to be an indispensable tool for both biological studies and biotechnological applications. Controlling gene overexpression becomes a critical issue when protein production is desired. In addition to several aspects regarding toxicity or plasmid instability, tight control of gene expression is an essential factor in biotechnological processes. Thus, the search for better-controlled circuits is an important issue among biotechnologists. Traditionally, expression systems involve a single regulatory protein operating over a target promoter. However, these circuits are limited on their induction ratios (e.g., by their restriction in the maximal expression capacity, by their leakiness under non-induced conditions). Due to these limitations, regulatory cascades, which are far more efficient, are necessary for biotechnological applications. Thus, regulatory circuits with two modules operating in cascade offer a significant advantage. In this review, we describe the regulatory cascade based on two salicylate-responsive transcriptional regulators of Pseudomonas putida (nahR/P(sal)::xylS2), its properties, and contribution to a tighter control over heterologous gene expression in different applications.Nowadays, heterologous expression has been proven to be an indispensable tool for tackling basic biological questions, as well as for developing biotechnological applications. As the nature of the protein of interest becomes more complex, biotechnologists find that a tight control of gene expression is a key factor which conditions the success of the downstream purification process, as well as the interpretation of the results in other type of studies. Fortunately, different expression systems can be found in the market, each of them with their own pros and cons. In this review we discuss the exploitation of prokaryotic expression systems based on a promising expression system, the salicylate-dependent control circuit encompassing nahR/P(sal)::xylS2, as well as some of the improvements that have been done on this system to exploit it more efficiently in the context of both biotechnological applications and basic research.
  • The missing link: Bordetella petrii is endowed with both the metabolic versatility of environmental bacteria and virulence traits of pathogenic Bordetellae

    Gross, Roy; Guzman, Carlos A; Sebaihia, Mohammed; Martins dos Santos, Vítor A; Pieper, Dietmar H; Koebnik, Ralf; Lechner, Melanie; Bartels, Daniela; Buhrmester, Jens; Choudhuri, Jomuna V; Ebensen, Thomas; Gaigalat, Lars; Herrmann, Stefanie; Khachane, Amit N; Larisch, Christof; Link, Stefanie; Linke, Burkhard; Meyer, Folker; Mormann, Sascha; Nakunst, Diana; Rückert, Christian; Schneiker-Bekel, Susanne; Schulze, Kai; Vorhölter, Frank-Jörg; Yevsa, Tetyana; Engle, Jacquelyn T; Goldman, William E; Pühler, Alfred; Göbel, Ulf B; Goesmann, Alexander; Blöcker, Helmut; Kaiser, Olaf; Martinez-Arias, Rosa (2008-09-30)
    Abstract Background Bordetella petrii is the only environmental species hitherto found among the otherwise host-restricted and pathogenic members of the genus Bordetella. Phylogenetically, it connects the pathogenic Bordetellae and environmental bacteria of the genera Achromobacter and Alcaligenes, which are opportunistic pathogens. B. petrii strains have been isolated from very different environmental niches, including river sediment, polluted soil, marine sponges and a grass root. Recently, clinical isolates associated with bone degenerative disease or cystic fibrosis have also been described. Results In this manuscript we present the results of the analysis of the completely annotated genome sequence of the B. petrii strain DSMZ12804. B. petrii has a mosaic genome of 5,287,950 bp harboring numerous mobile genetic elements, including seven large genomic islands. Four of them are highly related to the clc element of Pseudomonas knackmussii B13, which encodes genes involved in the degradation of aromatics. Though being an environmental isolate, the sequenced B. petrii strain also encodes proteins related to virulence factors of the pathogenic Bordetellae, including the filamentous hemagglutinin, which is a major colonization factor of B. pertussis, and the master virulence regulator BvgAS. However, it lacks all known toxins of the pathogenic Bordetellae. Conclusion The genomic analysis suggests that B. petrii represents an evolutionary link between free-living environmental bacteria and the host-restricted obligate pathogenic Bordetellae. Its remarkable metabolic versatility may enable B. petrii to thrive in very different ecological niches.

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