Biology subjects

Felgner, P.

Publications and source records attributed to Felgner, P..

7 recordsLinked to original sources

bioRxiv2026

Novel pESI-encoded autotransporter adhesin PeaP of epidemic Salmonella strains mediates adhesion, atypical biofilm formation, and poultry colonization

Salmonella enterica serovar Infantis (SIN) has rapidly become the dominant serovar in poultry worldwide, a success largely linked to the acquisition of the 285 kb megaplasmid pESI. While pESI-encoded antibiotic-resistance and iron-uptake systems are well characterized, pESI-mediated adhesion mechanisms remain poorly understood. Here we identify a novel pESI-encoded monomeric autotransporter adhesin, designated PeaP (pESI-encoded autotransporter protein), and demonstrate its pivotal role in atypical biofilm formation, interference with motility, and colonization of the chicken host. Biofilm assays revealed that pESI-harboring strain SIN 119944 forms robust biofilms at 37 {degrees}C and 42 {degrees}C, temperatures at which CsgD-dependent biofilm formation is negligible. Deletion of csgD did not impair this phenotype, whereas deletion of peaP abolished high-temperature biofilm development and restored motility to wild-type levels. Proteomic profiling of sessile versus planktonic cells highlighted PeaP as the most abundant pESI-derived protein in the biofilm fraction. AlphaFold-based modelling and negative-stain transmission electron microscopy showed that PeaP comprises a C-terminal {beta}-barrel and a 1,500 aa passenger domain with three tandem repeats, projecting filamentous appendages [~]37-nm from the outer membrane. Antibody blockade of PeaP reduced surface adhesion >6-fold, confirming its adhesive function. In an infection model of 2 day-old chicken, the peaP mutant displayed significantly lower colonization, indicating PeaP-mediated adhesion in vivo. Collectively, pESI-positive SIN deploys PeaP for CsgD-independent, temperature-tolerant biofilm formation and enhanced gastrointestinal colonization, providing a mechanistic basis for the epidemic spread of this multidrug-resistant pathogen in poultry.

microbiology↗

bioRxiv2026

Crystal structure of E. coli Nissle 1917 flagellin reveals novel features that modulate bacterial motility but not TLR5 recognition

The probiotic E. coli Nissle 1917 (EcN) strain is known to promote intestinal homeostasis via flagellin, the protomer of its motility apparatus, the flagellum. The flagellin of EcN shows atypical features, namely a hypervariable region (HVR), whose structure and significance have remained elusive. We therefore determined the crystal structure of the E. coli Nissle 1917 flagellin FliC at a resolution of 1.2 [A] which revealed an unusual domain architecture: the canonical D1 domain was found connected by an extended linker to an extensive HVR whose D2, D3 and D4 domains form an outer domain (OD) which surrounds the filament core comprised of conserved domains D0-D1. Using both recombinant proteins and gene-edited EcN strains expressing mutant flagellins, the functional requirement for these unique features was subsequently studied for effects on immune recognition on intestinal epithelial and immune cells, as well as on flagellar protein expression, assembly and bacterial motility. While human and mouse TLR5 immune recognition of flagellar proteins or intact bacteria was only moderately affected by removal of linker or D4, especially linker removal reduced protein stability and bacterial motility in both soft agar and liquid media swimming assays. Interestingly, depending on the environment, D4 or HVR removal had different effects on motility and surface structure. Finally, a site-directed mutagenesis approach highlighted that loss of TLR5 recognition strictly entails loss of motility but not vice versa. Our data indicate that specific HVRs/OD might be relevant for motility of E. coli Nissle 1917 in specialized environments, but not for immune recognition. Moreover, we find mutational tolerance is greater for immune recognition than for motility, providing new insights into bacterial adaptation to the host environment.

microbiology↗

bioRxiv2023

Repeated Plasmodium vivax blood stage infection provides sterile protection against homologous challenge in non-human primates

The malaria parasite Plasmodium vivax remains a major global public health challenge, causing major morbidity across tropical and subtropical regions. Several candidate vaccines are in preclinical and clinical trials, however no vaccine against P. vivax malaria is approved for use in humans. Here we assessed whether P. vivax strain-transcendent immunity can be achieved by repeated infection in Aotus monkeys. For this purpose, we repeatedly infected six animals with blood stages of the P. vivax Salvador 1 (SAL-1) strain until sterile immune, and then challenged with the AMRU-1 strain. Sterile immunity was achieved in 4/4 Aotus monkeys after two homologous infections with the SAL-1 strain, while partial protection against a heterologous AMRU-1 challenge (i.e., delay to infection and reduction in peak parasitemia compared to control) was achieved in 3/3 monkeys. IgG levels based on P. vivax lysate ELISA and protein microarray increased with repeated infections and correlated with the level of homologous protection. Analysis of parasite transcriptional profiles across inoculation levels provided no evidence of major antigenic switching upon homologous or heterologous challenge. In contrast, we observed significant transcriptional differences in the P. vivax core gene repertoire between SAL-1 and AMRU-1. Together with the strain-specific genetic diversity between SAL-1 and AMRU-1 these data suggest that the partial protection upon heterologous challenge is due to molecular differences between strains (at genome and transcriptome level) rather than immune evasion by antigenic switching. Our study demonstrates that sterile immunity against P. vivax can be achieved by repeated homologous blood stage infection in Aotus monkeys, thus providing a benchmark to test the efficacy of candidate blood stage P. vivax malaria vaccines. Author summaryPlasmodium vivax is the most widespread human malaria parasite. Elimination efforts are complicated due to the peculiar biology of P. vivax including dormant liver forms, cryptic reservoirs in bone marrow and spleen and a large asymptomatic infectious reservoir in affected populations. Currently there is no vaccine against malaria caused by P. vivax. Here we induce sterile immunity by repeated P. vivax infection with the SAL-1 strain in non-human primates. In contrast, heterologous challenge with the AMRU-1 strain only provided partial protection. Antibody levels against a crude antigen and a protein microarray correlated with the level of homologous protection. Parasite transcriptional profiles across inoculation levels failed to show major antigenic switching across SAL-1 infections or upon heterologous challenge, instead suggesting other mechanisms of immune evasion. Our study demonstrates that sterile immunity against P. vivax can be achieved by repeated blood stage infection in Aotus monkeys, thus providing a benchmark to test the efficacy of candidate blood stage P. vivax malaria vaccines.

microbiology↗

bioRxiv2021

Cytolytic memory CD4+ T cell clonotypes are expanded during Plasmodium falciparum infection

Plasmodium falciparum (Pf) malaria causes high rates of morbidity and mortality and lacks a sufficiently effective vaccine. Clinical immunity develops in residents of malaria endemic regions which confers reduced clinical symptoms during infection and protection against severe disease. We hypothesized that understanding the immune mechanisms of clinical immunity could inform vaccine design to improve efficacy. We compared the peripheral blood cellular and humoral immune responses during a mild episode of Pf malaria infection. Participants were classified as either clinically susceptible or clinically protected, based on the number of recurrent clinical infections over an 18-month longitudinal study in a malaria endemic region in Malawi. Susceptible participants had three or more recurrent clinical episodes while clinically immune individuals had one or none. Protected participants exhibited higher plasma immunoglobulin G (IgG) breadth and titers against Pf antigens, and greater antibody (Ab)-dependent Pf opsonization compared to susceptible participants. Using high dimensional mass cytometry (CyTOF), spectral flow cytometry and single-cell transcriptomic analyses, we identified expanded memory CD4+ T cell clones sharing identical T cell receptor clonotypes in the blood of protected participants during malaria infection. These cells express a strong cytolytic T helper 1 effector program with transcripts encoding granzymes (A, B, H, M), granulysin, NKG7 and the Zeb2 master transcriptional regulator of terminally differentiated effector T cells. Memory CD4+ T cells expressing Zeb2+ were CD39hiTIGIThi and expressed multiple chemotactic and checkpoint inhibitory receptors, although the cellular levels of several of these receptors were reduced in protected compared to susceptible individuals. We propose that clonally expanded Zeb2+ cytolytic memory CD4+ Th1 cells could represent essential contributors to clinical immunity against Pf malaria. One Sentence SummaryA population of cytolytic memory CD4+ T cells is clonally expanded in patients with Plasmodium falciparum malaria and has reduced chemotactic and inhibitory receptor expression in patients with naturally acquired clinical malaria immunity.

immunology↗

bioRxiv2020

Comprehensive single cell analyses of the nutritional environment of intracellular Salmonella enterica

The facultative intracellular pathogen Salmonella enterica Typhimurium (STM) resides in a specific membrane-bound compartment termed the Salmonella-containing vacuole (SCV). STM is able to obtain all nutrients required for rapid proliferation, although being separated from direct access to host cell metabolites. The formation of specific tubular membrane compartment, called Salmonella-induced filaments (SIFs) are known to provides bacterial nutrition by giving STM access to endocytosed material and enabling proliferation. Additionally, STM expresses a range of nutrient uptake system for growth in nutrient limited environments to overcome the nutrition depletion inside the host. By utilizing dual fluorescence reporters, we shed light on the nutritional environment of intracellular STM in various host cells and distinct intracellular niches. We showed that STM uses nutrients of the host cell and adapts uniquely to the different nutrient conditions. In addition, we provide further evidence for improved nutrient supply by SIF formation or presence in the cytosol of epithelial cells, and the correlation of nutrient supply to bacterial proliferation.

microbiology↗

bioRxiv2020

A Live Attenuated Vaccine Model Confers Cross-Protective Immunity Against Different Species of Leptospira spp.

Leptospirosis is the leading zoonotic disease in terms of morbidity and mortality worldwide with a great health and economic impact in both humans and animals. Effective prevention is urgently needed as rapid urbanization, climate change and drivers of disease transmission continue to intensify. The key challenge has been developing a widely-applicable vaccine that protects against the 13 different pathogenic species and >300 serovars that can cause leptospirosis, providing a major public health benefit and opportunity to leverage One Health approaches. Live attenuated mutants that can boost immunity and induce protection are enticing vaccine candidates and poorly explored in the field. We evaluated a recently characterized motility-deficient mutant lacking the expression of a flagellar protein, FcpA. Although the fcpA- mutant has lost its ability to penetrate mucous membranes and cause disease, a transient bacteremia prior to clearance by the host immune response was observed. In two animal models, immunization with a single dose of the fcpA- mutant was sufficient to induce robust anti-protein antibodies response that promoted protection against infection with different Leptospira spp. species. Furthermore, characterization of the immune response identified a small repertoire of biologically relevant proteins that are highly conserved among pathogenic Leptospira species and potential correlates of cross-protective immunity.

microbiology↗

bioRxiv2018

Protective immunity against severe malaria is associated with antibodies to a conserved repertoire of PfEMP1 variants

Extreme diversity of the major surface antigen and virulence determinant of the malaria parasite Plasmodium falciparum, Erythrocyte Membrane Protein-1 (PfEMP1), poses a major barrier to identifying targets of protective immunity. To overcome this problem, we developed a PfEMP1 protein microarray containing 456 DBL domains, which was used to characterize the immunome of a cohort of semi-immune children and to identify variants associated with protective immune responses. Children with high mean antibody levels to DBL group 2 had a 26-36% reduced risk of uncomplicated (clinical) malaria, however only 8 diverse DBL variants were weakly associated with protection from clinical malaria and had low predictive accuracy. On the other hand, children with high mean antibodies to DBL groups 1 and 2 (which are markers for pathogenic \"Type A\" PfEMP1) and elevated antibodies to 85 (18.6%) of individual DBL variants had a 70 -100% reduced risk of severe malaria. Of the top 20 predictive variants for severe disease protection, 17 were strongly associated with protection (86 - 100% reduction in risk of severe malaria) and had high predictive accuracy for severe disease risk. Many variants were conserved and had highly correlated antibody responses, including the three highest-ranking variants, which were linked to EPCR-binding CIDR domains. The results suggest that while immunity to uncomplicated malaria is characterised by antibodies to a diverse repertoire of PfEMP1, immunity to severe malaria requires antibodies to a limited subset of antigenically conserved variants. These findings provide new insights into antimalarial immunity and potential biomarkers for tracking disease risk.

microbiology↗