Impact and Control of Sugar Size in Glycoconjugate Vaccines

Glycoconjugate vaccines have contributed enormously to reducing and controlling encapsulated bacterial infections for over thirty years. Glycoconjugate vaccines are based on a carbohydrate antigen that is covalently linked to a carrier protein; this is necessary to cause T cell responses for optimal immunogenicity, and to protect young children. Many interdependent parameters affect the immunogenicity of glycoconjugate vaccines, including the size of the saccharide antigen. Here, we examine and discuss the impact of glycan chain length on the efficacy of glycoconjugate vaccines and report the methods employed to size polysaccharide antigens, while highlighting the underlying reaction mechanisms. A better understanding of the impact of key parameters on the immunogenicity of glycoconjugates is critical to developing a new generation of highly effective vaccines

Platelet-Induced Clumping of Plasmodium falciparum–Infected Erythrocytes from Malawian Patients with Cerebral Malaria—Possible Modulation In Vivo by Thrombocytopenia

Platelets may play a role in the pathogenesis of human cerebral malaria (CM), and they have been shown to induce clumping of Plasmodium falciparum–parasitized red blood cells (PRBCs) in vitro. Both thrombocytopenia and platelet-inducedPRBCclumping are associated with severe malaria and, especially, withCM.In the present study, we investigated the occurrence of the clumping phenomenon in patients with CM by isolating and coincubating their plasma and PRBCs ex vivo. Malawian children with CM all had low platelet counts, with the degree of thrombocytopenia directly proportional to the density of parasitemia. Plasma samples obtained from these patients subsequently induced weak PRBC clumping. When the assays were repeated, with the plasma platelet concentrations adjusted to within the physiological range considered to be normal, massive clumping occurred. The results of this study suggest that thrombocytopenia may, through reduction of platelet-mediated clumping of PRBCs, provide a protective mechanism for the host during CM

Host defense against malaria favors Salmonella.

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Vaccines against invasive Salmonella disease: Current status and future directions

Though primarily enteric pathogens, Salmonellae are responsible for a considerable yet under-appreciated global burden of invasive disease. In South and South-East Asia, this manifests as enteric fever caused by serovars Typhi and Paratyphi A. In sub-Saharan Africa, a similar disease burden results from invasive nontyphoidal Salmonellae, principally serovars Typhimurium and Enteritidis. The existing Ty21a live-attenuated and Vi capsular polysaccharide vaccines target S. Typhi and are not effective in young children where the burden of invasive Salmonella disease is highest. After years of lack of investment in new Salmonella vaccines, recent times have seen increased interest in the area led by emerging-market manufacturers, global health vaccine institutes and academic partners. New glycoconjugate vaccines against S. Typhi are becoming available with similar vaccines against other invasive serovars in development. With other new vaccines under investigation, including live-attenuated, protein-based and GMMA vaccines, now is an exciting time for the Salmonella vaccine field. © 2014 Landes Bioscience

Frontiers in Shigella Vaccine Development

In recent years, there has been a resurgence of interest in the development of vaccines against Shigella driven by the growing awareness of the impact of this pathogen on global health [...

The Shigella Vaccines Pipeline

Shigella is the leading cause of global diarrheal deaths that currently lacks a licensed vaccine. Shigellosis drives antimicrobial resistance and leads to economic impact through linear growth faltering. Today, there is a robust pipeline of vaccines in clinical development which are broadly divided into parenteral glycoconjugate vaccines, consisting of O-antigen conjugated to carrier proteins, and oral live attenuated vaccines, which incorporate targeted genetic mutations seeking to optimize the balance between reactogenicity, immunogenicity and ultimately protection. Proof of efficacy has previously been shown with both approaches but for various reasons no vaccine has been licensed to date. In this report, we outline the requirements for a Shigella vaccine and describe the current pipeline in the context of the many candidates that have previously failed or been abandoned. The report refers to papers from individual vaccine developers in this special supplement of Vaccines which is focused on Shigella vaccines. Once readouts of safety and immunogenicity from current trials of lead candidate vaccines among the target population of young children in low- and middle-income countries are available, the likely time to licensure of a first Shigella vaccine will become clearer

Meningococcal X polysaccharide quantification by high-performance anion-exchange chromatography with pulsed amperometric detection using synthetic N-acetylglucosamine-4-phosphate as standard

A method for meningococcal X (MenX) polysaccharide quantification by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) is described. The polysaccharide is hydrolysed by strong acidic treatment and the peak of glucosamine-4-phosphate (4P-GlcN) detected and measured after chromatography. In the selected conditions of hydrolysis, 4P-GlcN is the prevalent species formed, with GlcN detected for less than 5% in moles. As standard for the analysis, the commercially unavailable monomeric unit of MenX polysaccharide, N-acetylglucosamine-4-phosphate (4P-GlcNAc), was chemically synthesized according to an improved procedure. The method for MenX quantification proposed here is highly selective and sensitive and constitutes an important analytical tool for the development of a conjugate vaccine against MenX

HIV-associated bactericidal and inhibitory immunity to Salmonella in Africans

It has been recognized for almost thirty years that nontyphoidal strains of Salmonella are a leading cause of death in HIV-infected Africans. The mechanisms underlying this susceptibility to invasive Salmonella disease have only recently started to be identified. Firstly, it became apparent that antibody-induced complement-mediated bacterial killing can protect healthy Africans adults and children by preventing the spread of the Salmonella through the blood. More recently we reported that in many HIV-infected African adults, antibodies against Salmonella lipopolysaccharide (LPS) block the antibody-induced complement-mediated killing of these bacteria, thus allowing the extracellular spread of infection. Here, we show that the blocking effect of anti-LPS antibodies is a factor of antibody concentration, rather than antibody structure or specificity. Thus, antibodies from HIV-infected subjects that block killing of Salmonella at plasma concentrations induce complement-mediated killing of Salmonella in diluted plasma. Equally, bactericidal anti-LPS antibodies of all isotypes from individuals not infected with HIV inhibit the killing of Salmonella when concentrated to levels grossly above those found in plasma. These findings indicate that underlying susceptibility of HIV-infected individuals to Salmonella, and potentially other pathogens, relates to hypergammaglobulinemia associated with impaired immune homeostasis

Modulation of endotoxicity of Shigella Generalized Modules for Membrane Antigens (GMMA) by genetic lipid A modifications: relative activation of TLR4 and TLR2 pathways in different mutants

Outer membrane particles from Gram-negative bacteria are attractive vaccine candidates as they present surface antigens in their natural environment and orientation. We previously developed a high yield production process for genetically derived particles, called Generalized Modules for Membrane Antigens (GMMA), from Shigella. As GMMA are derived from the outer membrane and contain immune-stimulatory components, especially lipopolysaccharide (LPS), we examined ways of reducing their reactogenicity by modifying lipid A, the endotoxic part of LPS, through deletion of late acyltransferase genes msbB or htrB in GMMA-producing S. sonnei and S. flexneri strains. GMMA with resulting penta-acylated lipid A from the msbB mutants showed a 600-fold reduction, GMMA from the S. sonnei ΔhtrB mutant a 60,000-fold reduced ability compared to GMMA with wild-type lipid A to stimulated human Toll-like receptor 4 (TLR4) in a reporter cell line. In contrast, in the S. flexneri ΔhtrB mutant, a compensatory palmitoleoylation occurs resulting in hexa-acylated lipid A with approximately 10-fold higher activity than the penta-acylated lipid A. In human PBMC, GMMA with penta-acylated lipid A showed a marked reduction in induction of inflammatory cytokines (800-fold for S. sonnei ΔhtrB, 300-fold for the msbB mutants) compared to a 50-fold reduction observed for GMMA with palmitoleoylated lipid A from the S. flexneri ΔhtrB strain. We demonstrated that the residual activity of GMMA with penta-acylated lipid A is largely due to non-lipid A related TLR2 activation whereas GMMA with palmitoleoylated hexa-acylated lipid A predominantly activate TLR4. These results identify the relative activation of TLR4 and TLR2 pathways by GMMA

A scalable method for O-antigen purification applied to different Salmonella serovars

The surface lipopolysaccharide of Gram-negative bacteria is both a virulence factor and a B-cell antigen. Antibodies against O-antigen of lipopolysaccharide may confer protection against infection, and O-antigen conjugates have been designed against multiple pathogens. Here, we describe simplified methodology for extraction and purification of O-antigen-core portion of Salmonella lipopolysaccharide, suitable for large scale production. Lipopolysaccharide extraction and delipidation is performed by acetic acid hydrolysis of whole bacterial culture, and can take place directly in a bioreactor, without previous isolation and inactivation of bacteria. Further O-antigen-core purification consists of rapid filtration and precipitation steps, without using enzymes or hazardous chemicals. The process was successfully applied to different Salmonella enterica serovars (Paratyphi A, Typhimurium and Enteritidis), obtaining good yields of high quality material, suitable for conjugate vaccines preparations