Second language learners often produce language forms resembling those of children with Specific Language Impairment (SLI). At present, professionals working in language assessment and education have ...only limited diagnostic instruments to distinguish language impaired migrant children from those who will eventually catch up with their monolingual peers. This book presents a comprehensive set of tools for assessing the linguistic abilities of bilingual children. It aims to disentangle effects of bilingualism from those of SLI, making use of both models of bilingualism and models of language impairment. The book’s methods-oriented focus will make it an essential handbook for practitioners who look for measures which could be adapted to a variety of languages in diverse communities, as well as academic researchers.
Vaccines aim to efficiently and specifically activate the immune system
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a cascade of antigen uptake, processing, and presentation by antigen-presenting cells (APCs) to CD4 and CD8 T cells, which ...in turn drive humoral and cellular immune responses. The specific formulation of vaccine carriers can not only shield the antigens from premature sequestering before reaching APCs but also favorably promote intracellular antigen presentation and processing. This study compares two different acid-degradable polymeric nanoparticles that are capable of encapsulating a moderately immunogenic antigen, GFP, at nearly full efficacy
via
electrostatic interactions or molecular affinity between His tag and Ni-NTA-conjugated monomners. This resulted in GFP-encapsulating NPs composed of ketal monomers and crosslinkers (KMX/GFP NPs) and NTA-conjugated ketal monomers and crosslinkers (NKMX/GFP NPs), respectively. Encapsulated GFP was found to be released more rapidly from NKMX/GFP NPs (electrostatic encapsulation) than from KMX/GFP NPs (affinity-driven encapsulation).
In vivo
vaccination studies demonstrated that while repeated injections of either NP formulation resulted in poorer generation of anti-GFP antibodies than injections of the GFP antigen itself, sequential injections of NPs and GFP as prime and booster vaccines, respectively, restored the humoral response. We proposed that NPs primarily assist APCs in antigen presentation by T cells, and B cells need to be further stimulated by free protein antigens to produce antibodies. The findings of this study suggest that the immune response can be modulated by varying the chemistry of vaccine carriers and the sequences of vaccination with free antigens and antigen-encapsulating NPs.
Acid-degradable polymeric nanoparticles with a high capability of GFP encapsulation demonstrate efficient antibody production in combination with booster injection of free antigens.
The effects of adjuvants for increasing the immunogenicity of influenza vaccines are well known. However, the effect of adjuvants on increasing the breadth of cross-reactivity is less well ...understood. In this study we have performed a systematic screen of different toll-like receptor (TLR) agonists, with and without a squalene-in-water emulsion on the immunogenicity of a recombinant trimerized hemagglutinin (HA) vaccine in mice after single-dose administration. Antibody (Ab) cross-reactivity for other variants within and outside the immunizing subtype (homosubtypic and heterosubtypic cross-reactivity, respectively) was assessed using a protein microarray approach. Most adjuvants induced broad IgG profiles, although the response to a combination of CpG, MPLA and AddaVax (termed 'IVAX-1') appeared more quickly and reached a greater magnitude than the other formulations tested. Antigen-specific plasma cell labeling experiments show the components of IVAX-1 are synergistic. This adjuvant preferentially stimulates CD4 T cells to produce Th1>Th2 type (IgG2c>IgG1) antibodies and cytokine responses. Moreover, IVAX-1 induces identical homo- and heterosubtypic IgG and IgA cross-reactivity profiles when administered intranasally. Consistent with these observations, a single-cell transcriptomics analysis demonstrated significant increases in expression of IgG1, IgG2b and IgG2c genes of B cells in H5/IVAX-1 immunized mice relative to naïve mice, as well as significant increases in expression of the IFNγ gene of both CD4 and CD8 T cells. These data support the use of adjuvants for enhancing the breath and durability of antibody responses of influenza virus vaccines.
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Sustained signaling through the B cell antigen receptor (BCR) is thought to occur only when antigen(s) crosslink or disperse multiple BCR units, such as by multimeric antigens found on the surfaces ...of viruses or bacteria. B cell-intrinsic Toll-like receptor (TLR) signaling synergizes with the BCR to induce and shape antibody production, hallmarked by immunoglobulin (Ig) class switch recombination (CSR) of constant heavy chains from IgM/IgD to IgG, IgA or IgE isotypes, and somatic hypermutation (SHM) of variable heavy and light chains. Full B cell differentiation is essential for protective immunity, where class switched high affinity antibodies neutralize present pathogens, memory B cells are held in reserve for future encounters, and activated B cells also serve as semi-professional APCs for T cells. But the rules that fine-tune B cell differentiation remain partially understood, despite their being essential for naturally acquired immunity and for guiding vaccine development. To address this in part, we have developed a cell culture system using splenic B cells from naive mice stimulated with several biotinylated ligands and antibodies crosslinked by streptavidin reagents. In particular, biotinylated lipopolysaccharide (LPS), a Toll-like receptor 4 (TLR4) agonist, and biotinylated anti-IgM were pre-assembled (multimerized) using streptavidin, or immobilized on nanoparticles coated with streptavidin, and used to active B cells in this precisely controlled, high throughput assay. Using B cell proliferation and Ig class switching as metrics for successful B cell activation, we show that the stimuli are both synergistic and dose-dependent. Crucially, the multimerized immunoconjugates are most active over a narrow concentration range. These data suggest that multimericity is an essential requirement for B cell BCR/TLRs ligands, and clarify basic rules for B cell activation. Such studies highlight the importance in determining the choice of single vs multimeric formats of antigen and PAMP agonists during vaccine design and development.
Combining variant antigens into a multivalent vaccine is a traditional approach used to provide broad coverage against antigenically variable pathogens, such as polio, human papilloma and influenza ...viruses. However, strategies for increasing the breadth of antibody coverage beyond the vaccine are not well understood, but may provide more anticipatory protection. Influenza virus hemagglutinin (HA) is a prototypic variant antigen. Vaccines that induce HA-specific neutralizing antibodies lose efficacy as amino acid substitutions accumulate in neutralizing epitopes during influenza virus evolution. Here we studied the effect of a potent combination adjuvant (CpG/MPLA/squalene-in-water emulsion) on the breadth and maturation of the antibody response to a representative variant of HA subtypes H1, H5 and H7. Using HA protein microarrays and antigen-specific B cell labelling, we show when administered individually, each HA elicits a cross-reactive antibody profile for multiple variants within the same subtype and other closely-related subtypes (homosubtypic and heterosubtypic cross-reactivity, respectively). Despite a capacity for each subtype to induce heterosubtypic cross-reactivity, broader coverage was elicited by simply combining the subtypes into a multivalent vaccine. Importantly, multiplexing did not compromise antibody avidity or affinity maturation to the individual HA constituents. The use of adjuvants to increase the breadth of antibody coverage beyond the vaccine antigens may help future-proof vaccines against newly-emerging variants.
Vaccines are among the most cost-effective public health measures for controlling infectious diseases.
is the etiological agent of Q fever, a disease with a wide clinical spectrum that ranges from ...mild symptoms, such as fever and fatigue, to more severe disease, such as pneumonia and endocarditis. The formalin-inactivated whole-cell vaccine Q-VAX
contains hundreds of antigens and confers lifelong protection in humans, but prior sensitization from infection or vaccination can result in deleterious reactogenic responses to vaccination. Consequently, there is great interest in developing non-reactogenic alternatives based on adjuvanted recombinant proteins. In this study, we aimed to develop a multivalent vaccine that conferred protection with reduced reactogenicity. We hypothesized that a multivalent vaccine consisting of multiple antigens would be more immunogenic and protective than a monovalent vaccine owing to the large number of potential protective antigens in the
proteome. To address this, we identified immunogenic T and B cell antigens, and selected proteins were purified to evaluate with a combination adjuvant (IVAX-1), with or without
lipopolysaccharide (LPS) in immunogenicity studies
in mice and in a Hartley guinea pig intratracheal aerosol challenge model using
strain NMI RSA 493. The data showed that multivalent vaccines are more immunogenic than monovalent vaccines and more closely emulate the protection achieved by Q-VAX. Although six antigens were the most immunogenic, we also discovered that multiplexing beyond four antigens introduces detectable reactogenicity, indicating that there is an upper limit to the number of antigens that can be safely included in a multivalent Q-fever vaccine.
LPS also demonstrates efficacy as a vaccine antigen in conferring protection in an otherwise monovalent vaccine formulation, suggesting that its addition in multivalent vaccines, as demonstrated by a quadrivalent formulation, would improve protective responses.
Q fever is caused by the obligate intracellular bacterium,
, a designated potential agent of bioterrorism because of its route of transmission, resistance to disinfectants, and low infectious dose. ...The only vaccine licensed for human use is Q-VAX
(Seqirus, licensed in Australia), a formalin-inactivated whole-cell vaccine, which produces severe local and systemic reactogenic responses in previously sensitized individuals. Accordingly, the U.S. Food and Drug Administration and other regulatory bodies around the world, have been reluctant to approve Q-VAX for widespread use. To obviate these adverse reactions, we prepared recombinant protein subunit vaccine candidates containing purified CBU1910, CBU0307, CBU0545, CBU0612, CBU0891, and CBU1398 proteins and TLR triagonist adjuvants. TLR triagonist adjuvants combine different TLR agonists to enhance immune responses to vaccine antigens. We tested both the protective efficacy and reactogenicity of our vaccine candidates in Hartley guinea pigs using intratracheal infection with live
. While all of our candidates showed varying degrees of protection during challenge, local reactogenic responses were significantly reduced for one of our vaccine candidates when compared with a formalin-inactivated whole-cell vaccine. Our findings show that subunit vaccines combined with novel TLR triagonist adjuvants can generate protective immunity to
infection while reducing reactogenic responses.
Auranofin, a reprofiled FDA-approved drug originally designed to treat rheumatoid arthritis, has emerged as a promising anti-parasitic drug. It induces the accumulation of reactive oxygen species ...(ROS) in parasites, including
. We generated auranofin resistant
lines through chemical mutagenesis to identify the molecular target of this drug. Resistant clones were confirmed with a competition assay using wild-type
expressing yellow fluorescence protein (YFP) as a reference strain. The predicted auranofin target, thioredoxin reductase, was not mutated in any of our resistant lines. Subsequent whole genomic sequencing analysis (WGS) did not reveal a consensus resistance locus, although many have point mutations in genes encoding redox-relevant proteins such as superoxide dismutase (TgSOD2) and ribonucleotide reductase. We investigated the SOD2 L201P mutation and found that it was not sufficient to confer resistance when introduced into wild-type parasites. Resistant clones accumulated less ROS than their wild type counterparts. Our results demonstrate that resistance to auranofin in
enhances its ability to abate oxidative stress through diverse mechanisms. This evidence supports a hypothesized mechanism of auranofin anti-parasitic activity as disruption of redox homeostasis.
The arrival in 2015 and 2016 of over one million asylum seekers and refugees in Germany had major social consequences and gave rise to extensive debates about the nature of cultural diversity and ...collective life. This volume examines the responses and implications of what was widely seen as the most significant and contested social change since German reunification in 1990. It combines in-depth studies based on anthropological fieldwork with analyses of the longer trajectories of migration and social change. Its original conclusions have significance not only for Germany but also for the understanding of diversity and difference more widely.
Coxiella burnetii is an intracellular Gram-negative bacterium responsible for Q fever, a zoonotic disease with significant global health implications courtesy of its endemic status worldwide. This ...tier 2 select agent has the potential for use as a bioweapon due to low infectious dose and aerosol transmissibility. The development of a vaccine against C. burnetii is of great interest, particularly for people who are in close contact with ruminants and military personnel who may be exposed to the pathogen in the field. The Q-VAX vaccine, an inactivated whole-cell vaccine, has shown efficacy in preventing Q fever and has been used in high-risk populations. However, challenges remain in its widespread implementation due to limited availability and adverse reactions.Immunological studies have demonstrated the crucial role of macrophages in the host's defense against C. burnetii, with these cells playing a central role in phagocytosis, antigen presentation, and the production of pro-inflammatory cytokines. Interferon gamma (IFNγ), a key cytokine produced by T lymphocytes and natural killer cells, plays a significant role in controlling C. burnetii infection by activating macrophages and enhancing their antimicrobial activity.The use of nanoparticles as chemically programmable scaffolds in bead design shows promise in C. burnetii vaccine development. These nanoparticles can be engineered to display multiple proteins and epitopes, increasing immunogenicity and broadening the immune response. We showcased a high-throughput method of capturing expressed proteins onto bead scaffolds, which were directly administered to animals for in vivo evaluation. Additionally, the bacterial sacculus, the highly ordered peptidoglycan structure of bacteria, was explored as a vaccine scaffold and adjuvant. Muramyl dipeptide (MDP), the main component of the sacculus, acts as a pathogen-associated molecular pattern (PAMP) and activates innate immune responses through recognition by NOD-like receptors. The sacculus’s immunogenicity as an adjuvant was also evaluated in vivo.Adjuvants, such as toll-like receptor (TLR) agonists, are investigated here to enhance the immunogenicity of Q fever vaccines by increasing Th1 responses and IFNγ production. Challenge studies were conducted to evaluate subunit vaccine formulations containing downselected antigen candidates combined with optimized adjuvant combinations, including C. burnetii lipopolysaccharide.In conclusion, understanding the immunology of C. burnetii infection and the challenges associated with vaccine development is essential for effectively combating Q fever. Continued research into novel vaccine strategies, adjuvants, and delivery systems holds promise for improving vaccine efficacy, reducing the global burden of Q fever, and protecting at-risk individuals.
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