Biology subjects

Chopra, A. K.

Publications and source records attributed to Chopra, A. K..

8 recordsLinked to original sources

bioRxiv2025

Stress granule component TIA-1 is a specific negative regulator of the non-canonical NLRP3 inflammasome.

Inflammasomes are cytosolic signaling hubs assembled upon pathogen- or damage associated molecular patterns (PAMP and DAMP) sensing by innate immune pattern recognition receptors (PRR). Lipopolysaccharide (LPS) present on the cell wall of gram-negative bacteria is a PAMP that activates caspase 11 (CASP11) dependent nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 3 (NLRP3) inflammasome (known as non-canonical NLRP3 inflammasome) leading to pyroptosis. Several host factors are shown to promote non-canonical NLRP3 inflammasome activation by making LPS readily available for recognition by CASP11. Here, we report T-cell intracellular antigen-1 (TIA1), an RNA binding protein as a negative regulator of non-canonical NLRP3 inflammasome. Using bone marrow-derived macrophages (BMDMs), we demonstrated that the loss of TIA1 led to an increase in caspase-1 (CASP1) activity in response to cytosolic LPS. A previous study had demonstrated that mice lacking Tia1 are more susceptible to LPS mediated endotoxic shock. Our results provide a potential explanation for this observation by showing loss of TIA1 increases non-canonical NLRP3 inflammasome activation resulting in increased inflammation and pathogenesis during LPS mediated endotoxic shock. Further, TIA1 mediated inhibition of non-canonical NLRP3 inflammasome is independent of TIA1s regulatory role in gene transcription as well as its role in stress granule assembly. TIA1 is also dispensable for activation of the canonical NLRP3 inflammasome as well as AIM2 and NLRC4 inflammasomes. While, the exact mechanism by which TIA1 inhibits non-canonical inflammasome activation remains to be elucidated, our finding that TIA1 is a negative regulator indicates the presence of undiscovered regulatory mechanisms. Future studies will focus on unraveling these mechanisms for developing anti-inflammatory drugs that exploit non-canonical inflammasome activity modulation.

immunology↗

bioRxiv2024

Combination of live attenuated and adenovirus-based vaccines completely protects interferon gamma (IFNγ) knockout mice against pneumonic plague

Two live attenuated vaccines (LAVs), LMA and LMP, were evaluated alone or in combination with a trivalent adenoviral vector-based vaccine (Ad5-YFV) for their efficacy and immune responses in wild type (WT) and interferon gamma (IFN{gamma}) knockout (KO) mice in a C57BL/6 background. While LMA and LMP are triple deletion mutants of Yersinia pestis CO92 strain, Ad5-YFV incorporates three protective plague immunogens. An impressive 80-100% protection was observed in all vaccinated animals against highly lethal intranasal challenge doses of parental Y. pestis CO92. All vaccinated mice generated robust humoral and cellular immune responses. The immunized WT mice showed overall better antibody responses in both serum and bronchoalveolar lavage fluid with much higher percentages of polyfunctional T cell populations. On the other hand, vaccinated IFN{gamma} KO mice displayed better B cell activity in germinal centers with higher percentages of activated antigen specific T cells and memory T cells. In addition, depletion of IFN{gamma} and tumor necrosis factor alpha (TNF) from immunized WT mice prior to and during infection did not reduce protection against pulmonary Y. pestis CO92 challenge. These data demonstrated a dispensable nature of IFN{gamma} in mediating protection by the aforementioned vaccines. This is the first detailed immunogenicity study of two plague LAVs administered either alone or in combination with an Ad5-YFV vaccine in a prime-boost immunization strategy in IFN{gamma} KO mice. Further, by combining advantages of live-attenuated and adenovirus-based vaccines, augmentation of generalized immune responses were observed which could be beneficial in providing long-lasting immunity in the host.

immunology↗

bioRxiv2024

A Modular Bacteriophage T4 Nanoparticle Platform Enables Rapid Design of Completely Protective Multivalent Mucosal Dual Vaccine Against COVID-19 and Flu

A multivalent, rapidly deployable, mucosal vaccine platform is desperately needed to prevent the acquisition and transmission of respiratory infections during epidemics and pandemics. We present one such bacteriophage T4-based platform, and design of dual COVID-19-Flu mucosal vaccines by exploiting its unique architecture. These include: T4s natural affinity for nasal mucosa, flexible engineering to incorporate multiple antigens, and repeat and symmetric epitope presentation for enhanced B cell responses. Hundreds of SARS-CoV-2 spike trimers and nucleocapsid proteins, and influenza hemagglutinin trimers and M2e peptides, were incorporated into a single phage, creating the highest density nanoparticle presentation yet reported. Intranasal administration of adjuvant-free vaccine induced robust mucosal immunity in mice including, neutralizing antibody and secretory IgA, lung-resident CD4+/CD8+ T cells, diverse memory B cells, and complete protection against SARS-CoV-2 and influenza challenges. The noninfectious T4 phage offers an extraordinary platform to rapidly design potent mucosal vaccines against emerging bacterial and viral threats.

microbiology↗

bioRxiv2024

A Bacteriophage Cocktail Targeting Yersinia pestis Provides Strong Post-Exposure Protection in a Rat Pneumonic Plague Model

Yersinia pestis, one of the deadliest bacterial pathogens ever known, is responsible for three plague pandemics and several epidemics, with over 200 million deaths during recorded history. Due to high genomic plasticity, Y. pestis is amenable to genetic mutations as well as genetic engineering that can lead to the emergence or intentional development of pan-drug resistant strains. The dissemination of such Y. pestis strains could be catastrophic, with public health consequences far more daunting than those caused by the recent COVID-19 pandemic. Thus, there is an urgent need to develop novel, safe, and effective treatment approaches for managing Y. pestis infections. This includes infections by antigenically distinct strains for which vaccines, none FDA approved yet, may not be effective, and those that cannot be controlled by approved antibiotics. Lytic bacteriophages provide one such alternative approach. In this study, we examined post-exposure efficacy of a bacteriophage cocktail, YPP-401, to combat pneumonic plague caused by Y. pestis CO92. YPP-401 is a four-phage preparation with a 100% lytic activity against a panel of 68 genetically diverse Y. pestis strains. Using a pneumonic plague aerosol challenge model in gender-balanced Brown Norway rats, YPP-401 demonstrated [~]88% protection when delivered 18 hours post-exposure for each of two administration routes (i.e., intraperitoneal and intranasal) in a dose-dependent manner. Our studies suggest that YPP-401 could provide an innovative, safe, and effective approach for managing Y. pestis infections, including those caused by naturally occurring or intentionally developed strains that cannot be managed by vaccines in development and antibiotics.

microbiology↗

bioRxiv2022

A bacteriophage-based, highly efficacious, needle and adjuvant-free, mucosal COVID-19 vaccine

The authorized mRNA- and adenovirus-based SARS-CoV-2 vaccines are intramuscularly injected and effective in preventing COVID-19, but do not induce efficient mucosal immunity, or prevent viral transmission. We developed a bacteriophage T4-based, multicomponent, needle and adjuvant-free, mucosal vaccine by engineering spike trimers on capsid exterior and nucleocapsid protein in the interior. Intranasal administration of T4-COVID vaccine induced higher virus neutralization antibody titers against multiple variants, balanced Th1/Th2 antibody and cytokine responses, stronger CD4+ and CD8+ T cell immunity, and higher secretory IgA titers in sera and bronchoalveolar lavage with no effect on the gut microbiota, compared to vaccination of mice intramuscularly. The vaccine is stable at ambient temperature, induces apparent sterilizing immunity, and provides complete protection against original SARS-CoV-2 strain and its Delta variant with minimal lung histopathology. This mucosal vaccine is an excellent candidate for boosting immunity of immunized and/or as a second-generation vaccine for the unimmunized population.

microbiology↗

bioRxiv2021

Combinatorial viral vector-based and live-attenuated vaccines without an adjuvant to generate broader immune responses to effectively combat pneumonic plague

Mice immunized with a combination of an adenovirus vector (Ad5-YFV) and live-attenuated (LMA)-based vaccines were evaluated for protective efficacy against pneumonic plague. While the Ad5-YFV vaccine harbors a fusion cassette of three genes encoding YscF, F1, and LcrV, LMA represents a mutant of parental Yersinia pestis CO92 deleted for genes encoding Lpp, MsbB, and Ail. Ad5-YFV and LMA were either administered simultaneously (1-dose regimen) or 21 days apart in various order and route of administration combinations (2-dose regimen). The 2-dose regimen induced robust immune responses to provide full protection to animals against parental CO92 and its isogenic F1 (CAF-)-deletion mutant challenges during both short- and long-term studies. Mice intranasally (i.n.) immunized with Ad5-YFV first followed by LMA (i.n. or intramuscularly [i.m.]) had higher T- and B-cell proliferative responses and LcrV antibody titers than those in mice vaccinated with LMA (i.n. or i.m.) first ahead of Ad5-YFV (i.n.) during the long-term study. Specifically, the needle- and adjuvant-free vaccine combination (i.n.) is ideal for use in plague endemic regions. Conversely, with a 1-dose regimen, mice vaccinated with Ad5-YFV i.n. and LMA by the i.m. route provided complete protection to animals against CO92 and its CAF- mutant challenges, and elicited Th1/Th2, as well as Th17 responses, thus suitable for emergency vaccination during a plague outbreak or bioterrorist attack. This is a first study in which a viral vector-based and live-attenuated vaccines were effectively used in combination representing adjuvant- and/or needle- free immunization, with each vaccine triggering a distinct cellular immune response. ImportanceYersinia pestis, the causative agent of plague, is a Tier-1 select agent and a re-emerging human pathogen. A 2017 outbreak in Madagascar with >75% of cases being pneumonic and 8.6% causalities emphasized the importance of the disease. The World Health Organization has indicated an urgent need to develop new generation subunit and live-attenuated plague vaccines. We have developed a subunit vaccine including three components (YscF, F1, and LcrV) using an adenovirus platform (Ad5-YFV). In addition, we have deleted virulence genes of Y. pestis (e.g., lpp, msbB, and ail) to develop a live-attenuated vaccine (LMA). Both of these vaccines generated robust humoral and cellular immunity and were highly efficacious in several animal models. We hypothesized the use of a heterologous prime-boost strategy or administrating both vaccines simultaneously could provide an adjuvant- and/or a needle- free vaccine(s) which have attributes of both vaccines for use in endemic regions and during an emergency situation.

microbiology↗

bioRxiv2021

A Universal Bacteriophage T4 Nanoparticle Platform to Design Multiplex SARS-CoV-2 Vaccine Candidates by CRISPR Engineering

A "universal" vaccine design platform that can rapidly generate multiplex vaccine candidates is critically needed to control future pandemics. Here, using SARS-CoV-2 pandemic virus as a model, we have developed such a platform by CRISPR engineering of bacteriophage T4. A pipeline of vaccine candidates were engineered by incorporating various viral components into appropriate compartments of phage nanoparticle structure. These include: expressible spike genes in genome, spike and envelope epitopes as surface decorations, and nucleocapsid proteins in packaged core. Phage decorated with spike trimers is found to be the most potent vaccine candidate in mouse and rabbit models. Without any adjuvant, this vaccine stimulated robust immune responses, both TH1 and TH2 IgG subclasses, blocked virus-receptor interactions, neutralized viral infection, and conferred complete protection against viral challenge. This new type of nanovaccine design framework might allow rapid deployment of effective phage-based vaccines against any emerging pathogen in the future.

microbiology↗

bioRxiv2019

Gut-associated bacteria invade the midgut epithelium of Aedes aegypti and stimulate innate immunity and suppress Zika virus infection in cells.

Microbiota within mosquitoes influence nutrition, immunity, fecundity, and the capacity to transmit pathogens. Despite their importance, we have a limited understanding of host-microbiota interactions, especially at the cellular level. It is evident bacterial symbionts that are localized within the midgut also infect other organs within the mosquito; however, the route these symbionts take to colonize other tissues is unknown. Here, utilizing the gentamicin protection assay, we showed that the bacterial symbionts Cedecea and Serratia have the capacity to invade and reside intracellularly within mosquito cells. Symbiotic bacteria were found within a vacuole and bacterial replication was observed in mosquito cell by transmission electron microscopy, indicating bacteria were adapted to the intracellular milieu. Using gene silencing, we determined that bacteria exploited host factors, including actin and integrin receptors, to actively invade mosquito cells. As microbiota can affect pathogens within mosquitoes, we examined the influence of intracellular symbionts on Zika virus (ZIKV) infection. Mosquito cells harbouring intracellular bacteria had significantly less ZIKV compared to uninfected cells or cells exposed to non-invasive bacteria. Intracellular bacteria were observed to substantially upregulate the Toll and IMD innate immune pathways, providing a possible mechanism mediating these anti-viral effects. Examining mono-axenically infected mosquitoes using transmission electron and fluorescent microscopy revealed that bacteria occupied an intracellular niche in vivo. Our results provided evidence that bacteria that associate with the midgut of mosquitoes have intracellular lifestyles which likely have implications for mosquito biology and pathogen infection. This study expands our understanding of host-microbiota interactions in mosquitoes, which is important as symbiont microbes are being exploited for vector control strategies.

microbiology↗