Introduction: Gonorrhea is one of the most common sexually transmitted disease, caused by Gram negative diplococcus bacteria, Nisseria gonorrhoeae. Resistance has emerged to many antimicrobials; therefore, preventative vaccine for gonorrhea can be of great importance in the healthcare because of wide occurrence of the infection. Langerhans cells in skin, are phagocytic cells that signal T-cells. Upon activation, T cells and macrophages drain into nearby lymph nodes causing an increased immune response. Methods: Microparticles loaded with antigens, were prepared using spray drying method. The particulate vaccine formulation contains a biodegradable, biocompatible, non-antigenic and sustained releasing polymer components containing cross-linked albumin matrix and formalin-fixed inactivated whole-cell gonococci and adjuvants. The microparticles were characterized for percent yield, size, charge and poly dispersity index (PDI). The microparticulate vaccine was delivered via transdermal route using ablative laser (P.L.E.A.S.E.®). In-vivo efficacy of this vaccine was checked in 6-8 weeks old swiss webster mice. Mice were administered one prime dose at day 0 followed by two booster doses at week 2 and 4. There were five groups (n=6) in this study and animals were challenged with Gonorrhea bacteria at week 10, and sacrificed at week 12. Their lymph nodes and spleens were collected and levels of the immune cells such as CD4+ and CD8+ T cells in the collected spleens and lymph nodes were measured using BD Accuri™ C6

The measles virus (MV) vaccine lineage is a promising oncolytic but prior exposure to the measles vaccine or wild-type MV strains limits treatment utility due to the presence of antimeasles antibodies. MV entry can be redirected by displaying a polypeptide ligand on the Hemagglutinin (H) C-terminus. We hypothesized that retargeted MV would escape neutralization by monoclonal antibodies (mAbs) recognizing the H receptor-binding surface and be less susceptible to neutralization by human antisera. Using chimeric H proteins, with and without mutations that ablate MV receptor binding, we show that retargeted MVs escape mAbs that target the H receptor-binding surface by virtue of mutations that ablate infection via SLAM and CD46. However, C-terminally displayed domains do not mediate virus entry in the presence of human antibodies that bind to the underlying H domain. In conclusion, utility of retargeted oncolytic measles viruses does not extend to evasion of human serum neutralization.

The development of a safe, immunogenic, globally effective and affordable vaccine may be useful in the control of HIV/AIDS. The recombinant vaccines developed by cloning of HIV genes using different vectors have not been found to be effective due to poor or moderate immunogenicity and/or safety. Semliki Forest virus (SFV), an alpha virus does not have pre-existing immunity, has the cytoplasmic but not nuclear expression of heterologous proteins and non-pathogenic in humans. Therefore HIV1 Indian subtype C gag, env, and polRT genes were individually cloned using SFV vector to develop recombinant SFV2gen replicon RNA constructs and subsequently constructed recombinant SFV2gen viral like replicon particles (VRP) designated as rSFV2gen/gag VRP, rSFV2gen/env VRP, and rSFV2gen/polRT VRP by coelectroporation with Helper RNA. In vitro studies demonstrated high levels of expression of respective HIV1 proteins and their localization in the cytosol and not nucleus from all three recombinant constructs following infection of BHK-21 cells. The recombinant RNA constructs and VRPs individually and in a combination of three constructs elicited significantly high cell-mediated immune responses as detected by INF gamma and IL2 Assay and humoral immune responses in mice. VRPs have been found to be more immunogenic as compared to RNA constructs. Studies demonstrated that all three recombinant SFV2gen based vaccine constructs of Indian subtype C gag, env, and polRT genes were highly immunogenic in the mice model and therefore promising preventive and therapeutic candidate vaccines and therefore may be effective in control and management of HIV/AIDS.

The increasing importance of viral vaccine manufacturing has driven the need for high cell density processes that allow for higher production levels of viral vectors in efficient and scalable processes. The present investigation evaluated recombinant Vesicular Stomatitis virus (rVSV) viral vector production in the Pall iCELLis Nano bioreactor. Vero cell growth within the fixed bedding system was monitored using a biomass sensor that measures conductivity and capacitance. Infection was performed upon optimal cell growth as measured by the biomass sensor. Viral vector harvest was triggered by the biomass readings, which occurred two days post-infection. All iCELLis runs within the campaign yielded an increase of 1 to 2.5 logs of virus production per mL when compared to virus production from standard flat-stock methods. This correlated to an overall increase of 4 logs of virus per batch. In terms of dosages, utilization of the iCELLis Nano resulted in an increase in the number of doses per campaign, when compared to standard flat-stock methods, which ultimately led to a reduction in the cost per dose. In summary, use of the iCELLis bioreactor platform allows for a cost-efficient and scalable process for viral vaccine production.

Vaccine-enhanced respiratory disease has thwarted the attempts to develop a vaccine for Respiratory Syncytial Virus (RSV) using the inactivated form of the virus. As a result, there is a need for a safe and effective vaccine for RSV. Fusion protein is a one of the major proteins present on the surface of the virus, which can be integrated into a virus-like particle (VLP), yielding a highly immunogenic non- virulent F-VLP antigen. This study aims at using a needle free transdermal route of administration to exploit the rich population of Langerhans cells present in the epidermis and dermis. We have used microneedles (AdminPatch®) as well as Precise Laser Epidermal System (P.L.E.A.S.E), a minimally invasive ablative fractional (Er: YAG) laser for creating micro-channels to deliver the antigen.

Bacteria, viruses, parasites, and fungi those are resistant to drug cause 700,000 death each year. By 2050 superbugs inured to treatments could cause up to 10 million deaths annually and costs the global economy US$100 trillion. AMR (antimicrobial) resistance is regarded nowadays as a major threat to global public health. The issue is receiving high-level political attention (G7 and G20 in 2017 for the first time). Pandemics, drug resistance and neglected diseases framing health as a “global security issue”. The list was drawn up in a bid to guide and promote research and development (R&D) of new antibiotics, as part of WHO’s efforts for AMR (27th Feb 2017). Tuberculosis (MDR/XDR) and latent tuberculosis represent a major issue to tackle attract global attention as witnessed by recent WHO and interministerial meeting in November 2017 and high-level UN meetings which have been held in September 2018. The problem of resistance gets worsened due to declining number of new antibiotics and the limited number of new classes. Antibiotic use influences the composition of microbiota in each individual. Similar trends are seen in drug development and use for the treatment of tuberculosis. Microbiota is a complex and diverse bacterial community specific to each individual involved in host health and immunity. Microbiota under 3 years old fluctuates substantially and is more impressionable to environmental factors than the adult microbiota. Antibiotics shape the ecology of the gut microbiota in profound ways, causing lasting changes. To illustrate, antibiotic use is one of the known risk factors for Clostridium difficile infections. There is no simple relationship between antibiotic-mediated depletion of the colonic microbiota and the induction of C. difficile spore germination with subsequent toxin production. Rather, antibiotic exposure might directly stimulate C. difficile proliferation (that is, cause the germination of spores, which are the usual type of cells that are acquired and can remain quiescent in the gut) and toxin production, which occurs in late log phase. Clostridium difficile is the leading cause of antibioticassociated diarrhea, both in healthcare facilities and in the community. This medical urgent issue triggered increased interest to look on new antibiotics preserving microbiota, antibiotic inactivators, and monoclonal antibodies, gut microbiota modulating therapies like fecal microbiota transplantation, fecal bacteriotherapy, probiotics (controversial feedback) and finally vaccines. Gut exposure to antibiotic is accompanied by risk to spread antimicrobial resistance genes. Antibiotic resistance genes can cause phenotypic resistance through a variety of mechanisms, including the enzymatic inactivation of the antibiotic, the modification of the antibiotic target and the prevention of the accumulation of lethalintracellular concentrations of the antibiotic through efflux pumps. Therefore, a multifaceted strategy to promote and prioritize highly potential alternatives to tackle AMR like vaccines development is required. As an example, vaccines like diphtheria and tetanus did not prompt resistance. In 1980 the smallpox vaccine had eradicated the naturally circulating virus worldwide without generating resistance. Additionally, the introduction of live vaccines like measles and BCG has been associated with a much larger reduction of morality that can be explained by the prevention of the targeted infections and recent research like LATV pertussis highlights the importance of “off-target” effects to be evaluated in depth. Thoughtful and innovative vaccines development taking into account host microbiota “superorganism” and immune crosstalk - immune system “training “ opens the large avenue for future development and vaccine research. Accurate diagnostic and surveillance with a better understanding of the genetic and immunologic background of  host-specific response and pathogen evolution drive successful and innovative research. Innovative vaccines, as a highly potent tool and valuable alternative from a long term perspective, are clearly recognized as a major tool for public health already. Further strong support to promote research on alternative tools to tackle antibiotic resistance needs joint endorsement including regulatory and economic stakeholders along with necessary partnerships at the global level.

The D’Souza Vaccine Nanotechnology Laboratory at Mercer’s College of Pharmacy has been working on the design and delivery of microparticulate vaccines for infectious diseases. This presentation will highlight the triumphs and tribulations of this rather innovative, in-expensive and painless method of vaccine delivery using microneedle technology. Our patented technology is rather broad based and can be used to administer multiple vaccines in a single microneedle vaccine patch applied to the skin much like a “band-aid-patch” We have been working on particulate vaccines for several infectious diseases such as a) universal influenza, b) HPV, c) RSV, d) meningitis, e) gonorrhea, and f) measles vaccines designed for delivery via microneedles. In this presentation, the results of these vaccine prototypes will be discussed. The vaccine antigen was formulated in bio-degradable and bio-compatible matrices to prepare nanoparticles or microparticles using a spray dryer. These nano-vaccine particles were administered using micro-needles via the transdermal route. Serum samples were obtained at regular intervals in-order to determine the antigen specific antibody responses (such as IgG, Igg1to assess the systemic immunity. Animals were challenges with live virus/bacteria to determine the level of protective immunity developed after vaccination. Further, we examined the expression of co-stimulatory molecules that impact antigen presentation in an in-vitro system using human macrophages pulsed with the antigen. In-order to determine the correlates of protection by the vaccine in-vivo, the total IgG and its subtypes IgG2a and IgG2b were measured after immunizing mice. The up regulation of other co-stimulatory molecules such as CD-40, CD-80 and CD-86 were also determined. In conclusion, the novel nano-vaccines are robustly taken up by antigen presenting cells and up-regulate co-stimulatory molecules that enhance antigen presentation, which is a pre-requisite for inducing adaptive immunity.

Introduction: Gonorrhea is one of the most common sexually transmitted disease, caused by Gram negative diplococcus bacteria, Nisseria gonorrhoeae. Currently there are no vaccines for Gonorrhea. Therefore, there is an urgent need for a vaccine. An effective and patient compliant option could be a film dosage form of the vaccine for buccal or sublingual administration. administration. Film dosage forms are easy to prepare, cost effective, and patient complaint. When films are administered in buccal or sublingual area, the vaccines are absorbed systematically. The systemic route can avoid the first pass effect and can increase the bioavailability of the vaccines. The film can be designed is in multilayer forms, where the outer layer can be a protective layer which can prevent the destruction or swallowing of the vaccine. Additionally, it is possible to use vaccines in either naked or particulate form.

Method: The particle form of the Gonorrhea vaccine was prepared using a spray dryer. The physical properties of the particle were evaluated. The content analysis of the particle was conducted by western blot. The film dosage forms of these particle vaccines were prepared by solvent casting method. Films were evaluated for physical properties such as thickness, disintegration time, plasticity etc. Thickness was measured using the Mitutoyo digital caliper. The permeability of these particulate vaccines was determined by Franz cell using silicone membrane.

Plants suffer with the attack of microorganisms the same way that animals do. Diseases caused by viruses, bacteria and fungi can lead to great economical losses in agriculture. Just like animals, plants also have an ‘immune’ system that helps defend themselves against these aggressors. The defense mechanisms can be structural or substances derived from the plant secondary metabolism; and they may be constitutive or induced only after the plant receives damage. And after the plant is attacked, the plant keeps a long-term ‘memory’ from the pathogen stimuli leading to a more robust defense in the case of a future attack. Not only the aggressor can trigger this ‘defense priming’, but also substances called elicitors are able to act as vaccines, thus preparing the plant and enhancing its capacity for rapid and effective activation of cellular defense responses in the presence of a challenge. In contrast to direct induced defenses, only minimal fitness cost is associated with priming. This long-term primed state is usually associated with alterations to gene responsiveness. Several chemical elicitors have been described and proven to be effective in priming plants, such as salicylic acid, methyl salicylate and chitosan. The introduction of priming plants with chemical elicitors against pathogens as an agricultural practice could minimize the scope of  agrochemicals and would contribute to a more sustainable agriculture.

Glycopeptide antibiotics were once considered as drug of choice of serious gram positive infections. These antibiotics interrupt bacterial cell wall synthesis to exert their antibacterial effects. They pose higher barrier for drug resistance development, as they target non-protein components of bacterial cell wall. However, these antibiotics are increasingly becoming less effective due to emergence of resistant strains. To address this issue, modification of glycopeptide antibiotics to enhance their activity is therefore a useful strategy to develop new compounds against drug-resistant strains. We explored an underutilized reactive site on the glycopeptide antibiotics and developed a simple yet highly efficient scheme to synthesize various analogs. Using this scheme, the C-terminal carboxyl group of vancomycin was reacted with amine compounds to yield carboxamide analogs some of which with improved antibacterial activity upto 100 times. Usually multiple chemical reactions are needed to prepare antibiotic analogs. Our single-step scheme provides a simple yet efficient methodology to develop potent analogs of vancomycin. Different analogs are synthesized by reacting series of diamines with vancomycin.