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New vaccine research and development (R&D) and production is of importance in combating against prevalent infectious diseases, as vaccines have prevented countless cases of disease and saved millions of lives. We formulate a multi-stage game comprised by one developer, one manufacturer and one government in the new vaccine R&D and production, considering R&D failure risk and capacity sharing. This paper examines the effects of different subsidy forms and two alliance modes. The result reveals that: (1) The government should consistently provide a combo of subsidy to achieve maximum social welfare. The developer has no preference for the form of subsidy in the manufacturer-initiated mode, whereas the manufacturer desires to get subsidised by the per-unit cost form regardless of alliance modes. (2) Being the initiator inside the alliance may not always be the optimal decision for players. The manufacturer should relinquish the role as the initiator if certain requirements are fulfilled. (3) When capacity sharing revenue is substantial or the failure risk is sufficiently low, the government should give priority to using per-unit cost subsidy instead of quality improvement subsidy, and the manufacturer would like to be the follower if per-unit cost subsidy is offered. [ABSTRACT FROM AUTHOR]

The aim of the research is to analyze and improve the performance of the vaccine quality control queuing system to reduce delays and achieve the firm’s long-term goal on vaccine production capacity. The research focuses on the Bacterial Vaccine Quality Control (BVQC) at the largest vaccine manufacturers in Indonesia and Southeast Asia. The vaccines handled by BVQC include TT, DTP, BCG, BioTT, BioTd, DT, Td, and DTP-Hb-Hib. The BVQC operates a queuing system with eight servers, each assigned a fixed task. The existing system experienced delays ranging from 13 % to 61 % from January to June 2022. After identifying the queuing characteristics of the existing system, improvement proposals were suggested by modifying the assignment of the servers. This proposal was then simulated in November 2022, resulting in improved performance with no delays, a reduction in the length of the queue in the system (Lq) from 2.88 to 2.59, and a reduction in the average time spent in the system (Ws) from 0.0099 to 0.0044. The research suggests that modifying server assignments can be an effective method for improving the performance of a queuing system in vaccine quality control. This can lead to reduced delays, optimized queue lengths, and improved overall efficiency, potentially enhancing the firm’s ability to meet vaccine demand in the future.

Background: The COVID-19 pandemic revealed vaccine supply chain (VSC) weaknesses and enabled post-pandemic analysis highlighting the growing importance of supply chain resilience. This study analyzes weaknesses and potentials for VSC resilience from an industry perspective. Insights from this study are aimed at supporting helping managers and policy-makers build a more resilient vaccine supply. Methods: A qualitative semi-structured interview study was conducted with 12 industry experts along the VSC. The interviews were assessed concerning the learnings from the pandemic in a two-step content analysis. Codes were assigned to key VSC concepts and variables and then linked to political, economic, social, technological, legal, and environmental (PESTLE) dimensions. The complex multi-stakeholder supply chain was visualized in a system overview, highlighting main actors, roles, constraints, and resilience. Results: The analysis resulted in 60 codes, categorized into the six PESTLE dimensions and three additional (sub)groups (mRNA, Supply chain resilience, and Solutions). The largest dimension was Economic, with 39 codes, including the Supply chain resilience subgroup. Twelve stakeholder groups were identified, with purchasers, manufacturers, suppliers, developers, and regulatory agencies being the most significant in emergency vaccine manufacturing situations. Conclusions: The system overview demonstrated the VSC as a complex network of actors with unaligned goals rather than a linear supply chain. This study shows that the VSC is characterized by uncertainty due to external factors, like the unpredictability of new emergencies, and internal factors like vaccine demand. The lack of transparency between industry stakeholders exacerbates VSC disruption. We conclude that infrastructures and management practices that enable increased transparency and collaboration between stakeholders hold the greatest potential for strengthening the VSC’s resilience to future pandemics.

Chinese Hamster Ovary (CHO) cells, employed primarily for manufacturing monoclonal antibodies and other recombinant protein (r-protein) therapeutics, are emerging as a promising host for vaccine antigen production. This is exemplified by the recently approved CHO cell-derived subunit vaccines (SUV) against respiratory syncytial virus (RSV) and varicella-zoster virus (VZV), as well as the enveloped virus-like particle (eVLP) vaccine against hepatitis B virus (HBV). Here, we summarize the design, production, and immunogenicity features of these vaccine and review the most recent progress of other CHO-derived vaccines in pre-clinical and clinical development. We also discuss the challenges associated with vaccine production in CHO cells, with a focus on ensuring viral clearance for eVLP products. [ABSTRACT FROM AUTHOR]

The paper argues that vaccines could be viewed as artifacts which communicate various social messages and are used as instruments for fulfilling different sociopolitical goals besides meeting public health needs. It further suggests that such social, cultural and political influences may have real effects on the choices of vaccine technologies or vaccine production, and aims to demonstrate their importance in the area which is normally seen as the domain of objective science. This is demonstrated by using the example of the locally produced oral polio vaccine (OPV) in Serbia during the socialist and post-socialist periods in the country’s history.

This paper examines biosafety management in high-biosafety-risk workshops for vaccine production in China, focusing on the context of SARS-CoV-2 inactivated vaccine production. It addresses various aspects, including biosafety management, personnel, virus seed, facilities, and equipment, and compliance with relevant biosafety laws and regulations. The aim is to promote the use of high-level biosafety facilities when dealing with emerging infectious diseases.

In Imunizarea. Controversele vaccinurilor [Immunization: How Vaccines Became Controversial] (original title), Stuart Blume provides an in-depth analysis of vaccines, emphasizing their role beyond medicine into social, political, and economic realms. The book traces the history of vaccine hesitancy, linking it to socio-political changes, such as neoliberalism and commercialization processes. Blume highlights the dual nature of vaccines as public health instruments and commercial products, and how this duality contributes to vaccine hesitancy. He challenges common views on vaccine disparities between developed and developing nations, and examines the evolution of public attitudes towards vaccination. The transition from public to private vaccine production and the focus on profitable vaccines, often at the expense of less developed countries' needs, are some of the key themes in this book. Blume also discusses the redefinition of health risks and diseases, influencing vaccination policies and public perceptions. The book delves into the historical and evolving nature of resistance to vaccination, ultimately arguing that vaccine hesitancy is deeply rooted in both the commercial and public health significance of vaccines.

Bioprocess development benefits from kinetic models in many aspects, including scale-up, optimization, and process understanding. However, current models are unable to simulate the production process of a coxsackievirus A6 (CVA6) virus-like particle (VLP) vaccine using Chinese hamster ovary cell culture. In this study, a novel kinetic model was constructed, correlating (1) cell growth, death, and lysis kinetics, (2) metabolism of major metabolites, and (3) CVA6 VLP production. To construct the model, two batches of a laboratory-scale 2 L bioreactor cell culture were prepared and various pH shift strategies were applied to examine the effect of pH shift. The proposed model described the experimental data under various conditions with high accuracy and quantified the effect of pH shift. Next, cell culture performance with various pH shift timings was predicted by the calibrated model. A trade-off relationship was found between product yield and quality. Consequently, multiple objective optimization was performed by integrating desirability methodology with model simulation. Finally, the optimal operating conditions that balanced product yield and quality were predicted. In general, the proposed model improved the process understanding and enabled in silico process development of a CVA6 VLP vaccine. [ABSTRACT FROM AUTHOR]

In Imunizarea. Controversele vaccinurilor [Immunization: How Vaccines Became Controversial] (original title), Stuart Blume provides an in-depth analysis of vaccines, emphasizing their role beyond medicine into social, political, and economic realms. The book traces the history of vaccine hesitancy, linking it to socio-political changes, such as neoliberalism and commercialization processes. Blume highlights the dual nature of vaccines as public health instruments and commercial products, and how this duality contributes to vaccine hesitancy. He challenges common views on vaccine disparities between developed and developing nations, and examines the evolution of public attitudes towards vaccination. The transition from public to private vaccine production and the focus on profitable vaccines, often at the expense of less developed countries’ needs, are some of the key themes in this book. Blume also discusses the redefinition of health risks and diseases, influencing vaccination policies and public perceptions. The book delves into the historical and evolving nature of resistance to vaccination, ultimately arguing that vaccine hesitancy is deeply rooted in both the commercial and public health significance of vaccines. [ABSTRACT FROM AUTHOR]

Abstract Background We developed a MARC-145 cell culture and porcine reproductive and respiratory syndrome (PRRS) vaccine production using a novel CelCradle bioreactor. CelCradle is a packed-bed bioreactor capable of both batch and perfusion culture, and the operating parameters are easy to optimize. Results In this study, CelCradle reached a maximum cell density of 8.94 × 105 cells/mL at 5 days post-seeding when seeded at 8.60 × 104 cells/mL (doubling time = 35.52 h). Inoculation of PRRS vaccine candidate, K418DM1.1, was performed at a multiplicity of infection (MOI) of 0.01 at 5 days post-seeding, which resulted in a high viral titer of 2.04 × 108 TCID50/mL and total viral load of 1.02 × 1011 TCID50/500 mL at 2 days post-infection (dpi). The multilayer cultivation system, BioFactory culture, yielded a higher doubling time (37.14 h) and lower viral titer (i.e., 8.15 × 107 TCID50/mL) compared to the CelCradle culture. Thus, the culture medium productivity of the CelCradle culture was 2-fold higher than that of the BioFactory culture. In the animal experiment, the CelCradle-produced vaccine induced high levels of neutralizing antibodies and effectively protected pigs against homologous challenge, as shown by the significantly lower levels of viremia at 1- and 7-days post-challenge (dpc) compared to the non-vaccinated pigs. Conclusions Overall, this study demonstrates that the CelCradle system is an economical platform for PRRS vaccine production.

Background: We developed a MARC-145 cell culture and porcine reproductive and respiratory syndrome (PRRS) vaccine production using a novel CelCradle bioreactor. CelCradle is a packed-bed bioreactor capable of both batch and perfusion culture, and the operating parameters are easy to optimize. Results: In this study, CelCradle reached a maximum cell density of 8.94 × 105 cells/mL at 5 days post-seeding when seeded at 8.60 × 104 cells/mL (doubling time = 35.52 h). Inoculation of PRRS vaccine candidate, K418DM1.1, was performed at a multiplicity of infection (MOI) of 0.01 at 5 days post-seeding, which resulted in a high viral titer of 2.04 × 108 TCID50/mL and total viral load of 1.02 × 1011 TCID50/500 mL at 2 days post-infection (dpi). The multilayer cultivation system, BioFactory culture, yielded a higher doubling time (37.14 h) and lower viral titer (i.e., 8.15 × 107 TCID50/mL) compared to the CelCradle culture. Thus, the culture medium productivity of the CelCradle culture was 2-fold higher than that of the BioFactory culture. In the animal experiment, the CelCradle-produced vaccine induced high levels of neutralizing antibodies and effectively protected pigs against homologous challenge, as shown by the significantly lower levels of viremia at 1- and 7-days post-challenge (dpc) compared to the non-vaccinated pigs. Conclusions: Overall, this study demonstrates that the CelCradle system is an economical platform for PRRS vaccine production. [ABSTRACT FROM AUTHOR]

Classical swine fever (CSF) live vaccine used in Japan, GPE-strain, is produced using guinea pig kidney (GPK)-derived primary culture cells. This means that a large number of guinea pigs are used to generate the primary GPK cells needed to produce the CSF live vaccine, and alternative solution is desired. Hence, we established two GPK cell lines capable of culturing the GPE- strain: spontaneously immortalized GPK (GPK-SI) cells were generated by repeated passaging of primary GPK cells, and the other cell line, artificially immortalized GPK (GPK-AI) cells, were obtained by introducing the SV40 large T antigen gene into primary GPK cells. Both cell lines were susceptible to the GPE- virus, and the virus grew more efficiently in GPK-SI cells at 37°C. When the culture temperature was set to 30°C, the virus titer reached 104.8 50% Tissue Culture Infectious Dose (TCID50)/mL in GPK-SI cells 7 days after virus inoculation at a multiplicity of infection (MOI) of 1, which was equivalent to that in cells cultured at 37°C. When the virus was inoculated at MOI <1, the virus titer 7 days after inoculation was higher when cultured at 30°C than when cultured at 37°C in both cell lines, reaching 105.63 TCID50/mL in GPK-SI cells. These results indicate that GPK-SI and GPK-AI cells can potentially replace primary GPK cells for the production of CSF live vaccines. This could also contribute to stable CSF vaccine production and animal welfare. [ABSTRACT FROM AUTHOR]

ABSTRACT: The mammalian-derived MDCK cells are the most widely used for avian virus vaccine production at present. The use of heterologous cell systems for avian virus preparation may cause security risks. An avian cell line is available for avian virus vaccines urgently needed. In this study, a peacock immortalized fibroblast cell line that is suitable for avian virus vaccine production was generated. The primary peacock fibroblast cells were prepared, and the immortal cells PEF-1 were obtained by transferring hTERT into the primary cells and screening with G418. The PEF-1 has high cell viability and expresses exogenous TERT protein. More importantly, the virus replication ability was stronger in PEF-1 than in MDCK cells as evaluated by virus fluorescence and TCID50, after being infected with NDV-GFP, VSV-GFP, and AIV. In conclusion, the peacock immortalized PEF cells are expected to be used for the production of peacock and other avian virus vaccines.

Coronavirus disease-2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus (SARS-CoV)-2 spread globally and creates an alarming situation. Following the SARS-CoV-2 paradigm, therapeutic efficacy is achieved via repurposing several antiviral, antibacterial, and antimalarial drugs. Innate and adaptive immune cells work close to combat infection through the intricate production of antibodies (Abs) and inflammatory cytokines. As an essential component of the immune system, Abs play an important role in eliminating viruses and maintaining homeostasis. B lymphocytes (B cells) are effector cells, stringent to produce neutralizing Abs to combat infection. After recognizing SARS-CoV-2 antigens by a surface receptor called B cell receptors (BCRs) on the plasma membrane, the BCRs transmembrane signal transduction and immune activation results in Ab production and development of immune memory. Thus, it ensures that plasma B cells can quickly start an intricate immune response to generate efficient protective Abs to clear the pathogen. Nevertheless, considering therapeutic challenges in the context of the new coronavirus pandemic, this review addresses the molecular mechanism of the immune activation and function of novel SARS-CoV-2 specific B cells in the production of SARS-CoV-2 specific Abs. Additionally, these studies highlighted the Ab-mediated pathogenesis, the intriguing role of nano-scale signaling subunits, non-structural proteins during COVID-19 infection, and structural insights of SARS-CoV-2 specific Abs.

The influenza virus is a cause of seasonal epidemic disease and enormous economic injury. The best way to control influenza outbreaks is through vaccination. The Madin-Darby canine kidney cell line (MDCK) is currently approved to manufacture influenza vaccines. However, the viral load from cell-based production is limited by host interferons (IFN). Interferon regulating factor 7 (IRF7) is a transcription factor for type-I IFN that plays an important role in regulating the anti-viral mechanism and eliminating viruses. We developed IRF7 knock-out MDCK cells (IRF7-/- MDCK) using CRISPR/Cas9 technology. The RNA expression levels of IRF7 in the IRF7-/- MDCK cells were reduced by 94.76% and 95.22% under the uninfected and infected conditions, respectively. Furthermore, the IRF7 protein level was also significantly lower in IRF7-/- MDCK cells for both uninfected (54.85% reduction) and viral infected conditions (32.27% reduction) compared to WT MDCK. The differential expression analysis of IFN-related genes demonstrated that the IRF7-/- MDCK cell had a lower interferon response than wildtypeMDCKunder the influenza-infected condition. Gene ontology revealed down-regulation of the defense response against virus and IFN-gamma production in IRF7-/- MDCK. The evaluation of influenza viral titers by RT-qPCR and hemagglutination assay (HA) revealed IRF7-/- MDCK cells had higher viral titers in cell supernatant, including A/pH1N1 (4 to 5-fold) and B/Yamagata (2-fold). Therefore, the IRF7-/- MDCK cells could be applied to cell-based influenza vaccine production with higher capacity and efficiency. [ABSTRACT FROM AUTHOR]

The influenza virus is a cause of seasonal epidemic disease and enormous economic injury. The best way to control influenza outbreaks is through vaccination. The Madin-Darby canine kidney cell line (MDCK) is currently approved to manufacture influenza vaccines. However, the viral load from cell-based production is limited by host interferons (IFN). Interferon regulating factor 7 (IRF7) is a transcription factor for type-I IFN that plays an important role in regulating the anti-viral mechanism and eliminating viruses. We developed IRF7 knock-out MDCK cells (IRF7−/ − MDCK) using CRISPR/Cas9 technology. The RNA expression levels of IRF7 in the IRF7−/ − MDCK cells were reduced by 94.76% and 95.22% under the uninfected and infected conditions, respectively. Furthermore, the IRF7 protein level was also significantly lower in IRF7−/ − MDCK cells for both uninfected (54.85% reduction) and viral infected conditions (32.27% reduction) compared to WT MDCK. The differential expression analysis of IFN-related genes demonstrated that the IRF7−/ − MDCK cell had a lower interferon response than wildtype MDCK under the influenza-infected condition. Gene ontology revealed down-regulation of the defense response against virus and IFN-gamma production in IRF7−/ − MDCK. The evaluation of influenza viral titers by RT-qPCR and hemagglutination assay (HA) revealed IRF7−/ − MDCK cells had higher viral titers in cell supernatant, including A/pH1N1 (4 to 5-fold) and B/Yamagata (2-fold). Therefore, the IRF7−/ − MDCK cells could be applied to cell-based influenza vaccine production with higher capacity and efficiency.

Seasonal Influenza H3N2 virus poses a great threat to public health, but its vaccine efficacy remains suboptimal. One critical step in influenza vaccine production is the viral passage in embryonated eggs. Recently, the strength of egg passage adaptation was found to be rapidly increasing with time driven by convergent evolution at a set of functionally important codons in the hemagglutinin (HA1). In this study, we aim to take advantage of the negative correlation between egg passage adaptation and vaccine effectiveness (VE) and develop a computational tool for selecting the best candidate vaccine virus (CVV) for vaccine production. Using a probabilistic approach known as mutational mapping, we characterized the pattern of sequence evolution driven by egg passage adaptation and developed a new metric known as the adaptive distance (AD) which measures the overall strength of egg passage adaptation. We found that AD is negatively correlated with the influenza H3N2 vaccine effectiveness (VE) and ~75% of the variability in VE can be explained by AD. Based on these findings, we developed a computational package that can Measure the Adaptive Distance and predict vaccine Effectiveness (MADE). MADE provides a powerful tool for the community to calibrate the effect of egg passage adaptation and select more reliable strains with minimum egg-passaged changes as the seasonal A/H3N2 influenza vaccine. [ABSTRACT FROM AUTHOR]

The genus flavivirus of the Flaviridae family includes several human pathogens, like dengue, Zika, Japanese encephalitis, and yellow fever virus. These viruses continue to be a significant threat to human health. Vaccination remains the most useful approach to reduce the impact of flavivirus fever. However, currently available vaccines can induce severe side effects or have low effectiveness. An alternative is the use of recombinant vaccines, of which virus-like particles (VLP) and single-round infectious particles (SRIP) are of especial interest. VLP consist of the virus structural proteins produced in a heterologous system that self-assemble in a structure almost identical to the native virus. They are highly immunogenic and have been effective vaccines for other viruses for over 30 years. SRIP are promising vaccine candidates, as they induce both cellular and humoral responses, as viral proteins are expressed. Here, the state of the art to produce both types of particles and their use as vaccines against flaviviruses are discussed. We summarize the different approaches used for the design and production of flavivirus VLP and SRIP, the evidence for their safety and efficacy, and the main challenges for their use as commercial vaccines. [ABSTRACT FROM AUTHOR]

Since 2014, an Asian lineage of Zika virus has caused outbreaks, and it has been associated with neurological disorders in adults and congenital defects in newborns. The resulting threat of the Zika virus to human health has prompted the development of new vaccines, which have yet to be approved for human use. Vaccines based on the attenuated or chemically inactivated virus will require large‐scale production of the intact virus to meet potential global demands. Intact viruses are produced by infecting cultures of susceptible cells, a dynamic process that spans from hours to days and has yet to be optimized. Here, we infected Vero cells adhesively cultured in well‐plates with two Zika virus strains: a recently isolated strain from the Asian lineage, and a cell‐culture‐adapted strain from the African lineage. At different time points post‐infection, virus particles in the supernatant were quantified; further, microscopy images were used to quantify cell density and the proportion of cells expressing viral protein. These measurements were performed across multiple replicate samples of one‐step infections every four hours over 60 h and for multi‐step infections every four to 24 h over 144 h, generating a rich data set. For each set of data, mathematical models were developed to estimate parameters associated with cell infection and virus production. The African‐lineage strain was found to produce a 14‐fold higher yield than the Asian‐lineage strain in one‐step growth and a sevenfold higher titer in multi‐step growth, suggesting a benefit of cell‐culture adaptation for developing a vaccine strain. We found that image‐based measurements were critical for discriminating among different models, and different parameters for the two strains could account for the experimentally observed differences. An exponential‐distributed delay model performed best in accounting for multi‐step infection of the Asian strain, and it highlighted the significant sensitivity of virus titer to the rate of viral degradation, with implications for optimization of vaccine production. More broadly, this study highlights how image‐based measurements can contribute to the discrimination of virus‐culture models for the optimal production of inactivated and attenuated whole‐virus vaccines. [ABSTRACT FROM AUTHOR]

To present the recent update on the isolation, engineering techniques for extracellular vesicles, limitations associated with different isolation techniques, different biomedical applications, and challenges of engineered extracellular vesicles for the benefit of researchers from academic, industry, etc. Peer-reviewed articles from most recognized journals were collected, and presented information was analyzed to discuss collection, chemical, electroporation, cellular, and membrane surface engineering to design extracellular vesicles for various therapeutic applications. In addition, we present the applications and limitations of techniques for the collection of extracellular vesicles. There is a need for isolation techniques with the gold standard. However , advanced extracellular vesicle isolation techniques showed improved recovery, and purity of extracellular vesicles. Tumor therapy is a major part of the therapy section that illustrates the role of engineered extracellular vesicles in synergetic therapy such as phototherapy, theragnostic, and delivery of genetic materials. In addition, extracellular vesicles have shown their potential in the treatment of retinal disorders, neurodegenerative disease, tuberculosis, osteoporosis, inflammatory bowel disease, vaccine production, and wound healing. Engineered extracellular vesicles can deliver cargo to the specific cells, elicit an immune response and could be used for the development of the vaccines in the future. However, the progress is at the initial stage. Overall, this review will provide a comprehensive understanding and could serve as a reference for researchers in the clinical translation of engineered extracellular vesicles in different biomedical fields. [Display omitted] • Engineered EVs can be produced after modification to cells, gene(s), and EVs surface • Engineered EVs can bind to proteins and receptors on the cell surface • Engineered EVs display less immunogenicity and specific targeting [ABSTRACT FROM AUTHOR]

Classical swine fever virus (CSFV) causes an economically important and highly contagious disease of pigs, leading to economic losses around the world. Attenuated live vaccines with CSFV antigens have played an important role in the prevention and control of the disease. Porcine kidney 15 (PK15) cells have been widely used for the propagation of CSFV, but this cell line is not efficient or homogeneously susceptible to viral infection.

A segmented genome enables influenza virus to undergo reassortment when two viruses infect the same cell. Although reassortment is involved in the creation of pandemic influenza strains and is routinely used to produce influenza vaccines, our understanding of the factors that drive the emergence of dominant gene constellations during this process is incomplete. Recently, we defined a spectrum of interactions between the gene segments of the A/Udorn/307/72 (H3N2) (Udorn) strain that occur within virus particles, a major interaction being between the NA and PB1 gene segments. In addition, we showed that the Udorn PB1 is preferentially incorporated into reassortant viruses that express the Udorn NA. Here we use an influenza vaccine seed production model where eggs are coinfected with Udorn and the high yielding A/Puerto Rico/8/34 (H1N1) (PR8) virus and track viral genotypes through the reassortment process under antibody selective pressure to determine the impact of Udorn NA-PB1 co-selection. We discovered that 86% of the reassortants contained the PB1 from the Udorn parent after the initial co-infection and this bias towards Udorn PB1 was maintained after two further passages. Included in these were certain gene constellations containing Udorn HA, NA, and PB1 that confered low replicative fitness yet rapidly became dominant at the expense of more fit progeny, even when co-infection ratios of the two viruses favoured PR8. Fitness was not compromised, however, in the corresponding reassortants that also contained Udorn NP. Of particular note is the observation that relatively unfit reassortants could still fulfil the role of vaccine seed candidates as they provided high haemagglutinin (HA) antigen yields through co-production of non-infectious particles and/or by more HA molecules per virion. Our data illustrate the dynamics and complexity of reassortment and highlight how major gene segment interactions formed during packaging, in addition to antibody pressure, initially restrict the reassortant viruses that are formed.

Glycosylation is considered as a critical quality attribute for the production of recombinant biopharmaceuticals such as hormones, blood clotting factors, or monoclonal antibodies. In contrast, glycan patterns of immunogenic viral proteins, which differ significantly between the various expression systems, are hardly analyzed yet. The influenza A virus (IAV) proteins hemagglutinin (HA) and neuraminidase (NA) have multiple N‐glycosylation sites, and alteration of N‐glycan micro‐ and macroheterogeneity can have strong effects on virulence and immunogenicity. Here, we present a versatile and powerful glycoanalytical workflow that enables a comprehensive N‐glycosylation analysis of IAV glycoproteins. We challenged our workflow with IAV (A/PR/8/34 H1N1) propagated in two closely related Madin–Darby canine kidney (MDCK) cell lines, namely an adherent MDCK cell line and its corresponding suspension cell line. As expected, N‐glycan patterns of HA and NA from virus particles produced in both MDCK cell lines were similar. Detailed analysis of the HA N‐glycan microheterogeneity showed an increasing variability and a higher complexity for N‐glycosylation sites located closer to the head region of the molecule. In contrast, NA was found to be exclusively N‐glycosylated at site N73. Almost all N‐glycan structures were fucosylated. Furthermore, HA and NA N‐glycan structures were exclusively hybrid‐ and complex‐type structures, to some extent terminated with alpha‐linked galactose(s) but also with blood group H type 2 and blood group A epitopes. In contrast to the similarity of the overall glycan pattern, differences in the relative abundance of individual structures were identified. This concerned, in particular, oligomannose‐type, alpha‐linked galactose, and multiantennary complex‐type N‐glycans. [ABSTRACT FROM AUTHOR]

A segmented genome enables influenza virus to undergo reassortment when two viruses infect the same cell. Although reassortment is involved in the creation of pandemic influenza strains and is routinely used to produce influenza vaccines, our understanding of the factors that drive the emergence of dominant gene constellations during this process is incomplete. Recently, we defined a spectrum of interactions between the gene segments of the A/Udorn/307/72 (H3N2) (Udorn) strain that occur within virus particles, a major interaction being between the NA and PB1 gene segments. In addition, we showed that the Udorn PB1 is preferentially incorporated into reassortant viruses that express the Udorn NA. Here we use an influenza vaccine seed production model where eggs are coinfected with Udorn and the high yielding A/Puerto Rico/8/34 (H1N1) (PR8) virus and track viral genotypes through the reassortment process under antibody selective pressure to determine the impact of Udorn NA-PB1 co-selection. We discovered that 86% of the reassortants contained the PB1 from the Udorn parent after the initial co-infection and this bias towards Udorn PB1 was maintained after two further passages. Included in these were certain gene constellations containing Udorn HA, NA, and PB1 that confered low replicative fitness yet rapidly became dominant at the expense of more fit progeny, even when co-infection ratios of the two viruses favoured PR8. Fitness was not compromised, however, in the corresponding reassortants that also contained Udorn NP. Of particular note is the observation that relatively unfit reassortants could still fulfil the role of vaccine seed candidates as they provided high haemagglutinin (HA) antigen yields through co-production of non-infectious particles and/or by more HA molecules per virion. Our data illustrate the dynamics and complexity of reassortment and highlight how major gene segment interactions formed during packaging, in addition to antibody pressure, initially restrict the reassortant viruses that are formed. [ABSTRACT FROM AUTHOR]

Seasonal Influenza H3N2 virus poses a great threat to public health, but its vaccine efficacy remains suboptimal. One critical step in influenza vaccine production is the viral passage in embryonated eggs. Recently, the strength of egg passage adaptation was found to be rapidly increasing with time driven by convergent evolution at a set of functionally important codons in the hemagglutinin (HA1). In this study, we aim to take advantage of the negative correlation between egg passage adaptation and vaccine effectiveness (VE) and develop a computational tool for selecting the best candidate vaccine virus (CVV) for vaccine production. Using a probabilistic approach known as mutational mapping, we characterized the pattern of sequence evolution driven by egg passage adaptation and developed a new metric known as the adaptive distance (AD) which measures the overall strength of egg passage adaptation. We found that AD is negatively correlated with the influenza H3N2 vaccine effectiveness (VE) and ~75% of the variability in VE can be explained by AD. Based on these findings, we developed a computational package that can Measure the Adaptive Distance and predict vaccine Effectiveness (MADE). MADE provides a powerful tool for the community to calibrate the effect of egg passage adaptation and select more reliable strains with minimum egg-passaged changes as the seasonal A/H3N2 influenza vaccine.

Trypsin is a reagent widely used in the manufacture of biological medicinal products (BMPs). Until recently, pancreata of cattle, pigs and poultry were the main sources of trypsin preparations. The discovery of the disease called «transmissive spongiform encephalopathy» or «cow rabies» (TSE) in cattle in the late 1980s showed a clear need for limiting the use of this source. Given the potential risk of using trypsin obtained from cattle, porcine trypsin became more commonly used in the production of biological medicinal products. Enzymes obtained from raw materials of animal origin can be contaminated with circoviruses, parvo- and pestiviruses, and mycoplasmas that are common to pigs. Due to high resistance to physical and chemical treatment, these contaminants pose a potential risk to recipients of vaccines, as well as to other biological medicinal products. Prevention of contamination requires measures aimed at detection, reduction and inactivation of foreign agents, both in raw materials and during BMP production. The article considers the most common types of porcine trypsin contamination, methods of its detection, reduction and elimination. The article also contains information on the Russian and international requirements for the quality and safety of porcine trypsin used in the production of biological medicinal products.

The production of biological medicinal products used for prophylactic, diagnostic and therapeutic purposes includes the use of inherently variable biological processes and materials. In order to ensure the quality of the finished biological product it is necessary to determine the source, nature and fitness for use of the starting materials, including reagents, culture media, buffer solutions, sera, and enzymes. The article summarises literature data on the structure, properties and mode of action of the animal-derived reagent trypsin — proteolytic enzyme widely used in the production of biological medicinal products. The article dwells upon the sources of trypsin, methods of its production, requirements for trypsin products of different purity grades intended for human and veterinary use as well as for use as reagents in the production of vaccines, advanced therapy medicinal products and genetically engineered products. The article describes the role that trypsin plays in the human and animal intestinal digestive enzyme systems, in dissociation of cells in cultures in the process of cell passaging, in the mechanism of proteolitic activation and inactivation of a wide range of viruses, and in the examination of proteins primary structure.

With the COVID-19 pandemic now ongoing for close to a year, people all over the world are still waiting for a vaccine to become available. The initial focus of accelerated global research and development efforts to bring a vaccine to market as soon as possible was on novel platform technologies that promised speed but had limited history in the clinic. In contrast, recombinant protein vaccines, with numerous examples in the clinic for many years, missed out on the early wave of investments from government and industry. Emerging data are now surfacing suggesting that recombinant protein vaccines indeed might offer an advantage or complement to the nucleic acid or viral vector vaccines that will likely reach the clinic faster. Here, we summarize the current public information on the nature and on the development status of recombinant subunit antigens and adjuvants targeting SARS-CoV-2 infections. [Display omitted] • Recombinant protein vaccines constitute a proven, well-established vaccine development platform • Multiple expression systems are available to scale-up production • Various adjuvants are being used to elicit the desired immune response • Recombinant protein vaccines are suitable for production in low-resource countries • Recombinant protein COVID-19 vaccinesmay offer a better solution in the long-term [ABSTRACT FROM AUTHOR]

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Reuse reduces the consumption of freshwater supplies and the negative environmental impact caused by the discharge of industrial effluents. Some industries have already adopted this practice; however, no studies were found in the literature regarding this subject in the pharmaceutical industry. This work investigated the potential reuse of effluent (concentrate) generated from the Reverse Osmosis/Electro-deionization System (RO/EDI) that is used for the production of purified water in a Brazilian pharmaceutical plant. This industrial complex consumed about 200,000 m3 of water per year between 2012 and 2013 to produce one million of doses of vaccines, i.e., 2 L of water per dose of vaccine produced. During this period, the RO/EDI produced 27,000 m3 of purified water annually and generated 24,000 m3 of effluent (concentrate). This amount of effluent could be used to supply the production of industry steam (boilers) and/or cold water (cooling towers) that annually consumed an average of 12,000 m3 and 40,000 m3, respectively. The reuse of this effluent would result in a gross financial savings of 96,000 USD per year, excluding the costs of installation and control. From what has been researched in the literature, this work showed for the first time the possibility of reuse of effluent from RO/EDI System in the pharmaceutical industry.

Cell culture-based vaccine technology is a flexible and convenient approach for vaccine production that requires adaptation of the vaccine strains to the new cells. Driven by the motivation to develop a broadly permissive cell line for infection with a wide range of viruses, we identified a set of the most relevant host receptors involved in viral attachment and entry. This identification was done through a review of different viral entry pathways and host cell lines, and in the context of the Baltimore classification of viruses. In addition, we indicated the potential technical problems and proposed some solutions regarding how to modify the host cell genome in order to meet industrial requirements for mass production of antiviral vaccines. Our work contributes to a finer understanding of the importance of breaking the host–virus recognition specificities for the possibility of creating a cell line feasible for the production of vaccines against a broad spectrum of viruses. [ABSTRACT FROM AUTHOR]

In this study, a novel tube-fixed-bed bioreactor which consists of a TubeSpin bioreactor 50 tube and 0.44 g macrocarriers was developed as the scale-down model of a fixed-bed bioreactor. The adherent Vero cell–based pseudorabies virus (PRV) production process was tested in this novel model. The Vero cells grew well in the tube-fixed-bed bioreactor, and the cell density reached 5.8 × 106 cells/mL after 7 days of culture. The PRV production parameters (time of infection, multiplicity of infection, and harvest process) were optimized in the tube-fixed-bed bioreactor. Then the optimized process (time of infection = 3 days, multiplicity of infection = 0.001 and multiple harvest process) was scaled up 25-fold to an Xcell 1-L laboratory-scale fixed-bed bioreactor and 125-fold to an Xcell 5-L fixed-bed bioreactor successfully. The total PRV harvest in the Xcell 1-L bioreactor at 5 days after infection (dpi) was 10.25 log 10 TCID 50 which corresponds to 177,827 doses of vaccine. The total PRV harvest in the Xcell 5-L bioreactor at 5 dpi was 11.13 log 10 TCID 50 which corresponded to 1,348,962 doses of vaccine. The comparable growth curve, metabolism, and PRV production profile of the scaled-up bioreactors confirmed the feasibility and scalability of the tube-fixed-bed bioreactor as a scale-down model of the fixed-bed bioreactor for virus production process development. [ABSTRACT FROM AUTHOR]

The use of vaccines has become increasingly popular in preventing the causes of death. The Malaysian government has implemented a national immunization program that requires children to receive 12 types of vaccines since birth. Besides, the government also encourages adults to take certain vaccines to control harmful diseases. However, certain vaccines that produced indicate substances such as embryos from eggs and mammalian cells, the production of vaccines using these substances is because there is no other alternative. Due to this matter, this big issue has caused concern for Muslims to produce vaccines because of the excrement substances in the vaccine. This study deductively analyzes the principles and principles of Syariah related to the use of excrement for treating diseases. Therefore, this study summarizes the status of the vaccine based on excrement substances of the perspective of maqasid syariah. This study concludes that the production of vaccines containing excrement elements today follows the principles of maqasid syariah and specific conditions. [ABSTRACT FROM AUTHOR]

Although seaweeds have been used for decades as food, medicinal plants, and nutritional supplements, their use as a vaccine production and delivery platform has been barely explored, despite the availability of several genetic tools for seaweed transformation. The present review introduces seaweeds and their valuable immunomodulatory compounds in perspective as vaccine production and mucosal delivery platform. Genetic engineering tools are reviewed for macroalgal transformation, and the recombinant proteins produced in this platform are summarized. Finally, the hurdles and opportunities on seaweed potential as a vaccine production platform and delivery vehicle are proposed. As expected, seaweeds can soon be exploited to produce vaccine candidates due to their biological characteristics, including accelerated growth, photosynthetic capacity, and bioactive compound production, as well as the genetic engineering tools available. With all these properties -plus the commercial seaweed industry- this new envisioned technology can be adopted to improve human and animal health. • Macroalgae are sources of compounds with immunomodulatory properties. • Several approaches for genetic transformation in seaweeds have been used. • The most genetically engineered seaweeds have been red algae, followed by brown and green algae. • Seaweeds have successfully expressed recombinant enzymes and therapeutic drugs. • A macroalgae-made vaccine prototype (HbsAg) has been produced against Hepatitis B. [ABSTRACT FROM AUTHOR]

Introduction: Bovine neonatal pancytopenia (BNP) is a hemorrhagic disease that emerged in calves across Europe in 2007. Its occurrence is attributed to immunization of the calf’s mother with a vaccine produced using an allogeneic cell line. Vaccine-induced alloantibodies specific for major-histocompatibility class I antigens are transferred from the mother to the calf via colostrum, leading to profound depletion of peripheral blood and bone marrow cells that is often fatal. Areas covered: Pubmed and Web of Science were used to search for literature relevant to BNP and the use of allogeneic vaccine cell lines. Following a review of the pathology and pathogenesis of this novel condition, we discuss potential risks associated with the use of allogeneic vaccine cell lines. Expert commentary: Although BNP is associated with a specific vaccine, it highlights safety concerns common to all vaccines produced using allogeneic cell lines. Measures to prevent similar vaccine-induced alloimmune-mediated adverse events in the future are discussed.

Africa has the highest incidence of mortality caused by infectious diseases, and remarkably does not have the capacity to manufacture vaccines that are essential to reduce mortality, improving life expectancy, and promoting economic growth. GAVI has significantly helped introduction of new vaccines in Africa but its sustainability is questionable, and new vaccines introduction post-graduation is rare. Conversely, Africa with its high population and economy growth is an increasing potential market for vaccines. This study aimed to investigate how investment for vaccine production in Africa could be triggered and in which way it could be affordable to most African governments or investors. The investigation was based on a literature review and supplemented by online questionnaires directed to global vaccine stakeholders, African governments and regulatory authorities. In-depth interviews with experts in manufacturing capacity implementation and regulatory capacity building in Africa complemented the study. We also developed business plan scenarios including facility costs calculations and a possible investment plan based on expert opinions and publicly available information from pertinent sources. We saw that, governments in Africa, show interest in vaccine production establishments but only with external support for investment. The common regulatory functionality gap was the quality control laboratories to test vaccine lots before regulatory release. The global vaccine stakeholders showed less preference in investment for vaccine production establishment in Africa. The diverse political ambitions among African governments make it difficult to predict and access the market, a prerequisite for competitive production. A feasible solution could be a small production facility that would use technologies with high yield at low costs of goods to cover the regional needs. A respective antigen production facility is estimated to cost USD 25 Million, an affordable dimension for investors or interested African governments. Attractiveness for the African market is deemed to be high when targeting diseases almost exclusively for Africa (e.g., malaria or invasive non-typhoidal salmonella). With a smart 5 years tangible implementation plan, marketing agreements within existing regional collaborations and with a strong political will, an African government alone or together with an investor could convince global vaccine stakeholders and investors to support.

We discuss the complex eco-social factors involved in the puzzle of the unexpected rapid viral spread in the ongoing Brazilian yellow fever (YF) outbreak, which has increased the reurbanisation risk of a disease without urban cases in Brazil since 1942. Indeed, this rapid spatial viral dissemination to the Southeast and South regions, now circulating in the Atlantic Forest fragments close to peri-urban areas of the main Brazilian megalopolises (São Paulo and Rio de Janeiro) has led to an exponential increase in the number of yellow fever cases. In less than 18 months, 1,833 confirmed cases and 578 deaths were recorded most of them reported in the Southeast region (99,9%). Large epizooties in monkeys and other non-human primates (NHPs) were communicated in the country with 732 YF virus (YFV) laboratory confirmed events only in the 2017/2018 monitoring period. We also discuss the peculiarities and similarities of the current outbreak when compared with previous great epidemics, examining several hypotheses to explain the recent unexpected acceleration of epizootic waves in the sylvatic cycle of the YFV together with the role of human, NHPs and mosquito mobility with respect to viral spread. We conclude that the most feasible hypothesis to explain this rapidity would be related to human behavior combined with ecological changes that promoted a significant increase in mosquito and NHP densities and their contacts with humans. We emphasize the urgent need for an adequate response to this outbreak such as extending immunisation coverage to the whole Brazilian population and developing novel strategies for immunisation of NHPs confined in selected reserve areas and zoos. Finally, we stress the urgent need to improve the quality of response in order to prevent future outbreaks and a catastrophic reurbanisation of the disease in Brazil and other South American countries. Continuous monitoring of YFV receptivity and vulnerability conditions with effective control of the urban vector Aedes aegypti and significant investments in YF vaccine production capacity and research and development for reduction of adverse effects are of the highest priority.