This paper aims to elucidate the importance of TdaP vaccines in the prevention of pertussis, highlight the increased risk of this disease in adults, and underscore the benefits of using acellular vaccines over whole-cell vaccines.
This text underscores the escalating threat of Pertussis (whooping cough) in adults and its lethal repercussions on infants with lesser immunity. This has led to innovations in vaccine development, particularly the emergence of acellular vaccines like the TdaP vaccine that effectively combats Tetanus, Diphtheria, and Pertussis. This vaccine offers the advantage of fewer side effects compared to whole-cell vaccines and contains purified antigenic components of Bordetella pertussis obtained through various purification techniques.
The increased risk of pertussis in adults have become an important public-health issue. This contagious disease spreads to infants who have lesser immunity resulting in their death. Increased death rate caused the inventions of vaccinations to prevent this disease. Concerns regarding the safety of whole-cell vaccines used for pertussis treatment lead to the development of acellular pertussis vaccines like TdaP vaccines. This combination vaccine is 71-85% effective in the treatment of adults suffering from diseases tetanus, diphtheria and pertussis. These vaccines have limited side effects when compared to whole-cell vaccines as they contain purified antigenic components of Bordet Ella pertussis (Pertussis toxin, Filamentous hemagglutinin and Pertactin) that are obtained by the purification techniques Tangential flow filtration, Column Chromatography and Dialysis. These components are then formulated for purified vaccine production.
Table of Contents
2. INTRODUCTION
2.1 VACCINE:
2.1.1 HOW VACCINES ARE USED:
2.1.2 VACCINATION:
2.2 BORDETELLA PERTUSSIS
2.2.1 ABOUT BORDETELLA PERTUSSIS:
2.2.2 CAUSE OF DISESASE THROGH BACTERIA:
2.2.3 TYPES OF PERTUSSIS VACCINE:
2.2.4 PURIFICATION OF VACCINE
2.3 PERTACTIN (PRN)
2.3.1ORIGIN OF PRN
2.3.2 STRUCTURE OF PRN
2.4 FILAMENTOUS HEMAGGLUTION(FHA)
2.4.1 ABOUT FHA
2.4.2 NUCLEOTIDE SEQUCENCE OF FHA
3. REVIEW OF LITERATURE
3.1 AIM OF REVIEW:
3.2 MICROBIOLOGY OF BORDETELLA SPECIES:
3.3 MECHANISMS OF PATHOGENESIS:
3.4 EPIDEMIOLOGY:
3.4.1 Global burden of Pertussis
3.4.2 BURDEN OF PERTUSSIS IN INFANTS AND TODDLERS
3.4.3 BURDEN OF PERTUSSIS IN ADOLESCENTS AND ADULTS
3.4.4 TRANSMISSION DYNAMICS OF PERTUSSIS
3.5 IMPACT OF PERTUSSIS
3.6 POSTEXPOSURE PROPHYLAXIS AND THERAPEUTICS
3.6.1 Nonmacrolide Treatments
3.7 IMMUNITY TO PERTUSSIS
3.8 VACCINES AND IMMUNIZATIONS
3.8.1 DTP and DTaP Vaccines for Infants and Young Children
3.8.2 DIAGNOSIS
3.8.3 TYPES OF VACCINES
3.9 PROTEIN PURIFICATION
3.9.1 AIM OF PROTEIN PURIFICATION
3.9.2 TYPES OF CHROMATOGRAPHIC SYSTEM
4. MATERIALS AND METHODS
4.1TANGENTIAL FLOW FILTRATION
4.2 COLUMN CHROMATOGRAPHIC SYSTEM
4.3 DIALYSIS
5. PROCEDURE
5.1 PROCESS FOR PT AND FHA PURIFICATION:
5.2 PROCESS FOR PRN PURIFICATION:
6. RESULTS
Objectives and Topics
This work aims to develop an efficient, large-scale purification process for the antigenic components of Bordetella pertussis—specifically pertussis toxin (PT), filamentous hemagglutinin (FHA), and pertactin (PRN)—to facilitate the production of safer acellular pertussis vaccines compared to traditional whole-cell alternatives.
- Pathophysiology and epidemiology of Bordetella pertussis.
- Mechanisms of virulence factors (PT, FHA, PRN).
- Methodologies for protein purification, including Tangential Flow Filtration and Column Chromatography.
- Optimization of vaccine components for acellular vaccine formulation.
Excerpt from the Book
PROCEDURE
Acellular pertussis vaccine requires fermentation, isolation and purification of antigenic components i.e. pertussis toxoid (PT), filamentous hemagglutinin (FHA), pertactin (69kD protein), fimbriae and The number of doses per fermentation batch of acellular pertussis vaccine is usually 20-25 times lower than whole cell pertussis vaccine.
1. A fermentation batch was harvested for the separation of supernatant and pellet
2. This supernatant was used for the extraction of PT and FHA; while the pellet was used to extract PRN.
5.1 PROCESS FOR PT AND FHA PURIFICATION:
The supernatant from the harvest was concentrated using Tangential Flow Filtration system. During the first stage of tangential flow filtration UF permeate and UF retentate were obtained. The UF retentate acquired was concentrated with phosphate buffer of required composition to procure DF retentate. Parallelly chromatographic system set up was done i.e., slurry preparation, column packing and the column was washed and equilibrated with the corresponding buffers before the sample(DF retentate) was loaded. To the DF retentate 10% of tween-20 was added to prevent the aggregation. This DF retentate(load) was loaded onto anion exchange column chromatographic. The chromatographic system was manually run and the proteins PT and FHA were eluted at different salt concentrations (PT was eluted first with the lower salt concentration and FHA at later stage with little higher salt concentration) and were collected separately in different tubes and they were analysed for protein content and SDS-PAGE. After the analysis of protein content the PT containing samples were pooled together and FHA containing samples were also pooled together respectively in a separate glass bottles.
Summary of Chapters
2. INTRODUCTION: Covers fundamental concepts of vaccines, the biology of Bordetella pertussis, including its role as the causative agent of whooping cough, and the structural properties of its key antigenic proteins.
3. REVIEW OF LITERATURE: Provides an overview of the history, microbiology, pathogenic mechanisms, epidemiological impact, and historical vaccine development strategies related to pertussis.
4. MATERIALS AND METHODS: Details the essential equipment, chemicals, and specific scientific techniques—such as Tangential Flow Filtration, Column Chromatography, and Dialysis—employed for protein purification.
5. PROCEDURE: Outlines the step-by-step laboratory process for extracting and purifying PT, FHA, and PRN from fermentation batches for acellular vaccine production.
6. RESULTS: Presents the outcomes of the purification process, including SDS-PAGE verification of protein purity and the development/optimization of the process flow.
Keywords
Bordetella pertussis, Acellular pertussis vaccine, Protein purification, Pertussis toxin, Filamentous hemagglutinin, Pertactin, Tangential flow filtration, Column chromatography, Pathogenesis, Immunization, Vaccination, SDS-PAGE, Fermentation, Antigenic components, Whooping cough
Frequently Asked Questions
What is the primary objective of this research?
The research focuses on the optimization and development of robust, large-scale purification methods for key antigenic proteins (PT, FHA, and PRN) from Bordetella pertussis to support the manufacturing of safer acellular pertussis vaccines.
Which specific pathogens are targeted in this study?
The study specifically targets the bacterium Bordetella pertussis, which is responsible for the contagious respiratory disease known as pertussis or whooping cough.
What scientific methods are utilized for purifying the bacterial components?
The study employs several key biochemical techniques, primarily Tangential Flow Filtration for concentration/diafiltration, various modes of Column Chromatography (including anion exchange and multi-modal ion exchange), and Dialysis for buffer exchange.
Why are acellular vaccines preferred over whole-cell vaccines?
Acellular vaccines are preferred because they contain purified antigenic components, which significantly reduces the adverse side effects associated with whole-cell vaccines while maintaining high levels of efficacy.
What does the main body of the text cover?
The body covers a literature review of pertussis epidemiology and pathogenesis, a detailed methodology section on protein purification techniques, the experimental procedure used to extract the antigens, and the analytical results verifying the process.
What are the primary keys to success in this process development?
The success is attributed to optimizing fermentation scales (15L and 30L) and establishing a consistent and robust chromatographic purification pipeline.
How is the purity of the antigens verified?
The purity of the extracted PT, FHA, and PRN proteins is analyzed and verified using SDS-PAGE (Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis), which allows for the visual assessment of protein separation.
What role does the BvgAS system play in the bacteria's virulence?
The BvgAS two-component signal transduction system acts as a key regulatory system that enables Bordetella pertussis to control the expression of various virulence factors depending on temperature and host environmental conditions.
- Quote paper
- Hyacinthe Tuyubahe Praveen Kumar Vemuri (Author), 2023, Guarding Against Pertussis. The Efficacy and Advantages of the TdaP Vaccine, Munich, GRIN Verlag, https://www.grin.com/document/1351272
