Rachel Chikwamba

• Council for Scientific and Industrial Research, South Africa

The hypersensitive response is elicited by Agrobacterium infiltration of Nicotiana benthamiana, including the induction and accumulation of pathogenesis-related proteins, such as proteases. This includes the induction of the expression of several cysteine proteases from the C1 (papain-like cysteine protease) and C13 (legumain-like cysteine protease...

Broadly neutralising antibodies (bNAbs) against human immunodeficiency virus type 1 (HIV-1), such as CAP256-VRC26 are being developed for HIV prevention and treatment. These Abs carry a unique but crucial post-translational modification (PTM), namely O-sulfated tyrosine in the heavy chain complementarity determining region (CDR) H3 loop. Several st...

Africa is one of the world’s fasted growing economies, with South Africa having the fifth highest worldwide pharmaceutical expenditure per capita. In recent years, several companies have considered regional pharmaceutical production but have failed to make the investment, in stark contrast to the massive growth in pharmaceutical production in other...

Rabies is an ancient and neglected zoonotic disease caused by the rabies virus, a neurotropic RNA virus that belongs to the Rhabdoviridae family, genus Lyssavirus. It remains an important public health problem as there are cost and health concerns imposed by the current human post exposure prophylaxis therapy. The use of monoclonal antibodies (mAbs...

SDS-PAGE showing mAbs after protein A purification. PageRuler Prestained Protein ladder that indicated molecular masses in kDa, was loaded in lane 1. The mAbs E559 and the 62-71-3 were loaded in lane 2 and 3, respectively. (TIF)

Far-UV CD spectra of E559 and 62-71-3 mAbs. Far-UV CD spectra of E559 (grey dotted line) and 62-71-3 (black solid line) at a concentration of 2 μM. E559 had a total of 1324 residues while 62-71-3 had 1328 residues. Readings were taken in a 1 mm cuvette at 20°C. (TIF)

Averages of the deuterium percentages and standard deviation for E559 and 62-71-3. Evaluation of the standard deviation between the deuterium up take for the E559 (A) plus 62-71-3 (B) peptides that were plotted in Fig 6 and E559 (C) plus 62-71-3 (D) peptides that were plotted in Fig 7, for Cycle 0,1,3,5 and 7. (TIF)

Supplimentary information. E559 peptide list, oxidation time points and total area. (CSV)

Supplimentary information_62-71-3. 62-71-3 oxidation spectral biospec library. (BLIB)

Ramachandran results for E559 and 62-71-3 mAb Fab and Fc regions. Ramachandran plot statistics and G-factor parameters (TIF)

Supplimentary information_E559. E559 oxidation spectral biospec library. (BLIB)

Effect of freeze-thawing on the stability profile of E559 and 62-71-3. The full (blue) molecular mass for E559 is 145.5 kDa and 145.4 kDa for 62-71-3. LMM (red) indicate sizes lower than the full mAb while HMM (green) indicates sizes higher than the full mAb. LMM (low molecular mass), HMM (high molecular mass). (TIF)

Computationally modelled Fab regions of E559 (A) and 62-71-3 (B) mAb and the Fc region (C). One unit of the E559 (A) and 62-71-3 (B) FAB region and Fc dimer region (C) for both antibodies was modelled. The β-sheets were coloured in yellow, helices were coloured red and the coils are in green. The schematic mAb illustrates how the individual regions...

Supplimentary information. 62-71-3 peptide list, oxidation time points and total area. (CSV)

Figure S1 GRFT accumulation in the apoplast induced an HR‐like cell death response. Figure S2 Schematic representation of MagnICON promodule vectors used for GRFT and NahG expression. Figure S3 GRFT accumulation in the apoplast induced an HR‐like cell death response using binary vector. Figure S4 NahG and GRFTlec‐ did not induce cell death in N....

Rabies is a significant neglected vaccine-preventable disease that is global in distribution. Multiple biologics are utilized in routine prevention and control of this zoonosis. Currently, rabies vaccines are used to interrupt a productive viral encephalitis before or after pathogen exposure in humans and animals. In addition, rabies immune globuli...

Griffithsin (GRFT) is an antiviral lectin, originally derived from a red alga, which is currently being investigated as a topical microbicide to prevent transmission of human immunodeficiency virus (HIV). Targeting GRFT to the apoplast for production in Nicotiana benthamiana resulted in necrotic symptoms associated with a hypersensitive response (H...

Enterotoxaemia (pulpy kidney) is a common bacterial disease of sheep caused by Clostridium perfringens type D epsilon toxin. It has mortality rates of up to 30% in non-vaccinated animals. Current vaccines from whole cell cultures are expensive to manufacture and can induce local inflammatory responses in sheep. They usually have reduced immunogenic...

Rabies is an acute viral encephalomyelitis in warm-blooded vertebrates, caused by viruses belonging to Rhabdovirus family and genus Lyssavirus. Although rabies is categorised as a neglected disease, the rabies virus (RABV) is the most studied amongst Lyssaviruses which show nearly identical infection patterns. In efforts to improving post-exposure...

Rabies is a neglected zoonotic disease that has no effective treatment after onset of illness. However the disease can be prevented effectively by prompt administration of post exposure prophylaxis which includes administration of passive immunizing antibodies (Rabies Immune Globulin, RIG). Currently, human RIG suffers from many restrictions includ...

Sorghum is particularly drought tolerant compared with other cereal crops and is favoured for subsistence farming in water scarce regions of the world. This study was conducted to identify South African sorghum landraces with superior drought tolerance compared with a drought-tolerant breeding line (P898012). Seedlings of 14 South African sorghum l...

Rabies post-exposure prophylaxis (PEP) currently comprises administration of rabies vaccine together with rabies immunoglobulin (RIG) of either equine or human origin. In the developing world, RIG preparations are expensive, often in short supply, and of variable efficacy. Therefore, we are seeking to develop a monoclonal antibody cocktail to repla...

Key message: Co-suppressing major kafirin sub-classes is fundamental to improved protein digestibility and nutritional value of sorghum. The improvement is linked to an irregularly invaginated phenotype of protein bodies. Abstract: The combined suppression of only two genes, γ kafirin-1 (25 kDa) and γ-kafirin-2 (50 kDa), significantly increases...

Recombinant subunit vaccines are an efficient strategy to meet the demands of a possible influenza pandemic, because of rapid and scalable production. However, vaccines made from recombinant hemagglutinin (HA) subunit protein are often of low potency, requiring high dose or boosting to generate a sustained immune response. We have improved the immu...

We examined the ability of HIV-1 subtype C to develop resistance to the inhibitory lectins, griffithsin (GRFT), cyanovirin-N (CV-N) and scytovirin (SVN), which bind multiple mannose-rich glycans on gp120. Four primary HIV-1 strains cultured under escalating concentrations of these lectins became increasingly resistant tolerating 2 to 12 times their...

Molecular Pharming, the production of recombinant pharmaceuticals through plant biotechnology, has the potential to transform the biologics sector of the pharmaceutical industry. More fascinating however, is how it might be used to improve access to modern medicines, and improve health of the poor in developing countries and emerging economies. Alt...

Physical and biochemical analysis of protein polymorphisms in seed storage proteins of a mutant population of sorghum revealed a mutant with redirected accumulation of kafirin proteins in the germ. The change in storage proteins was accompanied by an unusually high level accumulation of free lysine and other essential amino acids in the endosperm....

Differences between the coding sequences of two banana homologues of the Non-expressor of Pathogenesis-Related 1 (NPR1) genes (MNPR1A and MNPR1B) were investigated as a possible cause for the differential activity of the two genes. Each of the MNPR1 coding sequences were expressed under the control of the cauliflower mosaic virus 35S promoter/termi...

It is generally believed that during the sexual transmission of HIV-1, the glycan-specific DC-SIGN receptor binds the virus and mediates its transfer to CD4(+) cells. The lectins griffithsin (GRFT), cyanovirin-N (CV-N) and scytovirin (SVN) inhibit HIV-1 infection by binding to mannose-rich glycans on gp120. We measured the ability of these lectins...

Background: The lectins Griffithsin (GRFT), Cyanovirin-N (CVN) and Scytovirin (SVN) are potential microbicides that inhibit HIV-1 infection by binding to mannose-rich glycans on the envelope. Since the DC-SIGN receptor plays a key role in the sexual transmission of HIV-1, we investigated whether these lectins could inhibit HIV-1 transfer and infect...

The glycans on HIV-1 gp120 play an important role in shielding neutralization-sensitive epitopes from antibody recognition. They also serve as targets for lectins that bind mannose-rich glycans. In this study, we investigated the interaction of the lectin griffithsin (GRFT) with HIV-1 gp120 and its effects on exposure of the CD4-binding site (CD4bs...

The field of plant-made therapeutics in South Africa is well established in the form of exploitation of the country's considerable natural plant diversity, both in the use of native plants in traditional herbal medicines over many centuries, and in the more modern extraction of pharmacologically-active compounds from plants, including those known t...

Genetically modified organisms are expected to have a large impact on the ability of humanity to feed, fuel and heal itself in light of the growing global change, and adverse geo-climatic conditions anticipated as a result of climate change.

The thrust of biotechnology research in Zimbabwe is to promote already developed technologies, especially for commercialized crops such as tobacco, wheat, cotton and sugarcane. This involve tissue culture techniques for disease elimination and rapid propagation, development of transgenic plants (tobacco) and development of biopesticides and biofert...

Griffithsin (GRFT), Cyanovirin-N (CV-N) and Scytovirin (SVN) are lectins that inhibit HIV-1 infection by binding to multiple mannose-rich glycans on the HIV-1 envelope glycoproteins (Env). Here we show that these lectins neutralize subtype C primary virus isolates in addition to Env-pseudotyped viruses obtained from plasma and cervical vaginal lava...

The efficiency of the bialophos ( bar ) and phosphomannose isomerase ( pmi ) selectable markers in microprojectile mediated transformation of P898012, a sorghum inbred line generally considered amenable to in vitro manipulation was investigated. Two plasmids containing bar gene, encoding bialaphos resistance and manA gene, for phosphomannose isomer...

Southern Africa is faced with the complex challenge of achieving sustainable economic development and food and energy security while protecting the environment. The region is currently experiencing an energy crisis, a result in part because of global increases in crude oil prices and limited generating capacity in some countries. We examined the po...

The non-expressor of pathogenesis-related genes 1 (NPR1) is an essential positive regulator of salicylic acid (SA)-induced pathogenesis-related (PR) gene expression and systemic acquired resistance (SAR). Two novel full length NPR1-like genes; MNPR1A and MNPR1B, were isolated from banana by application of the PCR and rapid amplification of cDNA end...

Regeneration capacity of five sorghum genotypes on three different solid nutrient media viz., medium J, Tadesse`s and CAPD were investigated. The study analysed the ability of genotypes to form totipotent callus cultures and plantlets within the shortest culture period in vitro . We observed that regeneration of plants was mainly through somatic em...

In the past few years, plant biotechnology has gone beyond traditional agricultural production of food, feed and fibre, and moved to address more complex contemporary health, social and industrial challenges. The new era involves production of novel pharmaceutical products, speciality and fine chemicals, phytoremediation and production of renewable...

Plant-derived pharmaceuticals are poised to become the next major commercial development in biotechnology. The advantages they offer in terms of production scale and economy, product safety, ease of storage and distribution cannot be matched by any current commercial system; they also provide the most promising opportunity to supply low-cost drugs...

This chapter discusses the advances in the field of transgenic plant-derived mucosal vaccines, focusing on the results of human clinical trials and on orally delivered animal vaccines. The transgenic plant-derived vaccine antigens are known to be orally immunogenic in humans and several animals species. The choice of crop for production of vaccines...

IntroductionMucosal Immunization Vaccination Strategies for Infectious DiseasesMucosal Immunization vs. Parenteral ImmunizationMucosal Immunization and AdjuvantsPlant-derived Edible Vaccines Advantages of the Plant-based SystemTransient and Stable Systems for Production of Plant-derived ProteinsChoice of Plants and Plant TissuesPlant-expression Sys...

The B subunit of Escherichia coli heat labile enterotoxin (LT-B) is a potent oral immunogen with potential for use as a vaccine, a carrier molecule to deliver antigens to gut-associated lymphoid tissues, and possibly an adjuvant to make coadministered vaccines more effective. LT-B produced in plants was shown to be functional and immunogenic in ani...

We have produced transgenic maize plants containing a wheat Glu-1Dx5 gene encoding the high-molecular-weight glutenin subunit 1Dx5. Analysis by SDS-PAGE showed that a protein similar in size to the wheat 1Dx5 subunit accumulates in the endosperm of transgenic maize from four independent transformation events. This protein reacts with a monoclonal a...

We have produced a functional heat labile enterotoxin (LT-) B subunit of Escherichia coli in maize. LT-B is a multimeric protein that presents an ideal model for an edible vaccine, displaying stability in the gut and inducing mucosal and systemic immune responses. Transgenic maize was engineered to synthesize the LT-B polypeptides, which assembled...

We have achieved routine transformation of maize (Zea mays) using an Agrobacterium tumefaciens standard binary (non-super binary) vector system. Immature zygotic embryos of the hybrid line Hi II were infected with A. tumefaciens strain EHA101 harboring a standard binary vector and cocultivated in the presence of 400 mg L-1 L-cysteine. Inclusion of...

We have produced the B subunit of the enterotoxigenic Escherichia coli (ETEC) heat-labile enterotoxin (LT-B) in transgenic maize seed. LT-B is a model antigen that induces a strong immune response upon oral administration and enhances immune responses to conjugated and co-administered antigens. Using a synthetic LT-B gene with optimized codon seque...

The ability to transfer foreign genes to grain sorghum would potentially allow production of cultivars with improved insect resistance and grain quality (nutritional, baking and brewing). We have established an Agrobacterium-medmted transformation protocol, based predominantly on axenic seedlings of inbred lines. After inoculation of Agrobacterium...

Rabies, an important disease in Asia and Africa, is an acute viral disease of the central nervous system that affects humans and other mammals. Upon bites or contact with rabid animals patients are immediately immunized with antibodies followed by human diploid cell vaccine (HDCV). Lack of post-exposure treatment usually results in 100% fatality. M...

CSIR Research and Innovation Conference: 1st CSIR Biennial Conference, CSIR International Convention Centre Pretoria, 27-28 February 2006 The demand for health products such as protein therapeutics and sub-unit vaccines to control infectious diseases far outweighs the current global production capacity of these molecules. These products are current...

Bio2Biz Conference, 29 Septemner 2006 The presentation outlines the global status of food security and implications for meeting the Millennium Development Goals targets, with particular focus on Sub-Saharan Africa; trends in hunger, food availability, food distribution, consumption; underlying causes of food insecurity; and the role of technology i...

Plants are becoming increasingly important as a production system for biopharmaceuticals and industrially important proteins. The work presented in this dissertation showed that maize can be used as a source and delivery vehicle for oral vaccines. Antigentic proteins from two economically important pathogens, enterotoxigenic Escherichia coli (E. co...

CSIR 3rd Biennial Conference 2010. Science Real and Relevant. CSIR International Convention Centre, Pretoria, South Africa, 30 August – 01 September 2010 HIV prevalence is over 33 million worldwide with 68% of AIDS sufferers residing in sub-Saharan Africa (1). Currently the available HIV prevention tools are feasible but women cannot insist on thes...

CSIR 3rd Biennial Conference 2010. Science Real and Relevant. CSIR International Convention Centre, Pretoria, South Africa, 30 August – 01 September 2010 Due to the increasing demand for pharmaceuticals and the high cost of production in traditional fermentor systems, plants are being considered as alternative production platforms for therapeutic m...