In the past Sarah C. Gilbert has collaborated on articles with Christopher Thomas Evans and Colin Ratledge. One of their most recent publications is A rapid screening method for lipid-accumulating yeast using a replica-printing technique. Which was published in journal Journal of Microbiological Methods.

More information about Sarah C. Gilbert research including statistics on their citations can be found on their Copernicus Academic profile page.

Sarah C. Gilbert's Articles: (9)

A rapid screening method for lipid-accumulating yeast using a replica-printing technique

AbstractA replica-printing technique has been developed which enables the tentative identification of a high lipid-containing yeast colony on an agar plate. The method involves a simple permeabilization and staining procedure, followed by destaining, and is able to detect colonies that accumulate lipid (that is those from Candida curvata, Lipomyces starkeyi and Trichosporon cutaneum) whilst easily distinguishing the latter from a low-lipid containing colony (Saccharomyces cerevisiae). The technique could not be successfully applied to the red yeast, Rhodosporidium toruloides. The method is simple and rapid and so provides for efficient screening and identification of high lipid-producing hybrids (protoplast fusion, recombinant DNA), mutants and natural isolates.

Research letterCarnitine acetyltransferase activity in oleaginous yeasts

AbstractThe highest activities of carnitine acetyltransferase (CAT) were found in non-oleaginous yeasts (Candida utilis and Saccharomyces cerevisiae); lower activities, ranging from 50% down to 3% of the highest values, were found in various strains of oleaginous yeasts (Candida curvata, Lipomyces starkeyi, Rhodosporidium toruloides and Trichosporon cutaneum). Supply of acetyl units into the cytosol of the latter, but not of the former yeasts, was therefore necessarily reliant on the action of ATP: citrate lyase (ACL), which was present in all oleaginous yeasts. There was no correlation, however, between the amount of lipid in the oleaginous yeasts and the specific activities of either CAT or ACL. Activity of CAT was increased up to 30-fold by growing yeasts on a triacylglycerol.

ReviewVaccine platforms for the prevention of Lassa fever

Highlights•The epidemiological significance of Lassa fever in West Africa is discussed.•Viral ecology, pathology, and immunobiology of Lassa virus infection is described.•Multiple vaccine candidates have been tested in pre-clinical models.•Lassa fever vaccine candidates have yet to progress to clinical trials.•Five platform technologies have been selected for expedited development.

Short communicationRapid generation of markerless recombinant MVA vaccines by en passant recombineering of a self-excising bacterial artificial chromosome

AbstractThe non-replicating poxviral vector modified vaccinia virus Ankara (MVA) is currently a leading candidate for development of novel recombinant vaccines against globally important diseases. The 1980s technology for making recombinant MVA (and other poxviruses) is powerful and robust, but relies on rare recombination events in poxviral-infected cells. In the 21st century, it has become possible to apply bacterial artificial chromosome (BAC) technology to poxviruses, as first demonstrated by B. Moss’ lab in 2002 for vaccinia virus. A similar BAC clone of MVA was subsequently derived, but while recombination-mediated genetic engineering for rapid production was used of deletion mutants, an alternative method was required for efficient insertion of transgenes. Furthermore “markerless” viruses, which carry no trace of the selectable marker used for their isolation, are increasingly required for clinical trials, and the viruses derived via the new method contained the BAC sequence in their genomic DNA. Two methods are adapted to MVA–BAC to provide more rapid generation of markerless recombinants in weeks rather than months. “En passant” recombineering is applied to the insertion of a transgene expression cassette and the removal of the selectable marker in bacteria; and a self-excising variant of MVA–BAC is constructed, in which the BAC cassette region is rapidly and efficiently lost from the viral genome following rescue of the BAC into infectious virus. These methods greatly facilitate and accelerate production of recombinant MVA, including markerless constructs.

A prime-boost immunisation regimen using DNA followed by recombinant modified vaccinia virus Ankara induces strong cellular immune responses against the Plasmodium falciparum TRAP antigen in chimpanzees

AbstractTwo chimpanzees were vaccinated intramuscularly against malaria using plasmid DNA expressing the pre-erythrocytic antigens thrombospondin related adhesion protein (PfTRAP) and liver stage specific antigen-1 (PfLSA-1) of Plasmodium falciparum together with GM-CSF protein. A recombinant modified vaccinia virus Ankara (MVA) expressing PfTRAP was injected intramuscularly 6 weeks later to boost the immune response. This sequence of antigen delivery induced a specific and long-lasting T cell and antibody response to PfTRAP as detected by ELISPOT assay and ELISA. Antibody responses were detected after four DNA injections, and were boosted by injection of recombinant MVA expressing PfTRAP. Interferon-gamma secreting antigen-specific T cells were detected in both animals, but only after boosting with recombinant MVA. By screening a panel of PfTRAP-derived peptides, an epitope was identified that was recognized by cytotoxic T lymphocytes in one of the chimpanzees studied. T cells specific for this epitope were present in PBMCs and liver-infiltrating lymphocytes at a frequency of between 1 in 200 and 1 in 500. The high immunogenicity of this prime-boost regimen in chimpanzees supports further assessment of this delivery strategy for the induction of protection against P. falciparum malaria in humans.

Dendritic cells infected by recombinant modified vaccinia virus Ankara retain immunogenicity in vivo despite in vitro dysfunction

AbstractThe administration of recombinant vaccinia virus Ankara (MVA) encoding a CTL epitope (pb9) from a malaria antigen induced activation and maturation of splenic dendritic cells (DCs) in vivo. In contrast, incubation of immature dendritic cells (iDCs) with the MVA, in vitro, resulted in down-regulation of MHC class I molecules and reduced their T-cell stimulatory ability. However, the ability of the infected DC to induce an antigen-specific CTL response, in vivo, remained intact. Furthermore, the administration of recombinant MVA-infected DC, but not pb9 peptide-pulsed DC, boosted and expanded the anti-pb9 CTL response that was primed by pb9 peptide-pulsed DC. These data indicate that despite the ability of poxviruses to impair DC maturation in vivo, the important ability of MVA to boost CD8 T-cell response in vivo is mediated at the level of the infected dendritic cells.

Synergistic DNA–MVA prime-boost vaccination regimes for malaria and tuberculosis

AbstractT-cell-mediated responses against the liver-stage of Plasmodium falciparum are critical for protection in the human irradiated sporozoite model and several animal models. Heterologous prime-boost approaches, employing plasmid DNA and viral vector delivery of malarial DNA sequences, have proved particularly promising for maximising T-cell-mediated protection in animal models. The T-cell responses induced by this prime-boost regime, in animals and humans, are substantially greater than the sum of the responses induced by DNA or MVA vaccines used alone, leading to the term introduced here of “synergistic” prime-boost immunisation. The insert in our first generation clinical constructs is known as multiple epitope-thrombospondin-related adhesion protein (ME-TRAP). We have performed an extensive series of phase I/II trials evaluating various prime-boost combination regimens for delivery of ME-TRAP in over 500 malaria-naïve and malaria-exposed individuals. The three delivery vectors are DNA, modified vaccinia virus Ankara (MVA) and, more recently, fowlpox strain 9 (FP9). Administration was intra-epidermal and intramuscular for DNA and intradermal for MVA and FP9. Doses of DNA ranged from 4 μg to 2 mg. Doses of MVA were up to 1.5 × 108 plaque forming units (pfu) and of FP9, up to 1.0 × 108 pfu. Further trials employing bacille Calmette–Guérin (BCG) as the priming agent and MVA expressing antigen 85A of Mycobacterium tuberculosis as the boosting agent has extended the scope of synergistic prime-boost vaccination. In this review we summarise the safety, immunogenicity and efficacy results from these malaria and tuberculosis vaccine clinical trials.

Immunogenicity of the candidate malaria vaccines FP9 and modified vaccinia virus Ankara encoding the pre-erythrocytic antigen ME–TRAP in 1–6 year old children in a malaria endemic area

AbstractIn a phase 1 trial, 22 children in a malaria endemic area were immunised with candidate malaria vaccination regimes. The regimes used two recombinant viral vectors, attenuated fowlpox strain FP9 and modified vaccinia virus Ankara (MVA). Both encoded the pre-erythrocytic malaria antigen construct ME–TRAP. Strong T cell responses were detected by both ex vivo and cultured ELISpot assays.Data from phase 1 trials in adults on anti-vector responses raised by FP9 is presented. These responses partially cross-reacted with MVA, and detectably reduced the immunogenicity of vaccination with MVA. This prompted the comparison of half dose and full dose FP9 priming vaccinations in children. Regimes using half dose FP9 priming tended to be more immunogenic than full dose.The potential for enhanced immunogenicity with half doses of priming vectors warrants further investigation, and larger studies to determine protection against malaria in children are required.

Tailoring subunit vaccine immunogenicity: Maximizing antibody and T cell responses by using combinations of adenovirus, poxvirus and protein-adjuvant vaccines against Plasmodium falciparum MSP1☆

AbstractSubunit vaccination modalities tend to induce particular immune effector responses. Viral vectors are well known for their ability to induce strong T cell responses, while protein-adjuvant vaccines have been used primarily for induction of antibody responses. Here, we demonstrate in mice using a Plasmodium falciparum merozoite surface protein 1 (PfMSP1) antigen that novel regimes combining adenovirus and poxvirus vectored vaccines with protein antigen in Montanide ISA720 adjuvant can achieve simultaneous antibody and T cell responses which equal, or in some cases surpass, the best immune responses achieved by either the viral vectors or the protein vaccine alone. Such broad responses can be achieved either using three-stage vaccination protocols, or with an equally effective two-stage protocol in which viral vectors are admixed with protein and adjuvant, and were apparent despite the use of a protein antigen that represented only a portion of the viral vector antigen. We describe further possible advantages of viral vectors in achieving consistent antibody priming, enhanced antibody avidity, and cytophilic isotype skew. These data strengthen the evidence that tailored combinations of vaccine platforms can achieve desired combinations of immune responses, and further encourage the co-administration of antibody-inducing recombinant protein vaccines with T cell- and antibody-inducing recombinant viral vectors as one strategy that may achieve protective blood-stage malaria immunity in humans.

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