Generation of a Novel Bacteriophage Library displaying scFv antibody fragments from the natural Buffalo host to identify antigens from adult Schistosoma japonicum for diagnostic development
- Authors: Hosking, Christopher , McWilliam, Hamish , Driguez, Patrick , Piedrafita, David , Li, Yuesheng , McManus, Donald , Ilag, Leodevico , Meeusen, Els , De Veer, Michael
- Date: 2015
- Type: Text , Journal article
- Relation: PLoS Neglected Tropical Diseases Vol. 9, no. 12 (2015), p. 1-20
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- Description: The development of effective diagnostic tools will be essential in the continuing fight to reduce schistosome infection; however, the diagnostic tests available to date are generally laborious and difficult to implement in current parasite control strategies. We generated a series of single-chain antibody Fv domain (scFv) phage display libraries from the portal lymph node of field exposed water buffaloes, Bubalus bubalis, 11–12 days post challenge with Schistosoma japonicum cercariae. The selected scFv-phages showed clear enrichment towards adult schistosomes and excretory-secretory (ES) proteins by immunofluorescence, ELISA and western blot analysis. The enriched libraries were used to probe a schistosome specific protein microarray resulting in the recognition of a number of proteins, five of which were specific to schistosomes, with RNA expression predominantly in the adult life-stage based on interrogation of schistosome expressed sequence tags (EST). As the libraries were enriched by panning against ES products, these antigens may be excreted or secreted into the host vasculature and hence may make good targets for a diagnostic assay. Further selection of the scFv library against infected mouse sera identified five soluble scFv clones that could selectively recognise soluble whole adult preparations (SWAP) relative to an irrelevant protein control (ovalbumin). Furthermore, two of the identified scFv clones also selectively recognised SWAP proteins when spiked into naïve mouse sera. These host B-cell derived scFvs that specifically bind to schistosome protein preparations will be valuable reagents for further development of a cost effective point-of-care diagnostic test. © 2015 Hosking et al.
Using the local immune response from the natural buffalo host to generate an antibody fragment library that binds the early larval stages of Schistosoma japonicum
- Authors: Hosking, Christopher , Driguez, Patrick , McWilliam, Hamish , Ilag, Leodevico , Gladman, Simon , Li, Yuesheng , Piedrafita, David , McManus, Donald , Meeusen, Els , De Veer, Michael
- Date: 2015
- Type: Text , Journal article
- Relation: International Journal for Parasitology Vol. 45, no. 11 (2015), p. 729-740
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- Description: Antibodies isolated from the local draining inguinal lymph node of field exposed-water buffaloes following challenge with Schistosoma japonicum cercariae showed high reactivity towards S. japonicum antigen preparations and bound specifically to formaldehyde-fixed S. japonicum schistosomules. Using this specific local immune response we produced a series of single-chain antibody Fv domain libraries from the same lymph nodes. Removal of phage that cross reacted with epitopes on adult parasites yielded a single-chain antibody Fv domain-phage library that specifically bound to whole formaldehyde-fixed and live S. japonicum schistosomules. DNA sequencing indicated clear enrichment of the single-chain antibody Fv domain library for buffalo B-cell complementarity determining regions post-selection for schistosomule binding. This study also revealed that long heavy chain complementarity determining regions appear to be an important factor when selecting for antibody binding fragments against schistosomule proteins. The selected single-chain antibody Fv domain-phage were used to probe a schistosome-specific protein microarray, which resulted in the recognition of many proteins expressed across all schistosome life-cycle stages. Following absorption to adult worms, the single-chain antibody Fv domain-phage library showed significantly reduced binding to most proteins, whilst two proteins (NCBI GenBank accession numbers AY915878 and AY815196) showed increased binding. We have thus developed a unique set of host derived single-chain antibody Fv domains comprising buffalo B-cell variable regions that specifically bind to early S. japonicum life-stages. © 2015 Australian Society for Parasitology Inc..
The effect of different adjuvants on immune parameters and protection following vaccination of sheep with a larval-specific antigen of the gastrointestinal nematode, Haemonchus contortus
- Authors: Piedrafita, David , Preston, Sarah , Kemp, Joanna , De Veer, Michael , Sherrard, Jayne , Kraska, Troy , Elhay, Martin , Meeusen, Els
- Date: 2013
- Type: Text , Journal article
- Relation: PLoS ONE. Vol. 8, no. 10, (Art. no.e78357) (2013), p. 1-8
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- Description: It has recently been recognised that vaccine adjuvants play a critical role in directing the nature of a vaccine induced effector response. In the present study, several adjuvants were evaluated for their ability to protect sheep after field vaccination with the larval-specific Haemonchus contortus antigen, HcsL3. Using a suboptimal antigen dose, aluminium adjuvant was shown to reduce the cumulative faecal egg counts (cFEC) and worm burden by 23% and 25% respectively, in agreement with a previous study. The addition of Quil A to the aluminium-adjuvanted vaccine brought cFEC back to control levels. Vaccination with the adjuvant DEAE-dextran almost doubled the protection compared to the aluminium-adjuvanted vaccine resulting in 40% and 41% reduction in cFEC and worm counts compared to controls. Examination of skin responses following i.d. injection of exsheathed L3, revealed that cFEC was negatively correlated with wheal size and tissue eosinophils for the DEAE-dextran and aluminium-adjuvanted groups respectively. These studies have for the first time shown the potential of DEAE-dextran adjuvant for helminth vaccines, and discovered significant cellular correlates of vaccine-induced protection. Funding: Australian Research Council Linkage grant LP0668945 and the ARC-Centre of Excellence in Structural and Functional Microbial Genomics. The funders have no role in study design, data collection and analysis, decision to publish, or preparation of the manuscrip
Field vaccination of sheep with a larval-specific antigen of the gastrointestinal nematode, Haemonchus contortus, confers significant protection against an experimental challenge infection
- Authors: Piedrafita, David , De Veer, Michael , Lydall, Jayne , Kraska, Troy , Elhay, Martin , Meeusen, Els
- Date: 2012
- Type: Text , Journal article
- Relation: Vaccine Vol. 30, no. 50 (2012), p. 7199-7204
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- Description: The availability of effective vaccines would add a valuable tool to the management of gastrointestinal nematode infections in livestock. While some experimental vaccines have shown protection in laboratory trials, few have been tested in the field. In the present study, eight month old sheep kept on pasture were treated with anthelmintic 8 weeks before vaccination with a larval surface antigen of the nematode parasite, Haemonchus contortus, under a commercially acceptable protocol, i.e. 2 immunizations using a commercial adjuvant; they were then given a controlled challenge infection 4 weeks later in indoor pens. Vaccination of sheep with 4 increasing doses of antigen resulted in significant reductions of 61% and 27% in cumulative faecal egg counts in the two highest dose groups, and a 69% reduction in worm burden in the highest dose group. Blood loss, as determined by packed cell volume, was also significantly reduced in the highest dose group of sheep. One outlier sheep showed an unusual increase in egg count without a concomitant increase in worm burden compared to the control sheep, indicating a vaccine-induced stress response. Antigen-specific serum antibody levels steadily increased in sheep while on pasture and decreased when transported to indoor pens. No difference in antibody levels could be detected between vaccinated and unvaccinated sheep, but all showed increased antibody levels compared to uninfected control sheep kept in indoors pens for 2–3 months, suggesting sheep were sensitized to the larval antigen either from low dose pasture contamination or cross reaction with pasture-related antigens. The results of these studies confirm the protective properties of the larval surface antigen and its protective effect when vaccinations are performed in the field.