Showing items 1 - 8 of 8

A novel ex vivo immunoproteomic approach characterising Fasciola hepatica tegumental antigens identified using immune antibody from resistant sheep

- Cameron, Timothy, Cooke, Ira, Faou, Pierre, Toet, Hayley, Piedrafita, David, Young, Neil, Rathinasamy, Vignesh, Beddoe, Travis, Anderson, Glenn, Dempster, Robert, Spithill, Terry

  • Authors: Cameron, Timothy , Cooke, Ira , Faou, Pierre , Toet, Hayley , Piedrafita, David , Young, Neil , Rathinasamy, Vignesh , Beddoe, Travis , Anderson, Glenn , Dempster, Robert , Spithill, Terry
  • Date: 2017
  • Type: Text , Journal article
  • Relation: International Journal for Parasitology Vol. 47, no. 9 (2017), p. 555-567
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  • Description: A more thorough understanding of the immunological interactions between Fasciola spp. and their hosts is required if we are to develop new immunotherapies to control fasciolosis. Deeper knowledge of the antigens that are the target of the acquired immune responses of definitive hosts against both Fasciola hepatica and Fasciola gigantica will potentially identify candidate vaccine antigens. Indonesian Thin Tail sheep express a high level of acquired immunity to infection by F. gigantica within 4 weeks of infection and antibodies in Indonesian Thin Tail sera can promote antibody-dependent cell-mediated cytotoxicity against the surface tegument of juvenile F. gigantica in vitro. Given the high protein sequence similarity between F. hepatica and F. gigantica, we hypothesised that antibody from F. gigantica-infected sheep could be used to identify the orthologous proteins in the tegument of F. hepatica. Purified IgG from the sera of F. gigantica-infected Indonesian Thin Tail sheep collected pre-infection and 4 weeks p.i. were incubated with live adult F. hepatica ex vivo and the immunosloughate (immunoprecipitate) formed was isolated and analysed via liquid chromatography-electrospray ionisation-tandem mass spectrometry to identify proteins involved in the immune response. A total of 38 proteins were identified at a significantly higher abundance in the immunosloughate using week 4 IgG, including eight predicted membrane proteins, 20 secreted proteins, nine proteins predicted to be associated with either the lysosomes, the cytoplasm or the cytoskeleton and one protein with an unknown cellular localization. Three of the membrane proteins are transporters including a multidrug resistance protein, an amino acid permease and a glucose transporter. Interestingly, a total of 21 of the 38 proteins matched with proteins recently reported to be associated with the proposed small exosome-like extracellular vesicles of adult F. hepatica, suggesting that the Indonesian Thin Tail week 4 IgG is either recognising individual proteins released from extracellular vesicles or is immunoprecipitating intact exosome-like extracellular vesicles. Five extracellular vesicle membrane proteins were identified including two proteins predicted to be associated with vesicle transport/ exocytosis (VPS4, vacuolar protein sorting-associated protein 4b and the Niemann-Pick C1 protein). RNAseq analysis of the developmental transcription of the 38 immunosloughate proteins showed that the sequences are expressed over a wide abundance range with 21/38 transcripts expressed at a relatively high level from metacercariae to the adult life cycle stage. A notable feature of the immunosloughates was the absence of cytosolic proteins which have been reported to be secreted markers for damage to adult flukes incubated in vitro, suggesting that the proteins observed are not inadvertent contaminants leaking from damaged flukes ex vivo. The identification of tegument protein antigens shared between F. gigantica and F. hepatica is beneficial in terms of the possible development of a dual purpose vaccine effective against both fluke species. © 2017 Australian Society for Parasitology

Cloning, expression, purification and crystallographic studies of galectin-11 from domestic sheep (Ovis aries)

- Sakthivel, Dhanasekaran, Littler, Dene, Shahine, Adam, Troy, Sally, Johnson, Matthew, Rossjohn, Jamie, Piedrafita, David, Beddoe, Travis

  • Authors: Sakthivel, Dhanasekaran , Littler, Dene , Shahine, Adam , Troy, Sally , Johnson, Matthew , Rossjohn, Jamie , Piedrafita, David , Beddoe, Travis
  • Date: 2015
  • Type: Text , Journal article
  • Relation: Acta Crystallographica Section:F Structural Biology Communications Vol. 71, no. (2015), p. 993-997
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  • Description: Galectins are an evolutionarily conserved family of proteins that translate glycan recognition into cellular effects. Galectin-11 is a unique member of the galectin family that is only expressed in ruminants such as sheep, goat and cattle and that plays a critical role in several important biological processes, such as reproduction and parasite-mediated innate immune responses. Currently, these two areas are of major importance for the sustainability of ruminant livestock production. Despite the emerging biological significance of galectin-11, no structural information is available. It is expected that structural studies will unravel the functional mechanisms of galectin-11 activity. Here, the expression, purification and crystallization of the ruminant-specific galectin-11 from domestic sheep and the collection of X-ray data to 2.0 Å resolution are reported. © 2015.

Evaluation of LAMP for Fasciola hepatica detection from faecal samples of experimentally and naturally infected cattle

- Bari, Tanjina, Al Mamun, Md Abdullah, Toet, Hayley, Rathinasamy, Vignesh, Larkins, Jo-Ann, Beddoe, Travis, Spithill, Terry, Piedrafita, David, Greenhill, Andrew

  • Authors: Bari, Tanjina , Al Mamun, Md Abdullah , Toet, Hayley , Rathinasamy, Vignesh , Larkins, Jo-Ann , Beddoe, Travis , Spithill, Terry , Piedrafita, David , Greenhill, Andrew
  • Date: 2024
  • Type: Text , Journal article
  • Relation: Veterinary Parasitology Vol. 327, no. (2024), p.
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  • Description: Fasciola hepatica causes liver fluke disease in production animals and humans worldwide. Faecal egg counts (FEC) are the most common diagnostic tool for the diagnosis of liver fluke disease. However, FEC has low sensitivity and is often unreliable for the detection of patent infection. In this study, loop-mediated isothermal amplification (LAMP) was optimised and evaluated for the detection of Fasciola hepatica infection, with the aim of increased sensitivity and making it suitable for on-farm application. LAMP was initially conducted under laboratory conditions, optimised to enable visual detection using calcein dye. DNA extraction based on bead-beating was developed to enable on-farm application. LAMP results were compared to FEC and polymerase chain reaction (PCR). Under laboratory conditions, LAMP was conducted using two incubation methods: a conventional PCR thermocycler and a field-deployable LAMP instrument. When compared to a ‘rigorous’ FEC protocol consisting of multiple counts using a comparatively large volume of faeces and with infection confirmed post-mortem, LAMP was highly sensitive and specific (using silica membrane DNA extraction sensitivity 88 %, specificity 100 %; using sieving and beat-beating DNA extraction sensitivity 98.9 %, specificity 100 %). When applied on-farm, LAMP was compared to conventional FEC, which suggested high sensitivity but low specificity (sensitivity 97 %, specificity 37.5 %). However, further analysis, comparing field LAMP results to laboratory PCR, suggested that the low specificity was likely the outcome of the inability of conventional FEC to detect all true F. hepatica positive samples. Based on the high sensitivity and specificity of LAMP compared to a ‘rigorous’ FEC protocol and its ability to be used in field settings, the study demonstrates the potential of LAMP for diagnosing F. hepatica infection in agriculture. © 2024 The Authors

Evaluation of the role of galectins in parasite immunity

- Preston, Sarah, Dunphy, Jillian, Beddoe, Travis, Meeusen, Els, Young, Anna

  • Authors: Preston, Sarah , Dunphy, Jillian , Beddoe, Travis , Meeusen, Els , Young, Anna
  • Date: 2014
  • Type: Text , Book chapter
  • Relation: Galectins: Methods and Protocols (Methods in Molecular Biology series) Chapter 25 p. 371-395
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  • Description: Galectin-11 and galectin-14 are ruminant galectins involved in parasitic infections. Although their roles in parasite immunity are still being elucidated, its appears that their functions are parasite specific. In gastrointestinal infections with the nematode Haemonchus contortus, both galectin-11 and galectin-14 appear to be protective. However, in a chronic infection of liver fluke, Fasciola hepatica, these galectins may aid parasite survival. This chapter discusses the methods designed to study parasitic infections in sheep, which have provided us with insight into the functions of galectin-11 and galectin-14 during host–parasite interactions. These methods include parasite cultivation and infection, galectin staining of host and parasite tissue, surface staining of parasites with recombinant galectins and in vitro assays to monitor the effect of galectins on larval development. © Springer Science+Business Media New York 2015.

Evaluation of the role of galectins in parasite immunity

- Swan, Jaclyn, Sakthivel, Dhanasekaran, Beddoe, Travis, Stear, Michael, Piedrafita, David, Preston, Sarah

  • Authors: Swan, Jaclyn , Sakthivel, Dhanasekaran , Beddoe, Travis , Stear, Michael , Piedrafita, David , Preston, Sarah
  • Date: 2022
  • Type: Text , Book chapter
  • Relation: Galectins: Methods in Molecular Biology Chapter 7 p. 475-515
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  • Description: Galectin-11 (LGALS-11) and galectin-14 (LGALS-14) are ruminant specific galectins, first reported in sheep. Although their roles in parasite immunity are still being elucidated, it appears that they influence protection against parasites. In gastrointestinal infections with the nematode Haemonchus contortus, both galectin-11 and galectin-14 appear to be protective. However, in a chronic infection of liver fluke, Fasciola hepatica, these galectins may aid parasite survival. To unravel the structural, functional, and ligand profile of galectin-11 and galectin-14, recombinant production of these proteins is vital. Here we present the recombinant production of soluble galectin-11 and galectin-14 from domestic sheep for in vitro and structural biology studies. These methods include parasite cultivation and infection, galectin staining of host and parasite tissue, surface staining of parasites with recombinant galectins, pull-down assays to identify endogenous galectin binding proteins, and in vitro assays to monitor the effect of galectins on parasite development. © 2022, Springer Science+Business Media, LLC, part of Springer Nature.

Galectin-11 : A novel host mediator targeting specific stages of the gastrointestinal nematode parasite, Haemonchus contortus

- Preston, Sarah, Beddoe, Travis, Walkden-Brown, Stephen, Meeusen, Els, Piedrafita, David

  • Authors: Preston, Sarah , Beddoe, Travis , Walkden-Brown, Stephen , Meeusen, Els , Piedrafita, David
  • Date: 2015
  • Type: Text , Journal article
  • Relation: International Journal for Parasitology Vol. 45, no. 12 (2015), p. 791-796
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  • Description: Galectin-11 is released from epithelial cells of the gastrointestinal tract, specifically following infection with gastrointestinal parasites including the highly pathogenic nematode, Haemonchus contortus. The function(s) of galectin-11 are currently unknown but seem to be associated with the development of immunity by the host. The aim of the present study was to examine the interaction of galectin-11 with the different parasitic life cycle stages of H. contortus and determine any effects on parasite development. The results of this study showed that galectin-11 binds to the surface of the L4 and adult stages of the parasite but not to the exsheathed L3 stage. In addition, at a lower concentration, binding to the L4 was specifically localised to the pharynx region. Subsequent in vitro assays demonstrated significant inhibition of larval growth and development in the presence of recombinant galectin-11. These results indicate, to our knowledge for the first time, a functional role for galectin-11 in gastrointestinal nematode infection of ruminants and a mechanism of action of galectin-11, targeting the development and growth of the L4 and possibly the adult parasite stage. © 2015 .

Proteomic identification of galectin-11 and 14 ligands from Haemonchus contortus

- Sakthivel, Dhanasekaran, Swan, Jaclyn, Preston, Sarah, Shakif-Azam, MD, Faou, Pierre, Jiao, Yaqing, Downs, Rachael, Rajapaksha, Harinda, Gasser, Robin, Piedrafita, David, Beddoe, Travis

  • Authors: Sakthivel, Dhanasekaran , Swan, Jaclyn , Preston, Sarah , Shakif-Azam, MD , Faou, Pierre , Jiao, Yaqing , Downs, Rachael , Rajapaksha, Harinda , Gasser, Robin , Piedrafita, David , Beddoe, Travis
  • Date: 2018
  • Type: Text , Journal article
  • Relation: Peerj Vol. 6, no. 3 (2018), p. 1-19
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  • Description: Haemonchus contortus is the most pathogenic nematode of small ruminants. Infection in sheep and goats results in anaemia that decreases animal productivity and can ultimately cause death. The involvement of ruminant-specific galectin-11 (LGALS-11) and galectin-14 (LGALS-14) has been postulated to play important roles in protective immune responses against parasitic infection; however, their ligands are unknown. In the current study, LGALS-11 and LGALS-14 ligands in H. contortus were identified from larval (L4) and adult parasitic stages extracts using immobilised LGALS-11 and LGALS-14 affinity column chromatography and mass spectrometry. Both LGALS-11 and LGALS-14 bound more putative protein targets in the adult stage of H. contortus (43 proteins) when compared to the larval stage (two proteins). Of the 43 proteins identified in the adult stage, 34 and 35 proteins were bound by LGALS-11 and LGALS-14, respectively, with 26 proteins binding to both galectins. Interestingly, hematophagous stage-specific sperm-coating protein and zinc metalloprotease (M13), which are known vaccine candidates, were identified as putative ligands of both LGALS-11 and LGALS- 14. The identification of glycoproteins of H. contortus by LGALS-11 and LGALS-14 provide new insights into host-parasite interactions and the potential for developing new interventions.

Vaccines for fasciola : new thinking for an old problem

- Spithill, Terry, Toet, Hayley, Rathinasamy, Vignesh, Zerna, Gemma, Swan, Jaclyn, Cameron, Timothy, Smooker, Peter, Piedrafita, David, Dempster, Robert, Beddoe, Travis

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