Conference Agenda

Background and Aims

Fasciola hepatica is a globally distributed helminth of importance to both human and animal health. Critical information on how F. hepatica interacts with its mammalian host is lacking due to a reliance on animal models. This study aimed to validate a recently-developed HepG2-derived 3D spheroid co-culture model for F. hepatica newly excysted juveniles (NEJ) against in-vivo infection, using advanced microscopy methods.

Methods

In-vivo samples from C57BL/6 mice, infected with 175 metacercariae each, and NEJ grown in-vitro were collected at 12, 48, 120, 144, 168, 180, and 192 hours post infection. Comparison of the growth and development of external morphological (spines, sensory papillae, suckers) and internal anatomical (musculature, gut and uterine development) features between the two culture conditions were made using fluorescent confocal and scanning electron microscopy.

Results

The data obtained provides the first detailed morphological comparison of the growth and development of F. hepatica NEJ cultured in-vitro and in-vivo and serves as a benchmark to enhance future models.

Conclusion

The progression of animal-free models will enable exploration of the intricacies of early infection. Future work will incorporate spatial transcriptomic analysis to elucidate temporal shifts in F. hepatica development and the subsequent discovery of drug and vaccine targets.

Fasciola hepatica causes fasciolosis in livestock and humans worldwide, yet reliable tools to assess drug efficacy against the early developmental stages of this parasite are lacking. Here, we developed an automated miracidial motility assay (MMA) using the WMicroTracker ONE infrared detection system to quantify the sensitivity of F. hepatica miracidia to anthelmintic compounds including clorsulon (CLORS), closantel (CLOS), triclabendazole (TCBZ) and triclabendazole-sulphoxide (TCBZ-SO). Systematic optimisation of assay conditions, including inoculum size, observation window and solvent concentration yielded a reliable platform for evaluating the sensitivity of F. hepatica miracidia from diverse geographic isolates to these compounds. Our results demonstrated that three compounds (CLOS, TCBZ and TCBZ-SO) produced concentration-dependent motility inhibition, whereas CLORS had no effect. CLOS displayed the highest potency among isolates from New South Wales (NSW), Tasmania (TAS) and Victoria (VIC), whereas TCBZ and TCBZ-SO exhibited isolate-specific sensitivity patterns. Miracidial responses of the NSW and TAS isolates to TCBZ, TCBZ-SO and CLOS were also compared in vitro with those of newly excysted juveniles (NEJs) produced from the same isolates. Overall, the findings show that MMA provides a reproducible, host-independent and high-throughput phenotypic platform for assessing miracidial sensitivity to compounds.

Fasciolosis, caused by the liver fluke Fasciola hepatica, is a climate-sensitive parasitic disease that threatens livestock productivity and farm profitability, with broader implications for food security, rural livelihoods, and sustainable food systems. This study aimed to quantify historical trends and project future fasciolosis risk across Victoria under changing climate conditions. High-resolution (5 km²) climate data were retrieved for 40,504 grid points across Victoria to validate a Growing Degree Days (GDD) model for estimating fasciolosis risk over the past 50 years (1975–2024) and to project future risk centred at 2050 and 2090 under medium (RCP 4.5) and high (RCP 8.5) representative concentration pathways. Linear regression analysis demonstrated a strong relationship between observed fasciolosis prevalence and modelled risk values (R² = 0.94, p < 0.003). Historical analyses revealed substantial interannual variability, with consistently higher risk in eastern Victoria, followed by western and northern regions. Under future climate scenarios, risk increased spatially by up to two-fold in Barwon and three-fold in the Great South Coast, particularly under the 2090 RCP 8.5 scenario. These findings provide spatially explicit evidence to support climate-responsive surveillance, risk-based control strategies, and integrated animal health and environmental policy development.

Gastrointestinal nematodes impose economic burdens on the sheep industry, impacting on animal welfare and management costs. Species surveillance is essential, yet larval culture is inefficient for large‑scale monitoring. Nemabiome deep-amplicon sequencing allows for sensitive detection of nematode species directly from eggs without having to hatch eggs for 10-14 days like larval culture.

This study compared larval culture with nemabiome sequencing to speciate worms from bulk faecal samples from 27 farms. Spearman’s correlations showed significant positive correlations for Haemonchus contortus (r_s=0.56, p<0.01), and Teladorsagia circumcincta (r_s=0.58, p <0.01) and a moderate positive correlation for Trichostrongylus spp (r_s=0.34, p=0.07). Positive correlations between the two techniques provides some confidence that nemabiome sequencing is useful for large-scale surveillance of nematode species across Victoria, which is being completed on saleyard samples collected four times a year for a two-year period.

Initial findings on 100 samples show that Victoria’s most abundant species are Teladorsagia circumcincta (92%), Trichostrongylus vitrinus (78%) and Haemonchus contortus (54%). Given the high prevalence and pathogenicity of T. vitrinus compared to other Trichostrongylus species more research should focus on understanding this worm to enhance management strategies. Less common species included Trichostrongylus colubriformis, Chabertia ovina, Trichostrongylus axei, Oesophagostomum venulosum, Nematodirus spathiger and Trichurus ovis.

Increased drench resistance in sheep parasite populations necessitates alternative control measures. Unlike Barbervax, no vaccines exist for sheep scour worms. Detailed knowledge of protective immunity is essential for vaccine development designed to mimic natural host responses. In this study, 17-week-old parasite-naïve Merino lambs were exposed to weekly trickle infections of 6,000 infective Trichostrongylus colubriformis larvae for 16 weeks; controls remained parasite-free. After 16 weeks, mean faecal egg counts were 1518 and 0. Histologically, there was a time-dependent increase in intestinal goblet cells and decrease in jejunal mast cells in infected lambs. Flow cytometry at week 16 identified Ki67⁺GATA3⁺CD4⁺ T cells, indicating Th2 polarisation and expansion, with increased plasma and mucus anti-larval IgA and IgG. After drenching and challenge with 10,000 larvae, trickle-infected lambs rejected ~83% of larvae and had significantly lower worm burdens than controls. Anti-L3 T. colubriformis secretory IgA and IgG in bile may indicate portal immunoglobulin recycling from the gut. Additionally, in vitro antigen re-stimulation of cryopreserved intestinal cells showed increased Ki67⁺GATA3⁺CD4⁺ proliferating leukocytes with soluble L3 antigens. These results indicate 16 weeks of exposure creates a hostile intestinal environment for L3 establishment. Ongoing work assesses responses to L3/L4 antigens and associated ovine intestinal gene expression profiles.

Phenotypic screening readily identifies compounds that impair worm motility or development, the intrinsic biological processes underlying chemical sensitivity in parasitic nematodes remain poorly defined. Here, we identified a hit compound with a pyridyl scaffold from a phenotypic screen against the parasitic nematode, Haemonchus contortus, and using structure–activity optimisation we generated a potent chemical probe, WEHI-864. To uncover protein networks associated with the mechanism of action, thermal proteome profiling and time-resolved quantitative proteomics interrogated WEHI-864-induced perturbations in H. contortus. Across larval and adult stages of this major parasite of livestock, proteome integral solubility alteration (PISA) assay revealed reproducible alterations in proteins associated with cytoskeletal organisation and intracellular trafficking, including actin- and motor-related components. Complementary quantitative proteomics identified induction of an aspartyl protease and suppression of secretory CAP family proteins. Integrated analysis of these datasets supports a model in which chemical perturbation of cytoskeletal and trafficking proteins is associated with secondary modulation of proteolytic pathways, coinciding with rapid impairment of motility. These findings indicate that linked structural and proteolytic responses contribute to chemical sensitivity in H. contortus and demonstrate how integrative proteomics can resolve organism-level responses to chemical perturbation beyond single-target paradigms.