Alex C. Loukas

Description:Different agencies have emphasized the need to evaluate current serological methods for screening patients with suspected urogenital schistosomiasis. However, there is still a lack of evidence regarding the most appropriate methods for this purpose. Here we assessed the diagnostic efficacy of a newly developed serological technique that utilizes the recombinant protein Sh-TSP-2, applied to the urine and serum of migrants suspected of having urogenital schistosomiasis. The sensitivity, specificity, positive and negative predictive values of an in-house enzyme-linked immunosorbent assay (ELISA) using the recombinant protein Sh-TSP-2 were analysed and compared with other commercial serological methods. Due to the limitations of microscopy as a perfect reference method, a latent class analysis (LCA) and composite reference standard (CRS) approach was used to determine the sensitivity and specificity of each test. According to the LCA model, the commercial tests NovaLisa® and immunochromatography test (ICT) immunoglobulin G-immunoglobulin M (IgG-IgM) presented the highest sensitivity (100%), whereas the Sh-TSP-2 serum ELISA test had 79.2%. The Sh-TSP-2 urine and serum ELISA tests had the highest specificities among the serological methods (87.5 and 75%, respectively). CRS modelling showed that the ICT IgG-IgM, NovaLisa® and Sh-TSP-2 serum tests led in sensitivity at 97.1, 88.6 and 71.4%, respectively, with all tests except that the ICT IgG-IgM test having a specificity >90%. Sh-TSP-2 has been validated as a screening tool for patients suspected of having urogenital schistosomiasis. Although commercial serological tests have shown higher sensitivities, Sh-TSP-2 could be valuable for confirming results from tests with lower specificity. Nevertheless, further studies with larger patient cohorts are necessary to fully verify its potential.

Description:There are numerous examples of topological isomers in organic chemistry, but such isomers are rare in disulfide-rich peptides. Here we characterise two structurally well-defined topological isomers in a peptide (GRN-P4A) containing the mini-granulin fold. The mini-granulin fold is emerging as an important disulfide-rich structural motif with promising implications for the enhancement of wound healing strategies. The two topological isomers of GRN-P4A have well-defined structures that do not interconvert, and although they have the same disulfide bond connectivity and similar overall structures, they have structural differences related to the first inter-cysteine loop. These structural changes influence the bioactivity as the isomers have significant differences in their cell proliferation activity. Prediction of the structure using AlphaFold3 identified the correct disulfide bond connectivity, but the structure of loop 1 was similar to the less abundant isomer of GRN-P4A and did not indicate topological isomerisation. These topological isomers introduce significant complexity to the understanding of folding mechanisms in this class of peptides, and potentially other disulfide-rich peptides, offering valuable insights for protein design and engineering by presenting a novel topological fold-switching mechanism. Additionally, they hold practical implications for the production of GRN-P4A, given its promising potential as a wound-healing agent.

Description:Parasitic gastrointestinal worms (i.e., helminths) remain a significant global health and economic burden. The increasing inefficacy of current anthelmintic drugs against parasitic diseases necessitates the discovery of novel therapeutic options. This study investigated the anthelmintic properties and therapeutic potential of stonefish ichthyocrinotoxins (i.e., secreted skin toxins). xWORM (xCELLigence Worm Real-Time Motility Assay) was used to evaluate the anthelmintic activity of ichthyocrinotoxins from two stonefish species, Synanceia horrida (Estuarine Stonefish) and Synanceia verrucosa (Reef Stonefish), against the infective third-stage larvae of Nippostrongylus brasiliensis (Rodent Hookworm). Both toxins demonstrated potent anthelmintic effects, with S. horrida ichthyocrinotoxin exhibiting greater potency (IC50 = 196.0 µg/mL) compared to ichthyocrinotoxin from S. verrucosa (IC50 = 329.7 µg/mL). Fractionation revealed that the anthelmintic activity of S. verrucosa is likely driven by synergistic interactions between the large (>3 kDa) and small (<3 kDa) components. In contrast, the small components isolated from S. horrida ichthyocrinotoxin were responsible for the majority of the observed activity, making them a more attractive therapeutic candidate. Furthermore, despite the cytotoxicity of crude S. horrida ichthyocrinotoxin against human skin and bile duct cell lines, the isolated small components exhibited potent anthelmintic effects (IC50 = 70.5 µg/mL) with negligible cytotoxicity (<10% decrease in survival at 100 µg/mL). While further research is necessary to fully characterise these compounds and assess their clinical suitability, this study highlights the potential of stonefish ichthyocrinotoxins as a novel source of anthelmintic therapeutics.

Description:Hookworms infect about half a billion people worldwide and are responsible for the loss of more than two billion disability-adjusted life years. Mass drug administration (MDA) is the most popular preventive approach, but it does not prevent reinfection. An effective vaccine would be a major public health tool in hookworm-endemic areas. We highlight recent human studies where vaccination with irradiated larvae and repeated rounds of infection-treatment have induced partial protection. These studies have emphasized the importance of targeting the infective larvae to generate immunity to prevent adult worms from maturing in the gut. We summarize the current status of human and animal model vaccine trials. Hookworm infection is endemic in resource-poor developing regions where polyparasitism is common, and vaccine cold chain logistics are complex. Humans do not develop sterile immunity to hookworms, and the elderly are frequently overlooked in MDA campaigns. For all these reasons, a vaccine is essential to create long-lasting protection. The lack of a robust animal model to mimic human hookworm infections is a barrier to the discovery and development of a vaccine, however, there have been major recent advances in human challenge studies which will accelerate the process.

Description:Liver fluke infection is a major risk for cholangiocarcinoma (CCA). It has been established that the Asian liver flukes, Clonorchis sinensis and Opisthorchis viverrini secrete growth factors, digestive enzymes, and extracellular vesicles (EVs) which contribute to abnormal cell development in the bile ducts where the worms reside. These secretions - combined with aberrant inflammation and repeated cycles of chronic wounding at the site of parasite attachment and grazing on the epithelium - promote biliary hyperplasia and fibrosis and ultimately malignant transformation. Application of post-genomic and gene-editing tools to the study of liver fluke immunobiology and pathogenesis has accelerated the discovery of essential virulence factors to which targeted therapies and diagnostics can be directed.