Modifications of the Helmintex Method: Integration of Treatments With Tween 20, Cellulase, Percoll and Auramine‐O Staining for Detection of <scp> <i>Schistosoma mansoni</i> </scp> Eggs

Sensitive diagnostic tools are essential to support schistosomiasis control and elimination programs, yet most long-standing parasitological and serological tests lack sufficient accuracy in low-intensity settings. The Helmintex (HTX) method, based on magnetic isolation of Schistosoma mansoni eggs from large stool volumes, is highly sensitive and serves as a reference standard for evaluating other diagnostic approaches but remains labour-intensive. We evaluated modifications aimed at reducing workload while preserving or enhancing performance. Eggs were seeded into human faeces and processed using standard HTX versus three key modifications: (i) cellulase digestion in 20% Tween-20, (ii) density separation with 60% Percoll and (iii) replacement of BioMag beads with magnetite nanopowder. In addition, we standardised an auramine-O fluorescent staining protocol for batch processing of HTX sediments from naturally infected samples. Sequential cellulase and Percoll treatments reduced the final sediment volume by 42.9%. Magnetite at 25 mg/mL achieved an average egg recovery of 70.4% compared with 38.5% for BioMag beads. Auramine-O staining facilitated rapid visualisation of eggs and retained fluorescence for at least 90 days. Together, these refinements streamline the HTX method, reinforcing its utility as a reference standard and positioning it as a practical tool for certification of transmission interruption in schistosomiasis-endemic areas.