Free-living amoebae predation in nutrient-enriched treated wastewater: ecological potential and machine...

Area of Science:

• Environmental Microbiology
• Water Reuse Technologies
• Biocontrol Agents

Background:

• Intensified pressure on freshwater resources necessitates treated wastewater (TWW) reuse for irrigation.
• TWW irrigation risks spreading pathogenic microbes and antibiotic-resistant bacteria (ARB).
• Free-living amoebae (FLA) are soil predators with potential as biocontrol agents against bacteria.

Purpose of the Study:

• To investigate the impact of key TWW ions (phosphate, ammonium, sulphate) on FLA predation efficiency.
• To assess the biocontrol potential of FLA against bacterial pathogens in TWW.
• To develop an AI-based model for tracking FLA behavior.

Main Methods:

• Studied predation by three FLA species (Vermamoeba vermiformis, Acanthamoeba castellanii, Heterolobosea sp.) on GFP-tagged Escherichia coli and Enterococcus mundtii.
• Enriched bacterial cultures with specific ions (PO₄³⁻, NH₄⁺, SO₄²⁻) at 100 mg/L.
• Developed and validated a YOLOv5 deep-learning model to track FLA movement in microscopy videos.

Main Results:

• Phosphate (PO₄³⁻) or ammonium (NH₄⁺) enrichment significantly enhanced Vermamoeba predation on E. coli, achieving 7 log₁₀ reductions in 72 hours.
• Acanthamoeba and Heterolobosea showed weaker predatory effects compared to Vermamoeba under ion enrichment.
• A mixed FLA consortium demonstrated faster elimination of E. mundtii than individual species; YOLOv5 model reliably tracked FLA movement.

Conclusions:

• Tailoring nutrient levels in TWW, specifically phosphate and ammonium, can enhance the biocontrol efficacy of free-living amoebae.
• This eco-biological approach, integrating AI for behavior tracking, offers a promising strategy for safer TWW reuse in agriculture.
• Findings support the development of ecological and AI-informed strategies to mitigate pathogen persistence in irrigation systems.