Disease control and biosecurity

Measured reductions in aquaculture bacteria

Ozone nanobubbles produced large, fast reductions in culturable bacteria in laboratory and pond-water tests. Oxygen nanobubbles, where they were compared, produced much smaller ones. The studies below report counts before and after treatment; they measured bacteria in water, not disease outcomes on farms.

Vibrio parahaemolyticus

Certain V. parahaemolyticus strains cause early mortality syndrome, also called acute hepatopancreatic necrosis disease (EMS/AHPND), in shrimp. In laboratory tests, ozone nanobubble treatment reduced V. parahaemolyticus from 10⁶ CFU/mL to undetectable levels within 6 minutes. Oxygen nanobubbles, tested alongside, reduced it by 31% over one week Nghia et al., 2022.

A second study with an EMS/AHPND strain found that ozone nanobubbles reduced culturable V. parahaemolyticus in seawater by more than 99.99% within 1 minute of exposure. Researchers reported that the treatment delayed mortality in infected shrimp Imaizumi et al., 2018.

Streptococcus and Aeromonas in fresh water

In fresh water, a 10-minute exposure to ozone nanobubbles reduced Streptococcus agalactiae by 96.11% and Aeromonas veronii by 97.92%. Organic matter, as found in culture water, lowered the effect about 1.6-fold. Repeated ozone nanobubble treatments kept bacterial numbers down, and the authors reported the treatment as safe for Nile tilapia under the trial conditions Jhunkeaw et al., 2021.

Total bacteria in pond water

Ozone nanobubbles at 0.15 mg/L reduced total heterotrophic bacteria in pond water by 90.9% to 99.4% Huang et al., 2023. A reduction of that size can reset the microbial community. Whether that creates a useful window to reintroduce beneficial microbes (probiotics) before other species re-establish is an open question the cited work does not test.

How the reductions are explained

The explanation offered is oxidative, with a possible physical component. It is a proposed mechanism, drawn partly from a review of industrial wastewater treatment.

Ozone nanobubbles are associated with reactive oxygen species (ROS), in particular hydroxyl radicals (•OH). These radicals have a high redox potential (2.80 V) and react non-selectively: they oxidize polyunsaturated fatty acids in bacterial membranes (lipid peroxidation) and damage DNA, and cell contents leak out Mukherjee et al., 2023; Huang et al., 2023.

A physical contribution has also been proposed: shock waves from collapsing bubbles may rupture cell membranes. The clearest evidence for combined effects involved another energy source. In laboratory tests with Aeromonas hydrophila and V. parahaemolyticus, oxygen nanobubbles combined with ultrasonication gave more than a 6 log CFU/mL reduction Rafeeq et al., 2020.

Biofilm on surfaces and pipes

Biofilms on tank walls and pipes can harbor pathogens, and nanobubbles have been studied as a way to loosen them. The evidence comes from food-contact surfaces, water lines and filters rather than aquaculture tanks.

Biofilms are held together by extracellular polymeric substances (EPS). Nanobubbles, and ozone nanobubbles in particular, are thought to penetrate this matrix and generate hydroxyl radicals that break down its proteins and polysaccharides, so the biofilm loses structure and detaches Shiroodi et al., 2021. In laboratory tests, nanobubbles alone or combined with neutral electrolyzed water reduced E. coli O157:H7, V. parahaemolyticus and Listeria innocua biofilms on plastic and stainless steel Shiroodi et al., 2021.

Physical forces play a part as well. Nanobubbles lowered the contact angle of water on surfaces (greater wettability), and their movement and collapse create local shear forces that can scour biofilm layers Shiroodi et al., 2021. A study of a biological aerated filter examined how shear stress from nanobubble aeration shapes biofilm structure and the microbial community Xiao et al., 2021.

Nanobubbles also mitigated biofouling in agricultural water distribution systems. The explanation offered is that higher dissolved oxygen (DO) and ROS make stagnant zones less favorable for anaerobic biofilms Xiao et al., 2020.

Fish immune and stress markers

Researchers also measured how fish responded to ozone nanobubble exposure. These are gene-expression, enzyme and challenge results in single species, reported here as research findings.

In Nile tilapia, researchers reported that ozone nanobubble exposure raised expression of the cytokine genes TNF-α (tumor necrosis factor alpha), IL-1β and IL-2β in the gills, head kidney and spleen within 15 minutes. This pattern was read as a sign that ozone nanobubbles act as a mild stressor that primes the innate immune response. Tilapia pre-treated with ozone nanobubbles and then challenged with S. agalactiae showed a relative percent survival (RPS) of 60–70% compared with untreated controls Linh et al., 2021.

In jade perch, superoxide dismutase (SOD) activity rose one day after ozone nanobubble treatment, to 8.0 units against 4.5 in controls, and returned to baseline within 48 hours Huang et al., 2023.

A different gas gave a related result in a laboratory model. Researchers reported that hydrogen nanobubble water reduced ROS accumulation, lowered inflammatory markers and improved survival in zebrafish infected with a virus Li et al., 2022.