WATER PRODUCTION FOR AWARE (Total Bacteria and Antibiotic Resistance Genes):

The water production for AWARE main activities includes three stages – disinfection by ultraviolet C radiation (UVC), storage for24h 0m 0s-24h 0m 0s(according to water load and season) and ozonation. The water quality is monitored at these three stages, for the parameters indicated in Figure 1 below.

Sampling, Processing, and Analyses

Water samples are collected (see Figure 2) and processed within a6h 0m 0sinterval, before being shipped for the partner responsible for the analyses (Table 1). In case no processing is needed, samples are frozen and stored atTemperature-80°Cwithin3h 0m 0s.

For each sampling event, the date, day of the week and hour; the temperature and rain. Sampling points, indicated in Figure 2 were designated from A to I:

• Influent of primary treatment (A)

• Influent of biological treatment (activated sludge) (B)

• Treated secondary effluent (C)

• Sand filter effluent (D)

• UVC effluent (E)

• Storage for reuse tank effluent (F)

• Ozonation effluent (1 dos, O3) - 5mgITO3X technology - (G)

• Effluent of the vacuum UV oxidation (VUV) (H)

• Effluent of reactive storage / Influent of the recirculation aquaculture system (RAS) (I)

Total Bacteria and Antibiotic Resistance Genes: Total Bacteria and Antibiotic Resistance Genes:

Analysis: Culture-independent detection and/or quantification of bacteria and bacterial contaminants.

Observations: Samples were filtered within6h 0m 0safter collection the filtering membranes were immediately frozen and stored at-8°Ctill shipping in dry ice to the respective partner who proceeded for DNA extraction.

Using the EU Drinking Water Directive:

Mesophilic Bacteria in PCA (Plate Count Agar) – 0 CFU100mL

Total coliforms and Escherichia coli –Number /Amount100 mL (0 MPN/100mL)

Fecal enterococci –Number100mL (0 MPN100mL )

Viral concentration - There are no legal requirements for viruses. They are not included in any regulation now.

Parasite - EU legislation (2020/741)

Metals - DIRECTIVE 2008/105/EC OF THE EUROPEAN PARLIAMENT AND OF THE COUNCIL of 16 December 2008 on environmental quality standards in the field of water policy

Organic contaminants - DIRECTIVE 2008/105/EC OF THE EUROPEAN PARLIAMENT AND THE

COUNCIL of 16 December 2008 on environmental quality standards in the field of water policy.