L cysteine l cys supplementation

The Legionella culture technique was based on ISO 11731:2017 (20 ). The hot- and cold-water samples were sampled following the Italian National Unification and European Committee (UNI EN) ISO 19458:2006 (46 ) and Italian guidelines (19 ).
Different aliquots (from 0.2 to 0.1 mL) of the untreated, filtered, heated, and acid-treated samples were seeded on plates of the selective medium glycine-vancomycin-polymyxin B-cycloheximide (GVPC) (Thermo Fisher Scientific, Diagnostic, Ltd., Basingstoke, UK) and incubated at 35 ± 2°C with 2.5% CO₂ for a maximum of 15 days. Legionella growth was evaluated every 2 or 3 days.
To confirm the presence of the Legionella genus, suspected colonies were subcultured on buffered charcoal yeast extract (BCYE) agar with (cys) and without (cys) l-cysteine (l-cys) supplementation (Thermo Fisher Scientific, Diagnostic, Ltd., Basingstoke, UK). Moreover, as a negative control, the same isolates were spread on tryptone soy agar (TSA) with 5% sheep blood agar (Thermo Fisher Scientific, Diagnostic, Ltd., Basingstoke, UK) and incubated under the same conditions previously described, as Legionella is not able to grow on this medium. Only the colonies that grew on BCYE cys⁺ agar were considered for the next steps of the study.

The Legionella culture technique was based on ISO 11731:2017 (20 ). The hot- and cold-water samples were sampled following the Italian National Unification and European Committee (UNI EN) ISO 19458:2006 (46 ) and Italian guidelines (19 ).
Different aliquots (from 0.2 to 0.1 mL) of the untreated, filtered, heated, and acid-treated samples were seeded on plates of the selective medium glycine-vancomycin-polymyxin B-cycloheximide (GVPC) (Thermo Fisher Scientific, Diagnostic, Ltd., Basingstoke, UK) and incubated at 35 ± 2°C with 2.5% CO₂ for a maximum of 15 days. Legionella growth was evaluated every 2 or 3 days.
To confirm the presence of the Legionella genus, suspected colonies were subcultured on buffered charcoal yeast extract (BCYE) agar with (cys) and without (cys) l-cysteine (l-cys) supplementation (Thermo Fisher Scientific, Diagnostic, Ltd., Basingstoke, UK). Moreover, as a negative control, the same isolates were spread on tryptone soy agar (TSA) with 5% sheep blood agar (Thermo Fisher Scientific, Diagnostic, Ltd., Basingstoke, UK) and incubated under the same conditions previously described, as Legionella is not able to grow on this medium. Only the colonies that grew on BCYE cys⁺ agar were considered for the next steps of the study.