Journal: The Science of the total environment

Article Title: Verification and application of qPCR and viability-qPCR for Legionella monitoring in evaporative cooling systems complementing the conventional culture method.

doi: 10.1016/j.scitotenv.2024.176011

Figure Lengend Snippet: Fig. 1. Recovery of L. pneumophila in Evian water and artificial process water (aPW) using qPCR and viability-qPCR. Ten samples per concentration level and matrix were produced by spiking with cryopreserved L. pneumophila Sg 1 standards. Additionally, DNA of three aliquots of the stock suspension (105 Legionella/100 mL) were extracted using the direct method without filtration steps and used as a reference value for the calculation of the recovery of L. pneumophila.

Article Snippet: Respective matrices were spiked with defined concentrations of total or intact L. pneumophila Philadelphia-1 (DSMZ 7516) Sg 1, L. pneumophila Sg 4 (DSMZ 7514) or L. pneumophila Sg 6 (DSMZ 25182), using cryopreserved Legionella standards (Table 1) stored at − 80 ◦C and thawed at 37 ◦C for 5 min. Cryopreserved standards were prepared by rqmicro AG according to following protocol and shipped within 24 h on dry ice.

Techniques: Concentration Assay, Produced, Suspension, Filtration

Journal: The Science of the total environment

Article Title: Verification and application of qPCR and viability-qPCR for Legionella monitoring in evaporative cooling systems complementing the conventional culture method.

doi: 10.1016/j.scitotenv.2024.176011

Figure Lengend Snippet: Fig. 2. Verification of linearity for the Bio-Rad L. pneumophila qPCR kit using artificial samples based on Evian water or artificial process water (aPW). Matrices were spiked with L. pneumophila Sg 1, Sg 4 or Sg 6 cryopreserved standards over a range of seven concentrations (102, 5 × 102, 103, 5 × 103, 104, 5 × 104, 105 TLC/100 mL). Each concentration was spiked in triplicates for qPCR (T1 to T3) using dilutions based on one stock suspension (105 TLC/mL) and the resulting DNA extracts were run as triplicates. Results of the qPCR in genomic units (GU) for each triplicate (n = 3) are plotted against spiked concentrations (m) based on total Legionella count (TLC). The reference value is indicating the curve if TLC and GU were identical. For linear regression, results below the limit of quantification were omitted.

Article Snippet: Respective matrices were spiked with defined concentrations of total or intact L. pneumophila Philadelphia-1 (DSMZ 7516) Sg 1, L. pneumophila Sg 4 (DSMZ 7514) or L. pneumophila Sg 6 (DSMZ 25182), using cryopreserved Legionella standards (Table 1) stored at − 80 ◦C and thawed at 37 ◦C for 5 min. Cryopreserved standards were prepared by rqmicro AG according to following protocol and shipped within 24 h on dry ice.

Techniques: Concentration Assay, Suspension

Journal: The Science of the total environment

Article Title: Verification and application of qPCR and viability-qPCR for Legionella monitoring in evaporative cooling systems complementing the conventional culture method.

doi: 10.1016/j.scitotenv.2024.176011

Figure Lengend Snippet: Fig. 3. Verification of linearity for the Hygiena qPCR kit using artificial samples based on Evian water or artificial process water (aPW). Matrices were spiked with L. pneumophila Sg 1 cryopreserved standards over a range of seven concentrations (102, 5 × 102, 103, 5 × 103, 104, 5 × 104, 105 ILC/100 mL). Each concentration was spiked in duplicates per matrix for viability-qPCR (V1, V2) and once for qPCR (T) and all resulting DNA extracts were run as duplicates. Results of the viability-qPCR or qPCR in genomic units (GU) for each reaction duplicate (n = 2) are plotted against spiked concentrations (m = 6) based on intact Legionella count (ILC). The reference value is indicating the curve if ILC and GU were identical. For linear regression results below the limit of quantification were omitted.

Article Snippet: Respective matrices were spiked with defined concentrations of total or intact L. pneumophila Philadelphia-1 (DSMZ 7516) Sg 1, L. pneumophila Sg 4 (DSMZ 7514) or L. pneumophila Sg 6 (DSMZ 25182), using cryopreserved Legionella standards (Table 1) stored at − 80 ◦C and thawed at 37 ◦C for 5 min. Cryopreserved standards were prepared by rqmicro AG according to following protocol and shipped within 24 h on dry ice.

Techniques: Concentration Assay

Journal: The Science of the total environment

Article Title: Verification and application of qPCR and viability-qPCR for Legionella monitoring in evaporative cooling systems complementing the conventional culture method.

doi: 10.1016/j.scitotenv.2024.176011

Figure Lengend Snippet: Fig. 4. Verification of linearity for the culture method using artificial samples based on Evian water or artificial process water (aPW) spiked with L. pneumophila Sg 1 cryopreserved standards over a range of seven concentrations (102, 5 × 102, 103, 5 × 103, 104, 5 × 104, 105 ILC/100 mL). For each concentration the weighted mean of evaluable plating (on GVPC agar) of a series of five (2 × 0.1 mL, 2 × 0.5 mL, membrane filtration of 20 mL) was used for linear regression. Results of the culture method (colony forming units, CFU) are plotted against spiked concentrations (m = 7) based on intact Legionella count (ILC). One additional run (Run3) was performed in Evian mineral water due to bigger deviations between the first two runs compared to aPW.

Article Snippet: Respective matrices were spiked with defined concentrations of total or intact L. pneumophila Philadelphia-1 (DSMZ 7516) Sg 1, L. pneumophila Sg 4 (DSMZ 7514) or L. pneumophila Sg 6 (DSMZ 25182), using cryopreserved Legionella standards (Table 1) stored at − 80 ◦C and thawed at 37 ◦C for 5 min. Cryopreserved standards were prepared by rqmicro AG according to following protocol and shipped within 24 h on dry ice.

Techniques: Concentration Assay, Membrane, Filtration

Journal: The Science of the total environment

Article Title: Verification and application of qPCR and viability-qPCR for Legionella monitoring in evaporative cooling systems complementing the conventional culture method.

doi: 10.1016/j.scitotenv.2024.176011

Figure Lengend Snippet: Fig. 6. qPCR (A) and viability-qPCR (B) results for environmental samples of evaporative cooling systems spiked with cryopreserved L. pneumophila Sg 1 to con centrations of 103 and 104 ILC/100 mL. Bar chart on the left is showing determined GU/100 mL for samples 18 to 25 for Bio-Rad (BR) and Hygiena (HG) kit. Results below Limit of quantification (

Article Snippet: Respective matrices were spiked with defined concentrations of total or intact L. pneumophila Philadelphia-1 (DSMZ 7516) Sg 1, L. pneumophila Sg 4 (DSMZ 7514) or L. pneumophila Sg 6 (DSMZ 25182), using cryopreserved Legionella standards (Table 1) stored at − 80 ◦C and thawed at 37 ◦C for 5 min. Cryopreserved standards were prepared by rqmicro AG according to following protocol and shipped within 24 h on dry ice.

Techniques: Environmental Sampling, Concentration Assay