X Lizana1, A López2, S Benito1, G Agustí3, M Ríos4, N Piqué5, A M Marqués5, F Codony6. 1. Aconsa, SL, Avinguda del Pla del Vent, 11A, 08970 Sant Joan Despí, Barcelona, Spain. 2. Aconsa, SL, Avinguda del Pla del Vent, 11A, 08970 Sant Joan Despí, Barcelona, Spain; Department of Microbiology and Parasitology, Pharmacy Faculty, Universitat de Barcelona, Av. Joan XXIII s/n, 08028 Barcelona, Spain. 3. GenIUL, Carrer de la Ciutat d'Asunción, 08030 Barcelona, Spain. 4. Department of Statistics, Biology Faculty, Universitat de Barcelona, Av. Diagonal, 643, 08028 Barcelona, Spain. 5. Department of Microbiology and Parasitology, Pharmacy Faculty, Universitat de Barcelona, Av. Joan XXIII s/n, 08028 Barcelona, Spain. 6. GenIUL, Carrer de la Ciutat d'Asunción, 08030 Barcelona, Spain. Electronic address: fcodony@geniul.com.
Abstract
BACKGROUND: Viability quantitative Polymerase Chain Reaction (v-qPCR) is a recent analytical approach for only detecting live microorganisms by DNA amplification-based methods This approach is based on the use of a reagent that irreversibly fixes dead cells DNA. In this study, we evaluate the utility of v-qPCR versus culture method for Legionellosis risk management. METHODS: The present study was performed using 116 real samples. Water samples were simultaneously analysed by culture, v-qPCR and qPCR methods. Results were compared by means of a non-parametric test. RESULTS: In 11.6% of samples using both methods (culture method and v-qPCR) results were positive, in 50.0% of samples both methods gave rise to negative results. As expected, equivalence between methods was not observed in all cases, as in 32.1% of samples positive results were obtained by v-qPCR and all of them gave rise to negative results by culture. Only in 6.3% of samples, with very low Legionella levels, was culture positive and v-qPCR negative. In 3.5% of samples, overgrowth of other bacteria did not allow performing the culture. When comparing both methods, significant differences between culture and v-qPCR were in the samples belonging to the cooling towers-evaporative condensers group. The v-qPCR method detected greater presence and obtained higher concentrations of Legionella spp. (p<0.001). Otherwise, no significant differences between methods were found in the rest of the groups. CONCLUSIONS: The v-qPCR method can be used as a quick tool to evaluate Legionellosis risk, especially in cooling towers-evaporative condensers, where this technique can detect higher levels than culture. The combined interpretation of PCR results along with the ratio of live cells is proposed as a tool for understanding the sample context and estimating the Legionellosis risk potential according to 4 levels of hierarchy.
BACKGROUND: Viability quantitative Polymerase Chain Reaction (v-qPCR) is a recent analytical approach for only detecting live microorganisms by DNA amplification-based methods This approach is based on the use of a reagent that irreversibly fixes dead cells DNA. In this study, we evaluate the utility of v-qPCR versus culture method for Legionellosis risk management. METHODS: The present study was performed using 116 real samples. Water samples were simultaneously analysed by culture, v-qPCR and qPCR methods. Results were compared by means of a non-parametric test. RESULTS: In 11.6% of samples using both methods (culture method and v-qPCR) results were positive, in 50.0% of samples both methods gave rise to negative results. As expected, equivalence between methods was not observed in all cases, as in 32.1% of samples positive results were obtained by v-qPCR and all of them gave rise to negative results by culture. Only in 6.3% of samples, with very low Legionella levels, was culture positive and v-qPCR negative. In 3.5% of samples, overgrowth of other bacteria did not allow performing the culture. When comparing both methods, significant differences between culture and v-qPCR were in the samples belonging to the cooling towers-evaporative condensers group. The v-qPCR method detected greater presence and obtained higher concentrations of Legionella spp. (p<0.001). Otherwise, no significant differences between methods were found in the rest of the groups. CONCLUSIONS: The v-qPCR method can be used as a quick tool to evaluate Legionellosis risk, especially in cooling towers-evaporative condensers, where this technique can detect higher levels than culture. The combined interpretation of PCR results along with the ratio of live cells is proposed as a tool for understanding the sample context and estimating the Legionellosis risk potential according to 4 levels of hierarchy.
Authors: Pablo Casino; Asunción López; Sara Peiró; Martín Ríos; Santiago Ríos; Aldous Porta; Gemma Agustí; Daniel Asensio; Ana María Marqués; Núria Piqué Journal: Microbiol Spectr Date: 2022-03-22