title: Different methods to quantify Listeria monocytogenes biofilms cells showed different profile in their viability.
abstract: Listeria monocytogenes is a foodborne pathogen able to adhere and to form biofilms in several materials commonly present in food processing plants. The aim of this study was to evaluate the resistance of Listeria monocytogenes attached to abiotic surface, after treatment with sanitizers, by culture method, microscopy and Quantitative Real Time Polymerase Chain Reaction (qPCR). Biofilms of L. monocytogenes were obtained in stainless steel coupons immersed in Brain Heart Infusion Broth, under agitation at 37 °C for 24 h. The methods selected for this study were based on plate count, microscopic count with the aid of viability dyes (CTC-DAPI), and qPCR. Results of culture method showed that peroxyacetic acid was efficient to kill sessile L. monocytogenes populations, while sodium hypochlorite was only partially effective to kill attached L. monocytogenes (p < 0.05). When, viability dyes (CTC/DAPI) combined with fluorescence microscopy and qPCR were used and lower counts were found after treatments (p < 0.05). Selective quantification of viable cells of L. monocytogenes by qPCR using EMA revelead that the pre-treatment with EMA was not appropriate since it also inhibited amplification of DNA from live cells by ca. 2 log. Thus, the use of CTC counts was the best method to count viable cells in biofilms.
authors.affiliation: Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil.
authors.forename: Lizziane Kretli
authors.initials: LK
authors.lastname: Winkelströter
authors.affiliation: Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brazil.
authors.forename: Elaine C P
authors.initials: EC
authors.lastname: De Martinis
mesh_terms: D058491:Bacterial Load / Q000379:methods*; D018441:Biofilms / Q000187:drug effects* / Q000254:growth & development; D015169:Colony Count, Microbial; D004202:Disinfectants / Q000494:pharmacology*; D004783:Environmental Microbiology; D008089:Listeria monocytogenes / Q000187:drug effects* / Q000302:isolation & purification / Q000502:physiology*; D050296:Microbial Viability / Q000187:drug effects*; D008853:Microscopy; D060888:Real-Time Polymerase Chain Reaction; D013696:Temperature; D013995:Time
chemical_list: D004202:Disinfectants
keywords: L. monocytogenes; biofilms; peroxyacetic acid; qPCR; sodium hypochlorite