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Density of fecal coliforms (FC) such as Escherichia coli is the most commonly used indicator of fecal pathogen content of biosolids. When biosolids are disposed off or used for soil amendment, they pose public health risks. So far anaerobic digesters have been considered to be an effective treatment option for pathogen and FC reduction in biosolids. However, recent studies revealed that there is a significant re-growth and reactivation of indicator organisms in biosolids upon dewatering by centrifugation. Although the exact mechanism of FC reactivation is yet to be understood, a few extensive recent studies strongly suggest that FC go into a viable but non-culturable (VBNC) state during anaerobic digestion. Therefore, quantitative detection of live cells among the total in biosolids samples, without using culturing-based approaches, is highly critical from a public health risk assessment perspective. Since recent investigations proved the significant re-growth and reactivation of indicator organisms. Persistence of DNA in the environment after cell death in the range of days to weeks limits the application of DNA-based approaches for the detection of live bacteria. Using selective nucleic acid intercalating dyes such as ethidium monoazide (EMA) and propidium monoazide (PMA) is one of the alternative approaches to detect and quantify the viable cells by quantitative PCR. These compounds have the ability to penetrate only into dead cells with compromised membrane integrity. They intercalate in the DNA via photo-inducible azide groups and in turn inhibit DNA amplification during PCR reactions. PMA has been successfully used in different studies and microorganisms but it has not been evaluated sufficiently for the complex environmental samples such as biosolids. In this study Escherichia coli ATCC 25922 and uidA gene were used as model organism and as target sequence respectively in absolute quantification method with real-time PCR. Experiments with the known quantities of live and dead cell mixtures showed that PMA treatment inhibits PCR amplification from dead cells with over 99% efficiency. The results of this study conclusively demonstrated that PMA-modified PCR could be successfully applied to the biosolids when total suspended solid (TSS) concentration is 2000 mg/L or below.

En conditions hivernales rigoureuses, l'application des fondants routiers (principalement du chlorure de sodium - NaCl) permet d'assurer la sécurité des usagers de réseaux. Toutefois, son utilisation généralisée entraîne des risques de contamination des écosystèmes aquatiques et terrestres. En milieu routier, le Na+ et Cl- sont transportés par les eaux de ruissellement vers des ouvrages de traitement tels que les bassins de rétention-décantation. Or, ces bassins n'ont pas la capacité de traiter efficacement cet apport de fondants sous forme dissoute. L'objectif de cette thèse est d'étudier le transfert de ces fondants au sein du système routier et de déterminer leur rôle dans la libération des polluants en période hivernale. Les caractéristiques du bassin situé à Azerailles (Lorraine, France) sur une route nationale moyennement fréquentée ont été étudiées, ainsi que les conditions météorologiques, les salages, la composition des matières en suspension (MES) et la concentration en éléments traces métalliques (ETM). Les résultats ont permis de montrer que entre 50 et 90 % du Cl- ont été collectés par le bassin. Il joue un rôle de tampon qui permet une libération du Cl- dilué durant la période de ruissellement suivant la période de salage. La présence de NaCl dans les eaux a joué un rôle sur la diminution de la qualité des matières en suspension, en mettant en évidence la présence de polluants organiques en période de salage. Elle entraîne également une modification de la chimie des eaux et une augmentation des polluants métalliques (zinc). / In severe winter conditions, the application of deicer salts (mainly sodium chloride -NaCl) ensures the safety of network users. However, its spreading carries contamination risks of aquatic and terrestrial ecosystems. On the road, Na+ and Cl- are transported by runoff water to retention-decantation ponds. These ponds do not have the capacity to efficiently treat this flux of deicer salts in dissolved form. In addition, they allow the modification of water and soil chemistry, generating the mobility of metallic pollutants. The objective of this thesis is to study the deicer salt transfer and to determine its role in the release of pollutants during winter. The characteristics of the pond located in Azerailles (Lorraine, France) on a medium-frequented national road were studied, as well as the weather and salting conditions, the composition of the suspended matters and the concentration of traces metals. The results highlighted that between 50 and 90 % of Cl- was collected by the pond. The pond acts as a buffer which allows a release of the collected Cl- diluted during the runoff period following the salting period. The presence of NaCl in runoff water reduced the quality of suspended solids with the presence of organic markers of road pollution and the increase of zinc. It also influenced the increase of zinc in water, the chemical composition and the speciation of elements.