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Abstract(s)
A água é o recurso natural mais valioso do planeta, pelo que a sua conservação constitui um dos mais importantes pilares do desenvolvimento sustentável. Nas regiões em que a escassez de recursos hídricos constitui uma realidade natural e naquelas em que o crescimento demográfico e/ou as alterações climáticas perspetivam essa escassez, a gestão sustentável dos recursos hídricos implica a sua conservação e inclui, por conseguinte, a reutilização da água. Um exemplo é a reutilização de águas residuais (AR’s) na irrigação na agricultura, sendo uma alternativa sustentável, promovendo a redução da captação de água e das descargas de efluentes no meio ambiente. No entanto, este tipo de água pode estar contaminada com microrganismos resistentes aos tratamentos de desinfeção convencionais aplicados em estações de tratamento de águas residuais (ETAR’s), como por exemplo vírus patogénicos tais como, Norovírus (NoV), vírus da Hepatite A (HAV), vírus da Hepatite E (HEV), Adenovírus (AdV) e Polyomavírus (PyV), constituindo um risco para a saúde humana. Mais ainda a avaliação da qualidade microbiológica das AR’s é realizada com recurso a microrganismos indicadores, sendo o mais comumente usado para a indicação de poluição fecal, os coliformes fecais, como por exemplo: Escherichia coli, da família Enterobacteriacae, que não são os adequados para a deteção da presença/concentração de partículas virais. Desta forma, este estudo teve como principal objetivo detetar e quantificar a existência de seis genomas virais (NoV GI e GII, HAV, HEV, AdV e PyV) presentes em amostras de AR colhidas em diferentes pontos de duas ETAR’S de Lisboa (ETAR X e ETAR Y), através de técnicas moleculares de amplificação dirigida (qPCR) e amplificação não-dirigida (NGS – do inglês, next generation sequen-cing). Foram amostrados quatro pontos ao longo de cada uma das ETAR’s em estudo em duas estações do ano distintas, outono e primavera. De um modo geral todos os genomas virais em estudo foram detetados em concentrações elevadas (acima de 106 cópias de genoma/L) nos influentes e nos respetivos efluentes reutilizáveis, com algumas exceções. Para o caso do genoma de AdV e de HEV estes apresentaram variações nas suas concentrações consoante a estação do ano estudada, estando presentes na primavera, mas não no outono. Da avaliação por sequenciação de próxima geração verifica-se que há uma maior diversidade de famílias virais no influente da ETAR X comparativamente ao influente da ETAR Y, estando este facto provavelmente associado ao tipo de AR que cada ETAR trata. Mais ainda, observou-se um aumento da diversidade de famílias virais no efluente após o tratamento secundário em ambas as ETAR’s, sendo maioritariamente famílias associadas a bacteriófagos. Apesar da evolução dos métodos de tratamento das ETAR’s, a sua eficiência na remoção/inativação dos vírus é ainda baixa, o que poderá contribuir para um risco de saúde publica, uma vez que os efluentes tratados destas ETAR’s são lançados nos recursos hídricos para reutilização, podendo ser uma via de transporte destes vírus para população. Mais ainda este estudo veio corroborar a importância de se começar a utilizar vírus como indicadores de contaminação fecal nas águas, sendo que os indicadores bacterianos, não conseguem retratar com precisão os riscos para a saúde humana. Desta forma, apesar de ser impraticável monitorizar a presença de todos os agentes patogénicos virais relacionados com a poluição das AR’s, o desenvolvimento de um indicador viral preciso da contaminação dos esgotos é necessário para uma monitorização melhorada da qualidade da água.
Water is the most valuable natural resource on the planet, so its conservation is one of the most important pillars of sustainable development. In regions where water scarcity is a natural reality and where population growth and/or climate change prospect that scarcity, sustainable management of water resources involves the conservation of these resources and therefore includes water reuse. An example of water reuse is wastewater (WW) reuse in agriculture irrigation, being a sustainable alter-native, promoting the reduction of water collection and discharges of effluents into the environment. However, this type of water may be contaminated with microorganisms resistant to conventional disinfection treatments applied in wastewater treatment plants (WWTP's), such as pathogenic viruses like Norovirus (NoV), Hepatitis A virus (HAV), Hepatitis E virus (HEV), Adenovirus (AdV) and Polyomavirus (PyV), constituting a risk to human health. Furthermore, the assessment of the microbiological quality of WW is carried out using indicator microorganisms, the most commonly used for the indication of faecal pollution, is faecal coliforms, such as: Escherichia coli, from the Enterobacteriacae family, which are not suitable for detection of the presence / concentration of viral particles. Thus, this study aimed to detect and quantify the presence of six viral genomes (NoV GI and GII, HAV, HEV, AdV and PyV) present in samples of residual water collected at different points of two WWTP's in Lisbon (WWTP X and WWTP Y), using molecular techniques of directed amplification (qPCR) and non-directed amplification (next generation sequencing - NGS). Four points were sampled along each WWTP under study in two different seasons of the year, autumn and spring. In general, all studied viral genomes were detected in high concentrations (above 106 copies of genome/L) in the influents and in the respective reused effluents, with some exceptions. For AdV and HEV genomes case, these showed variations in their concentrations according to the season in study, being present in the spring, but not in the autumn. From the next generation sequencing assessment, it appears that there is a greater diversity of viral families in the influent of WWTP X compared to the influent of WWTP Y, this fact is probably associated with the type of WW that each WWTP treats. In addition, there was an increase in the viral families diversity in the effluent after secondary treatment of both WWTP’s, being most families associated with bacteriophages. Despite the evolution of treatment methods of the WWTPs, their efficiency in removing/ inactivating viruses is still low, which may contribute to a public health risk, since the treated effluents from these WWTPs are released in the water resources for reuse, which can be a way of bringing these viruses to the population. Furthermore, this study corroborates the importance of starting to use viruses as indicators of faecal contamination in waters, since bacterial indicators are not able to accurately portray the risks to human health. Thus, although it is impractical to monitor the presence of all viral pathogens related to WW pollution, the development of an accurate viral indicator of sewage contamination is necessary for an improved monitoring of water quality.
Water is the most valuable natural resource on the planet, so its conservation is one of the most important pillars of sustainable development. In regions where water scarcity is a natural reality and where population growth and/or climate change prospect that scarcity, sustainable management of water resources involves the conservation of these resources and therefore includes water reuse. An example of water reuse is wastewater (WW) reuse in agriculture irrigation, being a sustainable alter-native, promoting the reduction of water collection and discharges of effluents into the environment. However, this type of water may be contaminated with microorganisms resistant to conventional disinfection treatments applied in wastewater treatment plants (WWTP's), such as pathogenic viruses like Norovirus (NoV), Hepatitis A virus (HAV), Hepatitis E virus (HEV), Adenovirus (AdV) and Polyomavirus (PyV), constituting a risk to human health. Furthermore, the assessment of the microbiological quality of WW is carried out using indicator microorganisms, the most commonly used for the indication of faecal pollution, is faecal coliforms, such as: Escherichia coli, from the Enterobacteriacae family, which are not suitable for detection of the presence / concentration of viral particles. Thus, this study aimed to detect and quantify the presence of six viral genomes (NoV GI and GII, HAV, HEV, AdV and PyV) present in samples of residual water collected at different points of two WWTP's in Lisbon (WWTP X and WWTP Y), using molecular techniques of directed amplification (qPCR) and non-directed amplification (next generation sequencing - NGS). Four points were sampled along each WWTP under study in two different seasons of the year, autumn and spring. In general, all studied viral genomes were detected in high concentrations (above 106 copies of genome/L) in the influents and in the respective reused effluents, with some exceptions. For AdV and HEV genomes case, these showed variations in their concentrations according to the season in study, being present in the spring, but not in the autumn. From the next generation sequencing assessment, it appears that there is a greater diversity of viral families in the influent of WWTP X compared to the influent of WWTP Y, this fact is probably associated with the type of WW that each WWTP treats. In addition, there was an increase in the viral families diversity in the effluent after secondary treatment of both WWTP’s, being most families associated with bacteriophages. Despite the evolution of treatment methods of the WWTPs, their efficiency in removing/ inactivating viruses is still low, which may contribute to a public health risk, since the treated effluents from these WWTPs are released in the water resources for reuse, which can be a way of bringing these viruses to the population. Furthermore, this study corroborates the importance of starting to use viruses as indicators of faecal contamination in waters, since bacterial indicators are not able to accurately portray the risks to human health. Thus, although it is impractical to monitor the presence of all viral pathogens related to WW pollution, the development of an accurate viral indicator of sewage contamination is necessary for an improved monitoring of water quality.
Description
Tese de mestrado em Biologia Humana e Ambiente, Universidade de Lisboa, Faculdade de Ciências, 2020
Keywords
Caracterização viral de AR’s NGS qPCR Disseminação de vírus patogénicos Teses de mestrado - 2020