Padronização de método de concentração e extração de ácidos nucleicos em amostras de esgoto sanitário: uma ferramenta de baixo custo para ser utilizada na vigilância epidemiológica de SARS-CoV-2 / Standardization of the method of concentration and extraction of nucleic acids in wastewater samples: a low-cost tool to be used in epidemiological surveillance of SARS-CoV-2

RESUMO A vigilância da qualidade dos esgotos sanitários pode representar uma ferramenta complementar para monitoramento de doenças infecciosas e prevenção de surtos epidêmicos, especialmente quando a capacidade para testes clínicos é limitada. Dessa maneira, o presente estudo descreve o detalhamento técnico de um método de baixo custo para a concentração e extração de ácidos nucleicos de amostras de esgoto sanitário como etapa prévia para a detecção de vírus e outros agentes patogênicos. Para validar a metodologia proposta, após as etapas de concentração e extração, analisaram-se a presença do ácido ribonucleico do SARS-CoV-2 (COVID-19) nas amostras, por meio de reação em cadeia da polimerase em tempo real. O ácido ribonucleico do vírus foi detectado em 80% das amostras de esgoto sanitário analisadas, comprovando o êxito do procedimento metodológico adotado. A detecção precoce de um patógeno associado ao trabalho de equipes multidisciplinares possibilita a prática da vigilância epidemiológica, que auxilia na tomada de decisões na Saúde Única — união indissociável entre a saúde animal, humana e ambiental.

ABSTRACT Sewage quality surveillance can represent a complementary tool for monitoring infectious diseases and preventing epidemic outbreaks, especially when the capacity for clinical testing is limited. Thus, the present study describes the technical details of a low-cost method for concentrating and extracting nucleic acids from sewage samples, as a preliminary step for the detection of viruses and other pathogens. To validate the proposed methodology, after the concentration and extraction steps, the presence of the SARS coronavirus-2 (COVID-19) in the samples was analyzed using real-time polymerase chain reaction. The virus' ribonucleic acid was detected in 80% of the sewage samples analyzed, proving the success of the methodological procedure adopted. The early detection of a pathogen associated with the work of multidisciplinary teams allows the practice of epidemiological surveillance, which assists in making decisions about One Health — an inseparable union between animal, human, and environmental health.

Long-term monitoring of SARS-COV-2 RNA in wastewater in Brazil: A more responsive and economical approach.

SARS-CoV-2, the novel Coronavirus, was first detected in Wuhan, China, in December 2019, and has since spread rapidly, causing millions of deaths worldwide. As in most countries of the world, in Brazil, the consequences of the COVID-19 pandemic have been catastrophic. Several studies have reported the fecal shedding of SARS-CoV-2 RNA titers from infected symptomatic and asymptomatic individuals. Therefore, the quantification of SARS-CoV-2 in wastewater can be used to track the virus spread in a population. In this study, samples of untreated wastewater were collected for 44 weeks at five sampling sites in the ABC Region (São Paulo, Brazil), in order to evaluate the SARS-CoV-2 occurrence in the sewerage system. SARS-CoV-2 RNA titers were detected throughout the period, and the concentration ranged from 2.7 to 7.7 log10 genome copies.L-1, with peaks in the last weeks of monitoring. Furthermore, we observed a positive correlation between the viral load in wastewater and the epidemiological/clinical data, with the former preceding the latter by approximately two weeks. The COVID-19 prevalence for each sampling site was estimated via Monte-Carlo simulation using the wastewater viral load. The mean predicted prevalence ranged 0.05 to 0.38%, slightly higher than reported (0.016 ± 0.005%) in the ABC Region for the same period. These results highlight the viability of the wastewater surveillance for COVID-19 infection monitoring in the largest urban agglomeration in South America. This approach can be especially useful for health agencies and public decision-makers in predicting SARS-CoV-2 outbreaks, as well as in local tracing of infection clusters.

Simultaneous removal of organic matter and nitrogen compounds from landfill leachate by aerobic granular sludge.

This study aimed at investigating the treatment of landfill leachate using the aerobic granular sludge process in a lab-scale sequential batch reactor (SBR-AGS). The leachate from a giant sanitary landfill localized in the State of São Paulo (Brazil) exhibited high concentration of organic matter (COD 5,300 ± 78 mg L-1) and total nitrogen (TKN 2,630 ± 355 mg L-1). Comparatively, the leachate was added to wastewater in three different volumetric ratios (5, 10 and 20%) and the mixtures were characterized over treatment. The results indicated that there were no significant changes in the behaviour of the biological process even at the highest leachate ratio. The granulation of the aerobic sludge occurred after 90 days of operation and the granules had a diameter of 485-1585 µm. SBR-AGS exhibited removal efficiency of 87-89% for organic matter and at least 98% for total nitrogen, regardless of the leachate ratio. The treated effluent that received 20% of leachate showed 2.7 mg L-1 ammonia and 1.1 mg L-1 nitrate. This study shows that SBR-AGS was able to form large granules, thus promoting a simultaneous nitrification and denitrification (SND) process. We highlighted that SND occurred in low dissolved oxygen concentrations (< 1.5 mg L-1) for 120 days, without compromising aerobic granule integrity. These results suggest that the aerobic granular sludge process is a promising alternative for the co-treatment of landfill leachate and domestic wastewater under tropical climate conditions and its use should be encouraged.

Treatment of real domestic sewage in a pilot-scale aerobic granular sludge reactor: Assessing start-up and operational control.

Aerobic granular sludge (AGS) has been considered a breakthrough in the wastewater treatment sector given its key characteristics, such as excellent settleability, simultaneous removal of organic and nutrient pollutants, and compactness. However, the formation of granules often delays the start-up of granular-based systems, especially in large-scale settings. This study addressed the start-up of a pilot-scale AGS sequencing batch reactor (SBR) treating domestic sewage, monitored for over 280 days. The challenges faced during aerobic granulation using a mixture of activated sludge and anaerobic granular sludge as inoculum and the performance of the reactor on organic matter, nitrogen, and phosphorus removal were discussed. Results showed that robust and stable granules were formed after an initial period of around six months, with the settling time playing a key role on granules development. At least 80% of granules had a diameter greater than 0.2 mm and 60% >1 mm. In general, the reactor achieved high nitrogen removal efficiency, as well as satisfactory removal of soluble COD. However, total COD abatement was impaired by the various episodes of suspended solids loss with the effluent. Overall, this study demonstrated that the reactor was efficient in the treatment of domestic sewage, but its performance was adversely affected from sudden changes in the influent quality. PRACTITIONER POINTS: Aerobic granular sludge (AGS) applied to small-scale domestic sewage treatment. The control of sludge age in AGS can be a problem due to short sedimentation times. High DO to maintain aerobic granulation can economically make the process economically unfeasible in tropical countries. A sludge with excellent sedimentation properties was obtained. However, maintaining the granule over time is a challenge.

Influence of Landfill Leachate on Organic Matter Removal in Activated Sludge and Activated Sludge with Carrier Processes

Abstract This study assessed the performance of two pilot units, a conventional activated sludge (CAS) and an Integrated Fixed-Film Activated Sludge (IAS), in the treatment of leachate from sanitary landfill combined with domestic sewage, with the aim of removing organic substances. In order to assess the possible impacts on treatment, three experimental phases were designed using influent leachate contributions of 5%, 10%, and 20% of the total BOD load. Overall, the results showed that no significant changes in the behavior of biological processes. The IAS unit presented average BOD removal efficiencies of above 88%, 87% and 80% for the three leachate load contributions of 5%, 10% and 20% studied. The CAS unit presented lower average efficiency with a 5% leachate contribution (64%), but displayed a similar performance to the IAS unit in the other phases (82% and 80%); similar quantitative aspects were observed for both treatment processes with regards to biomass composition analysis. The kinetic coefficients related to heterotrophic metabolism did not indicate negative effects on treatment efficiency as a result of the increase in leachate addition, with values of 2.8; 2.8 and 2.7 d-1 obtained respectively for each phase with the CAS unit, and 2.0; 1.5 and 1.6 d-1 with the IAS unit, representing similar values to those found in the literature (1.5 a 5.0 d-1). The results reinforced the conclusion that the leachate load introduced did not cause inhibition or significant alterations to heterotrophic metabolism, and, therefore, to the removal of organic matter.

Lodo ativado com aeração prolongada operado sob baixa concentração de oxigênio dissolvido: comportamento cinético das bactérias heterotróficas e autotróficas nitrificantes / Extended aeration activated sludge process operated under low dissolved oxygen concentration: Kinetic behavior of nitrifying heterotrophic and autotrophic bacteria

RESUMO Este trabalho teve como objetivo a determinação dos parâmetros cinéticos que descrevem o crescimento das bactérias heterotróficas e autotróficas nitrificantes em três sistemas de lodos ativados com aeração prolongada, operados com baixa aeração, a fim de se obter a remoção de nitrogênio de esgoto sanitário pelo processo de nitrificação e desnitrificação simultâneas (NDS). Em relação às remoções de matéria orgânica e nitrogênio, aplicando-se idade do lodo na faixa de 12 a 20 dias e concentrações de oxigênio dissolvido (OD) nos reatores em torno de 0,5 mg O2.L-1, foi possível obter eficiências superiores a 90% e um esgoto tratado com concentrações de nitrato inferiores a 7,0 mg N.L-1. Os experimentos realizados comprovam que, nas condições operacionais adotadas durante os testes respirométricos, as bactérias heterotróficas e autotróficas presentes nos lodos ativados estavam ativos e, ainda que baixa, a concentração de OD não foi limitante para o desenvolvimento da cinética dos processos envolvidos na remoção de materiais orgânico e nitrogenado.

ABSTRACT The objective of this work was to determine the kinetic parameters that describe the growth of heterotrophic and autotrophic nitrifying bacteria in three systems of activated sludge with prolonged aeration, operated with low aeration, with the objective of obtaining nitrogen removal of sanitary sewage by simultaneous nitrification and denitrification (SND). In relation to the removal of organic matter and nitrogen, by using sludge ages ranging from 12 to 20 days and OD concentrations in the reactors around 0.5 mgO2.L-1, it was possible to obtain efficiencies higher than 90% and a sewer treated with nitrate concentrations below 7.0 mgN.L-1. The experiments carried out prove that, under the operating conditions adopted during the respirometric tests, heterotrophic and autotrophic bacteria present in activated sludge were active and, although the low concentration of OD was not limiting for the development of the kinetics of the processes involved in the removal of organic and nitrogenous material.

Remoção simultânea de material orgânico, nitrogênio e fósforo em um reator em bateladas sequenciais com biofilme de leito móvel operado pelo processo anaeróbio-anóxico-óxico / Simultaneous removal of organic material, nitrogen and phosphorus in a reactor in batches of mobile bed biofilm sequencing by the anaerobic-anoxic-oxic process

RESUMO Neste estudo, foi avaliado o desempenho de um reator em bateladas sequenciais com biofilme de leito móvel operado pelo processo anaeróbio-anóxico-óxico (A2O), aplicado à remoção simultânea de material orgânico, nitrogênio e fósforo de esgoto sanitário. Utilizaram-se dois reatores com volume útil de 36 L cada, sendo que um deles recebeu anéis plásticos móveis do tipo Kaldnes®, com área superficial específica estimada em 310 m2/m3 que ocuparam 50% do volume útil do reator (denominado aqui de MBSBBR). O estudo foi conduzido com tempo de retenção de sólidos (TRS) de 12 dias, tempo de retenção hidráulico (TRH) de 6 horas, taxa de alimento/microrganismo (A/M) de 0,4 a 0,5 gDQO/SSV.d, oxigênio dissolvido (OD) na faixa de 0,3-0,8 mgO2/L e taxas de aplicação de carga orgânica, nitrogênio e fósforo de 47,2 gDQO/L.d, 5,4 gNTK/L.d e 0,4 gP/L.d, respectivamente. Os resultados de eficiência de remoção de demanda química de oxigênio (DQO), N-NH3 e fósforo foram de 91,1, 90,5 e 85,4% para o reator MBSBBR e de 89,5, 91,6 e 84,4% para o reator RBS, respectivamente. Os resultados do teste estatístico de ANOVA foram aplicados às principais variáveis estudadas, e os valores convergiram para valores de f observado < f crítico . Dessa forma, considera-se que houve similaridade de desempenho nos reatores MBSBBR e RBS. Conclui-se que a introdução do meio suporte não trouxe benefícios ao tratamento em termos de remoção de nutrientes. No entanto, o estudo mostrou que é possível de se obter elevada remoção de nitrogênio, em baixas concentrações de OD, o que se torna vantajoso para o tratamento de esgoto com elevada concentração de nitrogênio.

ABSTRACT In this study, the performance of a reactor in sequential batch with mobile bed biofilm operated by the anaerobic-anoxic-oxic (A2O) process, applied to the simultaneous removal of organic material, nitrogen and phosphorus from sanitary sewage was evaluated. Two reactors with a useful volume of 36 liters each were used, one of them receiving Kaldnes® type mobile plastic rings, with a specific surface area estimated at 310 m2/m3, which occupied 50% of the reactor's useful volume (MBSBBR). The study was conducted with 12-day solid retention time (SRT), 6 hour hydraulic retention time (HRT), food/microorganism (F/M) ratio of 0.4 to 0.5 gCOD/VSS.d, dissolved oxygen (DO) in the range of 0.3-0.8 mgO2/L and application rates of organic load, nitrogen and phosphorus of 47.2 gCOD/L.d, 5.4 gTNK/L.d and 0.4 gP/L.d, respectively. The efficiency of chemical oxygen demand (COD), N-NH3 and Phosphorus removal was of 91.1%, 90.5% and 85.4% for the MBSBBR reactor and 89.5%, 91.6% and 84.4% for the RBS reactor, respectively. The results of the statistical ANOVA test were applied to the main variables studied and the values converged to values of f observed < f critical . In this way, it is considered that there was a similarity of performance between the MBSBBR and RBS reactors. It is concluded that the introduction of the support medium did not bring benefits to the treatment in terms of nutrient removal. However, the study showed that it is possible to obtain high nitrogen removal at low concentrations of DO, which is advantageous for the sewage treatment with high nitrogen concentration.

Nitrificação e desnitrificação simultânea em reator com biomassa em suspensão e fluxo contínuo de esgoto / Simultaneous nitrification and denitrification in reactor with biomass in suspension and continuous flow of sewage

Recentemente foi desenvolvido na Europa o processo conhecido por Nitrificação e Desnitrificação Simultânea (NDS), em que o lodo ativado opera com tempo de retenção de sólidos suficientemente altos para que a nitrificação ocorra mesmo com baixa concentração de oxigênio dissolvido nos tanques de aeração, situação em que é possível a manutenção de condições anóxicas no interior dos flocos e a ocorrência da desnitrificação. Em países de clima tropical, muitas estações de tratamento de esgoto foram construídas ou estão sendo projetadas para operarem sob as condições da variante do processo de lodo ativado conhecida por aeração prolongada, reconhecidamente eficiente na remoção de matéria orgânica e na nitrificação do esgoto. Tais estações podem ser adaptadas para estas novas condições, de forma a se obter o benefício técnico da desnitrificação e o ganho econômico relativo à redução no consumo de energia elétrica para aeração. Este estudo avaliou as condições de remoção de nitrogênio via NDS em um sistema de lodo ativado com aeração prolongada alimentada em fluxo contínuo com diferentes idades do lodo 40, 30 e 20 dias, tratando esgoto sanitário. O processo de tratamento permaneceu estável durante todo o período de observação. Os resultados experimentais demonstram que é possível manter baixas concentrações de OD no tanque de aeração em uma faixa relativamente estreita (0,3-0,8 mg.L-1), e, consequentemente desenvolver os processos de nitrificação e desnitrificação sob as mesmas condições (NDS) com alta eficiência e estabilidade. A remoção de DQO e DBO foi acima de 90 por cento . Os valores de crescimento específico máximo (m20) das bactérias autotróficas nitritantes e nitratantes, resultaram muito próximos aos descritos na literatura, sendo para a idade do lodo de 20 dias os valores de 0,68 d-1 para as bactérias nitritantes e de 0,62 d-1 para as bactérias nitratantes. Os valores das taxas de utilização máxima de amônia foram maiores do que as obtidas com o nitrito e, pode-se observar que o acúmulo de nitrato devido à diminuição da amônia foi mais rápido do que o com nitrito. Desse modo, conclui-se que a nitrificação foi governada pelas bactérias autotróficas nitritantes. A taxa de utilização de nitrato foi maior tendo como substrato material rapidamente biodegradável. Confirmando que a disponibilidade de carbono tem papel fundamental nas taxas de desnitrificação no processo de NDS.