ABSTRACT
This study aimed to evaluate, by molecular methods, the presence of influenza A virus (IAV) and coronavirus in non-hematophagous bats collected in the state of São Paulo, Brazil. Samples of lung tissue and small intestine from 105 bats belonging to three families (Phyllostomidae, Vespertilionidae, and Molossidae) were collected in 22 municipalities in the state of São Paulo. Genetic identification of bats species was performed by amplification and sequencing of a fragment of 710 bp of the mitochondrial COI gene. In the detection of IAV, genomes were performed by RT-PCR, aiming at the amplification of a 245-bp fragment of the IAV matrix (M) protein gene. For coronaviruses, two fragments of 602 and 440 bp corresponding to segments along the gene encoding the RNA-dependent RNA polymerase (RdRp) were targeted. The detection limit for each of the PCRs was also determined. All samples analyzed here were negative for both viruses, and the lower limit of detection of the PCRs for the amplification of influenza virus A and coronavirus was estimated at 3.5 × 103 and 4.59 genomic copies per microliter, respectively. Although bats have been shown to harbor a large number of pathogens, the results of the present study support the theory that virus circulation in bats in the wild often occurs at low viral loads and that our understanding of the complex infectious dynamics of these viruses in wild conditions is still limited.
ABSTRACT
Wastewater-based epidemiology (WBE) may be successfully used to comprehensively monitor and determine the scale and dynamics of some infections in the community. We monitored severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in raw wastewater samples from Porto Alegre, Southern Brazil. The samples were collected and analyzed every week between May 2020 to May 2021. Meanwhile, different social restrictions were applied according to the number of hospitalized patients in the region. Weekly samples were obtained from two wastewater treatment plants (WWTP) (named Navegantes and Serraria). To determine the SARS-CoV-2 RNA titers in wastewater, we performed RT-qPCR analysis targeting the N gene (N1). The highest titer of SARS-CoV-2 RNA was observed between epidemiological weeks (EW) 33-37 (August), 42-43 (October), 45-46 (November), 49-51 (December) in 2020, and 1-3 (January), 7-13 (February to March) in 2021, with viral loads ranging from 1 x 106 ∼ 3 x 106 genomic copies/Liter. An increase in positive confirmed cases followed such high viral loads. Depending on the sampling method used, positive cases increased in 6∼7 days and 15 days after the rise of viral RNA titers in wastewater, with composite sampling methods showing a lower time lag and a higher resolution on the analyses. The results showed a direct relation between strict social restrictions and the loads of detected RNA reduction in wastewater, corroborating the number of confirmed cases. Differences in viral loads between different sampling points and methods were observed, as composite samples showed more stable results during the analyzed period. Besides, viral loads obtained from samples collected at Serraria WWTP were consistently higher than the ones obtained at Navegantes WWTP, indicating differences in local dynamics of SARS-CoV-2 spread in different regions of Porto Alegre. In conclusion, wastewater sampling to monitor SARS-CoV-2 is a robust tool to evaluate the viral loads contributing to hospitalized patients' data and confirmed cases. In addition, SARS-CoV-2 detection in sewage may inform and alert the government when the number of asymptomatic or non/tested patients is in evidence. Graphical [Image 1].
ABSTRACT
Wastewater-based epidemiology (WBE) may be successfully used to comprehensively monitor and determine the scale and dynamics of some infections in the community. We monitored severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA in raw wastewater samples from Porto Alegre, Southern Brazil. The samples were collected and analyzed every week between May 2020 to May 2021. Meanwhile, different social restrictions were applied according to the number of hospitalized patients in the region. Weekly samples were obtained from two wastewater treatment plants (WWTP) (named Navegantes and Serraria). To determine the SARS-CoV-2 RNA titers in wastewater, we performed RT-qPCR analysis targeting the N gene (N1). The highest titer of SARS-CoV-2 RNA was observed between epidemiological weeks (EW) 33-37 (August), 42-43 (October), 45-46 (November), 49-51 (December) in 2020, and 1-3 (January), 7-13 (February to March) in 2021, with viral loads ranging from 1 x 106 ∼ 3 x 106 genomic copies/Liter. An increase in positive confirmed cases followed such high viral loads. Depending on the sampling method used, positive cases increased in 6∼7 days and 15 days after the rise of viral RNA titers in wastewater, with composite sampling methods showing a lower time lag and a higher resolution on the analyses. The results showed a direct relation between strict social restrictions and the loads of detected RNA reduction in wastewater, corroborating the number of confirmed cases. Differences in viral loads between different sampling points and methods were observed, as composite samples showed more stable results during the analyzed period. Besides, viral loads obtained from samples collected at Serraria WWTP were consistently higher than the ones obtained at Navegantes WWTP, indicating differences in local dynamics of SARS-CoV-2 spread in different regions of Porto Alegre. In conclusion, wastewater sampling to monitor SARS-CoV-2 is a robust tool to evaluate the viral loads contributing to hospitalized patients' data and confirmed cases. In addition, SARS-CoV-2 detection in sewage may inform and alert the government when the number of asymptomatic or non/tested patients is in evidence.