Scalable RT-LAMP-based SARS-CoV-2 testing for infection surveillance with applications in pandemic preparedness.

Throughout the SARS-CoV-2 pandemic, limited diagnostic capacities prevented sentinel testing, demonstrating the need for novel testing infrastructures. Here, we describe the setup of a cost-effective platform that can be employed in a high-throughput manner, which allows surveillance testing as an acute pandemic control and preparedness tool, exemplified by SARS-CoV-2 diagnostics in an academic environment. The strategy involves self-sampling based on gargling saline, pseudonymized sample handling, automated RNA extraction, and viral RNA detection using a semiquantitative multiplexed colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay with an analytical sensitivity comparable with RT-qPCR. We provide standard operating procedures and an integrated software solution for all workflows, including sample logistics, analysis by colorimetry or sequencing, and communication of results. We evaluated factors affecting the viral load and the stability of gargling samples as well as the diagnostic sensitivity of the RT-LAMP assay. In parallel, we estimated the economic costs of setting up and running the test station. We performed > 35,000 tests, with an average turnover time of < 6 h from sample arrival to result announcement. Altogether, our work provides a blueprint for fast, sensitive, scalable, cost- and labor-efficient RT-LAMP diagnostics, which is independent of potentially limiting clinical diagnostics supply chains.

Evaluating combustion kinetics and quantifying fuel-N conversion tendency of shoe manufacturing waste.

Combustion is an effective and cost-efficient thermochemical conversion method for solid waste, showing promise for the resource utilization of shoe manufacturing waste (SMW). However, SMW is generally composed of different components, which can lead to unstable combustion and excessive pollutant emissions, especially NOx. To date, combustion characteristics, reaction mechanism and fuel nitrogen (fuel-N) conversion of different SMW components remain unclear. In this work, the combustion behavior of typical SMW components combustion was investigated using Thermogravimetric coupled with Fourier transform infrared spectrum (TG-FTIR). A simplified single-step reaction mechanism was proposed according to the temperature interval to estimate reaction mechanism of SMW. Additionally, the relationship between fuel-N conversion tendency and fuel properties was established. The results indicate that the values for the comprehensive combustion performance index (S) and flammability index (C) range from 1.65 to 0.44 and 3.98 to 1.37, respectively. This demonstrates the significant variability in combustion behavior among different SMW components. Cardboard, leather and sponge have higher values of S and C, suggesting a better ignition characteristic and a stable combustion process. During the combustion of SMW, nitrogen oxides (NO and N2O) are the main nitrogen-containing compounds in the flue gases, with NO being the major contributor, accounting for over 82.97 % of the nitrogen oxides. NO has a negative correlation with nitrogen content, but it is opposite for N2O, HCN and NH3. Furthermore, the conversion of NO, N2O and NH3 is proportional to logarithmic values of O/N, while its conversion to HCN is proportional to logarithmic values of VM/N. These findings facilitate the prediction of the fuel-N conversion of solid waste combustion. This work might shed light on combustion optimization and in-situ pollutant emission control in solid waste combustion.

Genome-wide C-SWAT library for high-throughput yeast genome tagging.

Here we describe a C-SWAT library for high-throughput tagging of Saccharomyces cerevisiae open reading frames (ORFs). In 5,661 strains, we inserted an acceptor module after each ORF that can be efficiently replaced with tags or regulatory elements. We validated the library with targeted sequencing and tagged the proteome with bright fluorescent proteins to quantify the effect of heterologous transcription terminators on protein expression and to localize previously undetected proteins.

YeastRGB: comparing the abundance and localization of yeast proteins across cells and libraries.

The ability to measure the abundance and visualize the localization of proteins across the yeast proteome has stimulated hypotheses on gene function and fueled discoveries. While the classic C' tagged GFP yeast library has been the only resource for over a decade, the recent development of the SWAT technology has led to the creation of multiple novel yeast libraries where new-generation fluorescent reporters are fused at the N' and C' of open reading frames. Efficient access to these data requires a user interface to visualize and compare protein abundance, localization and co-localization across cells, strains, and libraries. YeastRGB (www.yeastRGB.org) was designed to address such a need, through a user-friendly interface that maximizes informative content. It employs a compact display where cells are cropped and tiled together into a 'cell-grid.' This representation enables viewing dozens of cells for a particular strain within a display unit, and up to 30 display units can be arrayed on a standard high-definition screen. Additionally, the display unit allows users to control zoom-level and overlay of images acquired using different color channels. Thus, YeastRGB makes comparing abundance and localization efficient, across thousands of cells from different strains and libraries.

Accurate mapping of Chinese coastal aquaculture ponds using biophysical parameters based on Sentinel-2 time series images.

Aquaculture plays a crucial role in the global food security and nutrition supply, where China accounts for the largest market share. Although there are some studies that focus on large-scale extraction of coastal aquaculture ponds from satellite images, they have often variable accuracies and encounter misclassification due to the similar geometric characteristics of various vivid water bodies. This paper proposes an efficient and novel method that integrates the spatial characteristics and three biophysical parameters (Chlorophyll-a, Trophic State Index, and Floating Algae Index) to map coastal aquaculture ponds at a national scale. These parameters are derived from bio-optical models based on the Google Earth Engine (GEE) cloud computing platform and time series of high-resolution Sentinel-2 images. Our proposed method effectively addresses the misclassification issue between the aquaculture ponds and rivers, lakes, reservoirs, and salt pans and achieves an overall accuracy of 91 % and a Kappa coefficient of 0.83 in the Chinese coastal zone. Our results indicate that the total area of Chinese coastal aquaculture ponds was 1,039,214 ha in 2019, mainly distributed in the Shandong and Guangdong provinces. The highest aquaculture intensity occurs within the 1 km coastal buffer zone, accounting for 22.4 % of the total area. Furthermore, more than half of the Chinese coastal aquaculture ponds are concentrated in the 0-5 km buffer zone. Our method is of general applicability and thus is suitable for large-scale aquaculture ponds mapping projects. Moreover, the biophysical parameters we employ can be considered as new indicators for the classification of various water bodies even with different aquaculture species.

Quantifying seasonal variations in microphytobenthos biomass on estuarine tidal flats using Sentinel-1/2 data.

Quantifying temporal and spatial changes in microphytobenthos (MPB) biomass is critical for understanding its ecological function in estuarine food web networks and carbon flows. However, tidal fluctuations and the complex composition of coastal sediment limit remote sensing applications for estimating MPB biomass seasonal variations in estuarine tidal flats. We integrated radar Sentinel-1 (S1) and optical Sentinel-2 (S2) remote sensing data to quantify the temporal and spatial variability in MPB biomass in the Changjiang estuary, China. Pixels of water bodies on the tidal flats were removed by dynamic threshold segmentation of the water index with the combined S1 and S2 data, and salt marsh pixels were masked with the first red-edge band in the S2 data. We used the continuum-removed spectral absorption depth feature to construct a regression model for estimating MPB biomass with a regression coefficient of 0.81. The results showed that spectral absorption continuum removal methods using broadband multispectral data for MPB estimation are a promising alternative to hyperspectral narrowband ratio operation. Compared with the widely used normalized difference vegetation index (NDVI), the scaled absorption depth feature was more stable for MPB estimation under a changeable sediment background. The produced seasonal map showed that the high biomass levels of the MPB in the study area are not limited to one season and one site, with an annual mean biomass of 14.39 mg chlorophyll a (Chl-a)·m-2 and 71% confirmed accuracy. The highest biomass levels occurred in summer in the supratidal zone (19.51 mg Chl-a·m-2) and in spring in the intertidal zone (17.10 mg Chl-a·m-2) in the Changjiang estuary. The relative shore height, derived from the tidal range here, is an important variable that shapes the MPB spatial distribution. This study demonstrates the potential of integrating high-spatial-resolution (10 m) S1 and S2 data for future large-scale estimation of intertidal MPB.

Effectiveness and cost-effectiveness of four different strategies for SARS-CoV-2 surveillance in the general population (CoV-Surv Study): a structured summary of a study protocol for a cluster-randomised, two-factorial controlled trial.

OBJECTIVES: In this cluster-randomised controlled study (CoV-Surv Study), four different "active" SARS-CoV-2 testing strategies for general population surveillance are evaluated for their effectiveness in determining and predicting the prevalence of SARS-CoV-2 infections in a given population. In addition, the costs and cost-effectiveness of the four surveillance strategies will be assessed. Further, this trial is supplemented by a qualitative component to determine the acceptability of each strategy. Findings will inform the choice of the most effective, acceptable and affordable strategy for SARS-CoV-2 surveillance, with the most effective and cost-effective strategy becoming part of the local public health department's current routine health surveillance activities. Investigating its everyday performance will allow us to examine the strategy's applicability to real time prevalence prediction and the usefulness of the resulting information for local policy makers to implement countermeasures that effectively prevent future nationwide lockdowns. The authors would like to emphasize the importance and relevance of this study and its expected findings in the context of population-based disease surveillance, especially in respect to the current SARS-CoV-2 pandemic. In Germany, but also in many other countries, COVID-19 surveillance has so far largely relied on passive surveillance strategies that identify individuals with clinical symptoms, monitor those cases who then tested positive for the virus, followed by tracing of individuals in close contact to those positive cases. To achieve higher effectiveness in population surveillance and to reliably predict the course of an outbreak, screening and monitoring of infected individuals without major symptoms (about 40% of the population) will be necessary. While current testing capacities are also used to identify such asymptomatic cases, this rather passive approach is not suitable in generating reliable population-based estimates of the prevalence of asymptomatic carriers to allow any dependable predictions on the course of the pandemic. To better control and manage the SARS-CoV-2 pandemic, current strategies therefore need to be complemented by an active surveillance of the wider population, i.e. routinely conducted testing and monitoring activities to identify and isolate infected individuals regardless of their clinical symptoms. Such active surveillance strategies will enable more effective prevention of the spread of the virus as they can generate more precise population-based parameters during a pandemic. This essential information will be required in order to determine the best strategic and targeted short-term countermeasures to limit infection spread locally. TRIAL DESIGN: This trial implements a cluster-randomised, two-factorial controlled, prospective, interventional, single-blinded design with four study arms, each representing a different SARS-CoV-2 testing and surveillance strategy. PARTICIPANTS: Eligible are individuals age 7 years or older living in Germany's Rhein-Neckar Region who consent to provide a saliva sample (all four arms) after completion of a brief questionnaire (two arms only). For the qualitative component, different samples of study participants and non-participants (i.e. eligible for study, but refuse to participate) will be identified for additional interviews. For these interviews, only individuals age 18 years or older are eligible. INTERVENTION AND COMPARATOR: Of the four surveillance strategies to be assessed and compared, Strategy A1 is considered the gold standard for prevalence estimation and used to determine bias in other arms. To determine the cost-effectiveness, each strategy is compared to status quo, defined as the currently practiced passive surveillance approach. Strategy A1: Individuals (one per household) receive information and study material by mail with instructions on how to produce a saliva sample and how to return the sample by mail. Once received by the laboratory, the sample is tested for SARS-CoV-2 using Reverse Transcription Loop-mediated Isothermal Amplification (RT-LAMP). Strategy A2: Individuals (one per household) receive information and study material by mail with instructions on how to produce their own as well as saliva samples from each household member and how to return these samples by mail. Once received by the laboratory, the samples are tested for SARS-CoV-2 using RT-LAMP. Strategy B1: Individuals (one per household) receive information by mail on how to complete a brief pre-screening questionnaire which asks about COVID-19 related clinical symptoms and risk exposures. Only individuals whose pre-screening score crosses a defined threshold, will then receive additional study material by mail with instructions on how to produce a saliva sample and how to return the sample by mail. Once received by the laboratory, the saliva sample is tested for SARS-CoV-2 using RT-LAMP. Strategy B2: Individuals (one per household) receive information by mail on how to complete a brief pre-screening questionnaire which asks about COVID-19 related clinical symptoms. Only individuals whose pre-screening score crosses a defined threshold, will then receive additional study material by mail with instructions how to produce their own as well as saliva samples from each household member and how to return these samples by mail. Once received by the laboratory, the samples are tested for SARS-CoV-2 using RT-LAMP. In each strategy, RT-LAMP positive samples are additionally analyzed with qPCR in order to minimize the number of false positives. MAIN OUTCOMES: The identification of the one best strategy will be determined by a set of parameters. Primary outcomes include costs per correctly screened person, costs per positive case, positive detection rate, and precision of positive detection rate. Secondary outcomes include participation rate, costs per asymptomatic case, prevalence estimates, number of asymptomatic cases per study arm, ratio of symptomatic to asymptomatic cases per study arm, participant satisfaction. Additional study components (not part of the trial) include cost effectiveness of each of the four surveillance strategies compared to passive monitoring (i.e. status quo), development of a prognostic model to predict hospital utilization caused by SARS-CoV-2, time from test shipment to test application and time from test shipment to test result, and perception and preferences of the persons to be tested with regard to test strategies. RANDOMISATION: Samples are drawn in three batches of three continuous weeks. Randomisation follows a two-stage process. First, a total of 220 sampling points have been allocated to the three different batches. To obtain an integer solution, the Cox-algorithm for controlled rounding has been used. Afterwards, sample points have been drawn separately per batch, following a probability proportional to size (PPS) random sample. Second, for each cluster the same number of residential addresses is randomly sampled from the municipal registries (self-weighted sample of individuals). The 28,125 addresses drawn per municipality are then randomly allocated to the four study arms A1, A2, B1, and B2 in the ratio 5 to 2.5 to 14 to 7 based on the expected response rates in each arm and the sensitivity and specificity of the pre-screening tool as applied in strategy B1 and B2. Based on the assumptions, this allocation should yield 2500 saliva samples in each strategy. Although a municipality can be sampled by multiple batches and the overall number of addresses per municipality might vary, the number of addresses contacted in each arm is kept constant. BLINDING (MASKING): The design is single-blinded, meaning the staff conducting the SARS-CoV-2 tests are unaware of the study arm assignment of each single participant and test sample. SAMPLE SIZES: Total sample size for the trial is 10,000 saliva samples equally allocated to the four study arms (i.e. 2,500 participants per arm). For the qualitative component, up to 60 in-depth interviews will be conducted with about 30 study participants (up to 15 in each arm A and B) and 30 participation refusers (up to 15 in each arm A and B) purposefully selected from the quantitative study sample to represent a variety of gender and ages to explore experiences with admission or rejection of study participation. Up to 25 asymptomatic SARS-CoV-2 positive study participants will be purposefully selected to explore the way in which asymptomatic men and women diagnosed with SARS-CoV-2 give meaning to their diagnosis and to the dialectic between feeling concurrently healthy and yet also being at risk for transmitting COVID-19. In addition, 100 randomly selected study participants will be included to explore participants' perspective on testing processes and implementation. TRIAL STATUS: Final protocol version is "Surveillance_Studienprotokoll_03Nov2020_v1_2" from November 3, 2020. Recruitment started November 18, 2020 and is expected to end by or before December 31, 2020. TRIAL REGISTRATION: The trial is currently being registered with the German Clinical Trials Register (Deutsches Register Klinischer Studien), DRKS00023271 ( https://www.drks.de/drks_web/navigate.do?navigationId=trial . HTML&TRIAL_ID=DRKS00023271). Retrospectively registered 30 November 2020. FULL PROTOCOL: The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest in expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol.

Effectiveness and cost-effectiveness of four different strategies for SARS-CoV-2 surveillance in the general population (CoV-Surv Study): study protocol for a two-factorial randomized controlled multi-arm trial with cluster sampling.

BACKGROUND: To achieve higher effectiveness in population-based SARS-CoV-2 surveillance and to reliably predict the course of an outbreak, screening, and monitoring of infected individuals without major symptoms (about 40% of the population) will be necessary. While current testing capacities are also used to identify such asymptomatic cases, this rather passive approach is not suitable in generating reliable population-based estimates of the prevalence of asymptomatic carriers to allow any dependable predictions on the course of the pandemic. METHODS: This trial implements a two-factorial, randomized, controlled, multi-arm, prospective, interventional, single-blinded design with cluster sampling and four study arms, each representing a different SARS-CoV-2 testing and surveillance strategy based on individuals' self-collection of saliva samples which are then sent to and analyzed by a laboratory. The targeted sample size for the trial is 10,000 saliva samples equally allocated to the four study arms (2500 participants per arm). Strategies differ with respect to tested population groups (individuals vs. all household members) and testing approach (without vs. with pre-screening survey). The trial is complemented by an economic evaluation and qualitative assessment of user experiences. Primary outcomes include costs per completely screened person, costs per positive case, positive detection rate, and precision of positive detection rate. DISCUSSION: Systems for active surveillance of the general population will gain more importance in the context of pandemics and related disease prevention efforts. The pandemic parameters derived from such active surveillance with routine population monitoring therefore not only enable a prospective assessment of the short-term course of a pandemic, but also a more targeted and thus more effective use of local and short-term countermeasures. TRIAL REGISTRATION: ClinicalTrials.gov DRKS00023271 . Registered November 30, 2020, with the German Clinical Trials Register (Deutsches Register Klinischer Studien).

Rapid Loss of Tidal Flats in the Yangtze River Delta since 1974.

As the home to national nature reserves and a Ramsar wetland, the tidal flats of the Yangtze River Delta are of great significance for ecological security, at both the local and global scales. However, a comprehensive understanding of the spatiotemporal conditions of the tidal flats in the Yangtze River Delta remains lacking. Here, we propose using remote sensing to obtain a detailed spatiotemporal profile of the tidal flats, using all available Landsat images from 1974 to 2018 with the help of the Google Earth Engine cloud platform. In addition, reclamation data were manually extracted from time series Landsat images for the same period. We found that approximately 40.0% (34.9-43.1%) of the tidal flats in the study area have been lost since 1980, the year in which the tidal flat area was maximal. The change in the tidal flat areas was consistent with the change in the riverine sediment supply. We also found that the cumulative reclamation areas totaled 816.6 km2 and 431.9 km2 in the Yangtze estuary zone and along the Jiangsu coast, respectively, between 1974 and 2018. Because of reclamation, some areas (e.g., the Hengsha eastern shoal and Pudong bank), which used to be quite rich, have lost most of their tidal flats. Currently, almost 70% of the remaining tidal flats are located in the shrinking branch (North Branch) and the two National Nature Reserves (Chongming Dongtan and Jiuduansha) in the Yangtze estuary zone. Consequently, the large-scale loss of tidal flats observed was primarily associated with reduced sediment supply and land reclamation at the time scale of the study. Because increasing demand for land and rising sea levels are expected in the future, immediate steps should be taken to prevent the further deterioration of this valuable ecosystem.