Comparison of Nanopore and Synthesis-Based Next-Generation Sequencing Platforms for SARS-CoV-2 Variant Monitoring in Wastewater

Shortly after the beginning of the SARS-CoV-2 pandemic, many countries implemented sewage sentinel systems to monitor the circulation of the virus in the population. A fundamental part of these surveillance programs is the variant tracking through sequencing approaches to monitor and identify new variants or mutations that may be of importance. Two of the main sequencing platforms are Illumina and Oxford Nanopore Technologies. Here, we compare the performance of MiSeq (Illumina) and MinION (Oxford Nanopore Technologies), as well as two different data processing pipelines, to determine the effect they may have on the results. MiSeq showed higher sequencing coverage, lower error rate, and better capacity to detect and accurately estimate variant abundances than MinION R9.4.1 flow cell data. The use of different variant callers (LoFreq and iVar) and approaches to calculate the variant proportions had a remarkable impact on the results generated from wastewater samples. Freyja, coupled with iVar, may be more sensitive and accurate than LoFreq, especially with MinION data, but it comes at the cost of having a higher error rate. The analysis of MinION R10.4.1 flow cell data using Freyja combined with iVar narrows the gap with MiSeq performance in terms of read quality, accuracy, sensitivity, and number of detected mutations. Although MiSeq should still be considered as the standard method for SARS-CoV-2 variant tracking, MinION’s versatility and rapid turnaround time may represent a clear advantage during the ongoing pandemic

Dynamics of SARS-CoV-2 Alpha (B.1.1.7) variant spread: The wastewater surveillance approach

Wastewater based epidemiology (WBE) offers an overview of the SARS-CoV-2 variants circulating among the population thereby serving as a proper surveillance method. The variant of concern (VOC) Alpha was first identified in September 2020 in the United Kingdom, and rapidly became dominant across Europe. Our objective was to elucidate the Alpha VOC outcompetition rate and identify mutations in the spike glycoprotein (S) gene, indicative of the circulation of the Alpha VOC and/or other variants in the population through wastewater analysis. In the period covered by this study (November 2020-April 2021), forteen wastewater treatment plants (WWTPs) were weekly sampled. The total number of SARS-CoV-2 genome copies per L (GC/L) was determined with a Real-Time qPCR, targeting the N gene. Surveillance of the Alpha VOC circulation was ascertained using a duplex RT-qPCR, targeting and discriminating the S gene. Our results showed that in a period of 6 weeks the Alpha VOC was present in all the studied WWTPs, and became dominant in 11 weeks on average. The outcompetition rates of the Alpha VOC were estimated, and their relationship with different parameters statistically analyzed. The rapid spread of the Alpha VOC was influenced by its initial input and by the previous circulation of SARS-COV-2 in the population. This latter point could be explained by its higher transmissibility, particularly advantadgeous when a certain degree of herd immunity exists. Moreover, the presence of signature mutations of SARS-COV-2 variants were established by deep-sequencing of the complete S gene. The circulation of the Alpha VOC in the area under study was confirmed, and additionally two combinations of mutations in the S glycoprotein (T73A and D253N, and S477N and A522S) that could affect antibody binding were identified

Wastewater based epidemiology beyond SARS-CoV-2: Spanish wastewater reveals the current spread of Monkeypox virus

Besides nasopharyngeal swabs, monkeypox virus (MPXV) DNA has been detected in a variety of samples such as saliva, semen, urine and fecal samples. Using the environmental surveillance network previously developed in Spain for the routine wastewater surveillance of SARS-CoV-2 (VATar COVID-19), we have analyzed the presence of MPXV DNA in wastewater from different areas of Spain. Samples (n=312) from 24 different wastewater treatment plants were obtained between May 9 (week 22_19) and August 4 (week 22_31), 2022. Following concentration of viral particles by flocculation, a qPCR procedure allowed us to detect MPXV DNA in 63 wastewater samples collected from May 16 to August 4, 2022, with values ranging between 2.2 per 103 to 8.7 per 104 genome copies (gc)/L. This study shows that MPXV DNA can be reproducibly detected by qPCR in longitudinal samples collected from different Spanish wastewater treatment plants. According to data from the National Epidemiological Surveillance Network (RENAVE) in Spain a total of 6,119 cases have been confirmed as of August 19, 2022. However, and based on the wastewater data, the reported clinical cases seem to be underestimated and asymptomatic infections may be more frequent than expected.his research was supported by the European Commission NextGenerationEU fund, through CSIC's Global Health Platform (PTI Salud Global) and samples were obtained from the COVID-19 wastewater surveillance project (VATar COVID-19) funded by the Spanish Ministry for the Ecological Transition and the Demographic Challenge and the Spanish Ministry of Health. IGG is recipient of a predoctoral contract from the Generalitat Valenciana (ACIF/2021/181) and AP-C was supported by a postdoctoral fellowship (APOSTD/2021/292). PT is holding a Ramon y Cajal contract from the Ministerio de Ciencia e Innovacion and AC is recipient of a predoctoral contract FI-SDUR from the Generalitat de CatalunyaN

The Catalan Surveillance Network of SARS-CoV-2 in Sewage: design, implementation, and performance

Wastewater-based epidemiology has shown to be an efficient tool to track the circulation of SARS-CoV-2 in communities assisted by wastewater treatment plants (WWTPs). The challenge comes when this approach is employed to help Health authorities in their decision-making. Here, we describe the roadmap for the design and deployment of SARSAIGUA, the Catalan Surveillance Network of SARS-CoV-2 in Sewage. The network monitors, weekly or biweekly, 56 WWTPs evenly distributed across the territory and serving 6 M inhabitants (80% of the Catalan population). Each week, samples from 45 WWTPs are collected, analyzed, results reported to Health authorities, and finally published within less than 72 h in an online dashboard ( https://sarsaigua.icra.cat ). After 20 months of monitoring (July 20-March 22), the standardized viral load (gene copies/day) in all the WWTPs monitored fairly matched the cumulative number of COVID-19 cases along the successive pandemic waves, showing a good fit with the diagnosed cases in the served municipalities (Spearman Rho = 0.69). Here we describe the roadmap of the design and deployment of SARSAIGUA while providing several open-access tools for the management and visualization of the surveillance data.The authors wish to thank the staff from all the WWTPs monitored for their help and technical support during the sampling campaigns. The authors acknowledge the funding received from the ACA and the ASPCAT from the Catalan Government (Generalitat de Catalunya). ICRA authors acknowledge the funding provided by the Generalitat de Catalunya through the Consolidated Research Group grants ICRA-ENV 2017 SGR 1124 and ICRA-TiA 2017 SGR 1318. ICRA researchers also thank the funding from the CERCA program of the Catalan Government.Peer reviewe

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

Concurrent respiratory motion correction of abdominal PET and dynamic contrast-enhanced-MRI using a compressed sensing approach

We present an approach for concurrent reconstruction of respiratory motion-compensated abdominal dynamic contrast-enhanced (DCE)-MRI and PET data in an integrated PET/MR scanner. The MR and PET reconstructions share the same motion vector fields derived from radial MR data; the approach is robust to changes in respiratory pattern and does not increase the total acquisition time. Methods: PET and DCE-MRI data of 12 oncologic patients were simultaneously acquired for 6 min on an integrated PET/MR system after administration of 18F-FDG and gadoterate meglumine. Goldenangle radial MR data were continuously acquired simultaneously with PET data and sorted into multiple motion phases on the basis of a respiratory signal derived directly from the radial MR data. The resulting multidimensional dataset was reconstructed using a compressed sensing approach that exploits sparsity among respiratory phases. Motion vector fields obtained using the full 6-min (MC6-min) and only the last 1 min (MC1-min) of data were incorporated into the PET reconstruction to obtain motion-corrected PET images and in an MR iterative reconstruction algorithm to produce a series of motion-corrected DCE-MR images (moco-GRASP). The motioncorrection methods (MC6-min and MC1-min) were evaluated by qualitative analysis of the MR images and quantitative analysis of SUVmax and SUVmean, contrast, signal-to-noise ratio (SNR), and lesion volume in the PET images. Results: Motion-corrected MC6-min PET images demonstrated 30%, 23%, 34%, and 18% increases in average SUVmax, SUVmean, contrast, and SNR and an average 40% reduction in lesion volume with respect to the non-motion-corrected PET images. The changes in these figures of merit were smaller but still substantial for the MCMC1-min protocol: 19%, 10%, 15%, and 9% increases in average SUVmax, SUVmean, contrast, and SNR; and a 28% reduction in lesion volume. Moco-GRASP images were deemed of acceptable or better diagnostic image quality with respect to conventional breath-hold Cartesian volumetric interpolated breath-hold examination acquisitions. Conclusion: We presented a method that allows the simultaneous acquisition of respiratory motion-corrected diagnostic quality DCE-MRI and quantitatively accurate PET data in an integrated PET/MR scanner with negligible prolongation in acquisition time compared with routine PET/DCE-MRI protocols

PET/MRI in the presence of metal implants: Completion of the attenuation map from PET emission data

We present a novel technique for accurate whole-body attenuation correction in the presence of metallic endoprosthesis, on integrated non-time-of-flight (non-TOF) PET/MRI scanners. The proposed implant PET-based attenuationmap completion (IPAC) method performs a joint reconstruction of radioactivity and attenuation from the emission data to determine the position, shape, and linear attenuation coefficient (LAC) of metallic implants. Methods: The initial estimate of the attenuation map was obtained using the MR Dixon method currently available on the Siemens Biograph mMR scanner. The attenuation coefficients in the area of the MR image subjected to metal susceptibility artifacts are then reconstructed fromthe PET emission data using the IPAC algorithm. The method was tested on 11 subjects presenting 13 different metallic implants, who underwent CT and PET/MR scans. Relative mean LACs and Dice similarity coefficients were calculated to determine the accuracy of the reconstructed attenuation values and the shape of the metal implant, respectively. The reconstructed PET images were compared with those obtained using the reference CT-based approach and the Dixon-based method. Absolute relative change (aRC) images were generated in each case, and voxel-based analyses were performed. Results: The error in implant LAC estimation, using the proposed IPAC algorithm, was 15.7%± 7.8%, which was significantly smaller than the Dixon- (100%) and CT-(39%) derived values. A mean Dice similarity coefficient of 73% ± 9% was obtained when comparing the IPAC- with the CT-derived implant shape. The voxel-based analysis of the reconstructed PET images revealed quantification errors (aRC) of 13.2% ± 22.1% for the IPACwith respect to CT-corrected images. The Dixon-based method performed substantially worse, with a mean aRC of 23.1% ± 38.4%. Conclusion: We have presented a non-TOF emission-based approach for estimating the attenuation map in the presence of metallic implants, to be used for whole-body attenuation correction in integrated PET/MR scanners. The Graphics Processing Unit implementation of the algorithm will be included in the open-source reconstruction toolbox Occiput.io

Risdiplam-Treated Infants with Type 1 Spinal Muscular Atrophy versus Historical Controls

Background Type 1 spinal muscular atrophy (SMA) is a progressive neuromuscular disease characterized by an onset at 6 months of age or younger, an inability to sit without support, and deficient levels of survival of motor neuron (SMN) protein. Risdiplam is an orally administered small molecule that modifies SMN2 pre-messenger RNA splicing and increases levels of functional SMN protein in blood. Methods We conducted an open-label study of risdiplam in infants with type 1 SMA who were 1 to 7 months of age at enrollment. Part 1 of the study (published previously) determined the dose to be used in part 2 (reported here), which assessed the efficacy and safety of daily risdiplam as compared with no treatment in historical controls. The primary end point was the ability to sit without support for at least 5 seconds after 12 months of treatment. Key secondary end points were a score of 40 or higher on the Children's Hospital of Philadelphia Infant Test of Neuromuscular Disorders (CHOP-INTEND; range, 0 to 64, with higher scores indicating better motor function), an increase of at least 4 points from baseline in the CHOP-INTEND score, a motor-milestone response as measured by Section 2 of the Hammersmith Infant Neurological Examination (HINE-2), and survival without permanent ventilation. For the secondary end points, comparisons were made with the upper boundary of 90% confidence intervals for natural-history data from 40 infants with type 1 SMA. Results A total of 41 infants were enrolled. After 12 months of treatment, 12 infants (29%) were able to sit without support for at least 5 seconds, a milestone not attained in this disorder. The percentages of infants in whom the key secondary end points were met as compared with the upper boundary of confidence intervals from historical controls were 56% as compared with 17% for a CHOP-INTEND score of 40 or higher, 90% as compared with 17% for an increase of at least 4 points from baseline in the CHOP-INTEND score, 78% as compared with 12% for a HINE-2 motor-milestone response, and 85% as compared with 42% for survival without permanent ventilation (P<0.001 for all comparisons). The most common serious adverse events were pneumonia, bronchiolitis, hypotonia, and respiratory failure. Conclusions In this study involving infants with type 1 SMA, risdiplam resulted in higher percentages of infants who met motor milestones and who showed improvements in motor function than the percentages observed in historical cohorts. Longer and larger trials are required to determine the long-term safety and efficacy of risdiplam in infants with type 1 SMA. (Funded by F. Hoffmann-La Roche; FIREFISH ClinicalTrials.gov number, .)Small-Molecule SMN2 Modifier in Type 1 SMA The pre-mRNA SMN2 splicing modifier risdiplam was administered orally to 41 infants with type 1 spinal muscular atrophy. After 12 months of treatment, 12 infants were able to sit without support, and most had better scores on motor-performance scales than the upper limit of confidence intervals from historical controls