Two Years of Genomic Surveillance in Belgium during the SARS-CoV-2 Pandemic to Attain Country-Wide Coverage and Monitor the Introduction and Spread of Emerging Variants.

An adequate SARS-CoV-2 genomic surveillance strategy has proven to be essential for countries to obtain a thorough understanding of the variants and lineages being imported and successfully established within their borders. During 2020, genomic surveillance in Belgium was not structurally implemented but performed by individual research laboratories that had to acquire the necessary funds themselves to perform this important task. At the start of 2021, a nationwide genomic surveillance consortium was established in Belgium to markedly increase the country's genomic sequencing efforts (both in terms of intensity and representativeness), to perform quality control among participating laboratories, and to enable coordination and collaboration of research projects and publications. We here discuss the genomic surveillance efforts in Belgium before and after the establishment of its genomic sequencing consortium, provide an overview of the specifics of the consortium, and explore more details regarding the scientific studies that have been published as a result of the increased number of Belgian SARS-CoV-2 genomes that have become available.

Nationwide Harmonization Effort for Semi-Quantitative Reporting of SARS-CoV-2 PCR Test Results in Belgium.

From early 2020, a high demand for SARS-CoV-2 tests was driven by several testing indications, including asymptomatic cases, resulting in the massive roll-out of PCR assays to combat the pandemic. Considering the dynamic of viral shedding during the course of infection, the demand to report cycle threshold (Ct) values rapidly emerged. As Ct values can be affected by a number of factors, we considered that harmonization of semi-quantitative PCR results across laboratories would avoid potential divergent interpretations, particularly in the absence of clinical or serological information. A proposal to harmonize reporting of test results was drafted by the National Reference Centre (NRC) UZ/KU Leuven, distinguishing four categories of positivity based on RNA copies/mL. Pre-quantified control material was shipped to 124 laboratories with instructions to setup a standard curve to define thresholds per assay. For each assay, the mean Ct value and corresponding standard deviation was calculated per target gene, for the three concentrations (107, 105 and 103 copies/mL) that determine the classification. The results of 17 assays are summarized. This harmonization effort allowed to ensure that all Belgian laboratories would report positive PCR results in the same semi-quantitative manner to clinicians and to the national database which feeds contact tracing interventions.

Accuracy and Reproducibility of Low-Dose Submillisievert Chest CT for the Diagnosis of COVID-19.

PURPOSE: To demonstrate the accuracy and reproducibility of low-dose submillisievert chest CT for the diagnosis of coronavirus disease 2019 (COVID-19) infection in patients in the emergency department. MATERIALS AND METHODS: This was a Health Insurance Portability and Accountability Act-compliant, institutional review board-approved retrospective study. From March 14 to 24, 2020, 192 patients in the emergency department with symptoms suggestive of COVID-19 infection were studied by using low-dose chest CT and real-time reverse transcription polymerase chain reaction (RT-PCR). Image analysis included the likelihood of COVID-19 infection and the semiquantitative extent of lung involvement. CT images were analyzed by two radiologists blinded to the RT-PCR results. Reproducibility was assessed using the McNemar test and intraclass correlation coefficient. Time between CT acquisition and report was measured. RESULTS: When compared with RT-PCR, low-dose submillisievert chest CT demonstrated excellent sensitivity, specificity, positive predictive value, negative predictive value, and accuracy for diagnosis of COVID-19 (86.7%, 93.6%, 91.1%, 90.3%, and 90.2%, respectively), in particular in patients with clinical symptoms for more than 48 hours (95.6%, 93.2%, 91.5%, 96.5%, and 94.4%, respectively). In patients with a positive CT result, the likelihood of disease increased from 43.2% (pretest probability) to 91.1% or 91.4% (posttest probability), while in patients with a negative CT result, the likelihood of disease decreased to 9.6% or 3.7% for all patients or those with clinical symptoms for >48 hours. The prevalence of alternative diagnoses based on chest CT in patients without COVID-19 infection was 17.6%. The mean effective radiation dose was 0.56 mSv ± 0.25 (standard deviation). Median time between CT acquisition and report was 25 minutes (interquartile range: 13-49 minutes). Intra- and interreader reproducibility of CT was excellent (all intraclass correlation coefficients ≥ 0.95) without significant bias in the Bland-Altman analysis. CONCLUSION: Low-dose submillisievert chest CT allows for rapid, accurate, and reproducible assessment of COVID-19 infection in patients in the emergency department, in particular in patients with symptoms lasting longer than 48 hours. Chest CT has the additional advantage of offering alternative diagnoses in a significant subset of patients.© RSNA, 2020.

Bacterial and eukaryotic biodiversity patterns in terrestrial and aquatic habitats in the Sør Rondane Mountains, Dronning Maud Land, East Antarctica.

The bacterial and microeukaryotic biodiversity were studied using pyrosequencing analysis on a 454 GS FLX+ platform of partial SSU rRNA genes in terrestrial and aquatic habitats of the Sør Rondane Mountains, including soils, on mosses, endolithic communities, cryoconite holes and supraglacial and subglacial meltwater lenses. This inventory was complemented with Denaturing Gradient Gel Electrophoresis targeting Chlorophyta and Cyanobacteria. OTUs belonging to the Rotifera, Chlorophyta, Tardigrada, Ciliophora, Cercozoa, Fungi, Bryophyta, Bacillariophyta, Collembola and Nematoda were present with a relative abundance of at least 0.1% in the eukaryotic communities. Cyanobacteria, Proteobacteria, Bacteroidetes, Acidobacteria, FBP and Actinobacteria were the most abundant bacterial phyla. Multivariate analyses of the pyrosequencing data revealed a general lack of differentiation of both eukaryotes and prokaryotes according to habitat type. However, the bacterial community structure in the aquatic habitats was dominated by the filamentous cyanobacteria Leptolyngbya and appeared to be significantly different compared with those in dry soils, on mosses, and in endolithic habitats. A striking feature in all datasets was the detection of a relatively large amount of sequences new to science, which underscores the need for additional biodiversity assessments in Antarctic inland locations.

Bacterial diversity assessment in Antarctic terrestrial and aquatic microbial mats: a comparison between bidirectional pyrosequencing and cultivation.

The application of high-throughput sequencing of the 16S rRNA gene has increased the size of microbial diversity datasets by several orders of magnitude, providing improved access to the rare biosphere compared with cultivation-based approaches and more established cultivation-independent techniques. By contrast, cultivation-based approaches allow the retrieval of both common and uncommon bacteria that can grow in the conditions used and provide access to strains for biotechnological applications. We performed bidirectional pyrosequencing of the bacterial 16S rRNA gene diversity in two terrestrial and seven aquatic Antarctic microbial mat samples previously studied by heterotrophic cultivation. While, not unexpectedly, 77.5% of genera recovered by pyrosequencing were not among the isolates, 25.6% of the genera picked up by cultivation were not detected by pyrosequencing. To allow comparison between both techniques, we focused on the five phyla (Proteobacteria, Actinobacteria, Bacteroidetes, Firmicutes and Deinococcus-Thermus) recovered by heterotrophic cultivation. Four of these phyla were among the most abundantly recovered by pyrosequencing. Strikingly, there was relatively little overlap between cultivation and the forward and reverse pyrosequencing-based datasets at the genus (17.1-22.2%) and OTU (3.5-3.6%) level (defined on a 97% similarity cut-off level). Comparison of the V1-V2 and V3-V2 datasets of the 16S rRNA gene revealed remarkable differences in number of OTUs and genera recovered. The forward dataset missed 33% of the genera from the reverse dataset despite comprising 50% more OTUs, while the reverse dataset did not contain 40% of the genera of the forward dataset. Similar observations were evident when comparing the forward and reverse cultivation datasets. Our results indicate that the region under consideration can have a large impact on perceived diversity, and should be considered when comparing different datasets. Finally, a high number of OTUs could not be classified using the RDP reference database, suggesting the presence of a large amount of novel diversity.