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Here we describe, the longest microbial time-series analyzed to date using high-resolution 16S rRNA tag pyrosequencing of samples taken monthly over 6 years at a temperate marine coastal site off Plymouth, UK. Data treatment effected the estimation of community richness over a 6-year period, whereby 8794 operational taxonomic units (OTUs) were identified using single-linkage preclustering and 21 130 OTUs were identified by denoising the data. The Alphaproteobacteria were the most abundant Class, and the most frequently recorded OTUs were members of the Rickettsiales (SAR 11) and Rhodobacteriales. This near-surface ocean bacterial community showed strong repeatable seasonal patterns, which were defined by winter peaks in diversity across all years. Environmental variables explained far more variation in seasonally predictable bacteria than did data on protists or metazoan biomass. Change in day length alone explains >65% of the variance in community diversity. The results suggested that seasonal changes in environmental variables are more important than trophic interactions. Interestingly, microbial association network analysis showed that correlations in abundance were stronger within bacterial taxa rather than between bacteria and eukaryotes, or between bacteria and environmental variables.

Introduction: Primary immunodeficiencies (PIDs) are a heterogeneous group of disorders characterized by increased susceptibility to infections, immune dysregulation, and/or malignancy. Genetic studies, especially during the last decade, led to a better understanding of the pathogenesis of primary immunodeficiencies and contributed to their classification into distinct monogenic disorders falling under one of the >430 currently known inborn errors of immunity (IEI). The growing availability of molecular genetic testing resulted in the increasing identification of patients with IEI. Here, we evaluated the diagnostic yield and the clinical consequences of targeted next-generation sequencing (tNGS) in a cohort of 294 primary immunodeficiency patients, primarily consisting of cases with sporadic primary antibody deficiency. Method: We have custom designed a tNGS panel to sequence a cohort of PID patients. Agilent's HaloPlex Target Enrichment System for Illumina was used for DNA target enrichment. Results: tNGS identified a definite or predicted pathogenic variant in 15.3% of patients. The highest diagnostic rate was observed among patients with combined immunodeficiency or immune dysregulation, for whom genetic diagnosis may affect therapeutic decision-making. Conclusion: Next-generation sequencing has changed diagnostic assignment and paved the way for targeted therapeutic intervention with agents directed at reverting the disease-causing molecular abnormality or its pathophysiological consequences. Therefore, such targeted therapies and identifying the genetic basis of PID can be essential for patients with manifested immune dysregulation as conventional immunomodulatory regimens may exert an immunosuppressive effect, aggravating their immunodeficiency or may only inadequately control autoimmune or lymphoproliferative manifestations. [ABSTRACT FROM AUTHOR]

Infection with human cytomegalovirus (HCMV) can cause severe complications in immunocompromised individuals and congenitally infected children. Characterizing heterogeneous viral populations and their evolution by high-throughput sequencing of clinical specimens requires the accurate assembly of individual strains or sequence variants and suitable variant calling methods. However, the performance of most methods has not been assessed for populations composed of low divergent viral strains with large genomes, such as HCMV. In an extensive benchmarking study, we evaluated 15 assemblers and 6 variant callers on 10 lab-generated benchmark data sets created with two different library preparation protocols, to identify best practices and challenges for analyzing such data. Most assemblers, especially metaSPAdes and IVA, performed well across a range of metrics in recovering abundant strains. However, only one, Savage, recovered low abundant strains and in a highly fragmented manner. Two variant callers, LoFreq and VarScan2, excelled across all strain abundances. Both shared a large fraction of false positive variant calls, which were strongly enriched in T to G changes in a 'G.G' context. The magnitude of this context-dependent systematic error is linked to the experimental protocol. We provide all benchmarking data, results and the entire benchmarking workflow named QuasiModo, Quasispecies Metric determination on omics, under the GNU General Public License v3.0 (https://github.com/hzi-bifo/Quasimodo), to enable full reproducibility and further benchmarking on these and other data. [ABSTRACT FROM AUTHOR]

Late in 2020, two genetically-distinct clusters of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) with mutations of biological concern were reported, one in the United Kingdom and one in South Africa. Using a combination of data from routine surveillance, genomic sequencing and international travel we track the international dispersal of lineages B.1.1.7 and B.1.351 (variant 501Y-V2). We account for potential biases in genomic surveillance efforts by including passenger volumes from location of where the lineage was first reported, London and South Africa respectively. Using the software tool grinch (global report investigating novel coronavirus haplotypes), we track the international spread of lineages of concern with automated daily reports, Further, we have built a custom tracking website (cov-lineages.org/global_report.html) which hosts this daily report and will continue to include novel SARS-CoV-2 lineages of concern as they are detected., Competing Interests: No competing interests were disclosed., (Copyright: © 2021 O'Toole Á et al.)

Reactivation and shedding of human cytomegalovirus (HCMV) in breast milk during lactation is highly frequent in HCMV-seropositive mothers. This represents a key transmission route for postnatal HCMV infection and can lead to severe disease in preterm neonates. Little is known about HCMV strain composition or longitudinal intrahost viral population dynamics in breast milk from immunocompetent women. We performed HCMV-specific target enrichment and high-throughput sequencing of 38 breast milk samples obtained in Germany between days 10 and 60 postpartum from 15 mothers with HCMV DNA lactia, and assembled HCMV consensus sequences de novo . The genotype distribution and number of HCMV strains present in each sample were determined by quantifying genotype-specific sequence motifs in 12 hypervariable viral genes, revealing a wide range of genotypes (82/109) for these genes in the cohort and a unique, longitudinally stable strain composition in each mother. Reactivation of up to three distinct HCMV strains was detected in 8/15 of mothers, indicating that a representative subset of the woman's HCMV reservoir might be locally reactivated early during lactation. As described previously, nucleotide diversity of samples with multiple strains was much higher than that of samples with single strains. Breast milk as a main source of postnatal mother-to-infant transmission may serve as a repository for viral diversity and thus play an essential role in the natural epidemiology of HCMV., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Götting, Lazar, Suárez, Steinbrück, Rabe, Goelz, Schulz, Davison, Hamprecht and Ganzenmueller.)