Antigenic Characterization of Circulating and Emerging SARS-CoV-2 Variants in the U.S. throughout the Delta to Omicron Waves.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has evolved into numerous lineages with unique spike mutations and caused multiple epidemics domestically and globally. Although COVID-19 vaccines are available, new variants with the capacity for immune evasion continue to emerge. To understand and characterize the evolution of circulating SARS-CoV-2 variants in the U.S., the Centers for Disease Control and Prevention (CDC) initiated the National SARS-CoV-2 Strain Surveillance (NS3) program and has received thousands of SARS-CoV-2 clinical specimens from across the nation as part of a genotype to phenotype characterization process. Focus reduction neutralization with various antisera was used to antigenically characterize 143 SARS-CoV-2 Delta, Mu and Omicron subvariants from selected clinical specimens received between May 2021 and February 2023, representing a total of 59 unique spike protein sequences. BA.4/5 subvariants BU.1, BQ.1.1, CR.1.1, CQ.2 and BA.4/5 + D420N + K444T; BA.2.75 subvariants BM.4.1.1, BA.2.75.2, CV.1; and recombinant Omicron variants XBF, XBB.1, XBB.1.5 showed the greatest escape from neutralizing antibodies when analyzed against post third-dose original monovalent vaccinee sera. Post fourth-dose bivalent vaccinee sera provided better protection against those subvariants, but substantial reductions in neutralization titers were still observed, especially among BA.4/5 subvariants with both an N-terminal domain (NTD) deletion and receptor binding domain (RBD) substitutions K444M + N460K and recombinant Omicron variants. This analysis demonstrated a framework for long-term systematic genotype to antigenic characterization of circulating and emerging SARS-CoV-2 variants in the U.S., which is critical to assessing their potential impact on the effectiveness of current vaccines and antigen recommendations for future updates.

Genomic Surveillance for SARS-CoV-2 Variants: Circulation of Omicron Lineages - United States, January 2022-May 2023.

CDC has used national genomic surveillance since December 2020 to monitor SARS-CoV-2 variants that have emerged throughout the COVID-19 pandemic, including the Omicron variant. This report summarizes U.S. trends in variant proportions from national genomic surveillance during January 2022-May 2023. During this period, the Omicron variant remained predominant, with various descendant lineages reaching national predominance (>50% prevalence). During the first half of 2022, BA.1.1 reached predominance by the week ending January 8, 2022, followed by BA.2 (March 26), BA.2.12.1 (May 14), and BA.5 (July 2); the predominance of each variant coincided with surges in COVID-19 cases. The latter half of 2022 was characterized by the circulation of sublineages of BA.2, BA.4, and BA.5 (e.g., BQ.1 and BQ.1.1), some of which independently acquired similar spike protein substitutions associated with immune evasion. By the end of January 2023, XBB.1.5 became predominant. As of May 13, 2023, the most common circulating lineages were XBB.1.5 (61.5%), XBB.1.9.1 (10.0%), and XBB.1.16 (9.4%); XBB.1.16 and XBB.1.16.1 (2.4%), containing the K478R substitution, and XBB.2.3 (3.2%), containing the P521S substitution, had the fastest doubling times at that point. Analytic methods for estimating variant proportions have been updated as the availability of sequencing specimens has declined. The continued evolution of Omicron lineages highlights the importance of genomic surveillance to monitor emerging variants and help guide vaccine development and use of therapeutics.

Urine eosinophil-derived neurotoxin: A potential marker of activity in select eosinophilic disorders.

BACKGROUND: Biomarkers of eosinophilic disease activity, especially in the context of novel therapies that reduce blood eosinophil counts, are an unmet need. Absolute eosinophil count (AEC) does not accurately reflect tissue eosinophilia or eosinophil activation. Therefore, the aims of this study were to compare the reliability of plasma and urine eosinophil major basic protein 1, eosinophil cationic protein, eosinophil-derived neurotoxin (EDN), and eosinophil peroxidase measurement and to evaluate the usefulness of eosinophil granule protein (EGP) measurement for the assessment of disease activity in patients with eosinophil-associated diseases treated with mepolizumab, benralizumab, or dexpramipexole. METHODS: Eosinophil granule protein concentrations were measured in serum, plasma, and urine from healthy volunteers and patients with hypereosinophilic syndrome (HES), eosinophilic granulomatosis with polyangiitis (EGPA), and eosinophilic asthma using a multiplex assay. RESULTS: Urine EGP concentrations remained stable, whereas serum and plasma EGP concentrations increased significantly with delayed processing. Plasma (p) EDN, but not urine (u) EDN, concentration correlated with AEC and negatively correlated with prednisone dose. Both pEDN and uEDN decreased significantly following treatment of HES patients with benralizumab and EGPA patients with mepolizumab. uEDN appeared to increase with clinical relapse in both patient groups. CONCLUSIONS: Measurement of EGP in urine is noninvasive and unaffected by cellular lysis. Although plasma and urine EDN concentrations showed a similar pattern following benralizumab and mepolizumab treatment, the lack of correlation between AEC or prednisone dose and uEDN concentrations suggests that measurement of uEDN may provide a potential biomarker of disease activity in patients with HES and EGPA.

Genomic Surveillance for SARS-CoV-2 Variants: Predominance of the Delta (B.1.617.2) and Omicron (B.1.1.529) Variants - United States, June 2021-January 2022.

Genomic surveillance is a critical tool for tracking emerging variants of SARS-CoV-2 (the virus that causes COVID-19), which can exhibit characteristics that potentially affect public health and clinical interventions, including increased transmissibility, illness severity, and capacity for immune escape. During June 2021-January 2022, CDC expanded genomic surveillance data sources to incorporate sequence data from public repositories to produce weighted estimates of variant proportions at the jurisdiction level and refined analytic methods to enhance the timeliness and accuracy of national and regional variant proportion estimates. These changes also allowed for more comprehensive variant proportion estimation at the jurisdictional level (i.e., U.S. state, district, territory, and freely associated state). The data in this report are a summary of findings of recent proportions of circulating variants that are updated weekly on CDC's COVID Data Tracker website to enable timely public health action.† The SARS-CoV-2 Delta (B.1.617.2 and AY sublineages) variant rose from 1% to >50% of viral lineages circulating nationally during 8 weeks, from May 1-June 26, 2021. Delta-associated infections remained predominant until being rapidly overtaken by infections associated with the Omicron (B.1.1.529 and BA sublineages) variant in December 2021, when Omicron increased from 1% to >50% of circulating viral lineages during a 2-week period. As of the week ending January 22, 2022, Omicron was estimated to account for 99.2% (95% CI = 99.0%-99.5%) of SARS-CoV-2 infections nationwide, and Delta for 0.7% (95% CI = 0.5%-1.0%). The dynamic landscape of SARS-CoV-2 variants in 2021, including Delta- and Omicron-driven resurgences of SARS-CoV-2 transmission across the United States, underscores the importance of robust genomic surveillance efforts to inform public health planning and practice.

Resilience effects on student performance and well-being: the role of self-efficacy, self-set goals, and anxiety.

Universities prepare students to become contributing members to the workplace and to society. However, with rising tuition costs and other increasing time and resource demands, students face substantial adversity. Students' ability to cope with that adversity influences successful completion of academic coursework and retention in degree programs, ultimately providing a source of potential effective future employees. Previous research has demonstrated numerous direct relationships between dispositional resilience and pivotal outcomes, such as performance, life satisfaction, and subjective well-being. However, research has failed to explore underlying mechanisms through which resilience may affect these outcomes, especially in academic contexts. The purpose of the current study was to use self-regulation theory as a framework for examining the effects of students' resilience on outcomes. Using a sample of undergraduate students from a Midwestern university in the U.S. (N = 141), we proposed and tested a path model addressing self-efficacy, self-set goals, and state anxiety as mechanisms through which resilience influences performance and subjective well-being. Our results provided evidence supporting a structural model involving resilience, such that student resilience (a) has an indirect effect on performance through self-efficacy and self-set goals, (b) has an indirect effect on state anxiety through self-efficacy, and (c) accounts for unique variance in subjective well-being after controlling for state anxiety. Implications, limitations, and directions for future research are discussed.

Circular Permutation Obscures Universality of a Ribosomal Protein.

Functions, origins, and evolution of the translation system are best understood in the context of unambiguous and phylogenetically based taxonomy and nomenclature. Here, we map ribosomal proteins onto the tree of life and provide a nomenclature for ribosomal proteins that is consistent with phylogenetic relationships. We have increased the accuracy of homology relationships among ribosomal proteins, providing a more informative picture of their lineages. We demonstrate that bL33 (bacteria) and eL42 (archaea/eukarya) are homologs with common ancestry and acute similarities in sequence and structure. Their similarities were previously obscured by circular permutation. The most likely mechanism of permutation between bL33 and eL42 is duplication followed by fusion and deletion of both the first and last ß-hairpins. bL33 and eL42 are composed of zinc ribbon protein folds, one of the most common zinc finger fold-groups of, and most frequently observed in translation-related domains. Bacterial-specific ribosomal protein bL33 and archaeal/eukaryotic-specific ribosomal protein eL42 are now both assigned the name of uL33, indicating a universal ribosomal protein. We provide a phylogenetic naming scheme for all ribosomal proteins that is based on phylogenetic relationships to be used as a tool for studying the systemics, evolution, and origins of the ribosome.

Translation: The Universal Structural Core of Life.

The Universal Gene Set of Life (UGSL) is common to genomes of all extant organisms. The UGSL is small, consisting of <100 genes, and is dominated by genes encoding the translation system. Here we extend the search for biological universality to three dimensions. We characterize and quantitate the universality of structure of macromolecules that are common to all of life. We determine that around 90% of prokaryotic ribosomal RNA (rRNA) forms a common core, which is the structural and functional foundation of rRNAs of all cytoplasmic ribosomes. We have established a database, which we call the Sparse and Efficient Representation of the Extant Biology (the SEREB database). This database contains complete and cross-validated rRNA sequences of species chosen, as far as possible, to sparsely and efficiently sample all known phyla. Atomic-resolution structures of ribosomes provide data for structural comparison and validation of sequence-based models. We developed a similarity statistic called pairing adjusted sequence entropy, which characterizes paired nucleotides by their adherence to covariation and unpaired nucleotides by conventional conservation of identity. For canonically paired nucleotides the unit of structure is the nucleotide pair. For unpaired nucleotides, the unit of structure is the nucleotide. By quantitatively defining the common core of rRNA, we systematize the conservation and divergence of the translational system across the tree of life, and can begin to understand the unique evolutionary pressures that cause its universality. We explore the relationship between ribosomal size and diversity, geological time, and organismal complexity.

Eukaryotic Ribosomal Expansion Segments as Antimicrobial Targets.

Diversity in eukaryotic rRNA structure and function offers possibilities of therapeutic targets. Unlike ribosomes of prokaryotes, eukaryotic ribosomes contain species-specific rRNA expansion segments (ESs) with idiosyncratic structures and functions that are essential and specific to some organisms. Here we investigate expansion segment 7 (ES7), one of the largest and most variable expansions of the eukaryotic ribosome. We hypothesize that ES7 of the pathogenic fungi Candida albicans (ES7CA) could be a prototypic drug target. We show that isolated ES7CA folds reversibly to a native-like state. We developed a fluorescence displacement assay using an RNA binding fluorescent probe, F-neo. F-neo binds tightly to ES7CA with a Kd of 2.5 × 10-9 M but binds weakly to ES7 of humans (ES7HS) with a Kd estimated to be greater than 7 µM. The fluorescence displacement assay was used to investigate the affinities of a library of peptidic aminosugar conjugates (PAs) for ES7CA. For conjugates with highest affinities for ES7CA (NeoRH, NeoFH, and NeoYH), the lowest dose needed to induce mortality in C. albicans (minimum inhibitory concentration, MIC) was determined. PAs with the lowest MIC values were tested for cytotoxicity in HEK293T cells. Molecules with high affinity for ES7CA in vitro induce mortality in C. albicans but not in HEK293T cells. The results are consistent with the hypothesis that ESs represent useful targets for chemotherapeutics directed against eukaryotic pathogens.

Frozen in Time: The History of Proteins.

The ribosome is imprinted with a detailed molecular chronology of the origins and early evolution of proteins. Here we show that when arranged by evolutionary phase of ribosomal evolution, ribosomal protein (rProtein) segments reveal an atomic level history of protein folding. The data support a model in which aboriginal oligomers evolved into globular proteins in a hierarchical step-wise process. Complexity of assembly and folding of polypeptide increased incrementally in concert with expansion of rRNA. (i) Short random coil proto-peptides bound to rRNA, and (ii) lengthened over time and coalesced into ß-ß secondary elements. These secondary elements (iii) accreted and collapsed, primarily into ß-domains. Domains (iv) accumulated and gained complex super-secondary structures composed of mixtures of α-helices and ß-strands. Early protein evolution was guided and accelerated by interactions with rRNA. rRNA and proto-peptide provided mutual protection from chemical degradation and disassembly. rRNA stabilized polypeptide assemblies, which evolved in a stepwise process into globular domains, bypassing the immense space of random unproductive sequences. Coded proteins originated as oligomers and polymers created by the ribosome, on the ribosome and for the ribosome. Synthesis of increasingly longer products was iteratively coupled with lengthening and maturation of the ribosomal exit tunnel. Protein catalysis appears to be a late byproduct of selection for sophisticated and finely controlled assembly.

Yeast rRNA Expansion Segments: Folding and Function.

Divergence between prokaryotic and eukaryotic ribosomal RNA (rRNA) and among eukaryotic ribosomal RNAs is focused in expansion segments (ESs). Eukaryotic ribosomes are significantly larger than prokaryotic ribosomes partly because of their ESs. We hypothesize that larger rRNAs of complex organisms could confer increased functionality to the ribosome. Here, we characterize the binding partners of Saccharomyces cerevisiae expansion segment 7 (ES7), which is the largest and most variable ES of the eukaryotic large ribosomal subunit and is located at the surface of the ribosome. In vitro RNA-protein pull-down experiments using ES7 as a bait indicate that ES7 is a binding hub for a variety of non-ribosomal proteins essential to ribosomal function in eukaryotes. ES7-associated proteins observed here cluster into four groups based on biological process, (i) response to abiotic stimulus (e.g., response to external changes in temperature, pH, oxygen level, etc.), (ii) ribosomal large subunit biogenesis, (iii) protein transport and localization, and (iv) transcription elongation. Seven synthetases, Ala-, Arg-, Asp-, Asn-, Leu-, Lys- and TyrRS, appear to associate with ES7. Affinities of AspRS, TyrRS and LysRS for ES7 were confirmed by in vitro binding assays. The results suggest that ES7 in S. cerevisiae could play a role analogous to the multi-synthetase complex present in higher order organisms and could be important for the appropriate function of the ribosome. Thermal denaturation studies and footprinting experiments confirm that isolated ES7 is stable and maintains a near-native secondary and tertiary structure.

History of the ribosome and the origin of translation.

We present a molecular-level model for the origin and evolution of the translation system, using a 3D comparative method. In this model, the ribosome evolved by accretion, recursively adding expansion segments, iteratively growing, subsuming, and freezing the rRNA. Functions of expansion segments in the ancestral ribosome are assigned by correspondence with their functions in the extant ribosome. The model explains the evolution of the large ribosomal subunit, the small ribosomal subunit, tRNA, and mRNA. Prokaryotic ribosomes evolved in six phases, sequentially acquiring capabilities for RNA folding, catalysis, subunit association, correlated evolution, decoding, energy-driven translocation, and surface proteinization. Two additional phases exclusive to eukaryotes led to tentacle-like rRNA expansions. In this model, ribosomal proteinization was a driving force for the broad adoption of proteins in other biological processes. The exit tunnel was clearly a central theme of all phases of ribosomal evolution and was continuously extended and rigidified. In the primitive noncoding ribosome, proto-mRNA and the small ribosomal subunit acted as cofactors, positioning the activated ends of tRNAs within the peptidyl transferase center. This association linked the evolution of the large and small ribosomal subunits, proto-mRNA, and tRNA.

Evolution of the ribosome at atomic resolution.

The origins and evolution of the ribosome, 3-4 billion years ago, remain imprinted in the biochemistry of extant life and in the structure of the ribosome. Processes of ribosomal RNA (rRNA) expansion can be "observed" by comparing 3D rRNA structures of bacteria (small), yeast (medium), and metazoans (large). rRNA size correlates well with species complexity. Differences in ribosomes across species reveal that rRNA expansion segments have been added to rRNAs without perturbing the preexisting core. Here we show that rRNA growth occurs by a limited number of processes that include inserting a branch helix onto a preexisting trunk helix and elongation of a helix. rRNA expansions can leave distinctive atomic resolution fingerprints, which we call "insertion fingerprints." Observation of insertion fingerprints in the ribosomal common core allows identification of probable ancestral expansion segments. Conceptually reversing these expansions allows extrapolation backward in time to generate models of primordial ribosomes. The approach presented here provides insight to the structure of pre-last universal common ancestor rRNAs and the subsequent expansions that shaped the peptidyl transferase center and the conserved core. We infer distinct phases of ribosomal evolution through which ribosomal particles evolve, acquiring coding and translocation, and extending and elaborating the exit tunnel.

Differential mismatch recognition specificities of eukaryotic MutS homologs, MutSα and MutSß.

In eukaryotes, the recognition of the DNA postreplication errors and initiation of the mismatch repair is carried out by two MutS homologs: MutSα and MutSß. MutSα recognizes base mismatches and 1 to 2 unpaired nucleotides whereas MutSß recognizes longer insertion-deletion loops (IDLs) with 1 to 15 unpaired nucleotides as well as certain mismatches. Results from molecular dynamics simulations of native MutSß:IDL-containing DNA and MutSα:mismatch DNA complexes as well as complexes with swapped DNA substrates provide mechanistic insight into how the differential substrate specificities are achieved by MutSα and MutSß, respectively. Our simulations results suggest more extensive interactions between MutSß and IDL-DNA and between MutSα and mismatch-containing DNA that suggest corresponding differences in stability. Furthermore, our simulations suggest more expanded mechanistic details involving a different degree of bending when DNA is bound to either MutSα or MutSß and a more likely opening of the clamp domains when noncognate substrates are bound. The simulation results also provide detailed information on key residues in MutSß and MutSα that are likely involved in recognizing IDL-DNA and mismatch-containing DNA, respectively.

Lack of detectable HIV-1 molecular evolution during suppressive antiretroviral therapy.

A better understanding of changes in HIV-1 population genetics with combination antiretroviral therapy (cART) is critical for designing eradication strategies. We therefore analyzed HIV-1 genetic variation and divergence in patients' plasma before cART, during suppression on cART, and after viral rebound. Single-genome sequences of plasma HIV-1 RNA were obtained from HIV-1 infected patients prior to cART (N = 14), during suppression on cART (N = 14) and/or after viral rebound following interruption of cART (N = 5). Intra-patient population diversity was measured by average pairwise difference (APD). Population structure was assessed by phylogenetic analyses and a test for panmixia. Measurements of intra-population diversity revealed no significant loss of overall genetic variation in patients treated for up to 15 years with cART. A test for panmixia, however, showed significant changes in population structure in 2/10 patients after short-term cART (<1 year) and in 7/10 patients after long-term cART (1-15 years). The changes consisted of diverse sets of viral variants prior to cART shifting to populations containing one or more genetically uniform subpopulations during cART. Despite these significant changes in population structure, rebound virus after long-term cART had little divergence from pretherapy virus, implicating long-lived cells infected before cART as the source for rebound virus. The appearance of genetically uniform virus populations and the lack of divergence after prolonged cART and cART interruption provide strong evidence that HIV-1 persists in long-lived cells infected before cART was initiated, that some of these infected cells may be capable of proliferation, and that on-going cycles of viral replication are not evident.