Identification and genetic analysis of a pervasive 'needle-eye' sperm phenotype in Drosophila sterile hybrid males.

Interspecies hybrid sterility has been extensively studied, especially in the genus Drosophila. Hybrid sterility is more often found in the heterogametic (XY or ZW) sex, a trend called Haldane's rule. Although this phenomenon is pervasive, identification of a common genetic mechanism remains elusive, with modest support found for a range of potential theories. Here, we identify a single precise morphological phenotype, which we call 'needle-eye sperm', that is associated with hybrid sterility in three separate species pairs that span the Drosophila genus. The nature of the phenotype indicates a common point of meiotic failure in sterile hybrid males. We used 10 generations of backcross selection paired with whole-genome pooled sequencing to genetically map the regions underlying the needle-eye (NE) sperm phenotype. Surprisingly, the sterility phenotype was present in ~50% of males even after 10 generations of backcrossing, and only a single region of the X chromosome was associated with sterility in one direction of backcross. Owing to the common phenotype among sterile male hybrids, and the strong effect of individual loci, further exploration of these findings may identify a universal mechanism for the evolution of hybrid sterility.

Genetic and transcriptomic landscape of colonic diverticulosis.

OBJECTIVE: Colonic diverticulosis is a prevalent condition among older adults, marked by the presence of thin-walled pockets in the colon wall that can become inflamed, infected, haemorrhage or rupture. We present a case-control genetic and transcriptomic study aimed at identifying the genetic and cellular determinants underlying this condition and the relationship with other gastrointestinal disorders. DESIGN: We conducted DNA and RNA sequencing on colonic tissue from 404 patients with (N=172) and without (N=232) diverticulosis. We investigated variation in the transcriptome associated with diverticulosis and further integrated this variation with single-cell RNA-seq data from the human intestine. We also integrated our expression quantitative trait loci with genome-wide association study using Mendelian randomisation (MR). Furthermore, a Polygenic Risk Score analysis gauged associations between diverticulosis severity and other gastrointestinal disorders. RESULTS: We discerned 38 genes with differential expression and 17 with varied transcript usage linked to diverticulosis, indicating tissue remodelling as a primary diverticula formation mechanism. Diverticula formation was primarily linked to stromal and epithelial cells in the colon including endothelial cells, myofibroblasts, fibroblasts, goblet, tuft, enterocytes, neurons and glia. MR highlighted five genes including CCN3, CRISPLD2, ENTPD7, PHGR1 and TNFSF13, with potential causal effects on diverticulosis. Notably, ENTPD7 upregulation was confirmed in diverticulosis cases. Additionally, diverticulosis severity was positively correlated with genetic predisposition to diverticulitis. CONCLUSION: Our results suggest that tissue remodelling is a primary mechanism for diverticula formation. Individuals with an increased genetic proclivity to diverticulitis exhibit a larger numbers of diverticula on colonoscopy.

How well do existing surveys track fish community performance measures in the St. Clair-Detroit River System?

The St. Clair-Detroit River System (SCDRS) connects Lake Huron to Lake Erie and provides important habitats for many fishes of economic and ecological importance. Portions of the SCDRS are designated as Great Lakes Areas of Concern and fish production and conservation may be compromised. Efforts to address beneficial use impairments have focused on restoring habitat for native fishes and improving aquatic ecosystem health. Considerable site-specific research and long-term, annual fish surveys have examined responses to habitat improvements. However, there is uncertainty surrounding whether individual studies and surveys can assess (1) population-level benefits of habitat enhancements and (2) whether management objectives are being met. To identify monitoring gaps and inform long-term monitoring program development, we compared outputs from SCDRS fish monitoring surveys (based on discussions with regional agencies) with performance measures specified in management plans (obtained through gray literature searches). Performance measures for harvested species aligned well with outputs of existing surveys. In contrast, at-risk fishes often had objectives and performance measures that reflected knowledge gaps and study needs. Although harvested species were well-monitored relative to specified performance measures, at-risk fishes were less reliably collected by existing surveys, except for lake sturgeon Acipenser fulvescens. Effective evaluation of restoration efforts for at-risk fishes may require additional survey efforts that target species-specific habitat use and life history characteristics.

Oral SARS-CoV-2 host responses predict the early COVID-19 disease course.

Objectives: Oral fluids provide ready detection of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and host responses. This study sought to determine relationships between oral virus, oral anti-SARS-CoV-2-specific antibodies, and symptoms. Methods: Saliva/throat wash (saliva/TW) were collected from asymptomatic and symptomatic, nasopharyngeal (NP) SARS-CoV-2 RT-qPCR+, subjects (n=47). SARS-CoV-2 RT-qPCR, N-antigen detection by immunoblot and lateral flow assay (LFA) were performed. RT-qPCR targeting viral subgenomic RNA (sgRNA) was sequence confirmed. SARS-CoV-2-anti-S protein RBD LFA assessed IgM and IgG responses. Structural analysis identified host salivary molecules analogous to SARS-CoV-2-N-antigen. Statistical analyses were performed. Results: At baseline, LFA-detected N-antigen was immunoblot-confirmed in 82% of TW. However, only 3/17 were saliva/TW qPCR+. Sixty percent of saliva and 83% of TW demonstrated persistent N-antigen at 4 weeks. N-antigen LFA signal in three negative subjects suggested potential cross-detection of 4 structurally analogous salivary RNA binding proteins (alignment 19-29aa, RMSD 1-1.5 Angstroms). At entry, symptomatic subjects demonstrated replication-associated sgRNA junctions, were IgG+ (94%/100% in saliva/TW), and IgM+ (75%/63%). At 4 weeks, SARS-CoV-2 IgG (100%/83%) and IgM (80%/67%) persisted. Oral IgG correlated 100% with NP+PCR status. Cough and fatigue severity (p=0.0008 and 0.016), and presence of nausea, weakness, and composite upper respiratory symptoms (p=0.005, 0.037 and 0.017) were negatively associated with oral IgM. Female oral IgM levels were higher than male (p=0.056). Conclusion: Important to transmission and disease course, oral viral replication and persistence showed clear relationships with select symptoms, early Ig responses, and gender during early infection. N-antigen cross-reactivity may reflect mimicry of structurally analogous host proteins.

Oral SARS-CoV-2 host responses predict the early COVID-19 disease course.

Objectives: Oral fluids provide ready detection of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and host responses. This study sought to determine relationships between oral virus, oral anti-SARS-CoV-2-specific antibodies, and symptoms. Methods: Saliva/throat wash (saliva/TW) were collected from asymptomatic and symptomatic, nasopharyngeal (NP) SARS-CoV-2 RT-qPCR+, subjects (n=47). SARS-CoV-2 RT-qPCR, N-antigen detection by immunoblot and lateral flow assay (LFA) were performed. RT-qPCR targeting viral subgenomic RNA (sgRNA) was sequence confirmed. SARS-CoV-2-anti-S protein RBD LFA assessed IgM and IgG responses. Structural analysis identified host salivary molecules analogous to SARS-CoV-2-N-antigen. Statistical analyses were performed. Results: At baseline, LFA-detected N-antigen was immunoblot-confirmed in 82% of TW. However, only 3/17 were saliva/TW qPCR+. Sixty percent of saliva and 83% of TW demonstrated persistent N-antigen at 4 weeks. N-antigen LFA signal in three negative subjects suggested potential cross-detection of 4 structurally analogous salivary RNA binding proteins (alignment 19-29aa, RMSD 1-1.5 Angstroms). At entry, symptomatic subjects demonstrated replication-associated sgRNA junctions, were IgG+ (94%/100% in saliva/TW), and IgM+ (75%/63%). At 4 weeks, SARS-CoV-2 IgG (100%/83%) and IgM (80%/67%) persisted. Oral IgG correlated 100% with NP+PCR status. Cough and fatigue severity (p=0.0008 and 0.016), and presence of nausea, weakness, and composite upper respiratory symptoms (p=0.005, 0.037 and 0.017) were negatively associated with oral IgM. Female oral IgM levels were higher than male (p=0.056). Conclusion: Important to transmission and disease course, oral viral replication and persistence showed clear relationships with select symptoms, early Ig responses, and gender during early infection. N-antigen cross-reactivity may reflect mimicry of structurally analogous host proteins.

Empirical Evidence That Complexity Limits Horizontal Gene Transfer.

Horizontal gene transfer (HGT) is a major contributor to bacterial genome evolution, generating phenotypic diversity, driving the expansion of protein families, and facilitating the evolution of new phenotypes, new metabolic pathways, and new species. Comparative studies of gene gain in bacteria suggest that the frequency with which individual genes successfully undergo HGT varies considerably and may be associated with the number of protein-protein interactions in which the gene participates, that is, its connectivity. Two nonexclusive hypotheses have emerged to explain why transferability should decrease with connectivity: the complexity hypothesis (Jain R, Rivera MC, Lake JA. 1999. Horizontal gene transfer among genomes: the complexity hypothesis. Proc Natl Acad Sci U S A. 96:3801-3806.) and the balance hypothesis (Papp B, Pál C, Hurst LD. 2003. Dosage sensitivity and the evolution of gene families in yeast. Nature 424:194-197.). These hypotheses predict that the functional costs of HGT arise from a failure of divergent homologs to make normal protein-protein interactions or from gene misexpression, respectively. Here we describe genome-wide assessments of these hypotheses in which we used 74 existing prokaryotic whole genome shotgun libraries to estimate rates of horizontal transfer of genes from taxonomically diverse prokaryotic donors into Escherichia coli. We show that 1) transferability declines as connectivity increases, 2) transferability declines as the divergence between donor and recipient orthologs increases, and that 3) the magnitude of this negative effect of divergence on transferability increases with connectivity. These effects are particularly robust among the translational proteins, which span the widest range of connectivities. Whereas the complexity hypothesis explains all three of these observations, the balance hypothesis explains only the first one.

Influence of Grafting on Rootstock Rhizosphere Microbiome Assembly in Rosa sp. 'Natal Brier'.

The root microbiome is vital in plant development and health and is highly influenced by crop cultural practices. Rose (Rosa sp.) is the most popular cut flower worldwide. Grafting in rose production is a standard practice to increase yield, improve flower quality, or reduce root-associated pests and diseases. 'Natal Brier' is a standard rootstock used in most commercial operations in Ecuador and Colombia, leading countries in producing and exporting ornamentals. It is known that the rose scion genotype affects root biomass and the root exudate profile of grafted plants. However, little is known about the influence of the rose scion genotype on the rhizosphere microbiome. We examined the influence of grafting and scion genotype on the rhizosphere microbiome of the rootstock 'Natal Brier'. The microbiomes of the non-grafted rootstock and the rootstock grafted with two red rose cultivars were assessed using 16S rRNA and ITS sequencing. Grafting changed microbial community structure and function. Further, analysis of grafted plant samples revealed that the scion genotype highly influences the rootstock microbiome. Under the presented experimental conditions, the rootstock 'Natal Brier' core microbiome consisted of 16 bacterial and 40 fungal taxa. Our results highlight that the scion genotype influences root microbe's recruitment, which might also influence the functionality of assembled microbiomes.

Plastid Genome Assembly Using Long-read data.

Although plastid genome (plastome) structure is highly conserved across most seed plants, investigations during the past two decades have revealed several disparately related lineages that experienced substantial rearrangements. Most plastomes contain a large inverted repeat and two single-copy regions, and a few dispersed repeats; however, the plastomes of some taxa harbour long repeat sequences (>300 bp). These long repeats make it challenging to assemble complete plastomes using short-read data, leading to misassemblies and consensus sequences with spurious rearrangements. Single-molecule, long-read sequencing has the potential to overcome these challenges, yet there is no consensus on the most effective method for accurately assembling plastomes using long-read data. We generated a pipeline, plastid Genome Assembly Using Long-read data (ptGAUL), to address the problem of plastome assembly using long-read data from Oxford Nanopore Technologies (ONT) or Pacific Biosciences platforms. We demonstrated the efficacy of the ptGAUL pipeline using 16 published long-read data sets. We showed that ptGAUL quickly produces accurate and unbiased assemblies using only ~50× coverage of plastome data. Additionally, we deployed ptGAUL to assemble four new Juncus (Juncaceae) plastomes using ONT long reads. Our results revealed many long repeats and rearrangements in Juncus plastomes compared with basal lineages of Poales. The ptGAUL pipeline is available on GitHub: https://github.com/Bean061/ptgaul.

Social experience and pheromone receptor activity reprogram gene expression in sensory neurons.

Social experience and pheromone signaling in olfactory neurons affect neuronal responses and male courtship behaviors in Drosophila. We previously showed that social experience and pheromone signaling modulate chromatin around behavioral switch gene fruitless, which encodes a transcription factor necessary and sufficient for male sexual behaviors. Fruitless drives social experience-dependent modulation of courtship behaviors and physiological sensory neuron responses to pheromone; however, the molecular mechanisms underlying this modulation of neural responses remain less clear. To identify the molecular mechanisms driving social experience-dependent changes in neuronal responses, we performed RNA-seq from antennal samples of mutants in pheromone receptors and fruitless, as well as grouped or isolated wild-type males. Genes affecting neuronal physiology and function, such as neurotransmitter receptors, ion channels, ion and membrane transporters, and odorant binding proteins are differentially regulated by social context and pheromone signaling. While we found that loss of pheromone detection only has small effects on differential promoter and exon usage within fruitless gene, many of the differentially regulated genes have Fruitless-binding sites or are bound by Fruitless in the nervous system. Recent studies showed that social experience and juvenile hormone signaling co-regulate fruitless chromatin to modify pheromone responses in olfactory neurons. Interestingly, genes involved in juvenile hormone metabolism are also misregulated in different social contexts and mutant backgrounds. Our results suggest that modulation of neuronal activity and behaviors in response to social experience and pheromone signaling likely arise due to large-scale changes in transcriptional programs for neuronal function downstream of behavioral switch gene function.

Untangling the Effects of Plant Genotype and Soil Conditions on the Assembly of Bacterial and Fungal Communities in the Rhizosphere of the Wild Andean Blueberry (Vaccinium floribundum Kunth).

Microbial communities in the rhizosphere influence nutrient acquisition and stress tolerance. How abiotic and biotic factors impact the plant microbiome in the wild has not been thoroughly addressed. We studied how plant genotype and soil affect the rhizosphere microbiome of Vaccinium floribundum, an endemic species of the Andean region that has not been domesticated or cultivated. Using high-throughput sequencing of the 16S rRNA and ITS region, we characterized 39 rhizosphere samples of V. floribundum from four plant genetic clusters in two soil regions from the Ecuadorian Highlands. Our results showed that Proteobacteria and Acidobacteria were the most abundant bacterial phyla and that fungal communities were not dominated by any specific taxa. Soil region was the main predictor for bacterial alpha diversity, phosphorous and lead being the most interesting edaphic factors explaining this diversity. The interaction of plant genotype and altitude was the most significant factor associated with fungal diversity. This study highlights how different factors govern the assembly of the rhizosphere microbiome of a wild plant. Bacterial communities depend more on the soil and its mineral content, while plant genetics influence the fungal community makeup. Our work illustrates plant-microbe associations and the drivers of their variation in a unique unexplored ecosystem from the Ecuadorian Andes.

Diverse MarR bacterial regulators of auxin catabolism in the plant microbiome.

Chemical signalling in the plant microbiome can have drastic effects on microbial community structure, and on host growth and development. Previously, we demonstrated that the auxin metabolic signal interference performed by the bacterial genus Variovorax via an auxin degradation locus was essential for maintaining stereotypic root development in an ecologically relevant bacterial synthetic community. Here, we dissect the Variovorax auxin degradation locus to define the genes iadDE as necessary and sufficient for indole-3-acetic acid (IAA) degradation and signal interference. We determine the crystal structures and binding properties of the operon's MarR-family repressor with IAA and other auxins. Auxin degradation operons were identified across the bacterial tree of life and we define two distinct types on the basis of gene content and metabolic products: iac-like and iad-like. The structures of MarRs from representatives of each auxin degradation operon type establish that each has distinct IAA-binding pockets. Comparison of representative IAA-degrading strains from diverse bacterial genera colonizing Arabidopsis plants show that while all degrade IAA, only strains containing iad-like auxin-degrading operons interfere with auxin signalling in a complex synthetic community context. This suggests that iad-like operon-containing bacterial strains, including Variovorax species, play a key ecological role in modulating auxins in the plant microbiome.

Pharmacodynamic determinants of hangover: An intravenous alcohol self-administration study in non-dependent drinkers.

Alcohol hangover refers to the combination of negative mental and physical symptoms that can be experienced after an episode of alcohol consumption, typically emerging as blood alcohol concentration (BAC) approaches zero. Hangover has been associated with heavy drinking and may be relevant in the transition to alcohol use disorder (AUD). Our aim was to examine hangover prevalence and associated symptoms following intravenous alcohol self-administration (IV-ASA), and to identify possible predictors of hangover in non-dependent drinkers. Ninety-five drinkers without AUD completed an IV-ASA session. Pharmacodynamic measures of alcohol consumption included peak and average breath alcohol concentrations. Subjective measures of alcohol response included the Drug Effects Questionnaire and Biphasic Effects of Alcohol Scale. The Alcohol Hangover Scale assessed hangover symptoms from the end of the session until the following morning. 78% of participants endorsed at least one hangover symptom following IV-ASA. There was no association between hangover scores and IV-ASA measures of alcohol consumption. Additional mediation and moderation analysis revealed that self-reported intoxication was a significant mediator of the relationship between recent drinking and hangover symptoms. Hangover symptoms may be an early marker of the relationship between subjective response to alcohol and heavy drinking for those with no prior history of AUD. In particular, the effects of hangover go beyond exposure to alcohol and the individual's subjective response to this exposure is associated with their experience of hangover. Future studies should further characterize the determinants of hangover across different populations of drinkers to better understand the risk for AUD and inform prevention methods.

Prediction model to estimate overall survival benefit of postoperative radiotherapy for resected major salivary gland cancers.

OBJECTIVES: To develop and validate a prediction model to estimate overall survival (OS) with and without postoperative radiotherapy (PORT) for resected major salivary gland (SG) cancers. MATERIALS AND METHODS: Adults in the National Cancer Database diagnosed with invasive non-metastatic major SG cancer between 2004 and 2015 were identified. Exclusion criteria included prior malignancy, pT1N0 or unknown stage, no or unknown surgery, and neoadjuvant therapy. Cox proportional hazards models evaluated the effect of covariates on OS. A multivariate regression model was utilized to predict 2-, 5-, and 10-year OS. Internal cross-validation was performed using 50-50 hold-out and Harrell's concordance index. RESULTS: 18,400 subjects met inclusion criteria, including 9,721 (53%) who received PORT. Distribution of SG involvement was 86% parotid, 13% submandibular, and 1% sublingual. Median follow-up for living subjects was 4.9 years. PORT was significantly associated with improved OS for the following subgroups by log-rank test: pT3 (p < 0.001), pT4 (p < 0.001), high grade (p < 0.001), node-positive (p < 0.001), and positive margin (p < 0.001). The following variables were incorporated into a multivariate model: age, sex, Charlson-Deyo comorbidity score, involved SG, pathologic T-stage, grade, margin status, ratio of nodal positivity, and PORT. The resulting model based on data from 6,138 subjects demonstrated good accuracy in predicting OS, with Harrell's concordance index of 0.73 (log-rank p < 0.001). CONCLUSION: This cross-validated prediction model estimates 2-, 5-, and 10-year differences in OS based on receipt of PORT for resected major SG cancers using readily available clinicopathologic features. Clinicians can utilize this tool to aid personalized adjuvant therapy decisions.

Comparing Outcomes of Oligometastases Treated with Hypofractionated Image-Guided Radiotherapy (HIGRT) with a Simultaneous Integrated Boost (SIB) Technique versus Metastasis Alone: A Multi-Institutional Analysis.

PURPOSE: We previously reported on the clinical outcomes of treating oligometastases with radiation using an elective simultaneous integrated boost technique (SIB), delivering higher doses to known metastases and reduced doses to adjacent bone or nodal basins. Here we compare outcomes of oligometastases receiving radiation targeting metastases alone (MA) versus those treated via an SIB. METHODS: Oligometastatic patients with ≤5 active metastases treated with either SIB or MA radiation at two institutions from 2013 to 2019 were analyzed retrospectively for treatment-related toxicity, pain control, and recurrence patterns. Tumor metastasis control (TMC) was defined as an absence of progression in the high dose planning target volume (PTV). Marginal recurrence (MR) was defined as recurrence outside the elective PTV but within the adjacent bone or nodal basin. Distant recurrence (DR) was defined as any recurrence that is not within the PTV or surrounding bone or nodal basin. The outcome rates were estimated using the Kaplan-Meier method and compared between the two techniques using the log-rank test. RESULTS: 101 patients were treated via an SIB to 90 sites (58% nodal and 42% osseous) and via MA radiation to 46 sites (22% nodal and 78% osseous). The median follow-up among surviving patients was 24.6 months (range 1.4-71.0). Of the patients treated to MA, the doses ranged from 18 Gy in one fraction (22%) to 50 Gy in 10 fractions (50%). Most patients treated with an SIB received 50 Gy to the treated metastases and 30 Gy to the elective PTV in 10 fractions (88%). No acute grade ≥3 toxicities occurred in either cohort. Late grade ≥3 toxicity occurred in 3 SIB patients (vocal cord paralysis and two vertebral body compression), all related to the high dose PTV and not the elective volume. There was similar crude pain relief between cohorts. The MR-free survival rate at 2 years was 87% (95% CI: 70%, 95%) in the MA group and 98% (95% CI: 87%, 99%) in the SIB group (p = 0.07). The crude TMC was 89% (41/46) in the MA group and 94% (85/90) in the SIB group. There were no significant differences in DR-free survival (65% (95% CI: 55-74%; p = 0.24)), disease-free survival (60% (95% CI: 40-75%; p = 0.40)), or overall survival (88% (95% CI: 73-95%; p = 0.26)), between the MA and SIB cohorts. CONCLUSION: Both SIB and MA irradiation of oligometastases achieved high rates of TMC and similar pain control, with a trend towards improved MR-free survival for oligometastases treated with an SIB. Further investigation of this technique with prospective trials is warranted.

Genetic changes associated with relapse in favorable histology Wilms tumor: A Children's Oncology Group AREN03B2 study.

Over the last decade, sequencing of primary tumors has clarified the genetic underpinnings of Wilms tumor but has not affected therapy, outcome, or toxicity. We now sharpen our focus on relapse samples from the umbrella AREN03B2 study. We show that over 40% of relapse samples contain mutations in SIX1 or genes of the MYCN network, drivers of progenitor proliferation. Not previously seen in large studies of primary Wilms tumors, DIS3 and TERT are now identified as recurrently mutated. The analysis of primary-relapse tumor pairs suggests that 11p15 loss of heterozygosity (and other copy number changes) and mutations in WT1 and MLLT1 typically occur early, but mutations in SIX1, MYCN, and WTX are late developments in some individuals. Most strikingly, 75% of relapse samples had gain of 1q, providing strong conceptual support for studying circulating tumor DNA in clinical trials to better detect 1q gain earlier and monitor response.

Acute Leukemia Classification Using Transcriptional Profiles From Low-Cost Nanopore mRNA Sequencing.

PURPOSE: Most cases of pediatric acute leukemia occur in low- and middle-income countries, where health centers lack the tools required for accurate diagnosis and disease classification. Recent research shows the robustness of using unbiased short-read RNA sequencing to classify genomic subtypes of acute leukemia. Compared with short-read sequencing, nanopore sequencing has low capital and consumable costs, making it suitable for use in locations with limited health infrastructure. MATERIALS AND METHODS: We show the feasibility of nanopore mRNA sequencing on 134 cryopreserved acute leukemia specimens (26 acute myeloid leukemia [AML], 73 B-lineage acute lymphoblastic leukemia [B-ALL], 34 T-lineage acute lymphoblastic leukemia, and one acute undifferentiated leukemia). Using multiple library preparation approaches, we generated long-read transcripts for each sample. We developed a novel composite classification approach to predict acute leukemia lineage and major B-ALL and AML molecular subtypes directly from gene expression profiles. RESULTS: We demonstrate accurate classification of acute leukemia samples into AML, B-ALL, or T-lineage acute lymphoblastic leukemia (96.2% of cases are classifiable with a probability of > 0.8, with 100% accuracy) and further classification into clinically actionable genomic subtypes using shallow RNA nanopore sequencing, with 96.2% accuracy for major AML subtypes and 94.1% accuracy for major B-lineage acute lymphoblastic leukemia subtypes. CONCLUSION: Transcriptional profiling of acute leukemia samples using nanopore technology for diagnostic classification is feasible and accurate, which has the potential to improve the accuracy of cancer diagnosis in low-resource settings.

A prospective study of asymptomatic SARS-CoV-2 infection among individuals involved in academic research under limited operations during the COVID-19 pandemic.

BACKGROUND: Early in the pandemic, transmission risk from asymptomatic infection was unclear, making it imperative to monitor infection in workplace settings. Further, data on SARS-CoV-2 seroprevalence within university populations has been limited. METHODS: We performed a longitudinal study of University research employees on campus July-December 2020. We conducted questionnaires on COVID-19 risk factors, RT-PCR testing, and SARS-CoV-2 serology using an in-house spike RBD assay, laboratory-based Spike NTD assay, and standard nucleocapsid platform assay. We estimated prevalence and cumulative incidence of seroconversion with 95% confidence intervals using the inverse of the Kaplan-Meier estimator. RESULTS: 910 individuals were included in this analysis. At baseline, 6.2% (95% CI 4.29-8.19) were seropositive using the spike RBD assay; four (0.4%) were seropositive using the nucleocapsid assay, and 44 (4.8%) using the Spike NTD assay. Cumulative incidence was 3.61% (95% CI: 2.04-5.16). Six asymptomatic individuals had positive RT-PCR results. CONCLUSIONS: Prevalence and incidence of SARS-CoV-2 infections were low; however, differences in target antigens of serological tests provided different estimates. Future research on appropriate methods of serological testing in unvaccinated and vaccinated populations is needed. Frequent RT-PCR testing of asymptomatic individuals is required to detect acute infections, and repeated serosurveys are beneficial for monitoring subclinical infection.

Collaboration between Clinical and Academic Laboratories for Sequencing SARS-CoV-2 Genomes.

Genomic sequencing of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to provide valuable insight into the ever-changing variant makeup of the COVID-19 pandemic. More than three million SARS-CoV-2 genome sequences have been deposited in Global Initiative on Sharing All Influenza Data (GISAID), but contributions from the United States, particularly through 2020, lagged the global effort. The primary goal of clinical microbiology laboratories is seldom rooted in epidemiologic or public health testing, and many laboratories do not contain in-house sequencing technology. However, we recognized the need for clinical microbiologists to lend expertise, share specimen resources, and partner with academic laboratories and sequencing cores to assist in SARS-CoV-2 epidemiologic sequencing efforts. Here, we describe two clinical and academic laboratory collaborations for SARS-CoV-2 genomic sequencing. We highlight roles of the clinical microbiologists and the academic laboratories, outline best practices, describe two divergent strategies in accomplishing a similar goal, and discuss the challenges with implementing and maintaining such programs.

Nomogram Predicting Overall Survival Benefit of Stereotactic Ablative Radiotherapy for Early-Stage Non-Small Cell Lung Cancer.

OBJECTIVES: To develop and validate a nomogram that predicts overall survival (OS) for patients with early-stage non-small cell lung cancer (NSCLC) treated with stereotactic ablative radiotherapy (SABR) vs. observation. MATERIALS AND METHODS: Adults with biopsy-proven T1-T2N0 NCSLC treated with SABR (30-70 Gy in 1-10 fractions with biologically effective dose ≥100 Gy10) or observation between 2004 and 2015 in the National Cancer Database (NCDB) were identified. Propensity score was used to match SABR and observation cohorts on prognostic demographic and clinicopathologic factors identified by logistic regression. Using backward selection, a multivariable Cox proportional hazard was identified predicting 2- and 5-year OS via a nomogram. Model prediction accuracy was assessed by time-dependent receiver operating characteristic (ROC) curves and integrated area under the ROC curve (AUC) analysis. RESULTS: A total of 22,073 adults met inclusion criteria and 4418 matched pairs (total n = 8836) were identified for nomogram development. The factors most strongly associated with improved OS on multivariable analysis included younger age (HR 0.82 by decade, P < .001), female sex (HR 0.81, P < .001), lower comorbidity index (HR 0.65 for 0 vs. ≥3, P < .001), smaller tumor size (HR 0.60 for ≤3 cm vs. 5.1-7 cm, P < .001), adenocarcinoma histology (P < .001), and receipt of SABR (P < .001). Interaction between SABR and histology was significantly associated with OS (P = .017). Relative to adenocarcinoma, patients with squamous cell carcinoma who were observed (HR 1.44, 95% CI 1.33-1.56) or treated with SABR (HR 1.24, 95% CI 1.14-1.35) had significantly worse OS. The nomogram demonstrated fair accuracy for predicting OS, with an integrated time-dependent AUC of 0.694 over the entire follow-up period. CONCLUSION: This nomogram estimates OS at 2 and 5 years based on whether medically inoperable early-stage NSCLC patients receive SABR or elect for observation. Incorporation of other variables not captured within the NCDB may improve the model accuracy.