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Winogradsky columns were invented by Sergei Winogradsky in the 1880s and have commonly been used as a microbiology classroom learning tool in K-12 and collegiate education. However, they can be challenging to examine with microscopy. We scaled down Winogradsky columns into nuclear magnetic resonance (NMR) tubes and replaced the natural sediment with a transparent soil substitute toward the goal of observing the microbial growth under a bright-field microscope without column disassembly. Using this "Mini Winnie" approach, students can practice their microscopy skills while observing microbial growth inside the column after only days of incubation on the laboratory windowsill. Overall, we believe that the Mini Winnies provide a simple method for maximizing student engagement while giving them a greater understanding of how microorganisms interact in the environment., Competing Interests: The authors declare no conflict of interest.

Objective: Evaluate whether elevating the tympanic membrane from the malleus during endoscopic tympanoplasty may negatively affect postoperative hearing outcomes or perforation rates by comparing 2 similar endoscopic tympanoplasty techniques., Study Design: Retrospective cohort., Setting: Tertiary care center., Patients: Endoscopic over-under cartilage tympanoplasties age and gender matched to endoscopic underlay cartilage tympanoplasties between January 2015 and January 2019. Exclusion criteria included preoperative or intraoperative cholesteatoma, performance of mastoidectomy or ossicular chain reconstruction, and lack of postoperative audiogram., Interventions: Endoscopic cartilage tympanoplasty via over-under or underlay technique., Main Outcome Measures: Pre- and postoperative pure-tone average and word recognition score, graft success., Results: A total of 52 patients were evaluated: 26 endoscopic over-under cartilage tympanoplasties were matched to endoscopic underlay cartilage tympanoplasties. Both groups demonstrated a statistically significant improvement in air conduction hearing (9 dB [ P < 0.001] and 6 dB [ P < 0.01], respectively), and bone pure-tone average did not worsen in either group ( P < 0.001 and P < 0.05, respectively). Postoperative air conduction pure-tone average was statistically noninferior in the over-under group compared with the underlay group ( P < 0.05). Reperforation was present in 3 patients (12%) in the underlay group and none in the over-under group, but this difference was not statistically significant ( P = 0.24)., Conclusions: Endoscopic over-under cartilage tympanoplasty effectively closes tympanic membrane perforations and improves hearing, without greater risk than underlay tympanoplasty. Elevating the tympanic membrane from the malleus does not confer worsen hearing outcomes., Competing Interests: Book royalties, Elsevier (M.A.A.)., (Copyright © 2021 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of Otology & Neurotology, Inc.)

Background: The coronavirus disease 2019 pandemic spread to >200 countries in <6 months. To understand coronavirus spread, determining transmission rate and defining factors that increase transmission risk are essential. Most cases are asymptomatic, but people with asymptomatic infection have viral loads indistinguishable from those in symptomatic people, and they do transmit severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, asymptomatic cases are often undetected., Methods: Given high residence hall student density, the University of Colorado Boulder established a mandatory weekly screening test program. We analyzed longitudinal data from 6408 students and identified 116 likely transmission events in which a second roommate tested positive within 14 days of the index roommate., Results: Although the infection rate was lower in single-occupancy rooms (10%) than in multiple-occupancy rooms (19%), interroommate transmission occurred only about 20% of the time. Cases were usually asymptomatic at the time of detection. Notably, individuals who likely transmitted had an average viral load approximately 6.5-fold higher than individuals who did not (mean quantification cycle [Cq], 26.2 vs 28.9). Although students with diagnosed SARS-CoV-2 infection moved to isolation rooms, there was no difference in time to isolation between cases with or without interroommate transmission., Conclusions: This analysis argues that interroommate transmission occurs infrequently in residence halls and provides strong correlative evidence that viral load is proportional to transmission probability., (© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

We analyze data from the fall 2020 pandemic response efforts at the University of Colorado Boulder, where more than 72,500 saliva samples were tested for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) using qRT-PCR. All samples were collected from individuals who reported no symptoms associated with COVID-19 on the day of collection. From these, 1,405 positive cases were identified. The distribution of viral loads within these asymptomatic individuals was indistinguishable from what has been previously observed in symptomatic individuals. Regardless of symptomatic status, ∼50% of individuals who test positive for SARS-CoV-2 seem to be in noninfectious phases of the disease, based on having low viral loads in a range from which live virus has rarely been isolated. We find that, at any given time, just 2% of individuals carry 90% of the virions circulating within communities, serving as viral "supercarriers" and possibly also superspreaders., Competing Interests: Competing interest statement: Some of the authors of this study have financial ties to companies that offer commercial SARS-CoV-2 testing (Q.Y., N.R.M., C.L.P., and S.L.S. are cofounders of Darwin Biosciences; T.K.S., P.K.G., and E.L. are cofounders of TUMI Genomics). R.P. is a cofounder of Faze Medicines, and R.D.D. is a cofounder of Arpeggio Biosciences., (Copyright © 2021 the Author(s). Published by PNAS.)

Here, we develop a simple molecular test for SARS-CoV-2 in saliva based on reverse transcription loop-mediated isothermal amplification. The test has two steps: (1) heat saliva with a stabilization solution and (2) detect virus by incubating with a primer/enzyme mix. After incubation, saliva samples containing the SARS-CoV-2 genome turn bright yellow. Because this test is pH dependent, it can react falsely to some naturally acidic saliva samples. We report unique saliva stabilization protocols that rendered 295 healthy saliva samples compatible with the test, producing zero false positives. We also evaluated the test on 278 saliva samples from individuals who were infected with SARS-CoV-2 but had no symptoms at the time of saliva collection, and from 54 matched pairs of saliva and anterior nasal samples from infected individuals. The Saliva TwoStep test described herein identified infections with 94% sensitivity and >99% specificity in individuals with sub-clinical (asymptomatic or pre-symptomatic) infections., Competing Interests: QY, NM, CP Some of the authors of this study (NRM, QY, CLP, SLS) are founders of Darwin Biosciences, who licenses the Saliva TwoStep assay described herein. SC is an employee of Darwin Biosciences. WF, AG, AB, BH, EW, SW, DM, CD, TS, EL, PG, MF, KT, CH, JD, CO, GB, MM, LL, RP No competing interests declared, SS Senior editor, eLife, (© 2021, Yang et al.)

Multiple gram-negative bacteria encode type III secretion systems (T3SS) that allow them to inject effector proteins directly into host cells to facilitate colonization. To be secreted, effector proteins must be at least partially unfolded to pass through the narrow needle-like channel (diameter <2 nm) of the T3SS. Fusion of effector proteins to tightly packed proteins-such as GFP, ubiquitin, or dihydrofolate reductase (DHFR)-impairs secretion and results in obstruction of the T3SS. Prior observation that unfolding can become rate-limiting for secretion has led to the model that T3SS effector proteins have low thermodynamic stability, facilitating their secretion. Here, we first show that the unfolding free energy ([Formula: see text]) of two Salmonella effector proteins, SptP and SopE2, are 6.9 and 6.0 kcal/mol, respectively, typical for globular proteins and similar to published [Formula: see text] for GFP, ubiquitin, and DHFR. Next, we mechanically unfolded individual SptP and SopE2 molecules by atomic force microscopy (AFM)-based force spectroscopy. SptP and SopE2 unfolded at low force ( F unfold ≤ 17 pN at 100 nm/s), making them among the most mechanically labile proteins studied to date by AFM. Moreover, their mechanical compliance is large, as measured by the distance to the transition state (Δ x ‡ = 1.6 and 1.5 nm for SptP and SopE2, respectively). In contrast, prior measurements of GFP, ubiquitin, and DHFR show them to be mechanically robust ( F unfold > 80 pN) and brittle (Δ x ‡ < 0.4 nm). These results suggest that effector protein unfolding by T3SS is a mechanical process and that mechanical lability facilitates efficient effector protein secretion., Competing Interests: The authors declare no competing interest.

Objective: This study aimed to describe the microscopic over-under cartilage tympanoplasty technique, provide hearing results and detail clinically significant complications., Method: This was a retrospective case series chart review study of over-under cartilage tympanoplasty procedures performed by the senior author between January 2015 and January 2019 at three tertiary care centres. Cases were excluded for previous or intra-operative cholesteatoma, if a mastoidectomy was performed during the procedure or if ossiculoplasty was performed. Hearing results and complications were obtained., Results: Sixty-eight tympanoplasty procedures met the inclusion criteria. The median age was 13 years (range, 3-71 years). The mean improvement in pure tone average was 6 dB (95 per cent confidence interval 4-9 dB; p < 0.0001). The overall perforation closure rate was 97 per cent (n = 66). Revision surgery was recommended for a total of 6 cases (9 per cent) including 2 post-operative perforations, 1 case of middle-ear cholesteatoma and 3 cases of external auditory canal scarring., Conclusion: Over-under cartilage tympanoplasty is effective at improving clinically meaningful hearing with a low rate of post-operative complications.