Virology under the microscope—a call for rational discourse

Viruses have brought humanity many challenges: respiratory infection, cancer, neurological impairment and immunosuppression to name a few. Virology research over the last 60+ years has responded to reduce this disease burden with vaccines and antivirals. Despite this long history, the COVID-19 pandemic has brought unprecedented attention to the field of virology. Some of this attention is focused on concern about the safe conduct of research with human pathogens. A small but vocal group of individuals has seized upon these concerns – conflating legitimate questions about safely conducting virus-related research with uncertainties over the origins of SARS-CoV-2. The result has fueled public confusion and, in many instances, ill-informed condemnation of virology. With this article, we seek to promote a return to rational discourse. We explain the use of gain-of-function approaches in science, discuss the possible origins of SARS-CoV-2 and outline current regulatory structures that provide oversight for virological research in the United States. By offering our expertise, we – a broad group of working virologists – seek to aid policy makers in navigating these controversial issues. Balanced, evidence-based discourse is essential to addressing public concern while maintaining and expanding much-needed research in virology

Geochronology of granulitized eclogite from the Ama Drime Massif : implications for the tectonic evolution of the South Tibetan Himalaya

The Ama Drime Massif (ADM) is an elongate north-south trending antiformal feature that extends ∼70 km north across the crest of the South Tibetan Himalaya and offsets the position of the South Tibetan Detachment system. A detailed U(-Th)-Pb geochronologic study of granulitized mafic eclogites and associated rocks from the footwall of the ADM yields important insights into the middle to late Miocene tectonic evolution of the Himalayan orogen. The mafic igneous precursor to the granulitized eclogites is 986.6 ± 1.8 Ma and was intruded into the paleoproterozoic (1799 ± 9 Ma) Ama Drime orthogneiss, the latter being similar in age to rocks previously assigned to the Lesser Himalayan Series in the Himalayan foreland. The original eclogite-facies mineral assemblage in the mafic rocks has been strongly overprinted by granulite facies metamorphism at 750°C and 0.7–0.8 GPa. In the host Ama Drime orthogneiss, the granulite event is correlated with synkinematic sillimanite-grade metamorphism and muscovite dehydration melting. Monazite and xenotime ages indicate that the granulite metamorphism and associated anatexis occurred at <13.2 ± 1.4 Ma. High-grade metamorphism was followed by postkinematic leucogranite dyke emplacement at 11.6 ± 0.4 Ma. This integrated data set indicates that high-temperature metamorphism, decompression, and exhumation of the ADM postdates mid-Miocene south directed midcrustal extrusion and is kinematically linked to orogen-parallel extension

Origins of Replication Determine Plasmid Copy Number.

<p>The origins of replication used in the study are listed with their descriptions and part numbers in the Registry of Standard Biological Parts. The means and standard deviations of PCN values were determined by qPCR and yields of minipreps.</p

Results of Programmed Evolution.

<p>(A) The starting population with equal amounts of all 24 strains was spread on LB agar plates with the indicated antibiotic and a disk treated as indicated. (B) Top row: spots of cells on LB agar with ampicillin for all 24 starting strains (left) and examples of clones after Programmed Evolution (right). Middle row: Agarose gels with PCR products to determine PCN for all 24 strains (left) and examples after Programmed Evolution (right). The 750 bp band for the low copy origin and the 500 bp band for the high copy origin are indicated by arrows. Bottom row: Agarose gels with PCR products to chaperone genotype for all 24 strains (left) and examples after Programmed Evolution (right). (C) The graph shows relative frequency of each of the genotype before (top) and after (bottom) Programmed Evolution. The order of chaperone plasmids along the left to right horizontal axis is pG-Tf2, pTf16, pG-KJE8, pGro7, pKJE7, and no chaperone. The order of genotype combinations along the other horizontal axis from back to front is high strength promoter/RBS + high copy origin; high strength promoter/RBS + low copy origin; low strength promoter/RBS + high copy origin; and low strength promoter/RBS + low copy origin.</p

Relative Fitness of Genotypes as a Function of Theophylline Production.

<p>Theophylline production as measured by LC-MS analysis is listed for the three genotypes with the highest fitness and two genotypes with very low fitness.</p

Results of Programmed Evolution.

<p>The number and genotype of colonies analyzed after Programmed Evolution from three replicate plate experiments.</p

Starting Population for Programmed Evolution.

<p>(A) An ampicillin resistance plasmid carries variation in the strength of promoters and RBS elements as well as the low and high copy number origins of replication. (B) A chloramphenicol resistance plasmid carries chaperones DNA KJE, Trigger Factor, and Gro ESL chaperones individually and in two combinations (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118322#sec004" target="_blank">Methods</a> for details).</p

Biosensor and Fitness Modules.

<p>(A) The Biosensor Module contains a promoter, a riboswitch that binds to theophylline, and a GFP gene. (B) Cells with the indicated genotypes were incubated with caffeine or theophylline. Fluorescence of cells grown in theophylline or caffeine was divided by absorbance at 590 nm (relative fluorescence) to correct for variation in cell density. (C) Relative fluorescence as a function of time in cells with and without the biosensor grown in 2.5 mM theophylline. (D) The Fitness Module contains a promoter, a riboswitch that binds theophylline, and the tetracycline resistance gene (<i>tetA</i>). (E) Cell growth in media containing tetracycline and either theophylline or caffeine as indicated.</p