Robust Approaches to the Quantitative Analysis of Genome Formula Variation in Multipartite and Segmented Viruses.

When viruses have segmented genomes, the set of frequencies describing the abundance of segments is called the genome formula. The genome formula is often unbalanced and highly variable for both segmented and multipartite viruses. A growing number of studies are quantifying the genome formula to measure its effects on infection and to consider its ecological and evolutionary implications. Different approaches have been reported for analyzing genome formula data, including qualitative description, applying standard statistical tests such as ANOVA, and customized analyses. However, these approaches have different shortcomings, and test assumptions are often unmet, potentially leading to erroneous conclusions. Here, we address these challenges, leading to a threefold contribution. First, we propose a simple metric for analyzing genome formula variation: the genome formula distance. We describe the properties of this metric and provide a framework for understanding metric values. Second, we explain how this metric can be applied for different purposes, including testing for genome-formula differences and comparing observations to a reference genome formula value. Third, we re-analyze published data to illustrate the applications and weigh the evidence for previous conclusions. Our re-analysis of published datasets confirms many previous results but also provides evidence that the genome formula can be carried over from the inoculum to the virus population in a host. The simple procedures we propose contribute to the robust and accessible analysis of genome-formula data.

Evaluation of sequencing and PCR-based methods for the quantification of the viral genome formula.

Viruses show great diversity in their genome organization. Multipartite viruses package their genome segments into separate particles, most or all of which are required to initiate infection in the host cell. The benefits of such seemingly inefficient genome organization are not well understood. One hypothesised benefit of multipartition is that it allows for flexible changes in gene expression by altering the frequency of each genome segment in different environments, such as encountering different host species. The ratio of the frequency of segments is termed the genome formula (GF). Thus far, formal studies quantifying the GF have been performed for well-characterised virus-host systems in experimental settings using RT-qPCR. However, to understand GF variation in natural populations or novel virus-host systems, a comparison of several methods for GF estimation including high-throughput sequencing (HTS) based methods is needed. Currently, it is unclear how HTS-methods compare a golden standard, such as RT-qPCR. Here we show a comparison of multiple GF quantification methods (RT-qPCR, RT-digital PCR, Illumina RNAseq and Nanopore direct RNA sequencing) using three host plants (Nicotiana tabacum, Nicotiana benthamiana, and Chenopodium quinoa) infected with cucumber mosaic virus (CMV), a tripartite RNA virus. Our results show that all methods give roughly similar results, though there is a significant method effect on genome formula estimates. While the RT-qPCR and RT-dPCR GF estimates are congruent, the GF estimates from HTS methods deviate from those found with PCR. Our findings emphasize the need to tailor the GF quantification method to the experimental aim, and highlight that it may not be possible to compare HTS and PCR-based methods directly. The difference in results between PCR-based methods and HTS highlights that the choice of quantification technique is not trivial.

Towards evolutionary predictions: Current promises and challenges.

Evolution has traditionally been a historical and descriptive science, and predicting future evolutionary processes has long been considered impossible. However, evolutionary predictions are increasingly being developed and used in medicine, agriculture, biotechnology and conservation biology. Evolutionary predictions may be used for different purposes, such as to prepare for the future, to try and change the course of evolution or to determine how well we understand evolutionary processes. Similarly, the exact aspect of the evolved population that we want to predict may also differ. For example, we could try to predict which genotype will dominate, the fitness of the population or the extinction probability of a population. In addition, there are many uses of evolutionary predictions that may not always be recognized as such. The main goal of this review is to increase awareness of methods and data in different research fields by showing the breadth of situations in which evolutionary predictions are made. We describe how diverse evolutionary predictions share a common structure described by the predictive scope, time scale and precision. Then, by using examples ranging from SARS-CoV2 and influenza to CRISPR-based gene drives and sustainable product formation in biotechnology, we discuss the methods for predicting evolution, the factors that affect predictability and how predictions can be used to prevent evolution in undesirable directions or to promote beneficial evolution (i.e. evolutionary control). We hope that this review will stimulate collaboration between fields by establishing a common language for evolutionary predictions.

Unresolved advantages of multipartitism in spatially structured environments.

Multipartite viruses have segmented genomes and package each of their genome segments individually into distinct virus particles. Multipartitism is common among plant viruses, but why this apparently costly genome organization and packaging has evolved remains unclear. Recently Zhang and colleagues developed network epidemiology models to study the epidemic spread of multipartite viruses and their distribution over plant and animal hosts (Phys. Rev. Lett. 2019, 123, 138101). In this short commentary, we call into question the relevance of these results because of key model assumptions. First, the model of plant hosts assumes virus transmission only occurs between adjacent plants. This assumption overlooks the basic but imperative fact that most multipartite viruses are transmitted over variable distances by mobile animal vectors, rendering the model results irrelevant to differences between plant and animal hosts. Second, when not all genome segments of a multipartite virus are transmitted to a host, the model assumes an incessant latent infection occurs. This is a bold assumption for which there is no evidence to date, making the relevance of these results to understanding multipartitism questionable.

A 10-week exercise intervention can improve work posture but not neck/shoulder symptoms in dental health students: A pilot cohort study.

BACKGROUND: Dental workers including dental health students experience a high incidence of neck/shoulder symptoms. However, research into interventions has been scarce and largely focused on tools and technological modifications. OBJECTIVE: This study investigates the feasibility and benefits of a neck/shoulder exercise intervention on work posture and local symptoms in dental health students. METHODS: Twenty-three students participated in a 10-week resistance exercise program for 2 minutes daily, 5 days per week. Work posture (evaluated with the Rapid Upper Limb Assessment tool, RULA), muscle strength measures, self-rated symptom severity and function were collected. Feasibility of the intervention was determined with six items scored on a 5-point Likert scale. RESULTS: The exercise intervention improved: work posture score by 1.88 (95% CI 1.05-2.70, p < 0.01) points on the RULA; isometric maximal strength of neck flexor and extensor muscles by 17.2% (95% CI 6.9-27.6%, p < 0.01) and 23.2% (95% CI 10.2-36.3%, p < 0.01) respectively; and clinical performance (77%). Isometric strength of the shoulder muscles, and symptom severity at the neck/shoulder showed no improvement. Adherence to training was 77%. CONCLUSION: Ten weeks of exercise was feasible for dental health students, and likely beneficial with improvements in work posture and neck muscle strength.

3-Bromo-pyridin-2-amine.

In the crystal structure of the title compound, C(5)H(5)BrN(2), mol-ecules assemble via pairs of N-H⋯N hydrogen bonds into inversion dimers using only the syn H atom on the amine group. These dimers then assemble further into two-dimensional layers via type I C-Br⋯Br [Br⋯Br = 3.693 (s6) Å] halogen bonding along the (102) plane.