Phylogenetic analysis of Puumala virus strains from Central Europe highlights the need for a full-genome perspective on hantavirus evolution

Puumala virus (PUUV), carried by bank voles (Myodes glareolus), is the medically most important hantavirus in Central and Western Europe. In this study, a total of 523 bank voles (408 from Germany, 72 from Slovakia, and 43 from Czech Republic) collected between the years 2007–2012 were analyzed for the presence of hantavirus RNA. Partial PUUV genome segment sequences were obtained from 51 voles. Phylogenetic analyses of all three genome segments showed that the newfound strains cluster with other Central and Western European PUUV strains. The new sequences from ?umava (Bohemian Forest), Czech Republic, are most closely related to the strains from the neighboring Bavarian Forest, a known hantavirus disease outbreak region. Interestingly, the Slovak strains clustered with the sequences from Bohemian and Bavarian Forests only in the M but not S segment analyses. This well-supported topological incongruence suggests a segment reassortment event or, as we analyzed only partial sequences, homologous recombination. Our data highlight the necessity of sequencing all three hantavirus genome segments and of a broader bank vole screening not only in recognized endemic foci but also in regions with no reported human hantavirus disease cases.     

Clear-cell Renal Cell Carcinoma: Molecular Characterization of IMDC Risk Groups and Sarcomatoid Tumors

IntroductionRecent trials have suggested predictive biomarkers in advanced clear-cell renal cell carcinoma (accRCC): International Metastatic RCC Database Consortium (IMDC) good risk or angiogenic gene signature for sunitinib and IMDC intermediate/poor risk for ipilimumab-nivolumab and T-effector cell signature or sarcomatoid dedifferentiation for atezolizumab-bevacizumab. We hypothesized that earlier described molecular subtypes, ccrcc1 to ccrcc4, could provide similar information as a single generic biomarker and molecularly characterize the heterogeneous intermediate-risk group.Patients and MethodsPatients with accRCC treated with systemic therapies were included. We assessed associations between the 5 biomarkers and their impact on progression-free survival (PFS) and response rate (RR) on first-line sunitinib or pazopanib. The cutoff percentage of sarcomatoid dedifferentiation with optimal discriminative value was determined.ResultsIn total, 430 patients were included (163 with molecular data). The molecular ccrcc2 subtype identified tumors with higher angiogenic gene expression across IMDC risk groups: prevalence was high in IMDC good risk and low in IMDC poor risk (P < .001). Molecular subtype, IMDC, and angiogenic gene expression had comparable C-indices to predict PFS and RR (range, 60%-66%). The ccrcc2 subtype and angiogenic gene expression were positive predictors of PFS in IMDC intermediate-risk patients (P = .006; P = .04). Immune signature did not differ between IMDC groups, but was strongly correlated with molecular subtype (P = .8 and P = .0007). A cutoff value of 25% sarcomatoid differentiation discriminated tumors with distinct molecular characteristics and therapeutic sensitivity.ConclusionIn accRCC, molecular subtypes can explain differences in IMDC risk group, expression of angiogenesis and immune response genes, and sarcomatoid dedifferentiation. They can identify molecularly different patient populations within the heterogeneous IMDC intermediate group and select patients for systemic therapies.     

The Functional Living Index-Cancer: estimating its reliability based on clinical trial data

Purpose  

The Functional Living Index-Cancer was developed to measure quality of life in cancer trials as an adjunct to the usual clinical outcomes. The scale is considered conceptually good, since it covers a broad range of relevant aspects of quality of life, but the main criticism has been that its reliability has never been properly investigated. In this paper, we investigate the reliability of the FLIC.     

Human Diversity of Killer Cell Immunoglobulin-Like Receptors and Human Leukocyte Antigen Class I Alleles and Ebola Virus Disease Outcomes

We investigated the genetic profiles of killer cell immunoglobulin-like receptors (KIRs) in Ebola virus–infected patients. We studied the relationship between KIR–human leukocyte antigen (HLA) combinations and the clinical outcomes of patients with Ebola virus disease (EVD). We genotyped KIRs and HLA class I alleles using DNA from uninfected controls, EVD survivors, and persons who died of EVD. The activating 2DS4–003 and inhibitory 2DL5 genes were significantly more common among persons who died of EVD; 2DL2 was more common among survivors. We used logistic regression analysis and Bayesian modeling to identify 2DL2, 2DL5, 2DS4–003, HLA-B-Bw4-Thr, and HLA-B-Bw4-Ile as probably having a significant relationship with disease outcome. Our findings highlight the importance of innate immune response against Ebola virus and show the association between KIRs and the clinical outcome of EVD.     

Generalizability in nongaussian longitudinal clinical trial data based on generalized linear mixed models

This work investigates how generalizability, an extension of reliability, can be defined and estimated based on longitudinal data sequences resulting from, for example, clinical studies. Useful and intuitive approximate expressions are derived based on generalized linear mixed models. Data from four double-blind, randomized clinical trials into schizophrenia motivate the research and are used to estimate generalizability for a binary response parameter.     

Genomic basis of parallel adaptation varies with divergence in Arabidopsis and its relatives

Parallel adaptation provides valuable insight into the predictability of evolutionary change through replicated natural experiments. A steadily increasing number of studies have demonstrated genomic parallelism, yet the magnitude of this parallelism varies depending on whether populations, species, or genera are compared. This led us to hypothesize that the magnitude of genomic parallelism scales with genetic divergence between lineages, but whether this is the case and the underlying evolutionary processes remain unknown. Here, we resequenced seven parallel lineages of two Arabidopsis species, which repeatedly adapted to challenging alpine environments. By combining genome-wide divergence scans with model-based approaches, we detected a suite of 151 genes that show parallel signatures of positive selection associated with alpine colonization, involved in response to cold, high radiation, short season, herbivores, and pathogens. We complemented these parallel candidates with published gene lists from five additional alpine Brassicaceae and tested our hypothesis on a broad scale spanning ∼0.02 to 18 My of divergence. Indeed, we found quantitatively variable genomic parallelism whose extent significantly decreased with increasing divergence between the compared lineages. We further modeled parallel evolution over the Arabidopsis candidate genes and showed that a decreasing probability of repeated selection on the same standing or introgressed alleles drives the observed pattern of divergence-dependent parallelism. We therefore conclude that genetic divergence between populations, species, and genera, affecting the pool of shared variants, is an important factor in the predictability of genome evolution.

Evolution is driven by a complex interplay of deterministic and stochastic forces whose relative importance is a matter of debate (1). Being largely a historical process, we have limited ability to experimentally test for the predictability of evolution in its full complexity (i.e., in natural environments) (2). Distinct lineages that independently adapted to similar conditions by similar phenotype (termed parallel,” considered synonymous to “convergent” here) can provide invaluable insights into the issue (3, 4). An improved understanding of the probability of parallel evolution in nature may inform on constraints on evolutionary change and provide insights relevant for predicting the evolution of pathogens (5–7), pests (8, 9), or species in human-polluted environments (10, 11). Although the past few decades have seen an increasing body of work supporting the parallel emergence of traits by the same genes and even alleles, we know surprisingly little about what makes parallel evolution more likely and, by extension, what factors underlie evolutionary predictability (1, 12).A wealth of literature describes the probability of “genetic” parallelism, showing why certain genes are involved in parallel adaptation more often than others (13). There is theoretical and empirical evidence for the effect of pleiotropic constraints, availability of beneficial mutations or position in the regulatory network all having an impact on the degree of parallelism at the level of a single locus (3, 13–18). In contrast, we know little about causes underlying “genomic” parallelism (i.e., what fraction of the genome is reused in adaptation and why). Individual case studies demonstrate large variation in genomic parallelism, ranging from absence of any parallelism (19), similarity in functional pathways but not genes (20, 21), and reuse of a limited number of genes (22–24) to abundant parallelism at both gene and functional levels (25, 26). Yet, there is little consensus about what determines variation in the degree of gene reuse (fraction of genes that repeatedly emerge as selection candidates) across investigated systems (1).Divergence (the term used here to consistently describe both intra- and interspecific genetic differentiation) between the compared instances of parallelism appears as a potential driver of the variation in gene reuse (14, 27, 28). Phenotype-oriented meta-analyses suggest that both phenotypic convergence (28) and genetic parallelism underlying phenotypic traits (14) decrease with increasing time to the common ancestor. Although a similar targeted multiscale comparison is lacking at the genomic level, our brief review of published studies (29 cases, Dataset S1) suggests that also gene reuse tends to scale with divergence (Fig. 1A and SI Appendix, Fig. S1). Moreover, allele reuse (repeated sweep of the same haplotype that is shared among populations either via gene flow or from standing genetic variation) frequently underlies parallel adaptation between closely related lineages (29–32), while parallelism from independent de novo mutations at the same locus dominates between distantly related taxa (13). Similarly, previous studies reported a decreasing probability of hemiplasy (apparent convergence resulting from gene tree discordance) with divergence in phylogeny-based studies (33, 34). This suggests that the degree of allele reuse may be the primary factor underlying the hypothesized divergence-dependency of parallel genome evolution, possibly reflecting either weak hybridization barriers, widespread ancestral polymorphism between closely related lineages (35), or ecological reasons (lower niche differentiation and geographical proximity) (36, 37). However, the generally restricted focus of individual studies of genomic parallelism on a single level of divergence does not lend itself to a unified comparison across divergence scales. Although different ages of compared lineages affect a variety of evolutionary–ecological processes such as diversification rates, community structure, or niche conservatism (37), the hypothesis that genomic parallelism scales with divergence has not yet been systematically tested, and the underlying evolutionary processes remain poorly understood.Open in a separate windowFig. 1.Hypotheses regarding relationships between genomic parallelism and divergence and the Arabidopsis system used to address these hypotheses. (A) Based on our literature review, we propose that genetically closer lineages adapt to a similar challenge more frequently by gene reuse, sampling suitable variants from the shared pool (allele reuse), which makes their adaptive evolution more predictable. Color ramp symbolizes rising divergence between the lineages (∼0.02 to 18 Mya in this study); the symbols denote different divergence levels tested here using resequenced genomes of 22 Arabidopsis populations (circles) and meta-analysis of candidates in Brassicaceae (asterisks). (B) Spatial arrangement of lineages of varying divergence (neutral F_ST; bins only aid visualization; all tests were performed on a continuous scale) encompassing parallel alpine colonization within the two Arabidopsis outcrossers from central Europe: A. arenosa (diploid: aVT; autotetraploid: aNT, aZT, aRD, and aFG) and A. halleri (diploid: hNT and hFG). Note that only two of the ten between-species pairs (dark green) are shown to aid visibility. The color scale corresponds to the left part of the color ramp used in A. (C) Photos of representative alpine and foothill habitat. (D) Representative phenotypes of originally foothill and alpine populations grown in common garden demonstrating phenotypic convergence. Scale bar corresponds to 4 cm. (E) Morphological differentiation among 223 A. arenosa individuals originating from foothill (black) and alpine (gray) populations from four regions after two generations in a common garden. Principal component analysis was run using 16 morphological traits taken from ref. 45.Here, we aimed to test this hypothesis and investigate whether allele reuse is a major factor underlying the relationship. We analyzed replicated instances of adaptation to a challenging alpine environment, spanning a range of divergence from populations to tribes within the plant family Brassicaceae (38–43) (Fig. 1A). First, we took advantage of a unique naturally multireplicated setup in the plant model genus Arabidopsis that was so far neglected from a genomic perspective (Fig. 1B). Two predominantly foothill-dwelling Arabidopsis outcrossers (A. arenosa, A. halleri) exhibit scattered, morphologically distinct alpine occurrences at rocky outcrops above the timberline (Fig. 1C). These alpine forms are separated from the widespread foothill population by a distribution gap spanning at least 500 m of elevation. Previous genetic and phenotypic investigations and follow-up analyses presented here showed that the scattered alpine forms of both species represent independent alpine colonization in each mountain range, followed by parallel phenotypic differentiation (Fig. 1 D and E) (44–46). Thus, we sequenced genomes from seven alpine and adjacent foothill population pairs, covering all European lineages encompassing the alpine ecotype. We discovered a suite of 151 genes from multiple functional pathways relevant to alpine stress that were repeatedly differentiated between foothill and alpine populations. This points toward a polygenic, multifactorial basis of parallel alpine adaptation.We took advantage of this set of well-defined parallel selection candidates and tested whether the degree of gene reuse decreases with increasing divergence between the compared lineages (Fig. 1A). By extending our analysis to five additional alpine Brassicaceae species, we further tested whether there are limits to gene reuse above the species level. Finally, we inquired about possible underlying evolutionary processes by estimating the extent of allele reuse using a designated modeling approach. Overall, our empirical analysis provides a perspective to the ongoing discussion about the variability in the reported magnitude of parallel genome evolution and identifies allele reuse as an important evolutionary process shaping the extent of genomic parallelism between populations, species, and genera.