Multiple Immune and Genetic Mechanisms Contribute to Cmv5s-Driven Susceptibility and Tissue Damage during Acute Murine Cytomegalovirus Infection.

The MHC class I molecule H-2Dk conveys resistance to acute murine CMV infection in both C57L (H-2Dk transgenic) and MA/My mice. M.H2k/b mice are on an MA/My background aside from a C57L-derived region spanning the MHC (Cmv5s), which diminishes this resistance and causes significant spleen histopathology. To hone in on the effector elements within the Cmv5s interval, we generated several Cmv5-recombinant congenic mouse strains and screened them in vivo, allowing us to narrow the phenotype-associated interval >6-fold and segment the genetic mechanism to at least two independent loci within the MHC region. In addition, we sought to further characterize the Cmv5s-associated phenotypes in their temporal appearance and potential direct relationship to viral load. To this end, we found that Cmv5s histopathology and NK cell activation could not be fully mirrored in the MA/My mice with increased viral dose, and that marginal zone destruction was the first apparent Cmv5s phenotype, being reliably quantified as early as 2 d postinfection in the M.H2k/b mice, prior to divergence in viral load, weight loss, or NK cell phenotype. Finally, we further dissect NK cell involvement, finding no intrinsic differences in NK cell function, despite increased upregulation of activation markers and checkpoint receptors. In conclusion, these data dissect the genetic and immunologic underpinnings of Cmv5 and reveal a model in which polymorphism within the MHC region of the genome leads to the development of tissue damage and corrupts protective NK cell immunity during acute viral infection.

The Value of Interpersonal Network Continuity for College Students in Disruptive Times.

The COVID-19 pandemic-related social distancing practices that colleges implemented in Spring 2020 disrupted the typical mechanisms of propinquity (physical proximity) and homophily (shared characteristics) that physical institutions rely on to help students build and maintain relationships critical to learning and wellbeing. To explore how social distancing shaped students' academic and social networks and associated educational outcomes, we conceptualized it as a "network shock" and collected unique ego network data in April 2020. For participating students, maintaining interactions with the same set of individuals before and after social distancing was related to more positive outcomes across a range of self-reported wellbeing and learning indicators. On average, students experienced a loss of frequent academic contacts, while they maintained or replaced social interactions in their interpersonal networks after social distancing. Our investigation of the ways students experienced changes in their social and academic networks after a loss of physical proximity points to the role of interpersonal interaction network continuity for fostering wellbeing and learning in times of disruption, as well as the potential need for support in maintaining or rebuilding academic networks.

A directional spectrum evolution model for ship noise.

A radiation transport equation that describes the spatiotemporal evolution of the directional spectrum of underwater acoustic noise is presented and applied to ship noise. A ray-based algorithm is used to solve the transport equation and numerically simulate the evolution of the directional noise spectrum produced by a passing ship. The model described accounts for the transient and highly episodic nature of shipping noise, the strong anisotropy of the radiated shipping noise, the directional dependence of the resulting acoustic field, and the critical angle dependence of bottom-reflected energy. The model predicts time histories of sound pressure level and directional spectral energy density at distant locations if the ship track and the ship's radiated noise power are known. Simulations are shown to be in qualitatively good agreement with observations.

Boosting Live Malaria Vaccine with Cytomegalovirus Vector Can Prolong Immunity through Innate and Adaptive Mechanisms.

Vaccines to persistent parasite infections have been challenging, and current iterations lack long-term protection. Cytomegalovirus (CMV) chronic vaccine vectors drive protection against SIV, tuberculosis and liver-stage malaria correlated with antigen-specific CD8 T cells with a Tem phenotype. This phenotype is likely driven by a combination of antigen-specific and innate adjuvanting effects of the vector, though these mechanisms are less well understood. Sterilizing immunity from live Plasmodium chabaudi vaccination lasts less than 200 days. While P. chabaudi-specific antibody levels remain stable after vaccination, the decay of parasite-specific T cells correlates with loss of challenge protection. Therefore, we enlisted murine CMV as a booster strategy to prolong T cell responses against malaria. To study induced T cell responses, we included P. chabaudi MSP-1 epitope B5 (MCMV-B5). We found that MCMV vector alone significantly protected against a challenge P. chabaudi infection 40-60 days later, and that MCMV-B5 was able to make B5-specific Teff, in addition to previously-reported Tem, that survive to the challenge timepoint. Used as a booster, MCMV-B5 prolonged protection from heterologous infection beyond day 200, and increased B5 TCR Tg T cell numbers, including both a highly-differentiated Tem phenotype and Teff, both previously reported to protect. B5 epitope expression was responsible for maintenance of Th1 and Tfh B5 T cells. In addition, the MCMV vector had adjuvant properties, contributing non-specifically through prolonged stimulation of IFN-γ. In vivo neutralization of IFN-γ, but not IL-12 and IL-18, late in the course of MCMV, led to loss of the adjuvant effect. Mechanistically, sustained IFN-γ from MCMV increased CD8α+ dendritic cell numbers, and led to increased IL-12 production upon Plasmodium challenge. In addition, neutralization of IFN-γ before challenge reduced the polyclonal Teff response to challenge. Our findings suggest that, as protective epitopes are defined, an MCMV vectored booster can prolong protection through the innate effects of IFN-γ.

NK cells reduce anergic T cell development in early-stage tumors by promoting myeloid cell maturation.

Introduction: Studies of NK cells in tumors have primarily focused on their direct actions towards tumor cells. We evaluated the impact of NK cells on expression of homing receptor ligands on tumor vasculature, intratumoral T cell number and function, and T cell activation in tumor draining lymph node. Methods: Using an implantable mouse model of melanoma, T cell responses and homing receptor ligand expression on the vasculature were evaluated with and without NK cells present during the early stages of the tumor response by flow cytometry. Results: NK cells in early-stage tumors are one source of IFNγ that augments homing receptor ligand expression. More significantly, NK cell depletion resulted in increased numbers of intratumoral T cells with an anergic phenotype. Anergic T cell development in tumor draining lymph node was associated with increased T-cell receptor signaling but decreased proliferation and effector cell activity, and an incomplete maturation phenotype of antigen presenting cells. These effects of NK depletion were similar to those of blocking CD40L stimulation. Discussion: We conclude that an important function of NK cells is to drive proper APC maturation via CD40L during responses to early-stage tumors, reducing development of anergic T cells. The reduced development of anergic T cells resulting in improved tumor control and T cell responses when NK cells were present.

Altered-Self MHC Class I Sensing via Functionally Disparate Paired NK Cell Receptors Counters Murine Cytomegalovirus gp34-Mediated Immune Evasion.

The murine CMV (MCMV) immunoevasin m04/gp34 escorts MHC class I (MHC I) molecules to the surface of infected cells where these complexes bind Ly49 inhibitory receptors (IRs) and prevent NK cell attack. Nonetheless, certain self-MHC I-binding Ly49 activating and inhibitory receptors are able to promote robust NK cell expansion and antiviral immunity during MCMV infection. A basis for MHC I-dependent NK cell sensing of MCMV-infected targets and control of MCMV infection however remains unclear. In this study, we discovered that the Ly49R activation receptor is selectively triggered during MCMV infection on antiviral NK cells licensed by the Ly49G2 IR. Ly49R activating receptor recognition of MCMV-infected targets is dependent on MHC I Dk and MCMV gp34 expression. Remarkably, although Ly49R is critical for Ly49G2-dependent antiviral immunity, blockade of the activation receptor in Ly49G2-deficient mice has no impact on virus control, suggesting that paired Ly49G2 MCMV sensing might enable Ly49R+ NK cells to better engage viral targets. Indeed, MCMV gp34 facilitates Ly49G2 binding to infected cells, and the IR is required to counter gp34-mediated immune evasion. A specific requirement for Ly49G2 in antiviral immunity is further explained by its capacity to license cytokine receptor signaling pathways and enhance Ly49R+ NK cell proliferation during infection. These findings advance our understanding of the molecular basis for functionally disparate self-receptor enhancement of antiviral NK cell immunity.

Caspase-7 activates ASM to repair gasdermin and perforin pores.

Among the caspases that cause regulated cell death, a unique function for caspase-7 has remained elusive. Caspase-3 performs apoptosis, whereas caspase-7 is typically considered an inefficient back-up. Caspase-1 activates gasdermin D pores to lyse the cell; however, caspase-1 also activates caspase-7 for unknown reasons1. Caspases can also trigger cell-type-specific death responses; for example, caspase-1 causes the extrusion of intestinal epithelial cell (IECs) in response to infection with Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium)2,3. Here we show in both organoids and mice that caspase-7-deficient IECs do not complete extrusion. Mechanistically, caspase-7 counteracts gasdermin D pores and preserves cell integrity by cleaving and activating acid sphingomyelinase (ASM), which thereby generates copious amounts of ceramide to enable enhanced membrane repair. This provides time to complete the process of IEC extrusion. In parallel, we also show that caspase-7 and ASM cleavage are required to clear Chromobacterium violaceum and Listeria monocytogenes after perforin-pore-mediated attack by natural killer cells or cytotoxic T lymphocytes, which normally causes apoptosis in infected hepatocytes. Therefore, caspase-7 is not a conventional executioner but instead is a death facilitator that delays pore-driven lysis so that more-specialized processes, such as extrusion or apoptosis, can be completed before cell death. Cells must put their affairs in order before they die.

Rural college graduates: Who comes home?

Many rural communities throughout the United States have experienced brain drain, or the out-migration of educated young people. Explanations for why college-educated adults leave rural communities have relied on economic rationales; however, the effects of social identities, community context, and place attachment have also been shown to influence migration decisions. Using data from the National Longitudinal Study of Adolescent to Adult Health (Add Health), this study examines factors experienced during adolescence as well as postsecondary characteristics that promote college graduates' return to their rural communities when they are between the ages of 34 and 43. We find that among college graduates who had attended a rural public K-12 school, those who had higher levels of school attachment were significantly more likely to return home compared to graduates who had lower levels of school attachment. The findings also suggest that graduates who came from a lower college-educated community were more likely to return home than those from average or highly college-educated communities. By analyzing long-term outcomes, this study extends our understanding of the strengths of adolescent experiences and neighborhood context influencing the pull to return home and the support for policies strengthening rural communities as there may be long-term effects to returning home, even if youth leave for college.

Licensing Natural Killers for Antiviral Immunity.

Immunoreceptor tyrosine-based inhibitory motif (ITIM)-bearing receptors (IRs) enable discrimination between self- and non-self molecules on the surface of host target cells. In this regard, they have a vital role in self-tolerance through binding and activating intracellular tyrosine phosphatases which can inhibit cellular activation. Yet, self-MHC class I (MHC I)-specific IRs are versatile in that they can also positively impact lymphocyte functionality, as exemplified by their role in natural killer (NK) cell education, often referred to as 'licensing'. Recent discoveries using defined mouse models of cytomegalovirus (CMV) infection have revealed that select self-MHC I IRs can increase NK cell antiviral defenses as well, whereas other licensing IRs cannot, or instead impede virus-specific NK responses for reasons that remain poorly understood. This review highlights a role for self-MHC I 'licensing' IRs in antiviral immunity, especially in the context of CMV infection, their impact on virus-specific NK cells during acute infection, and their potential to affect viral pathogenesis and disease.

Data on the first functionally-annotated de novo transcriptome assembly for North American flying squirrels (genus Glaucomys).

We report the first functionally-annotated de novo transcriptome assembly for North American flying squirrels (genus Glaucomys). RNA was extracted from tissue samples obtained from two northern flying squirrels and two southern flying squirrels sampled from Ontario, Canada, and sequenced on an Illumina paired-end sequencing platform. We reconstructed 702,228 Glaucomys transcripts using 193,323,120 sequence read pairs and captured sequence homologies, protein domains, and gene function classifications. Introgressive hybridization between northern (Glaucomys sabrinus) and southern flying squirrels (G. volans) has been observed in some areas of North America. However, existing molecular markers lack the resolution to discriminate late-generation introgressants and describe the extent to which hybridization influences the Glaucomys gene pool. These genomic resources can increase the resolution of molecular techniques used to examine the dynamics of the Glaucomys hybrid zone.

Effects of cost surface uncertainty on current density estimates from circuit theory.

BACKGROUND: Conservation practitioners are often interested in developing land use plans that increase landscape connectivity, which is defined as the degree to which the landscape facilitates or impedes movement among resource patches. Landscape connectivity is often estimated with a cost surface that indicates the varying costs experienced by an organism in moving across a landscape. True, or absolute costs are rarely known however, and therefore assigning costs to different landscape elements is often a challenge in creating cost surface maps. As such, we consider it important to understand the sensitivity of connectivity estimates to uncertainty in cost estimates. METHODS: We used simulated landscapes to test the sensitivity of current density estimates from circuit theory to varying relative cost values, fragmentation, and number of cost classes (i.e., thematic resolution). Current density is proportional to the probability of use during a random walk. Using Circuitscape software, we simulated electrical current between pairs of nodes to create current density maps. We then measured the correlation of the current density values across scenarios. RESULTS: In general, we found that cost values were highly correlated across scenarios with different cost weights (mean correlation ranged from 0.87 to 0.92). Changing the spatial configuration of landscape elements by varying the degree of fragmentation reduced correlation in current density across maps. We also found that correlations were more variable when the range of cost values in a map was high. DISCUSSION: The low sensitivity of current density estimates to relative cost weights suggests that the measure may be reliable for land use applications even when there is uncertainty about absolute cost values, provided that the user has the costs correctly ranked. This finding should facilitate the use of cost surfaces by conservation practitioners interested in estimating connectivity and planning linkages and corridors.

Time-warping in underwater acoustic waveguides.

The traditional way to isolate fixed mode number contributions to a transient wavefield in an underwater acoustic waveguide involves measuring the wavefield on a dense water-column-spanning vertical array and exploiting orthogonality over depth of the modes at each frequency. Recently it has been demonstrated that essentially the same goal can be accomplished in an ideal shallow water waveguide using measurements made on an isolated receiver by employing a signal processing technique known as time-warping. Time-warping makes use of a special nonuniform temporal sampling of the measured signal for which contributions from individual mode numbers are isolated in the frequency spectrum of the time-warped signal. The time-warping transformation in a general underwater acoustic waveguide is derived here. The general time-warping transformation is shown to reduce to the ideal shallow water waveguide time-warping transform as a special case. Use of the general time-warping transformation is illustrated with simulations in both a mid-latitude deep ocean environment and a high-latitude environment.

Ly49R activation receptor drives self-MHC-educated NK cell immunity against cytomegalovirus infection.

Natural killer (NK) cells mediate vital control of cancer and viral infection. They rely on MHC class I (MHC I)-specific self-receptors to identify and lyse diseased cells without harming self-MHC I-bearing host cells. NK cells bearing inhibitory self-receptors for host MHC I also undergo education, referred to as licensing, which causes them to become more responsive to stimulation via activation receptor signaling. Previous work has shown that licensed NK cells selectively expand during virus infections and they are associated with improved clinical response in human patients experiencing certain chronic virus infections, including HIV and hepatitis C virus. However, the importance of inhibitory self-receptors in NK-mediated virus immunity is debated as they also limit signals in NK cells emanating from virus-specific activation receptors. Using a mouse model of MHC I-dependent (H-2Dk) virus immunity, we discovered that NK cells depend on the Ly49G2 inhibitory self-receptor to mediate virus control, which coincided with host survival during murine cytomegalovirus infection. This antiviral effect further requires active signaling in NK cells via the Ly49R activation receptor that also binds H-2Dk. In tandem, these functionally discordant Ly49 self-receptors increase NK cell proliferation and effector activity during infection, resulting in selective up-regulation of CD25 and KLRG1 in virus-specific Ly49R+ Ly49G2+ NK cells. Our findings establish that paired self-receptors act as major determinants of NK cell-mediated virus sensing and immunity.

Characterizing the seabed in the Straits of Florida by using acoustic noise interferometry and time warping.

Interferometry of ambient and shipping noise in the ocean provides a way to estimate physical parameters of the seafloor and the water column in an environmentally friendly manner without employing any controlled sound sources. With noise interferometry, two-point cross-correlation functions of noise serve as the probing signals and replace the Green's function measured in active acoustic remote sensing. The amount of environmental information that can be obtained with passive remote sensing and the robustness of the estimates of the seafloor parameters increase when contributions of individual normal modes are resolved in the noise cross-correlation function. Using the data obtained in the 2012 noise-interferometry experiment in the Straits of Florida, dispersion curves of the first four normal modes are obtained in this paper by application of the time-warping transform to noise cross correlations. The passively measured dispersion curves are inverted for unknown geoacoustic properties of the seabed. Resulting thickness of the sediment layer and sound speed are consistent with the geoacoustic models obtained earlier by other means.

Natural selection for killer receptors and their MHC class I ligands: In pursuit of gene pairs that fit well in tandem.

Our understanding of the genetic basis of host resistance to viral infection and disease has progressed significantly over the last century. Numerous genes coding for modifiers of immune functions have been identified, which impact a variety of critical cellular processes, including signaling via lymphocyte receptors and their ligands, signal transduction, cytokine signaling, production and release of cytotoxic effectors, transcriptional regulation, and proliferation. Genome-wide association studies implicate an important role for both highly polymorphic NK cell receptors and their MHC class I ligands in modifying host resistance. These findings indicate NK cells are critical mediators of viral control with considerable potential to affect morbidity and mortality outcomes. They further suggest that both stimulatory and inhibitory NK receptor polymorphisms alter NK cell sensing of MHC I ligands on viral targets, which influences how NK cells respond to infection. In many cases, however, the underlying causes associated with host outcomes remain elusive. Herein, we discuss several modes of NK cell sensing of MHC I and MHC I-like molecules on viral targets, and the role of genetic diversity in this evolutionarily dynamic process. We further suggest that natural selection for paired NK receptors with opposing function, but shared MHC I ligands may give rise to rare, but highly effective MHC I-dependent modes of NK cell sensing of viral targets.

Expiratory reactance abnormalities in patients with expiratory dynamic airway collapse: a new application of impulse oscillometry.

Expiratory dynamic airways collapse (EDAC) is a condition that affects the central airways; it is not well characterised physiologically, with relatively few studies. We sought to characterise impulse oscillometry (IOS) features of EDAC in patients with normal spirometry. Expiratory data were hypothesised to be the most revealing. In addition, we compared IOS findings in chronic obstructive pulmonary disease (COPD) patients with and without EDAC. EDAC was identified at bronchoscopy as 75-100% expiratory closure at the carina or bilateral main bronchi. Four patient groups were compared: controls with no EDAC and normal lung function; lone EDAC with normal lung function; COPD-only patients; and COPD patients with EDAC. 38 patients were studied. Mean IOS data z-scores for EDAC compared to controls showed significantly higher reactance (X) values including X at 5 Hz, resonance frequency and area under the reactance curve (AX). EDAC showed significantly greater expiratory/inspiratory differences in all IOS data compared to controls. Stepwise logistic regression showed that resonant frequency best discriminated between EDAC and normal control, whereas classification and regression tree analysis found AX ≥3.523 to be highly predictive for EDAC in cases with normal lung function (14 out of 15 cases, and none out of eight controls). These data show a new utility of IOS: detecting EDAC in patients with normal lung function.

Modulation of NKG2D, NKp46, and Ly49C/I facilitates natural killer cell-mediated control of lung cancer.

Natural killer (NK) cells play a critical role in controlling malignancies. Susceptibility or resistance to lung cancer, for example, specifically depends on NK cell function. Nevertheless, intrinsic factors that control NK cell-mediated clearance of lung cancer are unknown. Here we report that NK cells exposed to exogenous major histocompatibility class I (MHCI) provide a significant immunologic barrier to the growth and progression of malignancy. Clearance of lung cancer is facilitated by up-regulation of NKG2D, NKp46, and other activating receptors upon exposure to environmental MHCI. Surface expression of the inhibitory receptor Ly49C/I, on the other hand, is down-regulated upon exposure to tumor-bearing tissue. We thus demonstrate that NK cells exhibit dynamic plasticity in surface expression of both activating and inhibitory receptors based on the environmental context. Our data suggest that altering the activation state of NK cells may contribute to immunologic control of lung and possibly other cancers.

Multi-species genetic connectivity in a terrestrial habitat network.

BACKGROUND: Habitat fragmentation reduces genetic connectivity for multiple species, yet conservation efforts tend to rely heavily on single-species connectivity estimates to inform land-use planning. Such conservation activities may benefit from multi-species connectivity estimates, which provide a simple and practical means to mitigate the effects of habitat fragmentation for a larger number of species. To test the validity of a multi-species connectivity model, we used neutral microsatellite genetic datasets of Canada lynx (Lynx canadensis), American marten (Martes americana), fisher (Pekania pennanti), and southern flying squirrel (Glaucomys volans) to evaluate multi-species genetic connectivity across Ontario, Canada. RESULTS: We used linear models to compare node-based estimates of genetic connectivity for each species to point-based estimates of landscape connectivity (current density) derived from circuit theory. To our knowledge, we are the first to evaluate current density as a measure of genetic connectivity. Our results depended on landscape context: habitat amount was more important than current density in explaining multi-species genetic connectivity in the northern part of our study area, where habitat was abundant and fragmentation was low. In the south however, where fragmentation was prevalent, genetic connectivity was correlated with current density. Contrary to our expectations however, locations with a high probability of movement as reflected by high current density were negatively associated with gene flow. Subsequent analyses of circuit theory outputs showed that high current density was also associated with high effective resistance, underscoring that the presence of pinch points is not necessarily indicative of gene flow. CONCLUSIONS: Overall, our study appears to provide support for the hypothesis that landscape pattern is important when habitat amount is low. We also conclude that while current density is proportional to the probability of movement per unit area, this does not imply increased gene flow, since high current density tends to be a result of neighbouring pixels with high cost of movement (e.g., low habitat amount). In other words, pinch points with high current density appear to constrict gene flow.

Genome-Wide Exome Analysis of Cmv5-Disparate Mouse Strains that Differ in Host Resistance to Murine Cytomegalovirus Infection.

Host resistance to murine cytomegalovirus (MCMV) varies in different strains of laboratory mice due to differences in expression of determinants that control and clear viral infection. The major histocompatibility complex class I Dk molecule is one such determinant that controls MCMV through the action of natural killer (NK) cells. However, the extent of NK cell-mediated Dk-dependent resistance to infection varies in different mouse strains. The molecular genetic basis of this variation remains unclear. Previous work to examine the Dk effect on MCMV resistance in MA/My × C57L offspring discovered multiple quantitative trait loci (QTL) that may serve to modify NK cells or their capacity to respond during MCMV infection. One QTL in particular, Cmv5, was found to regulate the frequency of NK cells and secondary lymphoid organ structure in spleen during MCMV infection. Cmv5 alleles, however, have not been identified. We therefore sequenced and analyzed genome-wide exome (GWE) variants, including those aligned to the critical genetic interval, in Cmv5-disparate mouse strains. Their GWE variant profiles were compared to assess strain-specific sequence data integrity and to analyze mouse strain relatedness across the genome. GWE content was further compared against data from the Mouse Genomes Project. This approach was developed as a platform for using GWE variants to define genomic regions of divergence and similarity in different mouse strains while also validating the overall quality of GWE sequence data. Moreover, the analysis provides a framework for the selection of novel QTL candidate sequences, including at the Cmv5 critical region.