CD4 T Cell Immunity Is Critical for the Control of Simian Varicella Virus Infection in a Nonhuman Primate Model of VZV Infection

Primary infection with varicella zoster virus (VZV) results in varicella (more commonly known as chickenpox) after which VZV establishes latency in sensory ganglia. VZV can reactivate to cause herpes zoster (shingles), a debilitating disease that affects one million individuals in the US alone annually. Current vaccines against varicella (Varivax) and herpes zoster (Zostavax) are not 100% efficacious. Specifically, studies have shown that 1 dose of varivax can lead to breakthrough varicella, albeit rarely, in children and a 2-dose regimen is now recommended. Similarly, although Zostavax results in a 50% reduction in HZ cases, a significant number of recipients remain at risk. To design more efficacious vaccines, we need a better understanding of the immune response to VZV. Clinical observations suggest that T cell immunity plays a more critical role in the protection against VZV primary infection and reactivation. However, no studies to date have directly tested this hypothesis due to the scarcity of animal models that recapitulate the immune response to VZV. We have recently shown that SVV infection of rhesus macaques models the hallmarks of primary VZV infection in children. In this study, we used this model to experimentally determine the role of CD4, CD8 and B cell responses in the resolution of primary SVV infection in unvaccinated animals. Data presented in this manuscript show that while CD20 depletion leads to a significant delay and decrease in the antibody response to SVV, loss of B cells does not alter the severity of varicella or the kinetics/magnitude of the T cell response. Loss of CD8 T cells resulted in slightly higher viral loads and prolonged viremia. In contrast, CD4 depletion led to higher viral loads, prolonged viremia and disseminated varicella. CD4 depleted animals also had delayed and reduced antibody and CD8 T cell responses. These results are similar to clinical observations that children with agammaglobulinemia have uncomplicated varicella whereas children with T cell deficiencies are at increased risk of progressive varicella with significant complications. Moreover, our studies indicate that CD4 T cell responses to SVV play a more critical role than antibody or CD8 T cell responses in the control of primary SVV infection and suggest that one potential mechanism for enhancing the efficacy of VZV vaccines is by eliciting robust CD4 T cell responses

Comparisons between Tethyan Anorthosite-bearing Ophiolites and Archean Anorthosite-bearing Layered Intrusions: Implications for Archean Geodynamic Processes

Elucidating the petrogenesis and geodynamic setting(s) of anorthosites in Archean layered intrusions and Tethyan ophiolites has significant implications for crustal evolution and growth throughout Earth history. Archean anorthosite-bearing layered intrusions occur on every continent. Tethyan ophiolites occur in Europe, Africa, and Asia. In this contribution, the field, petrographic, petrological, and geochemical characteristics of 100 Tethyan anorthosite-bearing ophiolites and 155 Archean anorthosite-bearing layered intrusions are compared. Tethyan anorthosite-bearing ophiolites range from Devonian to Paleocene in age, are variably composite, contain anorthosites with highly calcic (An44-100) plagioclase and magmatic amphibole. These ophiolites formed predominantly at convergent plate margins, with some forming in mid-ocean ridge, continental rift, and mantle plume settings. The predominantly convergent plate margin tectonic setting of Tethyan anorthosite-bearing ophiolites is indicated by negative Nb and Ti anomalies and magmatic amphibole. Archean anorthosite-bearing layered intrusions are Eoarchean to Neoarchean in age, have megacrystic anorthosites with highly calcic (An20-100) plagioclase and magmatic amphibole and are interlayered with gabbros and leucogabbros and intrude pillow basalts. These Archean layered intrusions are interpreted to have predominantly formed at convergent plate margins, with the remainder forming in mantle plume, continental rift, oceanic plateau, post-orogenic, anorogenic, mid-ocean ridge, and passive continental margin settings. These layered intrusions predominantly crystallized from hydrous Ca- and Al-rich tholeiitic magmas. The field, petrographic and geochemical similarities between Archean and Tethyan anorthosites indicate that they were produced by similar geodynamic processes mainly in suprasubduction zone settings. We suggest that Archean anorthosite-bearing layered intrusions and spatially associated greenstone belts represent dismembered subduction-related Archean ophiolites

3D Reconstruction of VZV Infected Cell Nuclei and PML Nuclear Cages by Serial Section Array Scanning Electron Microscopy and Electron Tomography

Varicella-zoster virus (VZV) is a human alphaherpesvirus that causes varicella (chickenpox) and herpes zoster (shingles). Like all herpesviruses, the VZV DNA genome is replicated in the nucleus and packaged into nucleocapsids that must egress across the nuclear membrane for incorporation into virus particles in the cytoplasm. Our recent work showed that VZV nucleocapsids are sequestered in nuclear cages formed from promyelocytic leukemia protein (PML) in vitro and in human dorsal root ganglia and skin xenografts in vivo. We sought a method to determine the three-dimensional (3D) distribution of nucleocapsids in the nuclei of herpesvirus-infected cells as well as the 3D shape, volume and ultrastructure of these unique PML subnuclear domains. Here we report the development of a novel 3D imaging and reconstruction strategy that we term Serial Section Array-Scanning Electron Microscopy (SSA-SEM) and its application to the analysis of VZV-infected cells and these nuclear PML cages. We show that SSA-SEM permits large volume imaging and 3D reconstruction at a resolution sufficient to localize, count and distinguish different types of VZV nucleocapsids and to visualize complete PML cages. This method allowed a quantitative determination of how many nucleocapsids can be sequestered within individual PML cages (sequestration capacity), what proportion of nucleocapsids are entrapped in single nuclei (sequestration efficiency) and revealed the ultrastructural detail of the PML cages. More than 98% of all nucleocapsids in reconstructed nuclear volumes were contained in PML cages and single PML cages sequestered up to 2,780 nucleocapsids, which were shown by electron tomography to be embedded and cross-linked by an filamentous electron-dense meshwork within these unique subnuclear domains. This SSA-SEM analysis extends our recent characterization of PML cages and provides a proof of concept for this new strategy to investigate events during virion assembly at the single cell level

Two novel human cytomegalovirus NK cell evasion functions target MICA for lysosomal degradation

NKG2D plays a major role in controlling immune responses through the regulation of natural killer (NK) cells, αβ and γδ T-cell function. This activating receptor recognizes eight distinct ligands (the MHC Class I polypeptide-related sequences (MIC) A andB, and UL16-binding proteins (ULBP)1–6) induced by cellular stress to promote recognition cells perturbed by malignant transformation or microbial infection. Studies into human cytomegalovirus (HCMV) have aided both the identification and characterization of NKG2D ligands (NKG2DLs). HCMV immediate early (IE) gene up regulates NKGDLs, and we now describe the differential activation of ULBP2 and MICA/B by IE1 and IE2 respectively. Despite activation by IE functions, HCMV effectively suppressed cell surface expression of NKGDLs through both the early and late phases of infection. The immune evasion functions UL16, UL142, and microRNA(miR)-UL112 are known to target NKG2DLs. While infection with a UL16 deletion mutant caused the expected increase in MICB and ULBP2 cell surface expression, deletion of UL142 did not have a similar impact on its target, MICA. We therefore performed a systematic screen of the viral genome to search of addition functions that targeted MICA. US18 and US20 were identified as novel NK cell evasion functions capable of acting independently to promote MICA degradation by lysosomal degradation. The most dramatic effect on MICA expression was achieved when US18 and US20 acted in concert. US18 and US20 are the first members of the US12 gene family to have been assigned a function. The US12 family has 10 members encoded sequentially through US12–US21; a genetic arrangement, which is suggestive of an ‘accordion’ expansion of an ancestral gene in response to a selective pressure. This expansion must have be an ancient event as the whole family is conserved across simian cytomegaloviruses from old world monkeys. The evolutionary benefit bestowed by the combinatorial effect of US18 and US20 on MICA may have contributed to sustaining the US12 gene family

Modelling the Effects of Population Structure on Childhood Disease: The Case of Varicella

Realistic, individual-based models based on detailed census data are increasingly used to study disease transmission. Whether the rich structure of such models improves predictions is debated. This is studied here for the spread of varicella, a childhood disease, in a realistic population of children where infection occurs in the household, at school, or in the community at large. A methodology is first presented for simulating households with births and aging. Transmission probabilities were fitted for schools and community, which reproduced the overall cumulative incidence of varicella over the age range of 0–11 years old

Specialized Feed Induced Growth, Nuchal Hump Development and Histological Changes in Flowerhorn (Cichlosoma citrinella)

Ornamental fish trade is a multibillion-dollar industry worldwide. One of the highly-demanded ornamental fish is the bumpy head flowerhorn (FH). In spite of its popularity, scientific studies on the FH nuchal hump (NH) development are considerably limited. This inferential study aimed to determine the growth, development, and NH histology of FH treated with conventional and specialized feeds for 17 weeks. Two groups consisting of 12 fish were randomly selected and regularly observed. Quantitative and qualitative data were subjected to T-test and histological analysis, respectively. Results showed a significant increase in body length of the treated group on the fourth – seventh weeks (p= 0.04-0.01).  Likewise, the significant increase in weight started in the third week (p= 0.03-0.001) onwards. Nuchal humps’ vertical and horizontal components significantly increased at seventh week (p= 0.04) and fourteenth week (p=0.04), respectively. Histological analysis revealed greater fat deposits under stratum compactum, thicker dermis (203.58 μm), and bigger lateral line canal (84.4 μm). This study demonstrated that the specialized feed significantly contributed to the growth, nuchal development, and increase fat deposits in flower horn. Results provide vital information on the biology of flowerhorn and fish industry. Further study on the histological changes of other organs is recommended

Disruption of PML Nuclear Bodies Is Mediated by ORF61 SUMO-Interacting Motifs and Required for Varicella-Zoster Virus Pathogenesis in Skin

Promyelocytic leukemia protein (PML) has antiviral functions and many viruses encode gene products that disrupt PML nuclear bodies (PML NBs). However, evidence of the relevance of PML NB modification for viral pathogenesis is limited and little is known about viral gene functions required for PML NB disruption in infected cells in vivo. Varicella-zoster virus (VZV) is a human alphaherpesvirus that causes cutaneous lesions during primary and recurrent infection. Here we show that VZV disrupts PML NBs in infected cells in human skin xenografts in SCID mice and that the disruption is achieved by open reading frame 61 (ORF61) protein via its SUMO-interacting motifs (SIMs). Three conserved SIMs mediated ORF61 binding to SUMO1 and were required for ORF61 association with and disruption of PML NBs. Mutation of the ORF61 SIMs in the VZV genome showed that these motifs were necessary for PML NB dispersal in VZV-infected cells in vitro. In vivo, PML NBs were highly abundant, especially in basal layer cells of uninfected skin, whereas their frequency was significantly decreased in VZV-infected cells. In contrast, mutation of the ORF61 SIMs reduced ORF61 association with PML NBs, most PML NBs remained intact and importantly, viral replication in skin was severely impaired. The ORF61 SIM mutant virus failed to cause the typical VZV lesions that penetrate across the basement membrane into the dermis and viral spread in the epidermis was limited. These experiments indicate that VZV pathogenesis in skin depends upon the ORF61-mediated disruption of PML NBs and that the ORF61 SUMO-binding function is necessary for this effect. More broadly, our study elucidates the importance of PML NBs for the innate control of a viral pathogen during infection of differentiated cells within their tissue microenvironment in vivo and the requirement for a viral protein with SUMO-binding capacity to counteract this intrinsic barrier

Characteristics of the National Applicant Pool for Clinical Informatics Fellowships (2016-2017)

We conducted a national study to assess the numbers and diversity of applicants for 2016 and 2017 clinical informatics fellowship positions. In each year, we collected data on the number of applications that programs received from candidates who were ultimately successful vs. unsuccessful. In 2017, we also conducted an anonymous applicant survey. Successful candidates applied to an average of 4.2 and 5.5 programs for 2016 and 2017, respectively. In the survey, unsuccessful candidates reported applying to fewer programs. Assuming unsuccessful candidates submitted between 2-5 applications each, the total applicant pool numbered 42-69 for 2016 (competing for 24 positions) and 52-85 for 2017 (competing for 30 positions). Among survey respondents (n=33), 24% were female, 1 was black and none were Hispanic. We conclude that greater efforts are needed to enhance interest in clinical informatics among medical students and residents, particularly among women and members of underrepresented minority groups