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Supplementary Figure 1 from Antiestrogens Induce Transforming Growth Factor β–Mediated Immunosuppression in Breast Cancer

A small percentage of bladder cancers in the general population have been found to harbor DNA viruses. In contrast, up to 25% of tumors of solid organ transplant recipients, who are at an increased risk of developing bladder cancer and have overall poorer outcome, harbor BK polyomavirus (BKPyV). To better understand the biology of the tumors and the mechanisms of carcinogenesis from potential oncoviruses, we performed whole genome and transcriptome sequencing on bladder cancer specimens from 43 transplant patients. Nearly half of tumors from this patient population contained viral sequences. The most common were from BKPyV (N=9, 21%), JC polyomavirus (N=7, 16%), carcinogenic human papillomaviruses (N=3, 7%), and torque teno viruses (N=5, 12%). Immunohistochemistry revealed variable Large T antigen expression in BKPyV-positive tumors ranging from 100% positive staining of tumor tissue to less than 1%. In most cases of BKPyV-positive tumors, the viral genome appeared to be clonally integrated into the host chromosome consistent with microhomology-mediated end joining and coincided with focal amplifications of the tumor genome similar to other virus-mediated cancers. Significant changes in host gene expression consistent with the functions of BKPyV Large T antigen were also observed in these tumors. Lastly, we identified four mutation signatures in our cases with those attributable to APOBEC3 and SBS5 being the most abundant. Mutation signatures associated with the antiviral drug, ganciclovir, and aristolochic acid, a nephrotoxic compound found in some herbal medicines, were also observed. The results suggest multiple pathways to carcinogenesis in solid organ transplant recipients with a large fraction being virus-associated.Author SummarySolid organ transplant recipients are at a significantly increases risk for developing bladder cancer compared to the general population, suggesting a potential infectious origin to these tumors. This study identifies that BK polyomavirus, JC polyomavirus, human papillomaviruses, and anelloviruses are commonly found in bladder tumors of solid organ transplant recipients. In most cases when detected, BK polyomavirus is integrated into the tumor genome and associates with genomic structural changes and distinct gene expression through the activity of viral oncogenes. Additionally, mutational signature analysis suggests that a subset of tumors of solid organ transplant recipients develop through distinct mutagenic processes compared to the general population. Together these results indicate multiple distinct mechanisms of carcinogenesis in bladder cancers of solid organ transplant recipients that may have implications for prevention, treatment, and outcome.

The sense of smell has potent effects on appetite, but the underlying neural mechanisms are largely a mystery. The hypothalamic arcuate nucleus contains two subsets of neurons linked to appetite: AgRP (agouti-related peptide) neurons, which enhance appetite, and POMC (pro-opiomelanocortin) neurons, which suppress appetite. Here, we find that AgRP and POMC neurons receive indirect inputs from partially overlapping areas of the olfactory cortex, thus identifying their sources of odor signals. We also find neurons directly upstream of AgRP or POMC neurons in numerous other areas, identifying potential relays between the olfactory cortex and AgRP or POMC neurons. Transcriptome profiling of individual AgRP neurons reveals differential expression of receptors for multiple neuromodulators. Notably, known ligands of the receptors define subsets of neurons directly upstream of AgRP neurons in specific brain areas. Together, these findings indicate that higher olfactory areas can differentially influence AgRP and POMC appetite neurons, that subsets of AgRP neurons can be regulated by different neuromodulators, and that subsets of neurons upstream of AgRP neurons in specific brain areas use different neuromodulators, together or in distinct combinations to modulate AgRP neurons and thus appetite.

Fish papillomaviruses form a newly discovered group broadly recognized as the Secondpapillomavirinae subfamily. This study expands the documented genomes of the fish papillomaviruses from six to 16, including one from the Antarctic emerald notothen, seven from commercial market fishes, one from data mining of sea bream sequence data, and one from a western gull cloacal swab that is likely diet derived. The genomes of secondpapillomaviruses are ∼6 kilobasepairs (kb), which is substantially smaller than the ∼8 kb of terrestrial vertebrate papillomaviruses. Each genome encodes a clear homolog of the four canonical papillomavirus genes, E1, E2, L1, and L2. In addition, we identified open reading frames (ORFs) with short linear peptide motifs reminiscent of E6/E7 oncoproteins. Fish papillomaviruses are extremely diverse and phylogenetically distant from other papillomaviruses suggesting a model in which terrestrial vertebrate-infecting papillomaviruses arose after an evolutionary bottleneck event, possibly during the water-to-land transition.

Polyomaviruses are non–enveloped viruses with circular double-stranded DNA genomes that range in size from ∼4–7 kilobasepairs. Initially identified in mammals, polyomaviruses have now been identified in birds and a few fish species. Although fragmentary polyomavirus-like sequences have been detected as apparent ‘hitchhikers’ in shotgun genomics datasets for various arthropods, the possible diversity of these viruses in invertebrates remains unclear. In general, polyomaviruses are host-specific, showing strong evidence of host-virus co–evolution. Identification of polyomaviruses in a broader range of animals could shed useful light on the evolutionary history of this medically important group of viruses. Scorpions are predatory arachnids that are among the oldest terrestrial animals. Thus far, viromes of arachnids have been under–sampled and understudied. Here, high–throughput sequencing and traditional molecular techniques were used to explore the diversity of circular DNA viruses associated with bark scorpions (Centruroides sculpturatus) from the greater Phoenix area, Arizona, USA. The complete genomes of eight novel polyomaviruses were identified. Analysis of Centruroides transcriptomic datasets elucidated the splicing of the viral late gene array, which is more complex than that of vertebrate polyomaviruses. Phylogenetic analysis provides further evidence of co-divergence of polyomaviruses with their hosts, suggesting that at least one ancestral species of polyomaviruses was circulating amongst the primitive common ancestors of arthropods and chordates.

In 2019, there were about 100,000 kidney transplants globally, with more than a quarter of them performed in the United States. Unfortunately, some engrafted organs are lost to polyomavirus-associated nephropathy (PyVAN) caused by BK and JC viruses (BKPyV and JCPyV). Transplant patients are routinely monitored for BKPyV viremia, which is an accepted hallmark of nascent nephropathy. If viremia is detected, a reduction in immunosuppressive therapy is standard care, but the intervention comes with increased risk of immune rejection of the engrafted organ. Recent reports have suggested that transplant recipients with high levels of polyomavirus-neutralizing antibodies are protected against PyVAN. Virus-like particle (VLP) vaccines, similar to approved human papillomaviruses vaccines, have an excellent safety record and are known to induce high levels of neutralizing antibodies associated and long-lasting protection from infection. In this study, we demonstrate that VLPs representing BKPyV genotypes I, II, and IV, as well as JCPyV genotype 2 produced in insect cells elicit robust antibody titers. In rhesus macaques, all monkeys developed neutralizing antibody titers above a previously proposed protective threshold of 10,000. A second inoculation, administered 19 weeks after priming, boosted titers to a plateau of ≥25,000 that was maintained for almost two years. No vaccine-related adverse events were observed in any macaques. A multivalent BK/JC VLP immunogen did not show inferiority compared to the single-genotype VLP immunogens. Considering these encouraging results, we believe a clinical trial administering the multivalent VLP vaccine in patients waiting to receive a kidney transplant is warranted to evaluate its ability to reduce or eliminate PyVAN.HIGHLIGHTSRecombinant virus-like particle vaccine was safely administered to rhesus macaquesVaccination generated high-titer neutralizing antibody responsesMultivalent BK/JC polyomavirus vaccine was as effective as monovalent vaccinesHigh neutralizing titers were sustained for 92 weeks without appreciable decline

Polyomaviruses are oncogenic viruses that are generally thought to have co-evolved with their hosts. While primate and rodent polyomaviruses are increasingly well-studied, less is known about polyomaviruses that infect other mammals. In an effort to gain insight into polyomaviruses associated with carnivores, we surveyed fecal samples collected in the USA from bobcats (Lynx rufus), pumas (Puma concolor), Canada lynxes (Lynx canadensis), and grizzly bears (Ursus arctos). Using a viral metagenomic approach, we identified six novel polyomavirus genomes. Surprisingly, four of the six genomes showed a phylogenetic relationship to polyomaviruses found in prey animals. These included a putative rabbit polyomavirus from a bobcat fecal sample and two possible deer-trophic polyomaviruses from Canada lynx feces. One polyomavirus found in a grizzly bear sample was found to be phylogenetically distant from previously identified polyomaviruses. Further analysis of the grizzly bear fecal sample showed that it contained anelloviruses that are known to infect pigs, suggesting that the bear might have preyed on a wild or domestic pig. Interestingly, a polyomavirus genome identified in a puma fecal sample was found to be closely related both to raccoon polyomavirus 1 and to Lyon-IARC polyomavirus, the latter of which was originally identified in human saliva and skin swab specimens but has since been found in samples from domestic cats (Felis catus).

Scents have been employed for millennia to allay stress, but whether or how they might do so is largely unknown. Fear and stress induce increases in blood stress hormones controlled by hypothalamic corticotropin releasing hormone neurons (CRHNs). Here, we report that two common odorants block mouse stress hormone responses to three potent stressors: physical restraint, predator odor, and male–male social confrontation. One odorant inhibits restraint and predator odor activation of excitatory neurons upstream of CRHNs in the bed nucleus of the stria terminalis (BNSTa). In addition, both activate inhibitory neurons upstream of CRHNs in the hypothalamic ventromedial nucleus (VMH) and silencing of VMH inhibitory neurons hinders odor blocking of stress. Together, these findings indicate that odor blocking can occur via two mechanisms: (1) Inhibition of excitatory neurons that transmit stress signals to CRHNs and (2) activation of inhibitory neurons that act directly or indirectly to inhibit stressor activation of CRHNs.

Agricultural communication (AGCOM) has been known to aid in disseminating research-based agricultural information among Malawian farmers. In 1958 the Malawi Government, via the Ministry of Agriculture, established the Agricultural Communications Branch (ACB) in an attempt to increase access to and adoption of scientifically-proven technologies among farmers. Moreover, in 2000 the Malawi Government started implementing an agricultural extension policy that promoted pluralistic demand-driven extension, which led to the increased availability of non-governmental organizations providing AGCOM services to farmers. However, after several decades of using different communication tools to promote new technologies, low productivity in most small holder farms remains a challenge, with limited adoption of improved technology as one of the contributing factors. In this exploratory, convergent, mixed methods study, 30 Malawian farmers and six AGCOM officers who were selected using convenient and snow ball sampling respectively participated in key informant interviews. In addition, 64 AGCOM officers who were selected using simple random sampling were involved in a survey. The findings of the study revealed that information delivered to farmers does not address farmer needs in most cases. Specifically, existing policies, source and availability of funding, and the agricultural calendar influenced choice of information that was disseminated. For example, the existing policy does not allow AGCOM officers to disseminate local and innovative farmer practices unless they are tested and approved by scientists. Such policies, perpetuates a mindset among farmers that innovations originate from outside their communities, thereby making it hard for them to share their local, indigenous ideas with their colleagues. Moreover, it has contributed to the inability of AGCOM to be used as an innovation creation tool, hence AGCOMs’ limited impact. Keywords: adoption, improved technologies, agricultural communication, increased productivity, information dissemination

The mouse brain contains ~100 million neurons interconnected in a vast array of neural circuits. The identities and functions of individual neuronal components of most circuits are undefined. Here we describe a method, termed ‘Connect-seq’, which combines retrograde viral tracing and single cell transcriptomics to uncover the molecular identities of upstream neurons in a specific circuit and the signaling molecules they use to communicate. Connect-seq can generate a molecular map that can be superimposed on a neuroanatomical map to permit molecular and genetic interrogation of how the neuronal components of a circuit control its function. Application of this method to hypothalamic neurons controlling physiological responses to fear and stress reveal subsets of upstream neurons that express diverse constellations of signaling molecules and can be distinguished by their anatomical locations.

In 2014, the International Agency for Research on Cancer judged Merkel cell polyomavirus (MCPyV) to be a probable human carcinogen. BK polyomavirus (BKPyV, a distant cousin of MCPyV) was ruled a possible carcinogen. In this review, we argue that it has recently become reasonable to view both of these viruses as known human carcinogens. In particular, several complementary lines of evidence support a causal role for BKPyV in the development of bladder carcinomas affecting organ transplant patients. The expansion of inexpensive deep sequencing has opened new approaches to investigating the important question of whether BKPyV causes urinary tract cancers in the general population.

Polintons (also known as Mavericks) were initially identified as a widespread class of eukaryotic transposons named for their hallmark type B DNA polymerase and retrovirus-like integrase genes. It has since been recognized that many polintons encode possible capsid proteins and viral genome-packaging ATPases similar to those of a diverse range of double-stranded DNA viruses. This supports the inference that at least some polintons are actually viruses capable of cell-to-cell spread. At present, there are no polinton-associated capsid protein genes annotated in public sequence databases. To rectify this deficiency, we used a data-mining approach to investigate the distribution and gene content of polinton-like elements and related DNA viruses in animal genomic and metagenomic sequence datasets. The results define a discrete family-like clade of viruses with two genus-level divisions. We propose the family name Adintoviridae, connoting similarities to adenovirus virion proteins and the presence of a retrovirus-like integrase gene. Although adintovirus-class PolB sequences were detected in datasets for fungi and various unicellular eukaryotes, sequences resembling adintovirus virion proteins and accessory genes appear to be restricted to animals. Degraded adintovirus sequences are endogenized into the germlines of a wide range of animals, including humans.

Human papillomavirus (HPV) infections underlie a wide spectrum of both benign and malignant epithelial diseases. In this report, we describe the case of a young man who had encephalitis caused by herpes simplex virus during adolescence and currently presented with multiple recurrent skin and mucosal lesions caused by HPV. The patient was found to have a pathogenic germline mutation in the X-linked interleukin-2 receptor subunit gamma gene (IL2RG), which was somatically reverted in T cells but not in natural killer (NK) cells. Allogeneic hematopoietic-cell transplantation led to restoration of NK cytotoxicity, with normalization of the skin microbiome and persistent remission of all HPV-related diseases. NK cytotoxicity appears to play a role in containing HPV colonization and the ensuing HPV-related hyperplastic or dysplastic lesions. (Funded by the National Institutes of Health and the Herbert Irving Comprehensive Cancer Center Flow Cytometry Shared Resources.).

This exploratory qualitative study sought to gain initial insights into how farmers involved in dif­ferent production practices communicate with consumers. A thematic analysis of in-depth inter­views conducted with eight organic and 12 con­ventional farmers in Ohio indicated that organic farmers are proactive in communicating with the public about their production practices, unlike conventional farmers, who focus on improving productivity. Furthermore, the organic farmers reported using different communication channels such as Facebook, flyers, and YouTube when com­municating with consumers, while conventional farmers reported being busy working on their farms and not having time to communicate with consumers. Organic farmers’ involvement in com­munication activities with the public about their production practices and products was reported to stem from their beliefs and values toward sustain­able farming practices and environmental conser­vation. Furthermore, unlike conventional farmers, most organic farmers sold their produce directly to consumers, and as such, used communication as a marketing tool. The active involvement of organic farmers in communicating with consumers may be attributable in part to increased media coverage about the benefits of organic farming practices. On the other hand, limited involvement of conven­tional farmers in communicating with the public may be partially attributable to limited media cov­erage about the benefits of conventional farming. Therefore, to ensure that consumers make in­formed decisions, there is a need to start develop­ing standalone communication organizations and interventions committed to providing unbiased information about the benefits and disadvantages of the different farming practices.

Significance Mechanisms of many human chronic diseases involve abnormal action of the immune system and/or altered metabolism. The microbiome, an important regulator of metabolic and immune-related phenotypes, has been shown to be associated with or participate in the development of a variety of chronic diseases. Viruses of bacteria (i.e., “phages”) are ubiquitous and mysterious, and several studies have shown that phages exert great control over the behavior—and misbehavior—of their host bacteria. This study uses techniques to discover and analyze over 45,000 viruses associated with human bodies. The abundance of over 2,000 specific phages is found to correlate with a variety of common chronic diseases., Despite remarkable strides in microbiome research, the viral component of the microbiome has generally presented a more challenging target than the bacteriome. This gap persists, even though many thousands of shotgun sequencing runs from human metagenomic samples exist in public databases, and all of them encompass large amounts of viral sequence data. The lack of a comprehensive database for human-associated viruses has historically stymied efforts to interrogate the impact of the virome on human health. This study probes thousands of datasets to uncover sequences from over 45,000 unique virus taxa, with historically high per-genome completeness. Large publicly available case-control studies are reanalyzed, and over 2,200 strong virus–disease associations are found.

The effects of postemergence (POST) herbicides off-target movement on cotton has been evaluated, but no data is available evaluating simulated off-target movement of residual herbicides. Therefore, low-dose POST applications of flumioxazin were evaluated in cotton at the cotyledon, two- and four-leaf growth stages. Rates evaluated were 12.5, 25, and 50% of the labeled use rate of 72 g ai ha-1. Necrosis, cotton height and width reduction was observed. Cotyledon cotton was injured 69 to 86%, 80 to 91%, and 84 to 97% following the 12.5, 25, and 50% flumioxazin rates, respectively, 3 through 42 DAT. Injury of two-leaf cotton increased from 3 to 14 DAT for all flumioxazin rates with maximum injury of 40, 47, and 58% following the 12.5, 25, and 50% rates, respectively, 14 DAT, but injury decreased following the 14 DAT evaluation. Injury of four-leaf cotton was 46 to 58% 3 DAT and decreased over time regardless of rate. At 42 DAT, two- and four-leaf cotton was injured 14 to 33% and increased with flumioxazin rate. Cotton height and width averaged 40, 80, 86% of the nontreated following the cotyledon, two-, and four-leaf application timings, respectively, 42 DAT. In addition, height was more influenced by flumioxazin rate than cotton width. Yields were 24, 52, and 62% of the nontreated following the cotyledon, two-, and four-leaf applications timings, respectively. In addition, yields following the 12.5, 25, and 50% rates were 53, 45, and 40% of the nontreated. Low-doses of flumioxazin reduced revenue $1,172 to $2,344 ha-1 for lint and $212 to 423 ha-1 for cotton seed. Low-doses of flumioxazin POST can have negative effects on cotton growth and yield and could cause severe economic loss for a cotton producer.

All herbicides will move off-target to sensitive crops when not applied correctly. Therefore, low-dose applications of flumioxazin and metribuzin were evaluated in soybean at the unifoliate, V2, and V4 growth stages. Rates evaluated were 12.5%, 25%, and 50% of the labeled use rates of 72 and 316 g ai ha−1 of flumioxazin and metribuzin, respectively. Flumioxazin injury was characterized by necrosis and visible height and width reduction. Injury increased with rate 3 d after treatment (DAT), with unifoliate, V2, and V4 soybean injured 15% to 30%, 18% to 27%, and 5% to 8%, respectively. Unifoliate and V4 soybean were injured more than V4 soybean 3 to 14 DAT, but injury decreased to −1 and a revenue loss of 71 to 141 US$ ha−1. Metribuzin injury was primarily chlorosis with necrosis and a visible reduction in soybean height and width. Soybean at the V2 growth stage was injured 14% more than V4 soybean 3 DAT, regardless of metribuzin rate. Injury to V2 and V4 soybean was similar 14 DAT, with injury of 21% to 40% across rates. Soybean injury when treated at the V2 and V4 growth stages was 6% to 29% 42 DAT compared to unifoliate soybean at 0 to 17%. Soybean yields in the metribuzin study yields were 96% to 98% of the nontreated. However, a 2% to 4% reduction equates to a loss of 90 to 180 kg ha−1 and a revenue loss of 32 to 65 US$ ha−1. Unifoliate and V2 soybean are more sensitive to a low dose of flumioxazin POST, and V2 and V4 soybean are more sensitive to a low dose of metribuzin POST. Injury and the impact on soybean growth could potentially cause economic loss for a soybean producer.

Circular RNAs (circRNAs) are a conserved class of RNAs with diverse functions, including serving as messenger RNAs that are translated into peptides. Here we describe circular RNAs generated by human polyomaviruses (HPyVs), some of which encode variants of the previously described alternative large T antigen open reading frame (ALTO) protein. Circular ALTO RNAs (circALTOs) can be detected in virus positive Merkel cell carcinoma (VP-MCC) cell lines and tumor samples. CircALTOs are stable, predominantly located in the cytoplasm, and N6-methyladenosine (m6A) modified. The translation of MCPyV circALTOs into ALTO protein is negatively regulated by MCPyV-generated miRNAs in cultured cells. MCPyV ALTO expression increases transcription from some recombinant promoters in vitro and upregulates the expression of multiple genes previously implicated in MCPyV pathogenesis. MCPyV circALTOs are enriched in exosomes derived from VP-MCC lines and circALTO-transfected 293T cells, and purified exosomes can mediate ALTO expression and transcriptional activation in MCPyV-negative cells. The related trichodysplasia spinulosa polyomavirus (TSPyV) also expresses a circALTO that can be detected in infected tissues and produces ALTO protein in cultured cells. Thus, human polyomavirus circRNAs are expressed in human tumors and infected tissues and express proteins that have the potential to modulate the infectious and tumorigenic properties of these viruses., Author summary Human polyomaviruses (HPyV) have been linked to diseases including Merkel cell carcinoma (MCC), a skin cancer caused by Merkel cell polyomavirus (MCPyV), and skin and hair eruptions caused by trichodysplasia spinulosa polyomavirus (TSPyV). We discover that HPyVs generate single-stranded, circular RNAs (circRNAs) in affected tissues. Both MCPyV and TSPyV generate circRNAs encompassing part of the early region (circALTOs) that can be translated to the alternative large T antigen open reading frame (ALTO) protein in cultured cells. Previously described microRNAs (miRNAs) generated by MCPyV have the ability to inhibit the expression of ALTO. MCPyV circALTOs are enriched in extracellular vesicles produced by MCC cell lines and cells that express circALTO. Finally, we discover that MCPyV ALTO protein enhances transcription from some recombinant promoters and promotes the expression of multiple host cell genes previously implicated in MCPyV pathogenesis. Thus, HPyVs generate circRNAs, including circALTOs that are translated to ALTO protein, and MCPyV ALTO has the ability to regulate transcription.

Scents have been employed for millennia to allay fear and stress, but whether they do so is poorly understood. In response to fear and stress, hypothalamic corticotropin releasing hormone neurons (CRHNs) induce increases in blood stress hormones. Here, we find that certain structurally and perceptually dissimilar odorants can block mouse stress hormone responses to three potent stressors: physical restraint, predator odor, and male-male social confrontation. Both odorants activate GABAergic inhibitory neurons presynaptic to CRHNs in the hypothalamic ventromedial nucleus (VMH). Stimulation of those neurons inhibits restraint-induced activation of CRHNs and stress hormone increase, mimicking a blocking odorant. Conversely, their silencing prevents odorant blocking of both responses. Notably, we also observed odor blocking of stressor activation in neurons presynaptic to CRHNs in the bed nucleus of the stria terminalis (BNST). Together, these findings indicate that selected odorants can indeed block stress responses, and that odor blocking can occur via two routes: a direct route in which blocking odor signals directly inhibit CRHNs and an indirect route in which they inhibit stressor activation of neurons presynaptic to CRHNs and prevent them from transmitting stress signals to CRHNs.

An unusual feature of papillomaviruses is that their genomes are packaged into virions along with host histones. Viral minichromosomes were visualized as "beads on a string" by electron microscopy in the 1970s but, to date, little is known about the posttranslational modifications of these histones. To investigate this, we analyzed the histone modifications in HPV16/18 quasivirions, wart-derived bovine papillomavirus (BPV1), and wart-derived human papillomavirus type 1 (HPV1) using quantitative mass spectrometry. The chromatin from all three virion samples had abundant posttranslational modifications (acetylation, methylation, and phosphorylation). These histone modifications were verified by acid urea polyacrylamide electrophoresis and immunoblot analysis. Compared to matched host cell controls, the virion minichromosome was enriched in histone modifications associated with active chromatin and depleted for those commonly found in repressed chromatin. We propose that the viral minichromosome acquires specific histone modifications late in infection that are coupled to the mechanisms of viral replication, late gene expression, and encapsidation. We predict that, in turn, these same modifications benefit early stages of infection by helping to evade detection, promoting localization of the viral chromosome to beneficial regions of the nucleus, and promoting early transcription and replication.IMPORTANCE A relatively unique feature of papillomaviruses is that the viral genome is associated with host histones inside the virion. However, little is known about the nature of the epigenome within papillomavirions or its biological relevance to the infectious viral cycle. Here, we define the epigenetic signature of the H3 and H4 histones from HPV16 virions generated in cell culture and native human papillomavirus type 1 (HPV1) and bovine papillomavirus 1 (BPV1) virions isolated from bovine and human wart tissue. We show that native virions are enriched in posttranslational modifications associated with active chromatin and depleted with those associated with repressed chromatin compared to cellular chromatin. Native virions were also enriched in the histone variant H3.3 compared to the canonical histone H3.1. We propose that the composition of virion-packaged chromatin reflects the late stages of the viral life cycle and promotes the early stages of infection by being primed for viral transcription.

Viruses, despite their great abundance and significance in biological systems, remain largely mysterious. Indeed, the vast majority of the perhaps hundreds of millions of viral species on the planet remain undiscovered. Additionally, many viruses deposited in central databases like GenBank and RefSeq are littered with genes annotated as ‘hypothetical protein’ or the equivalent. Cenote-Taker 2, a virus discovery and annotation tool available on command line and with a graphical user interface with free high-performance computation access, utilizes highly sensitive models of hallmark virus genes to discover familiar or divergent viral sequences from user-input contigs. Additionally, Cenote-Taker 2 uses a flexible set of modules to automatically annotate the sequence features of contigs, providing more gene information than comparable tools. The outputs include readable and interactive genome maps, virome summary tables, and files that can be directly submitted to GenBank. We expect Cenote-Taker 2 to facilitate virus discovery, annotation, and expansion of the known virome.

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While there have been remarkable strides in microbiome research, the viral component of the microbiome has generally presented a more challenging target than the bacteriome. This is despite the fact that many thousands of shotgun sequencing runs from human metagenomic samples exist in public databases and all of them encompass large amounts of viral sequences. The lack of a comprehensive database for human-associated viruses, along with inadequate methods for high-throughput identification of highly divergent viruses in metagenomic data, has historically stymied efforts to characterize virus sequences in a comprehensive way. In this study, a new high-specificity and high-sensitivity bioinformatic tool, Cenote-Taker 2, was applied to thousands of human metagenome datasets, uncovering over 50,000 unique virus operational taxonomic units. Publicly available case-control studies were re-analyzed, and over 1,700 strong virus-disease associations were found.

Viruses, despite their great abundance and significance in biological systems, remain largely mysterious. Indeed, the vast majority of the perhaps hundreds of millions of viral species on the planet remain undiscovered. Additionally, many viruses deposited in central databases like GenBank and RefSeq are littered with genes annotated as “hypothetical protein” or the equivalent. Cenote-Taker2, a virus discovery and annotation tool available on command line and with a graphical user interface with free high-performance computation access, utilizes highly sensitive models of hallmark virus genes to discover familiar or divergent viral sequences from user-input contigs. Additionally, Cenote-Taker2 uses a flexible set of modules to automatically annotate the sequence features of contigs, providing more gene information than comparable tools. The outputs include readable and interactive genome maps, virome summary tables, and files that can be directly submitted to GenBank. We expect Cenote-Taker2 to facilitate virus discovery, annotation, and expansion of the known virome.

Animals exhibit instinctive behavioral and physiological responses to a variety of stressors to overcome danger and restore homeostasis. The physiological response to stress is governed by hypothalamic corticotropin-releasing hormone (CRH) neurons which regulate the hypothalamic-pituitary-adrenal axis to control blood levels of stress hormones. At present, the neural circuits and signaling mechanisms through which different stress signals are transmitted to CRH neurons are poorly understood. Here, we devised a new method, termed “Connect-Seq,” which couples single-cell transcriptomics and retrograde viral tracing to define the molecular identities of individual neurons in neural circuits. As a proof of concept, using Connect-Seq, we profiled single-cell transcriptomes of 124 brain neurons upstream of CRH neurons and identified subpopulations that are likely to communicate stress-related signals to CRH neurons. Analyses of single-cell transcriptomes for ‘fast-acting’ neurotransmitters revealed subsets of upstream neurons that expressed markers of inhibitory GABAergic neurons or excitatory glutamatergic neurons. Further analyses showed a number of other neuromodulators/neurotransmitters in upstream neurons, including acetylcholine, dopamine, histamine, and 43 different neuropeptides, each expressed in individual neurons or subsets of neurons. These findings reveal extreme molecular heterogeneity among upstream neurons and suggest the upstream neurons use diverse neurochemical messengers to transmit signals to CRH neurons. Many neurons coexpressed different neurotransmitters/neuromodulators, suggesting the co-release of neurochemical messengers. Dual labeling of brain sections verified expression of specific neuromodulators in virus-infected neurons upstream of CRH neurons in selected brain areas. Our results indicate that Connect-Seq can be applied to genetically dissect neural circuits and uncover molecular identities of neurons upstream of specific neuronal types of known function. Molecular markers identified in those neurons lay a foundation for the application of cell-specific genetic tools to investigate the functions and physiological significance of diverse neuronal subsets within complex neural circuits.

Increased renal complications have been suggested with suprarenal stent grafts, but long-term analyses have been limited. Therefore, the purpose of this study was to evaluate the effect of endograft choice on perioperative and long-term outcomes.We compared Medicare beneficiaries undergoing endovascular abdominal aortic aneurysms repair from 2005 to 2008 with endografts with infrarenal fixation and a single docking limb (AneuRx, Excluder) to those with suprarenal fixation and 2 docking limbs (Zenith), or a unibody configuration (Powerlink). Propensity score weighting accounted for differences in patient characteristics among the different graft formations, and perioperative mortality, complications, and length of stay and 4-year rates of survival, rupture, and reintervention were compared.Forty-six thousand one hundred seventy-one Medicare beneficiaries were identified including 11,002 (24%) with suprarenal fixation, 32,909 (71%) with infrarenal fixation, and 2,260 (5%) with a unibody graft. After propensity score weighting, there were no significant differences in patients' baseline clinical and demographic characteristics. The suprarenal fixation patients had higher rates of perioperative mortality (1.7% vs. 1.3%, P 0.01), renal failure (6.0% vs. 4.7%, P 0.001), and mesenteric ischemia (0.7% vs. 0.4%, P 0.01) and longer length of stay (3.4 days vs. 3.0 days, P 0.001) compared with patients with infrarenal fixation. Unibody grafts had higher rates of renal failure (5.9% vs. 4.7%, P 0.001), mesenteric ischemia (1.0% vs. 0.4%, P 0.001), and conversion to open repair (0.7% vs. 0.1%, P 0.001) compared to those with infrarenal fixation and single docking limbs. At 4 years, mortality remained slightly higher with suprarenal compared to infrarenal fixation (30% vs. 29%, P = 0.047), although these patients had fewer conversions to open repair (0.6% vs. 0.9%, P = 0.03) and aneurysm-related reinterventions (10% vs. 12%, P 0.01). At 4 years, unibody grafts had more aneurysm-related interventions compared to infrarenal fixation grafts (15% vs. 12%, P 0.01) but fewer conversions to open repair (0.4% vs. 0.9%, P = 0.02). Late rupture did not differ among the groups.Compared to infrarenal fixation devices, patients who underwent EVAR with suprarenal fixation had higher perioperative mortality and renal complications but fewer reinterventions including conversion, while the unibody graft had more perioperative complications and aneurysm-related reinterventions, but fewer conversions to open repair. Although these differences could be explained by selection bias, these data suggest that further comparative effectiveness analyses should be performed to understand the outcomes following EVAR with suprarenal fixation and unibody grafts.

Recent case series describe detection of BK polyomavirus (BKV) in urinary tract cancers in kidney transplant recipients, suggesting that BKV could contribute to the development of these cancers. We assessed risk for urinary tract cancers in kidney recipients with or without treatment for presumed BKV nephropathy (tBKVN) using data from the United States Transplant Cancer Match Study (2003-2013). Among 55 697 included recipients, 2015 (3.6%) were reported with tBKVN. Relative to the general population, incidence was similarly elevated (approximately 4.5-fold) for kidney cancer in recipients with or without tBKVN, and incidence was not increased in either group for prostate cancer. In contrast, for invasive bladder cancer, incidence was more strongly elevated in recipients with versus without tBKVN (standardized incidence ratios 4.5 vs. 1.7; N = 48 cases), corresponding to an incidence rate ratio (IRR) of 2.9 (95% confidence interval [CI] 1.0-8.2), adjusted for sex, age, transplant year, and use of polyclonal antibody induction. As a result, recipients with tBKVN had borderline increased incidence for all urothelial cancers combined (renal pelvis, ureter, and bladder cancers: adjusted IRR 2.2, 95% CI 0.9-5.4; N = 89 cases). Together with reports describing BKV detection in tumor tissues, these results support an association between BKV and urothelial carcinogenesis among kidney transplant recipients.

Summary: Progressive multifocal leukoencephalopathy (PML) is a lethal brain disease caused by uncontrolled replication of JC polyomavirus (JCV). JCV strains recovered from the brains of PML patients carry mutations that prevent the engagement of sialylated glycans, which are thought to serve as receptors for the infectious entry of wild-type JCV. In this report, we show that non-sialylated glycosaminoglycans (GAGs) can serve as alternative attachment receptors for the infectious entry of both wild-type and PML mutant JCV strains. After GAG-mediated attachment, PML mutant strains engage non-sialylated non-GAG co-receptor glycans, such as asialo-GM1. JCV-neutralizing monoclonal antibodies isolated from patients who recovered from PML appear to block infection by preventing the docking of post-attachment co-receptor glycans in an apical pocket of the JCV major capsid protein. Identification of the GAG-dependent/sialylated glycan-independent alternative entry pathway should facilitate the development of infection inhibitors, including recombinant neutralizing antibodies. : Geoghegan et al. show that JC polyomavirus strains that cause brain disease infect cells via a pathway involving a heparin-like attachment receptor and a non-sialylated co-receptor. Candidate therapeutic human monoclonal antibodies neutralize by blocking co-receptor engagement. Keywords: polyomavirus, JC, BK, SV40, progressive multifocal leukoencephalopathy, PML, monoclonal antibody, mAb, virus entry, receptor

Cotton growers commonly use glufosinate-based programs to control glyphosate-resistant Palmer amaranth. Palmer amaranth must be small (≤7.5 cm) for consistent control by glufosinate, and growers often miss the optimum application timing. XtendFlex™cotton may provide growers a tool to control larger Palmer amaranth. Glufosinate, dicamba, and glufosinate plus dicamba were compared for Palmer amaranth control in a rescue situation. Herbicides were applied to 16- to 23-cm weeds (POST-1) followed by a second application (POST-2) 12 d later. Glufosinate-ammonium at 590 g ai ha−1plus dicamba diglycolamine salt at 560 g ae ha−1POST-1 followed by glufosinate plus dicamba POST-2 was more effective than glufosinate at 880 g ha−1POST-1 followed by glufosinate at 590 g ha−1POST-2 or dicamba alone applied twice. Following a directed layby application of glyphosate, diuron, andS-metolachlor 14 d after POST-2, Palmer amaranth was controlled 99% by any system containing dicamba or glufosinate plus dicamba POST-1 followed by dicamba, glufosinate, or glufosinate plus dicamba POST-2 compared with 87% to 91% control by glufosinate alone applied twice. Cotton height and number of main stem nodes at layby were reduced in systems with dicamba only POST-1 followed by dicamba or glufosinate plus dicamba POST-2, presumably due to competition from the slowly dying Palmer amaranth with dicamba only POST-1. These treatments also delayed cotton maturity and reduced lint yield compared with systems containing glufosinate plus dicamba at POST-1.