The crystal structure of vaccinia virus protein E2 and perspectives on the prediction of novel viral protein folds

The morphogenesis of vaccinia virus (VACV, family Poxviridae), the smallpox vaccine, is a complex process involving multiple distinct cellular membranes and resulting in multiple different forms of infectious virion. Efficient release of enveloped virions, which promote systemic spread of infection within hosts, requires the VACV protein E2 but the molecular basis of E2 function remains unclear and E2 lacks sequence homology to any well-characterised family of proteins. We solved the crystal structure of VACV E2 to 2.3 Å resolution, revealing that it comprises two domains with novel folds: an N-terminal annular (ring) domain and a C-terminal globular (head) domain. The C-terminal head domain displays weak structural homology with cellular (pseudo)kinases but lacks conserved surface residues or kinase features, suggesting that it is not enzymatically active, and possesses a large surface basic patch that might interact with phosphoinositide lipid headgroups. Recent deep learning methods have revolutionised our ability to predict the three-dimensional structures of proteins from primary sequence alone. VACV E2 is an exemplar ‘difficult’ viral protein target for structure prediction, being comprised of multiple novel domains and lacking sequence homologues outside Poxviridae. AlphaFold2 nonetheless succeeds in predicting the structures of the head and ring domains with high and moderate accuracy, respectively, allowing accurate inference of multiple structural properties. The advent of highly accurate virus structure prediction marks a step-change in structural virology and beckons a new era of structurally-informed molecular virology.

Inter-organ transmission of hepatocellular senescence induces multi-organ dysfunction through the TGFβ signalling pathway.

Cellular senescence is associated with aging but also impacts various physiological and pathological processes such as embryonic development and wound healing. Factors secreted by senescent cells can affect their microenvironment, including local spreading of senescence. Acute severe liver disease is associated with hepatocyte senescence and frequently progresses to multi-organ failure. Why the latter occurs is poorly understood. Here, using genetic mouse models of hepatocyte-specific senescence, we demonstrate senescence development in extrahepatic organs and associated organ dysfunction in response to liver senescence. Additionally, we observe senescence-associated regeneration and reprogramming in the proximal tubules of the kidney. Using single cell transcriptomics and in vitro assays, we identify the Transforming Growth Factor β (TGFβ) pathway as a critical mediator of systemic spread of senescence. Lastly, TGFβ inhibition in our mouse models blocks senescence transmission to other organs and prevents renal dysfunction. Our results highlight the systemic consequences of organ-specific senescence which, independent of aging, contributes to multi-organ dysfunction.

Spread of Xanthomonas vasicola pv. musacearum within banana mats: implications for Xanthomonas wilt management

Xanthomonas wilt (XW) of banana caused by Xanthomonas vasicola pv. musacearum (Xvm) does not spread to all plants physically interconnected through the rhizome when one or a few are diseased. However, the factors behind this incomplete systemic spread of Xvm are not fully known yet could inform XW management. This study explored the effect of Xvm inoculum amounts, number and size of suckers, sucker positioning on mother plant corms and other mother plant corm attributes on sucker colonization. A shorter (p <0.05) incubation period (17.9 vs 21.1 days) and higher (p<.001) cumulative number of symptomatic leaves (5.2 vs 1.6 leaves) was observed when all (high inoculum) compared to two leaves (low inoculum) were inoculated. Xvm was recovered in corms at 29 days post inoculation (dpi) in both treatments with no differences (p >0.05) in proportions of corms with Xvm between the treatments. However, Xvm was recovered earlier and at a higher frequency in suckers when all leaves were inoculated. Lower Xvm recoveries occurred in the lower corm sections to which most suckers were attached relative to the middle and upper corm sections. Xvm incidence in corms increased with the number of attached maiden suckers, and the dpi while it declined with increasing mother plant and corm height. Thus, Xvm spread within mats is influenced by the amount of inoculum and the physiological stage (e.g., height) of the plant and attached suckers. The position of suckers, predominantly at the bottom of corms also protects them from infection. Measures that reduce Xvm inoculum build-up in mats are thus crucial for minimizing within mat XW spread.

Case Report: Safe Tourniquet Removal in Black Mamba (Dendroaspis polylepis) Bites

The black mamba is known for its notorious potent neurotoxic venom. For this reason, their bites are often erroneously treated in the field with the application of a tourniquet in the hope of delaying systemic spread of the venom. Observational studies have shown that inappropriate tourniquet application is a common, harmful practice. An arterial tourniquet is not a recommended first aid measure because of the risk of limb ischemia and gangrene. When inappropriately applied, the rapid removal of the tourniquet in the emergency department may precipitate a life-threatening venom and metabolic toxin rush, leading to respiratory arrest. We present two cases of black mamba bites in Gauteng, South Africa, where gradual tourniquet removal was used to avoid a venom rush and rapid respiratory paralysis. Venom and metabolic toxin rush with potentially fatal respiratory muscle paralysis may be averted by gradual, cautious removal of field-applied tourniquets with concomitant antivenom administration.

An Isoform of the Eukaryotic Translation Elongation Factor 1A (eEF1a) Acts as a Pro-Viral Factor Required for Tomato Spotted Wilt Virus Disease in Nicotiana benthamiana

The tripartite genome of the negative-stranded RNA virus Tomato spotted wilt orthotospovirus (TSWV) is assembled, together with two viral proteins, the nucleocapsid protein and the RNA-dependent RNA polymerase, into infectious ribonucleoprotein complexes (RNPs). These two viral proteins are, together, essential for viral replication and transcription, yet our knowledge on the host factors supporting these two processes remains limited. To fill this knowledge gap, the protein composition of viral RNPs collected from TSWV-infected Nicotiana benthamiana plants, and of those collected from a reconstituted TSWV replicon system in the yeast Saccharomyces cerevisiae, was analysed. RNPs obtained from infected plant material were enriched for plant proteins implicated in (i) sugar and phosphate transport and (ii) responses to cellular stress. In contrast, the yeast-derived viral RNPs primarily contained proteins implicated in RNA processing and ribosome biogenesis. The latter suggests that, in yeast, the translational machinery is recruited to these viral RNPs. To examine whether one of these cellular proteins is important for a TSWV infection, the corresponding N. benthamiana genes were targeted for virus-induced gene silencing, and these plants were subsequently challenged with TSWV. This approach revealed four host factors that are important for systemic spread of TSWV and disease symptom development.

Macrophage Selenoproteins Restrict Intracellular Replication of Francisella tularensis and Are Essential for Host Immunity

The essential micronutrient Selenium (Se) is co-translationally incorporated as selenocysteine into proteins. Selenoproteins contain one or more selenocysteines and are vital for optimum immunity. Interestingly, many pathogenic bacteria utilize Se for various biological processes suggesting that Se may play a role in bacterial pathogenesis. A previous study had speculated that Francisella tularensis, a facultative intracellular bacterium and the causative agent of tularemia, sequesters Se by upregulating Se-metabolism genes in type II alveolar epithelial cells. Therefore, we investigated the contribution of host vs. pathogen-associated selenoproteins in bacterial disease using F. tularensis as a model organism. We found that F. tularensis was devoid of any Se utilization traits, neither incorporated elemental Se, nor exhibited Se-dependent growth. However, 100% of Se-deficient mice (0.01 ppm Se), which express low levels of selenoproteins, succumbed to F. tularensis-live vaccine strain pulmonary challenge, whereas 50% of mice on Se-supplemented (0.4 ppm Se) and 25% of mice on Se-adequate (0.1 ppm Se) diet succumbed to infection. Median survival time for Se-deficient mice was 8 days post-infection while Se-supplemented and -adequate mice was 11.5 and &gt;14 days post-infection, respectively. Se-deficient macrophages permitted significantly higher intracellular bacterial replication than Se-supplemented macrophages ex vivo, corroborating in vivo observations. Since Francisella replicates in alveolar macrophages during the acute phase of pneumonic infection, we hypothesized that macrophage-specific host selenoproteins may restrict replication and systemic spread of bacteria. F. tularensis infection led to an increased expression of several macrophage selenoproteins, suggesting their key role in limiting bacterial replication. Upon challenge with F. tularensis, mice lacking selenoproteins in macrophages (TrspM) displayed lower survival and increased bacterial burden in the lung and systemic tissues in comparison to WT littermate controls. Furthermore, macrophages from TrspM mice were unable to restrict bacterial replication ex vivo in comparison to macrophages from littermate controls. We herein describe a novel function of host macrophage-specific selenoproteins in restriction of intracellular bacterial replication. These data suggest that host selenoproteins may be considered as novel targets for modulating immune response to control a bacterial infection.

Staphylococcal Scalded Skin Syndrome and Bullous Impetigo

Staphylococcal scalded skin syndrome (SSSS) and bullous impetigo are infections caused by Staphylococcus aureus. The pathogenesis of both conditions centers around exotoxin mediated cleavage of desmoglein-1, which results in intraepidermal desquamation. Bullous impetigo is due to the local release of these toxins and thus, often presents with localized skin findings, whereas SSSS is from the systemic spread of these toxins, resulting in a more generalized rash and severe presentation. Both conditions are treated with antibiotics that target S. aureus. These conditions can sometimes be confused with other conditions that result in superficial blistering; the distinguishing features are outlined below.

The crystal structure of vaccinia virus protein E2 and perspectives on the prediction of novel viral protein folds

The morphogenesis of vaccinia virus (VACV, family Poxviridae), the smallpox vaccine, is a complex process involving multiple distinct cellular membranes and resulting in multiple different forms of infectious virion. Efficient release of enveloped virions, which promote systemic spread of infection within hosts, requires the VACV protein E2 but the molecular basis of E2 function remains unclear and E2 lacks sequence homology to any well-characterised family of proteins. We solved the crystal structure of VACV E2 to 2.3 Å resolution, revealing that it comprises two domains with novel folds: an N-terminal annular (ring) domain and a C-terminal head domain. The C-terminal head domain displays weak structural homology with cellular (pseudo)kinases but lacks conserved surface residues or kinase features, suggesting that it is not enzymatically active, and possesses a large surface basic patch that might interact with phosphoinositide lipid headgroups. Recent deep learning methods have revolutionised our ability to predict the three-dimensional structures of proteins from primary sequence alone. VACV E2 is an exemplar 'difficult' viral protein target for structure prediction, being comprised of multiple novel domains and lacking sequence homologues outside Poxviridae. AlphaFold2 nonetheless succeeds in predicting the structures of the head and ring domains with high and moderate accuracy, respectively, allowing accurate inference of multiple structural properties. The advent of highly accurate virus structure prediction marks a step-change in structural virology and beckons a new era of structurally-informed molecular virology.

Suprasellar Melanocytoma with Leptomeningeal Seeding: An Aggressive Clinical Course for a Histologically Benign Tumor

Introduction. Meningeal melanocytoma (MM) is a very rare neuroectodermal neoplasm arising from the leptomeninges. Primary suprasellar melanocytomas are exceedingly rare, with only a handful of cases reported. The systemic spread of a nontransformed meningeal melanocytoma is an unusual occurrence. Herein, we report the first case of a primary sellar melanocytoma with cerebral and spinal meningeal seeding. Case Report. A 30-year-old male with no previous medical history presented to the endocrinology department with a loss of body hair. The endocrine workup concluded with isolated hypogonadotropic hypogonadism. The Magnetic Resonance Imaging (MRI) of the brain and sella revealed a large suprasellar mass continuous with the infundibulum of the pituitary gland. It was heterogeneously hyperintense on T1-, T2-, and FLAIR-weighted images and was enhanced with contrast, along with cerebral and spinal leptomeningeal spread. The patient was referred to the neurosurgery department, and a lumbar spine biopsy was indicated. The histopathological examination was suggestive of a grade I meningeal pigmented melanocytoma. Conclusion. Thus, primary sellar melanocytomas with leptomeningeal spread are an extremely rare phenomenon. Metastatic malignant melanoma should be ruled out. Being aware of differential diagnosis and the unusual behavior of meningeal melanocytoma will be necessary to manage the patient appropriately. Complete tumor resection is the best treatment whenever possible, and radiotherapy should be considered in case of unresectability or partial resection.

Extraintestinal Manifestation of Yersinia pseudotuberculosis Bacteremia as Acute Hepatitis: Case Report and Review of the Literature

Yersinia pseudotuberculosis is a causative agent of foodborne zoonosis that usually causes self-limiting pseudoappendicitis. Y. pseudotuberculosis infection also causes systemic spread or extraintestinal manifestations in patients with predisposing conditions. Here, we present a case of acute hepatitis with Y. pseudotuberculosis bacteremia in a 30-year-old man. He was previously healthy without significant medical history other than obesity and current smoking. At the time of admission, he presented with high fever accompanied by chills, jaundice, abdominal pain, and watery diarrhea. Laboratory studies revealed leukocytosis and elevated liver function parameters. A stool culture showed no causative pathogens. Empiric antibiotic therapy with ceftriaxone and metronidazole was administered. Y. pseudotuberculosis was later isolated from the initial blood culture performed on the day of admission using MALDI-TOF mass spectrometry. Antibiotic treatment was continued based on the susceptibility testing results from MALDI-TOF MS and VITEk®2, as well as clinical and laboratory improvements. The patient was discharged on the tenth day of admission and remained healthy with no recurrence during the 12-month follow-up. Here, we review the literature on the systemic infection caused by Y. pseudotuberculosis, including extraintestinal manifestations. This case highlights that Y. pseudotuberculosis may be considered a differential causative organism in patients with acute colitis and hepatitis.