Structural basis of substrate progression through the bacterial chaperonin cycle.

The bacterial chaperonin GroEL-GroES promotes protein folding through ATP-regulated cycles of substrate protein binding, encapsulation, and release. Here, we have used cryoEM to determine structures of GroEL, GroEL-ADP·BeF3, and GroEL-ADP·AlF3-GroES all complexed with the model substrate Rubisco. Our structures provide a series of snapshots that show how the conformation and interactions of non-native Rubisco change as it proceeds through the GroEL-GroES reaction cycle. We observe specific charged and hydrophobic GroEL residues forming strong initial contacts with non-native Rubisco. Binding of ATP or ADP·BeF3 to GroEL-Rubisco results in the formation of an intermediate GroEL complex displaying striking asymmetry in the ATP/ADP·BeF3-bound ring. In this ring, four GroEL subunits bind Rubisco and the other three are in the GroES-accepting conformation, suggesting how GroEL can recruit GroES without releasing bound substrate. Our cryoEM structures of stalled GroEL-ADP·AlF3-Rubisco-GroES complexes show Rubisco folding intermediates interacting with GroEL-GroES via different sets of residues.

Structural insights into the assembly and activation of the IL-27 signaling complex.

Interleukin 27 (IL-27) is a heterodimeric cytokine that elicits potent immunosuppressive responses. Comprised of EBI3 and p28 subunits, IL-27 binds GP130 and IL-27Rα receptor chains to activate the JAK/STAT signaling cascade. However, how these receptors recognize IL-27 and form a complex capable of phosphorylating JAK proteins remains unclear. Here, we used cryo electron microscopy (cryoEM) and AlphaFold modeling to solve the structure of the IL-27 receptor recognition complex. Our data show how IL-27 serves as a bridge connecting IL-27Rα (domains 1-2) with GP130 (domains 1-3) to initiate signaling. While both receptors contact the p28 component of the heterodimeric cytokine, EBI3 stabilizes the complex by binding a positively charged surface of IL-27Rα and Domain 1 of GP130. We find that assembly of the IL-27 receptor recognition complex is distinct from both IL-12 and IL-6 cytokine families and provides a mechanistic blueprint for tuning IL-27 pleiotropic actions.

Cerebral filariasis infection with Litomosoides in Molossus barnesi (Chiroptera: Molossidae) in the Brazilian eastern Amazon, with comments on Molossinema wimsatti Georgi, Georgi, Jiang and Fronguillo, 1987.

During bacterial and viral pathogen investigation of 30 specimens of bats captured in periurban forest areas in the city of Belém, Pará, Brazil, a case of cerebral filariasis was observed. In the course of histopathological examination, adult filariae were found in pseudocystic cavities brain of Molossus barnesi (Molossidae) and classified morphologically as Litomosoides by the shape of the spicules-left spicule with a handle longer than the blade; right spicule curved, with a sclerotized heel supporting a dorsal notch; the area rugosa constituted by a ventral band of small longitudinal crests; tail rounded in males; long esophagus with a slightly glandular distal portion; and a muscular bent vagina. All the specimens lack a stoma (buccal capsule). We compared our filarioids with the description of specimens of Molossinema wimsatti. Morphological characteristics of M. wimsatti resemble the genus Litomosoides. Thus, we believe that M. wimsatti is a synonym of L. molossi Esslinger, 1973, and filarioid specimens from material reported by Lichtenfels et al. (Trans Am Micros Soc 100:216-219, 1981) and from de Souto et al. (J. Helminthol 1195:e65, 2021) most probably correspond to Litomosoides. We suggest that the reduction of the buccal capsule may be attributable to the ectopic location. No evidence of tissue responses by the host was observed. This is the first record of Litomosoides infecting brain tissue of Molossus barnesi from Brazil, representing a record of a new host species. More specimens of bats should be examined in order to find filarioids in the brain and verify its taxonomic position using molecular techniques.

Tapping into natural history collections to assess latitudinal gradients of parasite diversity.

Parasites are key components of the biosphere not only due to their huge diversity, but also because they exert important influences on ecological processes. Nevertheless, we lack an understanding of the biogeographical patterns of parasite diversity. Here, we tap into the potential of biodiversity collections for understanding parasite biogeography. We assess species richness of supracommunities of helminth parasites infecting mammal assemblages in the Nearctic, and describe its relation to latitude, climate, host diversity, and land area. We compiled data from parasitology collections and assessed parasite diversity in Nearctic ecoregions for the entire parasite supracommunity of mammals in each ecoregion, as well as separately from carnivores and rodents to explore the effect of host taxonomic resolution on observed patterns. For carnivores, we found evidence of a negative latitudinal gradient, while parasites of rodents displayed no clear pattern. We found that parasite diversity was positively correlated with mean annual temperature and negatively correlated with seasonal precipitation. Parasite richness shows a diversity peak at intermediate host richness values and in carnivores correlates with temperature and seasonal precipitation. Rodent parasite diversity did not correlate with explored factors. Other researchers are encouraged to use parasitology collections to continue exploring patterns of parasite biogeography and macroecology.

Intercontinental comparisons of subterranean host-parasite communities using bipartite network analyses.

Rodents living in a subterranean ecotope face a unique combination of evolutionary and ecological pressures and while host species evolution may be driven by the selective pressure from the parasites they harbour, the parasites may be responding to the selective pressures of the host. Here, we obtained all available subterranean rodent host­parasite records from the literature and integrated these data by utilizing a bipartite network analysis to determine multiple critical parameters to quantify and measure the structure and interactions of the organisms present in host­parasite communities. A total of 163 species of subterranean rodent hosts, 174 parasite species and 282 interactions were used to create 4 networks with data well-represented from all habitable continents. The results show that there was no single species of parasite that infects subterranean rodents throughout all zoogeographical regions. Nevertheless, species representing the genera Eimeria and Trichuris were common across all communities of subterranean rodents studied. Based on our analysis of host­parasite interactions across all communities studied, the parasite linkages show that community connectance (due to climate change or other anthropogenic factors) appears to show degraded linkages in both the Nearctic and Ethiopian regions: in this case parasites are acting as bell-weather probes signalling the loss of biodiversity.

Less Is More: A Multimodal Pain Management Strategy Is Associated With Reduced Opioid Use in Hospitalized Trauma Patients.

INTRODUCTION: Adequate pain control is critical to the management and recovery of acutely injured patients. Opioids are associated with various adverse effects, and drug overdose is the leading cause of injury-related death in the United States. We hypothesized that a multimodal pain management protocol would reduce opioid use while still optimizing pain control. METHODS: The study included the preanalysis (August 2017-September 2018) and postanalysis (October 2018-August 2019) of a multimodal pain management strategy implemented in hospitalized adult patients admitted to the trauma service at a single American College of Surgeons-verified level-1 trauma center. Patients less than 18 y of age, pregnant patients, or imprisoned patients were excluded. The primary endpoint was opioid prescription on discharge (morphine milligram equivalent [MME]). The secondary endpoints included inpatient MMEs, nonopioid adjunct use, and pain scores. Subgroup analysis evaluating opioid use based on Injury Severity Score groups (mild, moderate, or severe) and by the Abbreviated Injury Scale body region was performed. RESULTS: There were 1755 patients in the PRE group and 1723 patients in the POST group. MMEs prescribed on discharge decreased from median 15 (interquartile range: 37.5) to 1.2 (interquartile range: 22.5) (P < 0.001). More patients in the POST group were discharged opioid-free (44% versus 37%, P < 0.001). There was a significant increase in the use of all nonopioid pain medications. Pain scores did not change. Subgroup analysis revealed a significant decrease in discharge MMEs in mild and moderate Injury Severity Score groups and in all injured body regions except the chest. CONCLUSIONS: The implementation of a multimodal pain management protocol in trauma patients targeting scheduled nonopioid medications and patient education is feasible and is associated with reduced opioid amount prescribed on discharge, without compromising pain control.

Characterization and influence of ipsilateral scapula fractures among patients who undergo surgical stabilization of sub-scapular rib fractures.

BACKGROUND: Current decision algorithms involving surgical stabilization of rib fractures (SSRF) do not consider either specific fracture locations or other chest wall bony injuries. Our objective was to characterize the impact of scapula fractures on morbidity among patients who underwent fixation of sub-scapular rib fractures. We hypothesized that an ipsilateral scapula fracture was associated with poor acute and long-term outcomes. METHODS: Retrospective review of two institutions' prospectively maintained SSRF databases (October 2010 to January 2019). Patients who underwent repair of ≥ 1 sub-scapular rib fracture were included. Patients were grouped by the presence of an ipsilateral scapula fracture. Outcomes were acute SSRF complications, long-term rib implant removal, and quality of life via phone survey. RESULTS: A total of 144 patients were analyzed; 53 (36.8%) had an ipsilateral scapula fracture. Patients with a scapula fracture had a higher injury severity score (p = 0.02), degree of pulmonary contusion (p < 0.01), and RibScore (p < 0.01). The overall incidence of both acute re-operation (n = 4, 2.8%) and long-term implant removal (n = 5, 3.8%) following SSRF was low and did not vary by the presence of a scapula fracture. Only twenty-one patients completed phone questionnaires a median of 38 months after SSRF; both shoulder and rib outcomes were excellent and did not vary by the presence of a scapula fracture. CONCLUSION: Ipsilateral scapula fractures are common in patients who undergo surgical stabilization of sub-scapular rib fractures. Despite higher injury severity, patients with an ipsilateral scapula fracture did not incur worse outcomes.

Wildfire and the ecological niche: Diminishing habitat suitability for an indicator species within semi-arid ecosystems.

Globally accelerating frequency and extent of wildfire threatens the persistence of specialist wildlife species through direct loss of habitat and indirect facilitation of exotic invasive species. Habitat specialists may be especially prone to rapidly changing environmental conditions because their ability to adapt lags behind the rate of habitat alteration. As a result, these populations may become increasingly susceptible to ecological traps by returning to suboptimal breeding habitats that were dramatically altered by disturbance. We demonstrate a multistage modeling approach that integrates habitat selection and survival during the key nesting life-stage of a bird species of high conservation concern, the greater sage-grouse (Centrocercus urophasianus; hereafter, sage-grouse). We applied these spatially explicit models to a spatiotemporally robust dataset of sage-grouse nest locations and fates across wildfire-altered sagebrush ecosystems of the Great Basin ecoregion, western United States. Female sage-grouse exhibited intricate habitat selection patterns that varied across regional gradients of ecological productivity among sagebrush communities, but often selected nest sites that disproportionately resulted in nest failure. For example, 23% of nests occurred in wildfire-affected habitats characterized by reduced sagebrush cover and greater composition of invasive annual grasses. We found survival of nests was negatively associated with wildfire-affected areas, but positively associated with higher elevations with increased ruggedness and overall shrub cover. Strong site fidelity likely drove sage-grouse to continue nesting in habitats degraded by wildfire. Hence, increasing frequency and extent of wildfire may contribute disproportionately to reduced reproductive success by creating ecological traps that act as population sinks. Identifying such habitat mismatches between selection and survival facilitates deeper understanding of the mechanisms driving reduced geographic niche space and population decline at broad spatiotemporal scales, while guiding management actions to areas that would be most beneficial to the species.

Health Disparities Research with American Indian Communities: The Importance of Trust and Transparency.

American Indian and Alaska Native (AI/AN) communities experience notable health disparities associated with substance use, including disproportionate rates of accidents/injuries, diabetes, liver disease, suicide, and substance use disorders. Effective treatments for substance use are needed to improve health equity for AI/AN communities. However, an unfortunate history of unethical and stigmatizing research has engendered distrust and reluctance to participate in research among many Native communities. In recent years, researchers have made progress toward engaging in ethical health disparities research by using a community-based participatory research (CBPR) framework to work in close partnership with community members throughout the research process. In this methodological process paper, we discuss the collaborative development of a quantitative survey aimed at understanding risk and protective factors for substance use among a sample of tribal members residing on a rural AI reservation with numerous systems-level barriers to recovery and limited access to treatment. By using a CBPR approach and prioritizing trust and transparency with community partners and participants, we were able to successfully recruit our target sample and collect quality data from nearly 200 tribal members who self-identified as having a substance use problem. Strategies for enhancing buy-in and recruiting a community sample are discussed.

Partnering with Native Communities to Develop a Culturally Grounded Intervention for Substance Use Disorder.

Studies have documented serious disparities in drug and alcohol-related morbidity and mortality among American Indians and Alaska Natives (AI/ANs) compared to other ethnic groups in the U.S. despite high rates of abstinence in these groups. Further complicating these health disparities are barriers to accessing evidence-based treatments that are culturally appropriate and acceptable. As part of a research program to promote health equity in rural communities, we developed an academic-community partnership to create a culturally grounded intervention for adults with substance use disorder (SUD) residing on a rural AI reservation. We describe the early phases of our long-term Community Based Participatory Research project and report findings from the first study we conducted. This key informant interview study consisted of in-depth qualitative interviews with 25 tribal members knowledgeable about substance use and recovery on the reservation. The goal was to understand social norms and cultural conceptualizations of substance use to inform the development of a sustainable, community-driven intervention. Participants reported that a holistic approach to recovery that emphasizes spiritual, cultural, and interpersonal harmony and connectedness was important to the community and would be necessary for the intervention to succeed. They also emphasized the need for a multi-level intervention targeting individuals, families, and the community as a whole. Through this initial study, we not only gained valuable information that will be used to guide future research and treatment efforts, but we also strengthened our partnership and built trust with the community. In this manuscript we tell the story of the development of our project and describe our shared vision for future directions.

A conservation planning tool for Greater Sage-grouse using indices of species distribution, resilience, and resistance.

Managers require quantitative yet tractable tools that identify areas for restoration yielding effective benefits for targeted wildlife species and the ecosystems they inhabit. As a contemporary example of high national significance for conservation, the persistence of Greater Sage-grouse (Centrocercus urophasianus) in the Great Basin is compromised by strongly interacting stressors of conifer expansion, annual grass invasion, and more frequent wildfires occurring in sagebrush ecosystems. Associated restoration treatments to a sagebrush-dominated state are often costly and may yield relatively little ecological benefit to sage-grouse if implemented without estimating how Sage-grouse may respond to treatments, or do not consider underlying processes influencing sagebrush ecosystem resilience to disturbance and resistance to invasive species. Here, we describe example applications of a spatially explicit conservation planning tool (CPT) to inform prioritization of: (1) removal of conifers (i.e., pinyon-juniper); and (2) wildfire restoration aimed at improving habitat conditions for the Bi-State Distinct Population Segment of Sage-grouse along the California-Nevada state line. The CPT measures ecological benefits to sage-grouse for a given management action through a composite index comprised of resource selection functions and estimates of abundance and space use. For pinyon-juniper removal, we simulated changes in land-cover composition following the removal of sparse trees with intact understories, and ranked treatments on the basis of changes in ecological benefits per dollar-unit of cost. For wildfire restoration, we formulated a conditional model to simulate scenarios for land cover changes (e.g., sagebrush to annual grass) given estimated fire severity and underlying ecosystem processes influencing resilience to disturbance and resistance to invasion by annual grasses. For both applications, we compared CPT rankings to land cover changes along with sagebrush resistance and resilience metrics. Model results demonstrated how the CPT can be an important step in identifying management projects that yield the highest quantifiable benefit to Sage-grouse while avoiding costly misallocation of resources, and highlight the importance of considering changes in sage-grouse ecological response and factors influencing sagebrush ecosystem resilience to disturbance and resistance to invasion. This unique framework can be adopted to help inform other management questions aimed at improving habitat for other species across sagebrush and other ecosystems.

Structural insights into IL-11-mediated signalling and human IL6ST variant-associated immunodeficiency.

IL-11 and IL-6 activate signalling via assembly of the cell surface receptor gp130; however, it is unclear how signals are transmitted across the membrane to instruct cellular responses. Here we solve the cryoEM structure of the IL-11 receptor recognition complex to discover how differences in gp130-binding interfaces may drive signalling outcomes. We explore how mutations in the IL6ST gene encoding for gp130, which cause severe immune deficiencies in humans, impair signalling without blocking cytokine binding. We use cryoEM to solve structures of both IL-11 and IL-6 complexes with a mutant form of gp130 associated with human disease. Together with molecular dynamics simulations, we show that the disease-associated variant led to an increase in flexibility including motion within the cytokine-binding core and increased distance between extracellular domains. However, these distances are minimized as the transmembrane helix exits the membrane, suggesting a stringency in geometry for signalling and dimmer switch mode of action.

Helminth and protozoan parasites of subterranean rodents (Chordata, Mammalia, Rodentia) of the world.

Published studies and ten new unpublished records included herein reveal that approximately 174 species of endoparasites (helminths and protozoans) are known from 65 of 163 species of rodents that occupy the subterranean ecotope globally. Of those, 94 endoparasite species were originally described from these rodents. A total of 282 host-parasite associations are summarized from four major zoogeographic regions including Ethiopian, Palearctic/Oriental, Nearctic, and Neotropical. Thirty-four parasite records from the literature have been identified to only the level of the genus. In this summary, ten new records have been added, and the most current taxonomic status of each parasite species is noted. Interestingly, there are no data on endoparasites from more than 68% of described subterranean rodents, which indicates that discovery and documentation are at an early stage and must continue.

Investigating relationships between climate controls and nutrient flux in surface waters, sediments, and subsurface pathways in an agricultural clay catchment of the Great Lakes Basin.

Water quality within agricultural catchments is governed by management practices and climate conditions that control the transport, storage, and exchange of nutrients between components of the hydrologic cycle. This study aims to improve knowledge of nitrogen (N) and phosphorus (P) transport in low permeability agricultural watersheds by considering spatial and temporal trends of surface water nutrient concentrations in relation to hydroclimatic drivers, sediment quality, shallow hyporheic exchange, groundwater quality, and tile drain discharge over a 14-month field study in a clay hydrosystem of the Lake Huron basin, one of the five Great Lakes. Results found that events of varying magnitude and intensity enhanced nutrient release from overland flow and subsurface pathways. Tile drain discharge was found to be a consistent and elevated source of P and N to surface waters when flowing, mobilizing both diffuse nutrients from fertilizer application and legacy stores in the vadose zone. Surface water quality was temporally variable at the seasonal and event scale. Targeted sampling following fertilization periods, snowmelt, and moderate precipitation events revealed catchment wide elevated nutrient concentrations, emphasizing the need for targeted sampling regimes. Controls other than discharge magnitude and overland flow were found to contribute to peak nutrient concentrations, including internal nitrate loading, soil-snowmelt interaction, catchment wetness, and freeze thaw cycles. Sediments were found to store P in calcium minerals and have a high P storage capacity. Instream mechanisms such as sediment P fixation and hyporheic exchange may play a role in mediating surface water quality, but currently have no discernable benefit to year-round surface water nutrient concentrations. Best management practices need to focus on reducing sources of agricultural nutrients (e.g., field phosphorus concentrations and tile drain discharge loading) at the watershed scale to reduce nutrient concentrations and export in flashy clay catchments.

Structural basis for membrane attack complex inhibition by CD59.

CD59 is an abundant immuno-regulatory receptor that protects human cells from damage during complement activation. Here we show how the receptor binds complement proteins C8 and C9 at the membrane to prevent insertion and polymerization of membrane attack complex (MAC) pores. We present cryo-electron microscopy structures of two inhibited MAC precursors known as C5b8 and C5b9. We discover that in both complexes, CD59 binds the pore-forming ß-hairpins of C8 to form an intermolecular ß-sheet that prevents membrane perforation. While bound to C8, CD59 deflects the cascading C9 ß-hairpins, rerouting their trajectory into the membrane. Preventing insertion of C9 restricts structural transitions of subsequent monomers and indirectly halts MAC polymerization. We combine our structural data with cellular assays and molecular dynamics simulations to explain how the membrane environment impacts the dual roles of CD59 in controlling pore formation of MAC, and as a target of bacterial virulence factors which hijack CD59 to lyse human cells.

Structure of the transmembrane protein 2 (TMEM2) ectodomain and its apparent lack of hyaluronidase activity.

Background: Hyaluronic acid (HA) is a major polysaccharide component of the extracellular matrix. HA has essential functions in tissue architecture and the regulation of cell behaviour. HA turnover needs to be finely balanced. Increased HA degradation is associated with cancer, inflammation, and other pathological situations. Transmembrane protein 2 (TMEM2) is a cell surface protein that has been reported to degrade HA into ~5 kDa fragments and play an essential role in systemic HA turnover. Methods: We produced the soluble TMEM2 ectodomain (residues 106-1383; sTMEM2) in human embryonic kidney cells (HEK293) and determined its structure using X-ray crystallography. We tested sTMEM2 hyaluronidase activity using fluorescently labelled HA and size fractionation of reaction products. We tested HA binding in solution and using a glycan microarray. Results: Our crystal structure of sTMEM2 confirms a remarkably accurate prediction by AlphaFold. sTMEM2 contains a parallel ß-helix typical of other polysaccharide-degrading enzymes, but an active site cannot be assigned with confidence. A lectin-like domain is inserted into the ß-helix and predicted to be functional in carbohydrate binding. A second lectin-like domain at the C-terminus is unlikely to bind carbohydrates. We did not observe HA binding in two assay formats, suggesting a modest affinity at best. Unexpectedly, we were unable to observe any HA degradation by sTMEM2. Our negative results set an upper limit for k cat of approximately 10 -5 min -1. Conclusions: Although sTMEM2 contains domain types consistent with its suggested role in TMEM2 degradation, its hyaluronidase activity was undetectable. HA degradation by TMEM2 may require additional proteins and/or localisation at the cell surface.

"Revisiting the past": a redescription of Physaloptera retusa (Nemata, Physalopteridae) from material deposited in museums and new material from Amazon lizards.

Physaloptera Rudolphi, 1819 is a genus of nematodes that includes approximately 100 species parasitic in vertebrates around the world. From these, approximately 30 occur in the Neotropical region, with nine reported from neotropical reptiles. Physaloptera spp. are recognized by their distinct morphology of the apical end and characters of the reproductive system. However, despite the fact that the morphological characters for species diagnosis have been firmly established, we frequently find identification problems regarding poorly detailed descriptions and poorly preserved specimens. These may lead to taxonomic incongruencies. Physaloptera retusa (Rudolphi, 1819) is the most common species of the genus and has been reported from several species of neotropical reptiles. Based on our reexaminations of nematode specimens identified as P. retusa from different museum collections, we provide a detailed redescription including the type material, voucher specimens and new specimens recovered currently and showed in this study with new morphological data obtained using light and scanning electron microscopy tools.

HOST-SWITCHING EVENTS IN LITOMOSOIDES CHANDLER, 1931 (FILARIOIDEA: ONCHOCERCIDAE) ARE NOT RAMPANT BUT CLADE DEPENDENT.

The genus Litomosoides Chandler, 1931, includes species that as adults occur in the thoracic and abdominal cavity of mammalian hosts and are presumably vectored by mites. The vertebrate hosts include a variety of Neotropical mammals such as phyllostomid and mormoopid bats; cricetid, sciurid, and hystricognath rodents; and didelphid marsupials. It has been suggested that Litomosoides is not a monophyletic group and that rampant horizontal transfer explains their presence in disparate groups of mammals. Herein we present a phylogenetic reconstruction including mitochondrial genes of 13 vouchered species. This phylogeny is used to reconstruct the evolutionary history of these parasites and the ancestral states of key characters used in species classification, namely, the configuration of the spicules. The historical association of these filarioids with 6 groups of mammals, as well as their ancestral geographic distributions, were reconstructed using Bayesian statistical approaches comparing alternative models of biogeography and evolution and fossil states in selected nodes of the phylogeny. The optimal reconstruction suggests a model of dispersal, extinction, and cladogenesis (DEC) driving the evolution of Litomosoides; the results suggest an origin of Litomosoides in South America and association of ancestors with phyllostomids, and strong evidence of at least 2 host-switching events: 1 of these involving cricetid rodents and the other mormoopid bats. The latter event included a simultaneous geographic expansion of the parasite lineage across South and North America. The host-switching event from phyllostomid bats into cricetid rodents occurred once these rodents diversified across South America; subsequent diversification of the latter clade resulted in 2 branches, each showing expansion of the parasites back into North America. This result suggests that both parasites and cricetid rodents established an association in South America, underwent diversification, and then dispersed into North America. Further, this clade of cricetid-dwelling species includes parasites featuring the "sigmodontis" spicule type. The identification of a single host-switching event involving the disparate lineages of Chiroptera and Rodentia offers a framework to reconstruct the gene evolution and diversification of this lineage after the host-switching event. This will help in predicting the ability of these parasites to infect sympatric mammals.