Nonlytic cellular release of hepatitis A virus requires dual capsid recruitment of the ESCRT-associated Bro1 domain proteins HD-PTP and ALIX.

Although picornaviruses are conventionally considered 'nonenveloped', members of multiple picornaviral genera are released nonlytically from infected cells in extracellular vesicles. The mechanisms underlying this process are poorly understood. Here, we describe interactions of the hepatitis A virus (HAV) capsid with components of host endosomal sorting complexes required for transport (ESCRT) that play an essential role in release. We show release of quasi-enveloped virus (eHAV) in exosome-like vesicles requires a conserved export signal located within the 8 kDa C-terminal VP1 pX extension that functions in a manner analogous to late domains of canonical enveloped viruses. Fusing pX to a self-assembling engineered protein nanocage (EPN-pX) resulted in its ESCRT-dependent release in extracellular vesicles. Mutational analysis identified a 24 amino acid peptide sequence located within the center of pX that was both necessary and sufficient for nanocage release. Deleting a YxxL motif within this sequence ablated eHAV release, resulting in virus accumulating intracellularly. The pX export signal is conserved in non-human hepatoviruses from a wide range of mammalian species, and functional in pX sequences from bat hepatoviruses when fused to the nanocage protein, suggesting these viruses are released as quasi-enveloped virions. Quantitative proteomics identified multiple ESCRT-related proteins associating with EPN-pX, including ALG2-interacting protein X (ALIX), and its paralog, tyrosine-protein phosphatase non-receptor type 23 (HD-PTP), a second Bro1 domain protein linked to sorting of ubiquitylated cargo into multivesicular endosomes. RNAi-mediated depletion of either Bro1 domain protein impeded eHAV release. Super-resolution fluorescence microscopy demonstrated colocalization of viral capsids with endogenous ALIX and HD-PTP. Co-immunoprecipitation assays using biotin-tagged peptides and recombinant proteins revealed pX interacts directly through the export signal with N-terminal Bro1 domains of both HD-PTP and ALIX. Our study identifies an exceptionally potent viral export signal mediating extracellular release of virus-sized protein assemblies and shows release requires non-redundant activities of both HD-PTP and ALIX.

In utero nicotine exposure affects murine palate development.

OBJECTIVES: Despite data linking smoking to increased risk of fetal morbidity and mortality, 11% of pregnant women continue to smoke or use alternative nicotine products. Studies confirm that nicotine exposure during pregnancy increases the incidence of birth defects; however, little research has focused on specific anatomic areas based on timing of exposure. We aim to determine critical in utero and postnatal periods of nicotine exposure that affect craniofacial development, specifically palate growth. Malformation of the palatal structures can result in numerous complications including facial growth disturbance, or impeding airway function. We hypothesized that both in utero and postnatal nicotine exposure will alter palate development. MATERIALS AND METHODS: We administered pregnant C57BL6 mice water supplemented with 100 µg/mL nicotine during early pregnancy, throughout pregnancy, during pregnancy and lactation, or lactation only. Postnatal day 15 pups underwent micro-computed tomography (µCT) analyses specific to the palate. RESULTS: Resultant pups revealed significant differences in body weight from lactation-only nicotine exposure, and µCT investigation revealed several dimensions affected by lactation-only nicotine exposure, including palate width, palate and cranial base lengths, and mid-palatal suture width. CONCLUSIONS: These results demonstrate the direct effects of nicotine on the developing palate beyond simple tobacco use. Nicotine exposure through tobacco alternatives, cessation methods, and electronic nicotine delivery systems (ENDS) may disrupt normal growth and development of the palate during development and the postnatal periods of breastfeeding. Due to the recent dramatic increase in the use of ENDS, future research will focus specifically on this nicotine delivery method.

In utero exposure to selective serotonin re-uptake inhibitor affects murine mandibular development.

OBJECTIVES: Antidepressants, specifically Selective Serotonin Re-uptake Inhibitors (SSRIs), that alter serotonin metabolism are currently the most commonly prescribed drugs for the treatment of depression. There is some evidence to suggest these drugs contribute to birth defects. As jaw development is often altered in craniofacial birth defects, the purpose of this study was to interrogate the effects of in utero SSRI exposure in a preclinical model of mandible development. MATERIALS AND METHODS: Wild-type C57BL6 mice were used to produce litters that were exposed in utero to an SSRI, Citalopram (500 µg/day). Murine mandibles from P15 pups were analysed for a change in shape and composition. RESULTS: Analysis indicated an overall shape change with total mandibular length and ramus height being shorter in exposed pups as compared to controls. Histomorphometric analysis revealed that first molar length was longer in exposed pups while third molar length was shorter in exposed as compared to control. Histological investigation of molars and surrounding periodontium revealed no change in collagen content of the molar in exposed pups, some alteration in collagen composition in the periodontium, increased alkaline phosphatase in molars and periodontium and decreased mesenchymal cell marker presence in exposed mandibles. CONCLUSION: The results of this study reveal SSRI exposure may interrupt mandible growth as well as overall dental maturation in a model of development giving insight into the expectation that children exposed to SSRIs may require orthodontic intervention.

In Utero Exposure to Maternal Electronic Nicotine Delivery System use Demonstrate Alterations to Craniofacial Development.

OBJECTIVE: Develop a model for the study of Electronic Nicotine Device (ENDS) exposure on craniofacial development. DESIGN: Experimental preclinical design followed as pregnant murine dams were randomized and exposed to filtered air exposure, carrier exposure consisting of 50% volume of propylene glycol and vegetable glycine (ENDS Carrier) respectively, or carrier exposure with 20 mg/ml of nicotine added to the liquid vaporizer (ENDS carrier with nicotine). SETTING: Preclinical murine model exposure using the SciReq exposure system. PARTICIPANTS: C57BL6 adult 8 week old female pregnant mice and exposed in utero litters. INTERVENTIONS: Exposure to control filtered air, ENDS carrier or ENDS carrier with nicotine added throughout gestation at 1 puff/minute, 4 h/day, five days a week. MAIN OUTCOME MEASURES: Cephalometric measures of post-natal day 15 pups born as exposed litters. RESULTS: Data suggests alterations to several facial morphology parameters in the developing offspring, suggesting electronic nicotine device systems may alter facial growth if used during pregnancy. CONCLUSIONS: Future research should concentrate on varied formulations and exposure regimens of ENDS to determine timing windows of exposures and ENDS formulations that may be harmful to craniofacial development.

Evaluating expert-based habitat suitability information of terrestrial mammals with GPS-tracking data.

Aim: Macroecological studies that require habitat suitability data for many species often derive this information from expert opinion. However, expert-based information is inherently subjective and thus prone to errors. The increasing availability of GPS tracking data offers opportunities to evaluate and supplement expert-based information with detailed empirical evidence. Here, we compared expert-based habitat suitability information from the International Union for Conservation of Nature (IUCN) with habitat suitability information derived from GPS-tracking data of 1,498 individuals from 49 mammal species. Location: Worldwide. Time period: 1998-2021. Major taxa studied: Forty-nine terrestrial mammal species. Methods: Using GPS data, we estimated two measures of habitat suitability for each individual animal: proportional habitat use (proportion of GPS locations within a habitat type), and selection ratio (habitat use relative to its availability). For each individual we then evaluated whether the GPS-based habitat suitability measures were in agreement with the IUCN data. To that end, we calculated the probability that the ranking of empirical habitat suitability measures was in agreement with IUCN's classification into suitable, marginal and unsuitable habitat types. Results: IUCN habitat suitability data were in accordance with the GPS data (> 95% probability of agreement) for 33 out of 49 species based on proportional habitat use estimates and for 25 out of 49 species based on selection ratios. In addition, 37 and 34 species had a > 50% probability of agreement based on proportional habitat use and selection ratios, respectively. Main conclusions: We show how GPS-tracking data can be used to evaluate IUCN habitat suitability data. Our findings indicate that for the majority of species included in this study, it is appropriate to use IUCN habitat suitability data in macroecological studies. Furthermore, we show that GPS-tracking data can be used to identify and prioritize species and habitat types for re-evaluation of IUCN habitat suitability data.

Response of barren-ground caribou to advancing spring phenology.

Phenological shifts are occurring in many ecosystems around the world. The capacity of species to adapt to changing phenology will be critical to their success under climate change scenarios. Failure to adjust migratory and reproductive timing to keep pace with the earlier onset of spring has led to negative demographic effects for populations of species across a variety of taxa. For caribou, there have been concerns that earlier spring green-up on calving areas might not be matched by earlier migration and parturition, potentially leading to a trophic mismatch with nutritional consequences for parturient and lactating caribou cows. However, there is limited evidence supporting these concerns. Here, we investigate the response of barren-ground caribou to changing spring phenology using data from telemetry and satellite imagery. From 2004 to 2016, we found that the average start of green-up on the calving area advanced by 7.25 days, while the start of migration advanced by 13.64 days, the end of migration advanced by 6.02 days, and the date of peak calving advanced by 9.42 days. Despite the advancing onset of green-up, we found no evidence for the development of a trophic mismatch because the advancing green-up coincided with earlier migration and calving by caribou. Changing snow cover on the late winter and migratory ranges was the most supported driver of advancing migratory behavior. The ability of caribou to adjust the timing of migratory and reproductive behavior in response to changing environmental conditions demonstrates the potential resilience of the species to some aspects of climate change.

Beaver (Castor canadensis) use of borrow pits in an industrial landscape in northwestern Alberta.

Borrow pits, dug by industry to provide substrate for infrastructure such as roads and well sites, are prevalent throughout the boreal forest of western Canada yet little is known about their use by wildlife. During field surveys in Rainbow Lake, Alberta, we found that beavers (Castor canadensis) used inundated borrow pits extensively for foraging and over wintering, suggesting that borrow pits increase beaver populations above their natural carrying capacity in industrial landscapes. We visited a random sample of 90 borrow pits in the field and categorized them as having active beaver lodges with caches (n = 16), inactive lodges (n = 13), cutting (n = 29), no activity (n = 13), or as not inundated or not a borrow pit (n = 19). We then used this sample to model where beavers established lodges versus where they were inactive or were only cutting. We found that borrow pits with active lodges had unique characteristics from other pits in our sample and were closer to streams, marshes, and swamps and had greater vegetation concealment from roads. These models can be used by managers to develop methods for increasing or decreasing the abundance of beavers living at borrow pits. Wildlife managers might want to use borrow pits to increase beaver abundance because of the positive effects beavers can have on ecosystems and because beavers are an important source of food for boreal carnivores. For example, we found that wolverines (Gulo gulo) used borrow pits as hunting grounds for beavers and one wolverine denned within a beaver lodge at a borrow pit. Conversely, managers might want to reduce beaver populations living at borrow pits because of the damage that beavers can cause to infrastructure and because subsidized beaver populations might alter predator-prey dynamics. To accomplish this, we suggest reducing available vegetation around borrow pits, isolating new borrow pits from natural beaver habitats, digging borrow pits where there is less chance of having surface water, filling in borrow pits with substrate, or not creating borrow pits and using a centralized gravel pit or mine for substrate.

Conservation Reserve Program is a key element for managing white-tailed deer populations at multiple spatial scales.

Understanding the underlying mechanisms driving population demographics such as species-habitat relationships and the spatial scale in which these relationships occur is essential for developing optimal management strategies. Here we evaluated how landscape characteristics and winter severity measured at three spatial scales (1 km2, 9 km2, and hunting unit) influenced white-tailed deer occurrence and abundance across North Dakota by using 10 years of winter aerial survey data and generalized linear mixed effects models. In general, forest, wetland, and Conservation Reserve Program (CRP) lands were the main drivers of deer occurrence and abundance in most of the spatial scales analyzed. However, the effects of habitat features vary between the home-range scale (9 km2) and the finer spatial scale (1 km2; i.e., within home ranges). While escape cover was the main factor driving white-tailed deer occurrence and abundance at broad spatial scales, at a fine spatial scale deer also selected for food (mainly residual winter cropland). With CRP appearing in nearly all top models, here we had strong evidence that this type of program will be fundamental to sustaining populations of white-tailed deer that can meet recreational demands. In addition, land managers should focus on ways to protect other escape covers (e.g., forest and wetland) on a broad spatial scale while encouraging landowners to supply winter resources at finer spatial scales. We therefore suggest a spatial multi-scale approach that involves partnerships among landowners and government agencies for effectively managing white-tailed deer.

Coexistence with Large Carnivores Supported by a Predator-Compensation Program.

Compensation programs are used globally to increase tolerance for and help offset economic loss caused by large carnivores. Compensation program funding comes from a variety of sources, and in Wyoming and Idaho, USA and Alberta, Canada this includes revenue from hunting and fishing license sales. We review the patterns of livestock depredation and compensation costs of Alberta's predator-compensation program, and compare Alberta's program to compensation programs in neighboring Canadian and American jurisdictions. Current compensation costs in Alberta are well below historic levels, but have been rapidly increasing in recent years due to an increase in depredation events coupled with increased cattle prices. That increase has caused push back from Alberta's hunting and fishing community that finances the compensation program, although less than 3.6% of Alberta's license levy dollars are used for predator compensation. Hunting effort in Alberta is highest on the same privately owned lands with livestock depredation problems, suggesting that private lands support habitats for hunted ungulate species as well as carnivores. Although compensation programs do not prevent depredation events themselves, compensation programs effectively can support the maintenance of wildlife habitats on private lands.

Inter-segment complementarity in orbiviruses: a driver for co-ordinated genome packaging in the Reoviridae?

The process by which eukaryotic viruses with segmented genomes select a complete set of genome segments for packaging into progeny virus particles is not understood. In this study a model based on the association of genome segments through specific RNA-RNA interactions driven by base pairing was formalized and tested in the Orbivirus genus of the Reoviridae family. A strategy combining screening of the genomic sequences for inter-segment complementarity with direct functional testing of inter-segment RNA-RNA interactions using reverse genetics is described in the type species of the Orbivirus genus, Bluetongue virus (BTV). Two examples, involving four of the ten BTV genomic segments, of specific inter-segment interaction motifs whose maintenance is essential for the generation of infectious virus, were identified. Equivalent inter-segment complementarities were found between the identified regions of the orthologous genome segments of all orbiviruses, including phylogenetically distant species. Specific interaction of the participating RNA segments was confirmed in vitro using electrophoretic mobility shift assays, with the interactions inhibited using oligonucleotides complementary to the interaction motif of one of the interacting partners, and also through mutagenesis of the motifs. In each example, the base pairing rather than the absolute sequence was critical to the formation of a functional inter-segment interaction, with mutations only being tolerated in rescued virus if compensating changes were made in the interacting partner to restore uninterrupted base pairing. The absolute sequence of the complementarity motifs varied between species, indicating that this newly identified phenomenon may contribute to the observed lack of reassortment between Orbivirus species.