Microbiomes, diet flexibility, and the spread of a beetle parasite of honey bees.

Invasive pests may disturb and destructively reformat the local ecosystem. The small hive beetle (SHB), Aethina tumida, originated in Africa and has expanded to America, Australia, Europe, and Asia. A key factor facilitating its fast global expansion is its ability to subsist on diverse food inside and outside honey bee colonies. SHBs feed on various plant fruits and exudates in the environment while searching for bee hives. After sneaking into a bee hive, they switch their diet to honey, pollen, and bee larvae. How SHBs survive on such a broad range of food remains unclear. In this study, we simulated the outside and within hive stages by providing banana and hive resources and quantified the SHB associated microbes adjusted by the diet. We found that SHBs fed on bananas were colonized by microbes coding more carbohydrate-active enzymes and a higher alpha diversity than communities from SHBs feeding on hive products or those collected directly from bee hives. SHBs fed on bananas and those collected from the hive showed high symbiont variance, indicated by the beta diversity. Surprisingly, we found the honey bee core symbiont Snodgrassella alvi in the guts of SHBs collected in bee hives. To determine the role of S. alvi in SHB biology, we inoculated SHBs with a genetically tagged culture of S. alvi, showing that this symbiont is a likely transient of SHBs. In contrast, the fungus Kodamaea ohmeri is the primary commensal of SHBs. Diet-based microbiome shifts are likely to play a key role in the spread and success of SHBs.

Promotion of Th17 Polarized Immunity via Co-Delivery of Mincle Agonist and Tuberculosis Antigen Using Silica Nanoparticles.

Adjuvants and immunomodulators that effectively drive a Th17-skewed immune response are not part of the standard vaccine toolkit. Vaccine adjuvants and delivery technologies that can induce Th17 or Th1/17 immunity and protection against bacterial pathogens, such as tuberculosis (TB), are urgently needed. Th17-polarized immune response can be induced using agonists that bind and activate C-type lectin receptors (CLRs) such as macrophage inducible C-type lectin (Mincle). A simple but effective strategy was developed for codelivering Mincle agonists with the recombinant Mycobacterium tuberculosis fusion antigen, M72, using tunable silica nanoparticles (SNP). Anionic bare SNP, hydrophobic phenyl-functionalized SNP (P-SNP), and cationic amine-functionalized SNP (A-SNP) of different sizes were coated with three synthetic Mincle agonists, UM-1024, UM-1052, and UM-1098, and evaluated for adjuvant activity in vitro and in vivo. The antigen and adjuvant were coadsorbed onto SNP via electrostatic and hydrophobic interactions, facilitating multivalent display and delivery to antigen presenting cells. The cationic A-SNP showed the highest coloading efficiency for the antigen and adjuvant. In addition, the UM-1098-adsorbed A-SNP formulation demonstrated slow-release kinetics in vitro, excellent stability over 12 months of storage, and strong IL-6 induction from human peripheral blood mononuclear cells. Co-adsorption of UM-1098 and M72 on A-SNP significantly improved antigen-specific humoral and Th17-polarized immune responses in vivo in BALB/c mice relative to the controls. Taken together, A-SNP is a promising platform for codelivery and proper presentation of adjuvants and antigens and provides the basis for their further development as a vaccine delivery platform for immunization against TB or other diseases for which Th17 immunity contributes to protection.

Vaccination with Mincle agonist UM-1098 and mycobacterial antigens induces protective Th1 and Th17 responses.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is one of the top infectious killers in the world. The only licensed vaccine against TB, Bacille Calmette-Guérin (BCG), provides variable protection against pulmonary TB, especially in adults. Hence, novel TB vaccine approaches are urgently needed. Both Th1 and Th17 responses are necessary for protection against TB, yet effective adjuvants and vaccine delivery systems for inducing robust Th1 and Th17 immunity are lacking. Herein we describe a synthetic Mincle agonist, UM-1098, and a silica nanoparticle delivery system that drives Th1/Th17 responses to Mtb antigens. Stimulation of human peripheral blood mononuclear cells (hPBMCs) with UM-1098 induced high levels of Th17 polarizing cytokines IL-6, IL-1ß, IL-23 as well as IL-12p70, IL-4 and TNF-α in vitro. PBMCs from both C57BL/6 and BALB/c mice responded with a similar cytokine pattern in vitro and in vivo. Importantly, intramuscular (I.M.) vaccination with UM-1098-adjuvanted TB antigen M72 resulted in significantly higher antigen-specific IFN-γ and IL-17A levels in C57BL/6 wt mice than Mincle KO mice. Vaccination of C57BL/6 wt mice with immunodominant Mtb antigens ESAT6/Ag85B or M72 resulted in predominantly Th1 and Th17 responses and induced antigen-specific serum antibodies. Notably, in a virulent Mtb challenge model, vaccination with UM-1098 adjuvanted ESAT6/Ag85B or M72 significantly reduced lung bacterial burden when compared with unvaccinated mice and protection occurred in the absence of pulmonary inflammation. These data demonstrate that the synthetic Mincle agonist UM-1098 induces strong Th1 and Th17 immunity after vaccination with Mtb antigens and provides protection against Mtb infection in mice.

The recent revision of the genera Nosema and Vairimorpha (Microsporidia: Nosematidae) was flawed and misleads the bee scientific community.

The genus Vairimorpha was proposed for several species of Nosema in 1976 (Pilley, 1976), almost 70 years after Nosema apis Zander (Zander, 1909). Tokarev and colleagues proposed the redefinition of 17 microsporidian species in four genera, Nosema, Vairimorpha, Rugispora, and Oligosporidium, based on phylogenetic trees of two genetic markers (SSU rRNA and RPB1) (Tokarev et al., 2020). Several issues should invalidate this new classification, leading to the synonymization of Vairimorpha within Nosema.

Adjuvanted Vaccine Induces Functional Antibodies against Pseudomonas aeruginosa Filamentous Bacteriophages.

Pseudomonas aeruginosa (Pa), a WHO priority 1 pathogen, resulted in approximately 559,000 deaths globally in 2019. Pa has a multitude of host-immune evasion strategies that enhance Pa virulence. Most clinical isolates of Pa are infected by a phage called Pf that has the ability to misdirect the host-immune response and provide structural integrity to biofilms. Previous studies demonstrate that vaccination against the coat protein (CoaB) of Pf4 virions can assist in the clearance of Pa from the dorsal wound model in mice. Here, a consensus peptide was derived from CoaB and conjugated to cross-reacting material 197 (CRM197). This conjugate was adjuvanted with a novel synthetic Toll-like receptor agonist (TLR) 4 agonist, INI-2002, and used to vaccinate mice. Mice vaccinated with CoaB-CRM conjugate and INI-2002 developed high anti-CoaB peptide-specific IgG antibody titers. Direct binding of the peptide-specific antibodies to whole-phage virus particles was demonstrated by ELISA. Furthermore, a functional assay demonstrated that antibodies generated from vaccinated mice disrupted the replicative cycle of Pf phages. The use of an adjuvanted phage vaccine targeting Pa is an innovative vaccine strategy with the potential to become a new tool targeting multi-drug-resistant Pa infections in high-risk populations.

Case Report: Emerging Losses of Managed Honey Bee Colonies.

United States commercial beekeepers prepare honey bee colonies for almond pollination in California each year in late January to early February. This represents the largest managed pollination event in the world and involves more than half of all U.S. honey bee colonies. In winter 2023, numerous colonies in Florida, which were graded as suitable for almonds (larger than ten frames of bees), dwindled suddenly or altogether died within several weeks, just prior to movement for almonds. The timing of these losses and the resulting morbidity caused severe economic harm to affected operations. This study reports interviews with affected stakeholders, their economic harm, and analyses of pathogens and parasites found in their colonies.

Susceptible and infectious states for both vector and host in a dynamic pathogen-vector-host system.

Deformed wing virus (DWV) is a resurgent insect pathogen of honeybees that is efficiently transmitted by vectors and through host social contact. Continual transmission of DWV between hosts and vectors is required to maintain the pathogen within the population, and this vector-host-pathogen system offers unique disease transmission dynamics for pathogen maintenance between vectors and a social host. In a series of experiments, we measured vector-vector, host-host and host-vector transmission routes and show how these maintain DWV in honeybee populations. We found co-infestations on shared hosts allowed for movement of DWV from mite to mite. Additionally, two social behaviours of the honeybee, trophallaxis and cannibalization of pupae, provide routes for horizontal transmission from bee to bee. Circulation of the virus solely among hosts through communicable modes provides a reservoir of DWV for naïve Varroa to acquire and subsequently vector the pathogen. Our findings illustrate the importance of community transmission between hosts and vector transmission. We use these results to highlight the key avenues used by DWV during maintenance and infection and point to similarities with a handful of other infectious diseases of zoonotic and medical importance.

Co-Delivery of a Novel Lipidated TLR7/8 Agonist and Hemagglutinin-Based Influenza Antigen Using Silica Nanoparticles Promotes Enhanced Immune Responses.

Co-delivery of antigens and adjuvants to the same antigen-presenting cells (APCs) can significantly improve the efficacy and safety profiles of vaccines. Here, we report amine-grafted silica nanoparticles (A-SNP) as a tunable vaccine co-delivery platform for TLR7/8 agonists along with the recombinant influenza antigen hemagglutinin H7 (H7) to APCs. A-SNP of two different sizes (50 and 200 nm) were prepared and coated with INI-4001 at different coating densities, followed by co-adsorption of H7. Both INI-4001 and H7 showed >90% adsorption to the tested A-SNP formulations. TNF-α and IFN-α cytokine release by human peripheral blood mononuclear cells as well as TNF-α, IL-6, and IL-12 release by mouse bone marrow-derived dendritic cells revealed that the potency of the INI-4001-adsorbed A-SNP (INI-4001/A-SNP) formulations was improved relative to aqueous formulation control. This improved potency was dependent on particle size and ligand coating density. In addition, slow-release profiles of INI-4001 were measured from INI-4001/A-SNP formulations in plasma with 30-50% INI-4001 released after 7 days. In vivo murine immunization studies demonstrated significantly improved H7-specific humoral and Th1/Th17-polarized T cell immune responses with no observed adverse reactions. Low-density 50 nm INI-4001/A-SNP elicited significantly higher IFN-γ and IL-17 induction over that of the H7 antigen-only group and INI-4001 aqueous formulation controls. In summary, this work introduces an effective and biocompatible SNP-based co-delivery platform that enhances the immunogenicity of TLR7/8 agonist-adjuvanted subunit influenza vaccines.

Host switch by honey bee parasitic mites leads to symbiont diversification.

Host-parasite co-evolution is a reciprocal genetic change; however, the parasite may switch to a novel host, deviating from conventional co-evolution. Varroa destructor is a native parasite of the honey bee Apis cerana, and the mite has established infestation in another honey bee, Apis mellifera, causing colony failure. When mites switched to the novel host, they formed a distinct population from mites that remained on the native host. Consequently, this led to divergence in the microbiota associated with mites in two host populations. The microbes were conserved at the species level reflected by alpha diversity, with substantial relative abundance variance. Microbes found in mites were distinct from the bee microbiota. They mainly were pathogenic with antibiotic resistance, while a few bacterial taxa were previously found in honey bees, including Klebsiella pneumoniae and Pseudomanas aeruginosa. These symbionts may transfer between the mites and honey bees.

Bacterial diversity using metagenomics of 16s rDNA in water kefir, an innovative source of probiotics for bee nutrition.

Water kefir is a sparkling, slightly acidic fermented beverage made from sugar, water, and water kefir grains, which are a mixture of yeast and bacteria. These grains produce a variety of fermentation compounds such as lactic acid, acetaldehyde, acetoin, ethanol and carbon dioxide. In this study, a high-throughput sequencing technique was used to characterize the bacterial composition of the original water kefir from which potential probiotics were obtained. We studied the bacterial diversity of both water kefir grains and beverages. DNA was extracted from three replicate samples of both grains and beverages using the Powerlyzer Microbial Kit. The hypervariable V1-V2 region of the bacterial 16S ribosomal RNA gene was amplified to prepare six DNA libraries. Between 1.4M and 2.4M base-pairs were sequenced for the library. In total, 28721971 raw reads were obtained from all the samples. Estimated species richness was higher in kefir beverage samples compared to grain samples. Moreover, a higher level of microbial alpha diversity was observed in the beverage samples. Particularly, the predominant bacteria in beverages were Anaerocolumna and Ralstonia, while in grains Liquorilactobacillus dominated, with lower levels of Leuconostoc and Oenococcus. Although the bacterial diversity in kefir grains was low because only three genera were the most represented, all of them are LAB bacteria with the potential to serve as probiotics in the artificial feeding of bees.

Mapping Respiratory Health Digital Interventions in South and Southeast Asia: Protocol for a Scoping Review.

BACKGROUND: The last 2 decades have been a time of exponential growth and maturation for digital health, while the global burden of respiratory disease continues to grow worldwide. Leveraging digital health interventions (DHIs) to manage and mitigate respiratory disease and its adverse health effects presents itself as an obvious path forward. OBJECTIVE: We aimed to understand the current digital landscape and enabling environment around respiratory health to reduce costs, avoid duplication, and understand the comprehensiveness of DHIs. METHODS: This study will follow a scoping review methodology as outlined by Arksey and O'Malley, the Joanna Briggs Institute, and the PRISMA-ScR (Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews) checklist. MEDLINE, Embase, CINAHL, PsycINFO, Cochrane Library, Web of Science, PakiMedNet, and MyMedR databases will be searched along with key websites, repositories, and gray literature databases. The terms "respiratory health," "digital health," "South Asia," and "Southeast Asia," as well as related terms will be searched. The results will be screened for duplicates and then against the inclusion and exclusion criteria. For the studies included, data will be extracted, collated, and analyzed. RESULTS: The scoping review was started in July 2023 and will be finalized by February 2024. Results will be presented following the World Health Organization's classification of DHIs to categorize interventions in a standardized format and the mobile health evidence reporting and assessment checklist to report on the effectiveness of interventions. Further exposition of the evidence extracted will be presented through narrative synthesis. CONCLUSIONS: As DHIs continue to proliferate, the need to understand the current landscape becomes more pertinent. In this scoping review, we will seek to more clearly understand what digital health tools and technologies are being used in the current landscape of digital health in South and Southeast Asia for respiratory health and to what extent they are addressing the respiratory health needs of the region. The results will inform recommendations on digital health tools for respiratory health in South and Southeast Asia will help funders and implementers of DHIs leverage existing technologies and accelerate innovations that address documented gaps in the studied countries. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/52517.

Prevalence and distribution of Varroa destructor and Nosema spp. in symptomatic honey bee colonies across the USA from 2015 to 2022.

USDA-ARS Bee Research Laboratory received symptomatic honey bee (Apis mellifera L.) samples across the United States for disease diagnosis. Here, we present a retrospective study and cartography of ectoparasite Varroa destructor and intracellular microsporidia parasite Nosema spp. These two major parasites were identified in the diseased honey bee samples between 2015 and 2022. Varroa infestation level (VIL) was examined by a wash technique (Mites/100 bees) and calculated as a percentage, while Nosema infection was quantified by microscopical spore count (Million Spores/Bee). Data were analyzed by month, year, state, and by nine geographical climate regions described in the U.S. Of adult bee samples (n = 4039) that were analyzed for Varroa mite infestation, the overall VIL in the U.S. ranged between 0.4 and 30.85%, with an overall national VIL and Varroa prevalence of 8.21% and 85.14%, respectively. Overall monthly data showed VIL constantly exceeded the critical level of 4% except from June to September and reached a maximum of 15% in January and December. Nationwide, VIL significantly (p < 0.001) increased from 2015 to 2018 (1.1-4.7%), plateaued from 2018 to 2021 (4.7-4.5%), followed by a significant decrease in 2022 (3.6%). Significant VIL differences (p < 0.001) were recorded among climate regions, with the highest mite infestation levels in the Upper Midwest region (13.9%) and the lowest in the West region (5.1%). Of adult bee samples (n = 2,994) that were analyzed for Nosema infection, Nosema spore count ranged between (1-16.8) million spores per bee among states, with a national average of 6.8 and a prevalence of 99.7%. The lowest and highest Nosema loads were respectively recorded in the South region (3.1) and Upper Midwest (10.5), a significant difference (p < 0.001). No statistical differences were recorded among the six other climate regions. Overall, VIL and Nosema infection correlated significantly (p < 0.001) with a regression coefficient of (R2 = 0.6). Our data, which originated from ailing bee colonies, showed significantly higher rates of maladies compared to data from healthy colonies obtained by the USDA-APHIS National Honey Bee Survey, demonstrating the role of bee diseases caused by Varroa mite and Nosema in honey bee population declines.

High temperatures augment inhibition of parasites by a honey bee gut symbiont.

Temperature affects growth, metabolism, and interspecific interactions in microbial communities. Within animal hosts, gut bacterial symbionts can provide resistance to parasitic infections. Both infection and populations of symbionts can be shaped by the host body temperature. However, the effects of temperature on the antiparasitic activities of gut symbionts have seldom been explored. The Lactobacillus-rich gut microbiota of facultatively endothermic honey bees is subject to seasonal and ontogenetic changes in host temperature that could alter the effects of symbionts against parasites. We used cell cultures of a Lactobacillus symbiont and an important trypanosomatid gut parasite of honey bees to test the potential for temperature to shape parasite-symbiont interactions. We found that symbionts showed greater heat tolerance than parasites and chemically inhibited parasite growth via production of acids. Acceleration of symbiont growth and acid production at high temperatures resulted in progressively stronger antiparasitic effects across a temperature range typical of bee colonies. Consequently, the presence of symbionts reduced both the peak growth rate and heat tolerance of parasites. Substantial changes in parasite-symbiont interactions were evident over a temperature breadth that parallels changes in diverse animals exhibiting infection-related fevers and the amplitude of circadian temperature variation typical of endothermic birds and mammals, implying the frequent potential for temperature to alter symbiont-mediated resistance to parasites in endo- and ectothermic hosts. Results suggest that the endothermic behavior of honey bees could enhance the impacts of gut symbionts on parasites, implicating thermoregulation as a reinforcer of core symbioses and possibly microbiome-mediated antiparasitic defense. IMPORTANCE Two factors that shape the resistance of animals to infection are body temperature and gut microbiota. However, temperature can also alter interactions among microbes, raising the question of whether and how temperature changes the antiparasitic effects of gut microbiota. Honey bees are agriculturally important hosts of diverse parasites and infection-mitigating gut microbes. They can also socially regulate their body temperatures to an extent unusual for an insect. We show that high temperatures found in honey bee colonies augment the ability of a gut bacterial symbiont to inhibit the growth of a common bee parasite, reducing the parasite's ability to grow at high temperatures. This suggests that fluctuations in colony and body temperatures across life stages and seasons could alter the protective value of bees' gut microbiota against parasites, and that temperature-driven changes in gut microbiota could be an underappreciated mechanism by which temperature-including endothermy and fever-alters animal infection.

Diamino Allose Phosphates: Novel, Potent, and Highly Stable Toll-like Receptor 4 Agonists.

Most known synthetic toll-like receptor 4 (TLR4) agonists are carbohydrate-based lipid-A mimetics containing several fatty acyl chains, including a labile 3-O-acyl chain linked to the C-3 position of the non-reducing sugar known to undergo cleavage impacting stability and resulting in loss of activity. To overcome this inherent instability, we rationally designed a new class of chemically more stable synthetic TLR4 ligands that elicit robust innate and adaptive immune responses. This new class utilized a diamino allose phosphate (DAP) scaffold containing a nonhydrolyzable 3-amide bond instead of the classical 3-ester. Accordingly, the DAPs have significantly improved thermostability in aqueous formulations and potency relative to other known natural and synthetic TLR4 ligands. Furthermore, the DAP analogues function as potent vaccine adjuvants to enhance influenza-specific antibodies in mice and provide protection against lethal influenza virus challenges. This novel set of TLR4 ligands show promise as next-generation vaccine adjuvants and stand-alone immunomodulators.

Self-Adjuvanting TLR7/8 Agonist and Fentanyl Hapten Co-Conjugate Achieves Enhanced Protection against Fentanyl Challenge.

Currently approved pharmacotherapies for opioid use disorders (OUDs) and overdose reversal agents are insufficient to slow the spread of OUDs due to the proliferation of fentanyl. This is evident in the 31% rise in drug overdose deaths from 2019 to 2022, with rates increasing from 21.6 to 28.3 overdoses per 100,000 deaths. Vaccines are a potential alternative or adjunct therapy for the treatment of several substance use disorders (nicotine, cocaine) but have shown limited clinical success due to suboptimal antibody titers. In this study, we demonstrate that coconjugation of a Toll-like receptor 7/8 (TLR7/8) agonist (UM-3006) alongside a fentanyl-based hapten (F1) on the surface of the carrier protein cross-reactive material 197 (CRM) significantly increased generation of high-affinity fentanyl-specific antibodies. This demonstrated enhanced protection against fentanyl challenges relative to an unconjugated (admix) adjuvant control in mice. Inclusion of aluminum hydroxide (alum) adjuvant further increased titers and enhanced protection, as determined by analysis of fentanyl concentration in serum and brain tissue. Collectively, our findings present a promising approach to enhance the efficacy of antiopioid vaccines, underscoring the need for extensive exploration of TLR7/8 agonist conjugates as a compelling strategy to combat opioid use disorders.

Widespread distribution of honey bee-associated pathogens in native bees and wasps: Trends in pathogen prevalence and co-occurrence.

Pollinators have experienced significant declines in the past decade, in part due to emerging infectious diseases. Historically, studies have primarily focused on pathogens in the Western honey bee, Apis mellifera. However, recent work has demonstrated that these pathogens are shared by other pollinators and can negatively affect their health. Here, we surveyed honey bees and 15 native bee and wasp species for 13 pathogens traditionally associated with honey bees. The native bee and wasp species included 11 species not previously screened for pathogens. We found at least one honey bee-associated pathogen in 53% of native bee and wasp samples. The most widely distributed and commonly detected pathogens were the microsporidian Nosema ceranae, the bacterium Melissococcus plutonius, and the viruses deformed wing virus and black queen cell virus. The prevalence of viruses was generally higher in honey bees than in native bees and wasps. However, the prevalence of M. plutonius and the brood fungus Ascosphaera apis was significantly higher in some native bee species than in honey bees. The data also reveal novel trends in the association between co-occurring pathogens in honey bees and native bees and wasps at the pathogen community level. These results can inform the assessment of risks that native pollinator species face from pathogen stress, and indicate that many non-viral pathogens, notably M. plutonius and N. ceranae, are far more widely distributed and commonly found in native bees and wasps than previously thought.

A lipidated TLR7/8 adjuvant enhances the efficacy of a vaccine against fentanyl in mice.

Opioid use disorders (OUD) and opioid-related fatal overdoses are a public health concern in the United States. Approximately 100,000 fatal opioid-related overdoses occurred annually from mid-2020 to the present, the majority of which involved fentanyl or fentanyl analogs. Vaccines have been proposed as a therapeutic and prophylactic strategy to offer selective and long-lasting protection against accidental or deliberate exposure to fentanyl and closely related analogs. To support the development of a clinically viable anti-opioid vaccine suitable for human use, the incorporation of adjuvants will be required to elicit high titers of high-affinity circulating antibodies specific to the target opioid. Here we demonstrate that the addition of a synthetic TLR7/8 agonist, INI-4001, but not a synthetic TLR4 agonist, INI-2002, to a candidate conjugate vaccine consisting of a fentanyl-based hapten, F1, conjugated to the diphtheria cross-reactive material (CRM), significantly increased generation of high-affinity F1-specific antibody concentrations, and reduced drug distribution to the brain after fentanyl administration in mice.

Incentivizing sustainable fire management in Australia's northern arid spinifex grasslands.

Fire management across Australia's fire-prone 1.2 M km2 northern savannas region has been transformed over the past decade supported by the inception of Australia's national regulated emissions reduction market in 2012. Today, incentivised fire management is undertaken over a quarter of that entire region, providing a range of socio-cultural, environmental, and economic benefits, including for remote Indigenous (Aboriginal and Torres Strait Islander) communities and enterprises. Building on those advances, here we explore the emissions abatement potential for expanding incentivised fire management opportunities to include a contiguous fire-prone region, extending to monsoonal but annually lower (<600 mm) and more variable rainfall conditions, supporting predominantly shrubby spinifex (Triodia) hummock grasslands characteristic of much of Australia's deserts and semi-arid rangelands. Adapting a standard methodological approach applied previously for assessing savanna emissions parameters, we first describe fire regime and associated climatic attributes for a proposed ∼850,000 km2 lower rainfall (600-350 mm MAR) focal region. Second, based on regional field assessments of seasonal fuel accumulation, combustion, burnt area patchiness, and accountable methane and nitrous oxide Emission Factor parameters, we find that significant emissions abatement is feasible for regional hummock grasslands. This applies specifically for more frequently burnt sites under higher rainfall conditions if substantial early dry season prescribed fire management is undertaken resulting in marked reduction in late dry season wildfires. The proposed Northern Arid Zone (NAZ) focal envelope is substantially under Indigenous land ownership and management, and in addition to reducing emissions impacts associated with recurrent extensive wildfires, development of commercial landscape-scale fire management opportunities would significantly support social, cultural and biodiversity management aspirations as promoted by Indigenous landowners. Combined with existing regulated savanna fire management regions, inclusion of the NAZ under existing legislated abatement methodologies would effectively provide incentivised fire management covering a quarter of Australia's landmass. This could complement an allied (non-carbon) accredited method valuing combined social, cultural and biodiversity outcomes from enhanced fire management of hummock grasslands. Although the management approach has potential application to other international fire-prone savanna grasslands, caution is required to ensure that such practice does not result in irreversible woody encroachment and undesirable habitat change.

Apis mellifera Solinvivirus-1, a Novel Honey Bee Virus That Remained Undetected for over a Decade, Is Widespread in the USA.

A metagenomic analysis of the virome of honey bees (Apis mellifera) from an apiary with high rates of unexplained colony losses identified a novel RNA virus. The virus, which was named Apis mellifera solinvivirus 1 (AmSV1), contains a 10.6 kb positive-strand genomic RNA with a single ORF coding for a polyprotein with the protease, helicase, and RNA-dependent RNA polymerase domains, as well as a single jelly-roll structural protein domain, showing highest similarity with viruses in the family Solinviviridae. The injection of honey bee pupae with AmSV1 preparation showed an increase in virus titer and the accumulation of the negative-strand of AmSV1 RNA 3 days after injection, indicating the replication of AmSV1. In the infected worker bees, AmSV1 was present in heads, thoraxes, and abdomens, indicating that this virus causes systemic infection. An analysis of the geographic and historic distribution of AmSV1, using over 900 apiary samples collected across the United States, showed AmSV1 presence since at least 2010. In the year 2021, AmSV1 was detected in 10.45% of apiaries (95%CI: 8.41-12.79%), mostly sampled in June and July in Northwestern and Northeastern United States. The diagnostic methods and information on the AmSV1 distribution will be used to investigate the connection of AmSV1 to honey bee colony losses.

A TLR7/8 agonist increases efficacy of anti-fentanyl vaccines in rodent and porcine models.

Opioid use disorders (OUD) and overdose are public health threats worldwide. Widespread access to highly potent illicit synthetic opioids such as fentanyl is driving the recent rise in fatal overdoses. Vaccines containing fentanyl-based haptens conjugated to immunogenic carrier proteins offer a long-lasting, safe, and cost-effective strategy to protect individuals from overdose upon accidental or deliberate exposure to fentanyl and its analogs. Prophylactic or therapeutic active immunization with an anti-fentanyl vaccine induces the production of fentanyl-specific antibodies that bind the drug in the blood and prevent its distribution to the brain, which reduces its reinforcing effects and attenuates respiratory depression and bradycardia. To increase the efficacy of a lead anti-fentanyl vaccine, this study tested whether the incorporation of synthetic toll-like receptor (TLR) 4 and TLR7/8 agonists as vaccine adjuvants would increase vaccine efficacy against fentanyl challenge, overdose, and self-administration in either rats or Hanford miniature pigs. Formulation of the vaccine with a nucleolipid TLR7/8 agonist enhanced its immunogenicity and efficacy in preventing fentanyl-induced respiratory depression, analgesia, bradycardia, and self-administration in either rats or mini-pigs. These studies support the use of TLR7/8 adjuvants in vaccine formulations to improve their clinical efficacy against OUD and potentially other substance use disorders (SUD).