Arctic ground squirrels (Urocitellus parryii) hibernate for several months without eating or drinking yet suffer no disuse atrophy. We are investigating the potential contributions of gut microorganisms to host nitrogen homeostasis, and here, we describe the genome assemblies of 35 isolated bacteria collected from gastrointestinal material and sequenced using Nanopore technology.

Neodothiora populina is a black yeast-like fungus in the family of Dothioraceae. It causes an aggressive canker disease of trembling aspen that results in widespread mortality of aspen across the boreal forest of Interior Alaska. Here we report a high-quality draft genome including functional annotation of this emerging fungal pathogen based on Nanopore Technology longread sequences. Our initial genome assembly totaled 23,960,169 bp and contained 18 contigs and we identified 7,343 genes. This resource announcement provides new genomic data useful long-term to improve our understanding of forest health in Alaska.

ABSTRACTArbuscular mycorrhizal fungi (AMF, phylum Glomeromycota) are essential to plant community diversity and ecosystem functioning. However, increasing human land use represents a major threat to native AMF globally. Characterizing the loss of AMF diversity remains challenging because many taxa are undescribed, resulting in poor documentation of their biogeography and family‐level disturbance sensitivity. We survey sites representing native and human‐altered ecosystems across the American continents—in Alaska, Kansas, and Brazil—to shed light on these gaps. Using a recently developed pipeline for phylogenetic placement of eDNA, we find evidence for three putative novel clades within the Glomeromycota, sister to Entrophosporaceae, Glomeraceae, and Archaeosporaceae, with evidence for geographic structuring. We further find that taxa in the Diversisporaceae, Glomeraceae, and Entrophosporaceae relatively high families are overrepresented and more diverse in temperate samples. By contrast, the diversity of taxa that cannot be placed into a family is higher in tropical samples, suggesting that tropical sites harbor relatively high undescribed AMF diversity. Moreover, we find evidence that Entrophosporaceae is more tolerant, while Glomeraceae is more sensitive to disturbance. These results underscore the vast undescribed diversity of AMF while highlighting a way forward to systematically improve our understanding of AMF biogeography and response to human disturbance.

Seasonal affective disorder (SAD) is a recurrent depression triggered by exposure to short photoperiods, with a subset of patients reporting hypersomnia, increased appetite, and carbohydrate craving. Dysfunction of the microbiota - gut - brain axis is frequently associated with depressive disorders, but its role in SAD is unknown. Nile grass rats (Arvicanthis niloticus) are potentially useful for exploring the pathophysiology of SAD, as they are diurnal and have been found to exhibit anhedonia and affective-like behavior in response to short photoperiods. Further, given grass rats have been found to spontaneously develop metabolic syndrome, they may be particularly susceptible to environmental triggers of metabolic dysbiosis. We conducted a 2 × 2 factorial design experiment to test the effects of short photoperiod (4 h:20 h Light:Dark (LD) vs. neutral 12:12 LD), access to a high concentration (8%) sucrose solution, and the interaction between the two, on activity, sleep, liver steatosis, and the gut microbiome of grass rats. We found that animals on short photoperiods maintained robust diel rhythms and similar subjective day lengths as controls in neutral photoperiods but showed disrupted activity and sleep patterns (i.e. a return to sleep after an initial bout of activity that occurs ~ 13 h before lights off). We found no evidence that photoperiod influenced sucrose consumption. By the end of the experiment, some grass rats were overweight and exhibited signs of non-alcoholic fatty liver disease (NAFLD) with micro- and macro-steatosis. However, neither photoperiod nor access to sucrose solution significantly affected the degree of liver steatosis. The gut microbiome of grass rats varied substantially among individuals, but most variation was attributable to parental effects and the microbiome was unaffected by photoperiod or access to sucrose. Our study indicates short photoperiod leads to disrupted activity and sleep in grass rats but does not impact sucrose consumption or exacerbate metabolic dysbiosis and NAFLD.

Genomes of a Campylobacter coli OR12-like strain ChP2023 (1,713,995 bp) isolated from broiler chicken and Campylobacter jejuni subsp. jejuni NCTC 11168R-like strain KF2023 (1,729,995 bp) isolated from turkey, from poultry production facilities in Ukraine in 2023, were sequenced using Oxford Nanopore Technologies. Both genomes included antibiotic resistance genes and other virulence factors.

Antimicrobial resistance (AMR) is a global public health threat. Environmental microbial communities act as reservoirs for AMR, containing genes associated with resistance, their precursors, and the selective pressures promoting their persistence. Genomic surveillance could provide insights into how these reservoirs change and impact public health. Enriching for AMR genomic signatures in complex microbial communities would strengthen surveillance efforts and reduce time-to-answer. Here, we tested the ability of nanopore sequencing and adaptive sampling to enrich for AMR genes in a mock community of environmental origin. Our setup implemented the MinION mk1B, an NVIDIA Jetson Xavier GPU, and Flongle flow cells. Using adaptive sampling, we observed consistent enrichment by composition. On average, adaptive sampling resulted in a target composition 4× higher than without adaptive sampling. Despite a decrease in total sequencing output, adaptive sampling increased target yield in most replicates. We also demonstrate enrichment in a diverse community using an environmental sample. This method enables rapid and flexible genomic surveillance.

African swine fever (ASF) is a viral disease, endemic to Africa, that causes high mortality when introduced into domestic pig populations. Since the emergence of p72-genotype II African swine fever virus (ASFV) in Georgia in 2007, an ASF epidemic has been spreading across Europe and many countries in Asia. The epidemic first reached Ukraine in 2012. To better understand the dynamics of spread of ASF in Ukraine, we analyzed spatial and temporal outbreak data reported in Ukraine between 2012 and mid-2023. The highest numbers of outbreaks were reported in 2017 (N = 163) and 2018 (N = 145), with overall peak numbers of ASF outbreaks reported in August (domestic pigs) and January (wild boars). While cases were reported from most of Ukraine, we found a directional spread from the eastern and northern borders towards the western and southern regions of Ukraine. Many of the early outbreaks (before 2016) were adjacent to the border, which is again true for more recent outbreaks in wild boar, but not for recent outbreaks in domestic pigs. Outbreaks prior to 2016 also occurred predominantly in areas with a below average domestic pig density. This new analysis suggests that wild boars may have played an important role in the introduction and early spread of ASF in Ukraine. However, in later years, the dynamic suggests human activity as the predominant driver of spread and a separation of ASF epizootics between domestic pigs and in wild boars. The decline in outbreaks since 2019 suggests that the implemented mitigation strategies are effective, even though long-term control or eradication remain challenging and will require continued intensive surveillance of ASF outbreak patterns.

Emerging RNA virus infections are a growing concern among domestic poultry industries due to the severe impact they can have on flock health and economic livelihoods. Avian paramyxoviruses (APMV; avulaviruses, AaV) are pathogenic, negative-sense RNA viruses that cause serious infections in the respiratory and central nervous systems. APMV was detected in multiple avian species during the 2017 wild bird migration season in Ukraine and studied using PCR, virus isolation, and sequencing. Of 4090 wild bird samples collected, mostly from southern Ukraine, eleven isolates were grown in ovo and identified for APMV serotype by hemagglutinin inhibition test as: APMV-1, APMV-4, APMV-6, and APMV-7. To build One Health’s capacity to characterize APMV virulence and analyze the potential risks of spillover to immunologically naïve populations, we sequenced virus genomes in veterinary research labs in Ukraine using a nanopore (MinION) platform. RNA was extracted and amplified using a multiplex tiling primer approach to specifically capture full-length APMV-1 (n = 5) and APMV-6 (n = 2) genomes at high read depth. All APMV-1 and APMV-6 fusion (F) proteins possessed a monobasic cleavage site, suggesting these APMVs were likely low virulence, annually circulating strains. Utilization of this low-cost method will identify gaps in viral evolution and circulation in this understudied but important critical region for Eurasia.

Alaska has the lowest population density in the United States (US) with a mix of urban centers and isolated rural communities. Alaska’s distinct population dynamics compared to the contiguous US may have contributed to unique patterns of SARS-CoV-2 variants observed in early 2021. Here we examined 2323 SARS-CoV-2 genomes from Alaska and 278,635 from the contiguous US collected from December 2020 through June 2021 because of the notable emergence and spread of lineage B.1.1.519 in Alaska. We found that B.1.1.519 was consistently detected from late January through June of 2021 in Alaska with a peak prevalence in April of 77.9% unlike the rest of the US at 4.6%. The earlier emergence of B.1.1.519 coincided with a later peak of Alpha (B.1.1.7) compared to the contiguous US. We also observed differences in variant composition over time between the two most populated regions of Alaska and a modest increase in COVID-19 cases during the peak incidence of B.1.1.519. However, it is difficult to disentangle how social dynamics conflated changes in COVID-19 during this time. We suggest that the viral characteristics, such as amino acid substitutions in the spike protein, likely contributed to the unique spread of B.1.1.519 in Alaska.

Alaska is a unique US state because of its large size, geographically disparate population density, and physical distance from the contiguous United States. Here, we describe a pattern of SARS-CoV-2 variant emergence across Alaska reflective of these differences. Using genomic data, we found that in Alaska, the Omicron sublineage BA.2.3 overtook BA.1.1 by the week of 27 February 2022, reaching 48.5% of sequenced cases. On the contrary, in the contiguous United States, BA.1.1 dominated cases for longer, eventually being displaced by BA.2 sublineages other than BA.2.3. BA.2.3 only reached a prevalence of 10.9% in the contiguous United States. Using phylogenetics, we found evidence of potential origins of the two major clades of BA.2.3 in Alaska and with logistic regression estimated how it emerged and spread throughout the state. The combined evidence is suggestive of founder events in Alaska and is reflective of how Alaska's unique dynamics influence the emergence of SARS-CoV-2 variants.

Microbes influence the surrounding environment and contribute to human health. Metagenomics can be used as a tool to explore the interactions between microbes. Metagenomic assemblies built using long read nanopore data depend on the read level accuracy. The read level accuracy of nanopore sequencing has made dramatic improvements over the past several years. However, we do not know if the increased read level accuracy allows for faster assemblers to make as accurate metagenomic assemblies as slower assemblers. Here, we present the results of a benchmarking study comparing three commonly used long read assemblers, Flye, Raven, and Redbean. We used a prepared DNA standard of seven bacteria as our input community. We prepared a sequencing library using a VolTRAX V2 and sequenced using a MinION mk1b. We basecalled with Guppy v5.0.7 using the super-accuracy model. We found that increasing read depth benefited each of the assemblers, and nearly complete community member chromosomes were assembled with as little as 10× read depth. Polishing assemblies using Medaka had a predictable improvement in quality. We found Flye to be the most robust across taxa and was the most effective assembler for recovering plasmids. Based on Flye’s consistency for chromosomes and increased effectiveness at assembling plasmids, we would recommend using Flye in future metagenomic studies.

Porcine circovirus type 2 (PCV2) is responsible for a number of porcine circovirus-associated diseases (PCVAD) that can severely impact domestic pig herds. For a non-enveloped virus with a small genome (1.7 kb ssDNA), PCV2 is remarkably diverse, with eight genotypes (a–h). New genotypes of PCV2 can spread through the migration of wild boar, which are thought to infect domestic pigs and spread further through the domestic pig trade. Despite a large swine population, the diversity of PCV2 genotypes in Ukraine has been under-sampled, with few PCV2 genome sequences reported in the past decade. To gain a deeper understanding of PCV2 genotype diversity in Ukraine, samples of blood serum were collected from wild boars (n = 107) that were hunted in Ukraine during the November–December 2012 hunting season. We found 34/107 (31.8%) prevalence of PCV2 by diagnostic PCR. For domestic pigs, liver samples (n = 16) were collected from a commercial market near Kharkiv in 2019, of which 6 out of 16 (37%) samples were positive for PCV2. We sequenced the genotyping locus ORF2, a gene encoding the PCV2 viral capsid (Cap), for 11 wild boar and six domestic pig samples in Ukraine using an Oxford Nanopore MinION device. Of 17 samples with resolved genotypes, the PCV2 genotype b was the most common in wild boar samples (10 out of 11, 91%), while the domestic pigs were infected with genotypes b and d. We also detected genotype b/d and b/a co-infections in wild boars and domestic pigs, respectively, and for the first time in Ukraine we detected genotype f in a wild boar from Poltava. Building a maximum-likelihood phylogeny, we identified a sublineage of PCV2 genotype b infections in both wild and domestic swine, suggesting a possible epizootic cluster and an ecological interaction between wild boar and domestic pig populations in northeastern Ukraine.

Permafrost, an important source of soil disturbance, is particularly vulnerable to climate change in Alaska where 85% of the land is underlained with discontinuous permafrost. Boreal forests, home to plants integral to subsistence diets of many Alaska Native communities, are not immune to the effects of climate change. Soil disturbance events, such as permafrost thaw, wildfires, and land use change can influence abiotic conditions, which can then affect active layer soil microbial communities. In a previous study, we found negative effects on boreal plants inoculated with microbes impacted by soil disturbance compared to plants inoculated with microbes from undisturbed soils. Here, we identify key shifts in microbial communities altered by soil disturbance using 16S rRNA gene sequencing and make connections between microbial community changes and previously observed plant growth. Additionally, we identify further community shifts in potential functional mechanisms using long read metagenomics. Across a soil disturbance gradient, microbial communities differ significantly based on the level of soil disturbance. Consistent with the earlier study, the family Acidobacteriaceae, which consists of known plant growth promoters, was abundant in undisturbed soil, but practically absent in most disturbed soil. In contrast, Comamonadaceae, a family with known agricultural pathogens, was overrepresented in most disturbed soil communities compared to undisturbed. Within our metagenomic data, we found that soil disturbance level is associated with differences in microbial community function, including mechanisms potentially involved in plant pathogenicity. These results indicate that a decrease in plant growth can be linked to changes in the microbial community and functional composition driven by soil disturbance and climate change. Together, these results build a genomic understanding of how shifting soil microbiomes may affect plant productivity and ecosystem health as the Arctic warms.

The complete genome of Erysipelothrix sp. strain Poltava, isolated from fatal acute septic erysipelas of swine in Ukraine, was assembled using Nanopore sequences. One circular chromosome of 1,794,858 bp ( N 50 , 1,794,858 bp) encodes 16 putative antibiotic resistance genes and secreted virulence factors, highlighting the risk of cross-species livestock and human infection.

Alaska is the largest geographic state in the United States with the lowest population density and a mix of urban centers and isolated rural communities. The differences in population dynamics in Alaska from the contiguous United States may have contributed to a unique pattern of emergence and spread of SARS-CoV-2 variants observed in early 2021. Here we examined 2,323 virus genomes from Alaska and 278,635 virus genomes from the contiguous United States collected between the first week of December 2020 through the last week of June 2021. We focused on this timeframe because of the notable emergence and spread of the SARS-CoV-2 lineage B.1.1.519 observed in Alaska. We found that this variant was consistently detected in Alaska from the end of January through June of 2021 with a peak prevalence in April of 77.9% unlike the rest of the United States with a peak prevalence of 4.6%. In Alaska, the earlier emergence of B.1.1.519 coincided with a later peak of Alpha (B.1.1.7) when compared to the rest of the United States. We also observed differences in the composition of lineages and variants over time between the two most populated regions of Alaska. Although there was a modest increase in COVID-19 cases during the peak incidence of B.1.1.519, it is difficult to disentangle how social dynamics conflated changes in COVID-19 during this time. We suggest that the viral characteristics, such as amino acid substitutions in the spike protein, and a founder effect likely contributed to the unique spread of B.1.1.519 in Alaska.

Recent advances in climate research have discovered that permafrost is particularly vulnerable to the changes occurring in the atmosphere and climate, especially in Alaska where 85% of the land is underlain by mostly discontinuous permafrost. As permafrost thaws, research has shown that natural and anthropogenic soil disturbance causes microbial communities to undergo shifts in membership composition and biomass, as well as in functional diversity. Boreal forests are home to many plants that are integral to the subsistence diets of many Alaska Native communities. Yet, it is unclear how the observed shifts in soil microbes can affect above ground plant communities that are relied on as a major source of food. In this study, we tested the hypothesis that microbial communities associated with permafrost thaw affect plant productivity by growing five plant species found in Boreal forests and Tundra ecosystems, including low-bush cranberry and bog blueberry, with microbial communities from the active layer soils of a permafrost thaw gradient. We found that plant productivity was significantly affected by the microbial soil inoculants. Plants inoculated with communities from above thawing permafrost showed decreased productivity compared to plants inoculated with microbes from undisturbed soils. We used metagenomic sequencing to determine that microbial communities from disturbed soils above thawing permafrost differ in taxonomy from microbial communities in undisturbed soils above intact permafrost. The combination of these results indicates that a decrease in plant productivity can be linked to soil disturbance driven changes in microbial community membership and abundance. These data contribute to an understanding of how microbial communities can be affected by soil disturbance and climate change, and how those community shifts can further influence plant productivity in Boreal forests and more broadly, ecosystem health.

Monitoring antibiotic resistance genes (ARGs) across ecological niches is critical for assessing the impacts distinct microbial communities have on the global spread of resistance. In permafrost-associated soils, climate and human driven disturbances augment near-surface thaw shifting the predominant bacteria that shape the resistome in overlying active layer soils. This thaw is of concern in Alaska, because 85% of land is underlain by permafrost, making soils especially vulnerable to disturbances. The goal of this study is to assess how soil disturbance, and the subsequent shift in community composition, will affect the types, abundance, and mobility of ARGs that compose the active layer resistome. We address this goal through the following aims: (1) assess resistance phenotypes through antibiotic susceptibility testing, and (2) analyze types, abundance, and mobility of ARGs through whole genome analyses of bacteria isolated from a disturbance-induced thaw gradient in Interior Alaska. We found a high proportion of isolates resistant to at least one of the antibiotics tested with the highest prevalence of resistance to ampicillin. The abundance of ARGs and proportion of resistant isolates increased with disturbance; however, the number of ARGs per isolate was explained more by phylogeny than isolation site. When compared to a global database of soil bacteria, RefSoil+, our isolates from the same genera had distinct ARGs with a higher proportion on plasmids. These results emphasize the hypothesis that both phylogeny and ecology shape the resistome and suggest that a shift in community composition as a result of disturbance-induced thaw will be reflected in the predominant ARGs comprising the active layer resistome.

The novel coronavirus SARS-CoV-2, the causative agent of COVID-19, emerged in the human population in December 2019 and spread worldwide within a few short months. Much of the public health focus for preventing and mitigating the spread of COVID-19 has been on individual and collective behaviors, such as social distancing, mask-wearing, and hygiene. It is important to recognize that these behaviors and health outcomes occur within broader social and environmental contexts, and factors within local communities such as regional policy, historical context, cultural beliefs, and natural- and built environmental characteristics affect underlying population health and the spread of disease. For example, the COVID-19 pandemic has renewed attention to the importance of secure water and sanitation services in protecting human health; many remote Alaskan communities are particularly vulnerable to infectious disease transmission because of inadequate water and sanitation services. In addition, there are a number of socio-economic, physical, and infrastructure factors in rural Alaska (e.g., remoteness, household overcrowding, climate change impacts, limited medical facilities, and high prevalence of chronic diseases) that contribute to the potential for more severe COVID-19 disease outcomes in these predominantly Alaska Native communities.

The complete genome sequence of Brucella abortus 68, isolated from an aborted sheep fetus in Luhansk, Ukraine, was assembled using Nanopore sequences. Two circular chromosomes totaling 3,281,317 bp (N-50, 2,124,943 bp) comprised the complete genome sequence. The strain encodes the fosfomycin antibiotic resistance gene fosX, highlighting the risk of cross-species livestock and human infection.