Insights into bear evolution from a Pleistocene polar bear genome.

The polar bear (Ursus maritimus) has become a symbol of the threat to biodiversity from climate change. Understanding polar bear evolutionary history may provide insights into apex carnivore responses and prospects during periods of extreme environmental perturbations. In recent years, genomic studies have examined bear speciation and population history, including evidence for ancient admixture between polar bears and brown bears (Ursus arctos). Here, we extend our earlier studies of a 130,000- to 115,000-y-old polar bear from the Svalbard Archipelago using a 10× coverage genome sequence and 10 new genomes of polar and brown bears from contemporary zones of overlap in northern Alaska. We demonstrate a dramatic decline in effective population size for this ancient polar bear's lineage, followed by a modest increase just before its demise. A slightly higher genetic diversity in the ancient polar bear suggests a severe genetic erosion over a prolonged bottleneck in modern polar bears. Statistical fitting of data to alternative admixture graph scenarios favors at least one ancient introgression event from brown bears into the ancestor of polar bears, possibly dating back over 150,000 y. Gene flow was likely bidirectional, but allelic transfer from brown into polar bear is the strongest detected signal, which contrasts with other published work. These findings may have implications for our understanding of climate change impacts: Polar bears, a specialist Arctic lineage, may not only have undergone severe genetic bottlenecks but also been the recipient of generalist, boreal genetic variants from brown bears during critical phases of Northern Hemisphere glacial oscillations.

Vertical stratification of the air microbiome in the lower troposphere.

The troposphere constitutes the final frontier of global ecosystem research due to technical challenges arising from its size, low biomass, and gaseous state. Using a vertical testing array comprising a meteorological tower and a research aircraft, we conducted synchronized measurements of meteorological parameters and airborne biomass (n = 480) in the vertical air column up to 3,500 m. The taxonomic analysis of metagenomic data revealed differing patterns of airborne microbial community composition with respect to time of day and height above ground. The temporal and spatial resolution of our study demonstrated that the diel cycle of airborne microorganisms is a ground-based phenomenon that is entirely absent at heights >1,000 m. In an integrated analysis combining meteorological and biological data, we demonstrate that atmospheric turbulence, identified by potential temperature and high-frequency three-component wind measurements, is the key driver of bioaerosol dynamics in the lower troposphere. Multivariate regression analysis shows that at least 50% of identified airborne microbial taxa (n = ∼10,000) are associated with either ground or height, allowing for an understanding of dispersal patterns of microbial taxa in the vertical air column. Due to the interconnectedness of atmospheric turbulence and temperature, the dynamics of microbial dispersal are likely to be impacted by rising global temperatures, thereby also affecting ecosystems on the planetary surface.

Elevated c-di-GMP Levels and Expression of the Type III Secretion System Promote Corneal Infection by Pseudomonas aeruginosa.

Pseudomonas aeruginosa is generally believed to establish biofilm-associated infections under the regulation of the secondary messenger c-di-GMP. To evaluate P. aeruginosa biofilm physiology during ocular infections, comparative transcriptomic analysis was performed on wild-type P. aeruginosa PAO1, a ΔwspF mutant strain (high c-di-GMP levels), and a plac-yhjH-containing strain (low c-di-GMP levels) from mouse corneal infection, as well as in vitro biofilm and planktonic cultures. The c-di-GMP content in P. aeruginosa during corneal infection was monitored using a fluorescent c-di-GMP reporter strain. Biofilm-related genes were induced in in vivo PAO1 compared to in vitro planktonic bacteria. Several diguanylate cyclases and phosphodiesterases were commonly regulated in in vivo PAO1 and in vitro biofilm compared to in vitro planktonic bacteria. Several exopolysaccharide genes and motility genes were induced and downregulated, respectively, in in vivo PAO1 and the in vivo ΔwspF mutant compared to the in vivo plac-yhjH-containing strain. Elevation of c-di-GMP levels in P. aeruginosa began as early as 2 h postinfection. The ΔwspF mutant was less susceptible to host clearance than the plac-yhjH-containing strain and could suppress host immune responses. The type III secretion system (T3SS) was induced in in vivo PAO1 compared to in vitro biofilm bacteria. A ΔwspF mutant with a defective T3SS was more susceptible to host clearance than a ΔwspF mutant with a functional T3SS. Our study suggests that elevated intracellular c-di-GMP levels and T3SS activity in P. aeruginosa are necessary for establishment of infection and modulation of host immune responses in mouse cornea.

Airborne SARS-CoV-2 surveillance in hospital environment using high-flowrate air samplers and its comparison to surface sampling.

Reliable methods to detect the presence of SARS-CoV-2 at venues where people gather are essential for epidemiological surveillance to guide public policy. Communal screening of air in a highly crowded space has the potential to provide early warning on the presence and potential transmission of SARS-CoV-2 as suggested by studies early in the epidemic. As hospitals and public facilities apply varying degrees of restrictions and regulations, it is important to provide multiple methodological options to enable environmental SARS-CoV-2 surveillance under different conditions. This study assessed the feasibility of using high-flowrate air samplers combined with RNA extraction kit designed for environmental sample to perform airborne SARS-CoV-2 surveillance in hospital setting, tested by RT-qPCR. The success rate of the air samples in detecting SARS-CoV-2 was then compared with surface swab samples collected in the same proximity. Additionally, positive RT-qPCR samples underwent viral culture to assess the viability of the sampled SARS-CoV-2. The study was performed in inpatient ward environments of a quaternary care university teaching hospital in Singapore housing active COVID-19 patients within the period of February to May 2020. Two types of wards were tested, naturally ventilated open-cohort ward and mechanically ventilated isolation ward. Distances between the site of air sampling and the patient cluster in the investigated wards were also recorded. No successful detection of airborne SARS-CoV-2 was recorded when 50 L/min air samplers were used. Upon increasing the sampling flowrate to 150 L/min, our results showed a high success rate in detecting the presence of SARS-CoV-2 from the air samples (72%) compared to the surface swab samples (9.6%). The positive detection rate of the air samples along with the corresponding viral load could be associated with the distance between sampling site and patient. The furthest distance from patient with PCR-positive air samples was 5.5 m. The airborne SARS-CoV-2 detection was comparable between the two types of wards with 60%-87.5% success rate. High prevalence of the virus was found in toilet areas, both on surfaces and in air. Finally, no successful culture attempt was recorded from the environmental air or surface samples.

Microbial communities in the tropical air ecosystem follow a precise diel cycle.

The atmosphere is vastly underexplored as a habitable ecosystem for microbial organisms. In this study, we investigated 795 time-resolved metagenomes from tropical air, generating 2.27 terabases of data. Despite only 9 to 17% of the generated sequence data currently being assignable to taxa, the air harbored a microbial diversity that rivals the complexity of other planetary ecosystems. The airborne microbial organisms followed a clear diel cycle, possibly driven by environmental factors. Interday taxonomic diversity exceeded day-to-day and month-to-month variation. Environmental time series revealed the existence of a large core of microbial taxa that remained invariable over 13 mo, thereby underlining the long-term robustness of the airborne community structure. Unlike terrestrial or aquatic environments, where prokaryotes are prevalent, the tropical airborne biomass was dominated by DNA from eukaryotic phyla. Specific fungal and bacterial species were strongly correlated with temperature, humidity, and CO2 concentration, making them suitable biomarkers for studying the bioaerosol dynamics of the atmosphere.

A synbiotic intervention modulates meta-omics signatures of gut redox potential and acidity in elective caesarean born infants.

BACKGROUND: The compromised gut microbiome that results from C-section birth has been hypothesized as a risk factor for the development of non-communicable diseases (NCD). In a double-blind randomized controlled study, 153 infants born by elective C-section received an infant formula supplemented with either synbiotic, prebiotics, or unsupplemented from birth until 4 months old. Vaginally born infants were included as a reference group. Stool samples were collected from day 3 till week 22. Multi-omics were deployed to investigate the impact of mode of delivery and nutrition on the development of the infant gut microbiome, and uncover putative biological mechanisms underlying the role of a compromised microbiome as a risk factor for NCD. RESULTS: As early as day 3, infants born vaginally presented a hypoxic and acidic gut environment characterized by an enrichment of strict anaerobes (Bifidobacteriaceae). Infants born by C-section presented the hallmark of a compromised microbiome driven by an enrichment of Enterobacteriaceae. This was associated with meta-omics signatures characteristic of a microbiome adapted to a more oxygen-rich gut environment, enriched with genes associated with reactive oxygen species metabolism and lipopolysaccharide biosynthesis, and depleted in genes involved in the metabolism of milk carbohydrates. The synbiotic formula modulated expression of microbial genes involved in (oligo)saccharide metabolism, which emulates the eco-physiological gut environment observed in vaginally born infants. The resulting hypoxic and acidic milieu prevented the establishment of a compromised microbiome. CONCLUSIONS: This study deciphers the putative functional hallmarks of a compromised microbiome acquired during C-section birth, and the impact of nutrition that may counteract disturbed microbiome development. TRIAL REGISTRATION: The study was registered in the Dutch Trial Register (Number: 2838 ) on 4th April 2011.

Metagenomics Reveals a Core Macrolide Resistome Related to Microbiota in Chronic Respiratory Disease.

Rationale: Long-term antibiotic use for managing chronic respiratory disease is increasing; however, the role of the airway resistome and its relationship to host microbiomes remains unknown.Objectives: To evaluate airway resistomes and relate them to host and environmental microbiomes using ultradeep metagenomic shotgun sequencing.Methods: Airway specimens from 85 individuals with and without chronic respiratory disease (severe asthma, chronic obstructive pulmonary disease, and bronchiectasis) were subjected to metagenomic sequencing to an average depth exceeding 20 million reads. Respiratory and device-associated microbiomes were evaluated on the basis of taxonomical classification and functional annotation including the Comprehensive Antibiotic Resistance Database to determine airway resistomes. Co-occurrence networks of gene-microbe association were constructed to determine potential microbial sources of the airway resistome. Paired patient-inhaler metagenomes were compared (n = 31) to assess for the presence of airway-environment overlap in microbiomes and/or resistomes.Measurements and Main Results: Airway metagenomes exhibit taxonomic and metabolic diversity and distinct antimicrobial resistance patterns. A "core" airway resistome dominated by macrolide but with high prevalence of ß-lactam, fluoroquinolone, and tetracycline resistance genes exists and is independent of disease status or antibiotic exposure. Streptococcus and Actinomyces are key potential microbial reservoirs of macrolide resistance including the ermX, ermF, and msrD genes. Significant patient-inhaler overlap in airway microbiomes and their resistomes is identified where the latter may be a proxy for airway microbiome assessment in chronic respiratory disease.Conclusions: Metagenomic analysis of the airway reveals a core macrolide resistome harbored by the host microbiome.

Environmental fungal sensitisation associates with poorer clinical outcomes in COPD.

INTRODUCTION: Allergic sensitisation to fungi such as Aspergillus are associated to poor clinical outcomes in asthma, bronchiectasis and cystic fibrosis; however, clinical relevance in COPD remains unclear. METHODS: Patients with stable COPD (n=446) and nondiseased controls (n=51) were prospectively recruited across three countries (Singapore, Malaysia and Hong Kong) and screened against a comprehensive allergen panel including house dust mites, pollens, cockroach and fungi. For the first time, using a metagenomics approach, we assessed outdoor and indoor environmental allergen exposure in COPD. We identified key fungi in outdoor air and developed specific-IgE assays against the top culturable fungi, linking sensitisation responses to COPD outcomes. Indoor air and surface allergens were prospectively evaluated by metagenomics in the homes of 11 COPD patients and linked to clinical outcome. RESULTS: High frequencies of sensitisation to a broad range of allergens occur in COPD. Fungal sensitisation associates with frequent exacerbations, and unsupervised clustering reveals a "highly sensitised fungal predominant" subgroup demonstrating significant symptomatology, frequent exacerbations and poor lung function. Outdoor and indoor environments serve as important reservoirs of fungal allergen exposure in COPD and promote a sensitisation response to outdoor air fungi. Indoor (home) environments with high fungal allergens associate with greater COPD symptoms and poorer lung function, illustrating the importance of environmental exposures on clinical outcomes in COPD. CONCLUSION: Fungal sensitisation is prevalent in COPD and associates with frequent exacerbations representing a potential treatable trait. Outdoor and indoor (home) environments represent a key source of fungal allergen exposure, amenable to intervention, in "sensitised" COPD.

Longitudinal assessment of antibiotic resistance gene profiles in gut microbiomes of infants at risk of eczema.

BACKGROUND: While there is increasing knowledge about the gut microbiome, the factors influencing and the significance of the gut resistome are still not well understood. Infant gut commensals risk transferring multidrug-resistant antibiotic resistance genes (ARGs) to pathogenic bacteria. The rapid spread of multidrug-resistant pathogenic bacteria is a worldwide public health concern. Better understanding of the naïve infant gut resistome may build the evidence base for antimicrobial stewardship in both humans and in the food industry. Given the high carriage rate of extended spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae in Asia, we aimed to evaluate community prevalence, dynamics, and longitudinal changes in antibiotic resistance gene (ARG) profiles and prevalence of ESBL-producing E. coli and K. pneumoniae in the intestinal microbiome of infants participating in the Growing Up in Singapore Towards Healthy Outcomes (GUSTO) study, a longitudinal cohort study of pregnant women and their infants. METHODS: We analysed ARGs in the first year of life among 75 infants at risk of eczema who had stool samples collected at multiple timepoints using metagenomics. RESULTS: The mean number of ARGs per infant increased with age. The most common ARGs identified confer resistance to aminoglycoside, beta-lactam, macrolide and tetracycline antibiotics; all infants harboured these antibiotic resistance genes at some point in the first year of life. Few ARGs persisted throughout the first year of life. Beta-lactam resistant Escherichia coli and Klebsiella pneumoniae were detected in 4 (5.3%) and 32 (42.7%) of subjects respectively. CONCLUSION: In this longitudinal cohort study of infants living in a region with high endemic antibacterial resistance, we demonstrate that majority of the infants harboured several antibiotic resistance genes in their gut and showed that the infant gut resistome is diverse and dynamic over the first year of life.

Long-read sequencing uncovers the adaptive topography of a carnivorous plant genome.

Utricularia gibba, the humped bladderwort, is a carnivorous plant that retains a tiny nuclear genome despite at least two rounds of whole genome duplication (WGD) since common ancestry with grapevine and other species. We used a third-generation genome assembly with several complete chromosomes to reconstruct the two most recent lineage-specific ancestral genomes that led to the modern U. gibba genome structure. Patterns of subgenome dominance in the most recent WGD, both architectural and transcriptional, are suggestive of allopolyploidization, which may have generated genomic novelty and led to instantaneous speciation. Syntenic duplicates retained in polyploid blocks are enriched for transcription factor functions, whereas gene copies derived from ongoing tandem duplication events are enriched in metabolic functions potentially important for a carnivorous plant. Among these are tandem arrays of cysteine protease genes with trap-specific expression that evolved within a protein family known to be useful in the digestion of animal prey. Further enriched functions among tandem duplicates (also with trap-enhanced expression) include peptide transport (intercellular movement of broken-down prey proteins), ATPase activities (bladder-trap acidification and transmembrane nutrient transport), hydrolase and chitinase activities (breakdown of prey polysaccharides), and cell-wall dynamic components possibly associated with active bladder movements. Whereas independently polyploid Arabidopsis syntenic gene duplicates are similarly enriched for transcriptional regulatory activities, Arabidopsis tandems are distinct from those of U. gibba, while still metabolic and likely reflecting unique adaptations of that species. Taken together, these findings highlight the special importance of tandem duplications in the adaptive landscapes of a carnivorous plant genome.

Complete Genome Sequence of Penicillium oxalicum Strain SGAir0226 Isolated from Outdoor Tropical Air in Singapore.

Penicillium oxalicum strain SGAir0226 was isolated from a tropical air sample collected in Singapore. The complete genome was assembled from long reads obtained from single-molecule real-time sequencing and was further polished and error corrected using short read sequencing data. The assembly comprises 20 contigs with a total length of 30.7 Mb. The genome was predicted to contain 8310 protein-coding genes, 237 tRNAs and 83 rRNAs.

Whole-Genome Sequencing of Aspergillus terreus Species Complex.

Aspergillus terreus species complex is an opportunistic fungal pathogen increasingly implicated in invasive infection, as well as chronic respiratory disease. Currently, an understanding of A. terreus pathogenicity is impeded by a limited number of whole-genome sequences of this fungal pathogen. We here describe a high-quality whole-genome assembly of European A. terreus clinical isolate M6925, derived by single-molecule real-time sequencing with short-read polishing.

Repurposing the anticancer drug cisplatin with the aim of developing novel Pseudomonas aeruginosa infection control agents.

Antibiotic resistance threatens effective treatment of microbial infections globally. This situation has spurred the hunt for new antimicrobial compounds in both academia and the pharmaceutical industry. Here, we report how the widely used antitumor drug cisplatin may be repurposed as an effective antimicrobial against the nosocomial pathogen Pseudomonas aeruginosa. Cisplatin was found to effectively kill strains of P. aeruginosa. In such experiments, transcriptomic profiling showed upregulation of the recA gene, which is known to be important for DNA repair, implicating that cisplatin could interfere with DNA replication in P. aeruginosa. Cisplatin treatment significantly repressed the type III secretion system (T3SS), which is important for the secretion of exotoxins. Furthermore, cisplatin was also demonstrated to eradicate in vitro biofilms and in vivo biofilms in a murine keratitis model. This showed that cisplatin could be effectively used to eradicate biofilm infections which were otherwise difficult to be treated by conventional antibiotics. Although cisplatin is highly toxic for humans upon systemic exposure, a low toxicity was demonstrated with topical treatment. This indicated that higher-than-minimal inhibitory concentration (MIC) doses of cisplatin could be topically applied to treat persistent and recalcitrant P. aeruginosa infections.

The quest to resolve recent radiations: Plastid phylogenomics of extinct and endangered Hawaiian endemic mints (Lamiaceae).

The Hawaiian mints (Lamiaceae), one of the largest endemic plant lineages in the archipelago, provide an excellent system to study rapid diversification of a lineage with a remote, likely paleohybrid origin. Since their divergence from New World mints 4-5 million years ago the members of this lineage have diversified greatly and represent a remarkable array of vegetative and reproductive phenotypes. Today many members of this group are endangered or already extinct, and molecular phylogenetic work relies largely on herbarium samples collected during the last century. So far a gene-by-gene approach has been utilized, but the recent radiation of the Hawaiian mints has resulted in minimal sequence divergence and hence poor phylogenetic resolution. In our quest to trace the reticulate evolutionary history of the lineage, a resolved maternal phylogeny is necessary. We applied a high-throughput approach to sequence 12 complete or nearly complete plastid genomes from multiple Hawaiian mint species and relatives, including extinct and rare taxa. We also targeted 108 hypervariable regions from throughout the chloroplast genomes in nearly all of the remaining Hawaiian species, and relatives, using a next-generation amplicon sequencing approach. This procedure generated ∼20Kb of sequence data for each taxon and considerably increased the total number of variable sites over previous analyses. Our results demonstrate the potential of high-throughput sequencing of historic material for evolutionary studies in rapidly evolving lineages. Our study, however, also highlights the challenges of resolving relationships within recent radiations even at the genomic level.

Mitogenomes reveal diversity of the European Lyme borreliosis vector Ixodes ricinus in Italy.

In Europe, the Ixodes ricinus tick is the most important vector of the etiological agents of Lyme borreliosis and several other emerging tick-borne diseases. Because tick-borne pathogens are dependent on their vectors for transmission, understanding the vector population structure is crucial to inform public health research of pathogen dynamics and spread. However, the population structure and dynamics of this important vector species are not well understood as most genetic studies utilize short mitochondrial and nuclear sequences with little diversity. Herein we obtained and analyzed complete mitochondrial genome (hereafter "mitogenome") sequences to better understand the genetic diversity and the population structure of I. ricinus from two long-standing tick-borne disease foci in northern Italy. Complete mitogenomes of 23 I. ricinus ticks were sequenced at high coverage. Out of 23 mitogenome sequences we identified 17 unique haplotypes composed of 244 segregating sites. Phylogenetic reconstruction using 18 complete mitogenome sequences revealed the coexistence of four highly divergent I. ricinus maternal lineages despite the narrow spatial scale over which these samples were obtained (100km). Notably, the estimated coalescence time of the 18 mitogenome haplotypes is ∼427 thousand years ago (95% HPD 330, 540). This divergence between I. ricinus lineages is consistent with the mitochondrial diversity of other arthropod vector species and indicates that long-term I. ricinus populations may have been less structured and larger than previously thought. Thus, this study suggests that a rapid and accurate retrieval of full mitochondrial genomes from this disease vector enables fine-resolution studies of tick intraspecies genetic relationships, population differentiation, and demographic history.

Purine and carbohydrate availability drive Enterococcus faecalis fitness during wound and urinary tract infections.

IMPORTANCE: Although E. faecalis is a common wound pathogen, its pathogenic mechanisms during wound infection are unexplored. Here, combining a mouse wound infection model with in vivo transposon and RNA sequencing approaches, we identified the E. faecalis purine biosynthetic pathway and galactose/mannose MptABCD phosphotransferase system as essential for E. faecalis acute replication and persistence during wound infection, respectively. The essentiality of purine biosynthesis and the MptABCD PTS is driven by the consumption of purine metabolites by E. faecalis during acute replication and changing carbohydrate availability during the course of wound infection. Overall, our findings reveal the importance of the wound microenvironment in E. faecalis wound pathogenesis and how these metabolic pathways can be targeted to better control wound infections.

Structure vs. chemistry: Alternate mechanisms for controlling leaf microbiomes.

The analysis of phyllosphere microbiomes traditionally relied on DNA extracted from whole leaves. To investigate the microbial communities on the adaxial (upper) and abaxial (lower) leaf surfaces, swabs were collected from both surfaces of two garden plants, Rhapis excelsa and Cordyline fruticosa. Samples were collected at noon and midnight and at five different locations to investigate if the phyllosphere microbial communities change with time and location. The abaxial surface of Rhapis excelsa and Cordyline fruticosa had fewer bacteria in contrast to its adaxial counterpart. This observation was consistent between noon and midnight and across five different locations. Our co-occurrence network analysis further showed that bacteria were found almost exclusively on the adaxial surface while only a small group of leaf blotch fungi thrived on the abaxial surface. There are higher densities of stomata on the abaxial surface and these openings are vulnerable ports of entry into the plant host. While one might argue about the settling of dust particles and microorganisms on the adaxial surface, we detected differences in reactive chemical activities and microstructures between the adaxial and abaxial surfaces. Our results further suggest that both plant species deploy different defence strategies to deter invading pathogens on the abaxial surface. We hypothesize that chemical and mechanical defence strategies evolved independently for harnessing and controlling phyllosphere microbiomes. Our findings have also advanced our understanding that the abaxial leaf surface is distinct from the adaxial surface and that the reduced microbial diversity is likely a consequence of plant-microbe interactions.

Shotgun metagenomic sequencing analysis of ocular surface microbiome in Singapore residents with mild dry eye.

The ocular surface microbiome has implications for ocular surface inflammation and immunology. Previous shotgun metagenomics analyses were performed in China, showing results that differed according to environment and age. Patients with Sjogren's syndrome were reported to have altered conjunctival microbiome, but such studies have not been done in milder dry eye. The aim of this study is to describe the conjunctival microbiome in people with mild dry eye in Singapore. Samples were collected from 14 participants with mild dry eye and 10 age-matched comparison participants recruited from Singapore National Eye Centre (SNEC) clinics. Shotgun metagenomic sequencing analysis was employed to evaluate the conjunctival microbiome composition. Proteobacteria formed the predominant phylum in the conjunctiva. As in a study from a coastal city in China, Achromobacter spp. was numerically most abundant. Compared to age-matched controls, the conjunctival microbial composition in mild dry eye was similar. Several microorganisms, including Streptococcus spp. increased in representation with age, and the abundance of Staphylococcus correlated with Schirmer readings. In addition, when cultured corneal epithelial cells were exposed to three strains of Achromobacter xylosoxidans, cytokines such as TNF-α and IL-6 were upregulated in the cell lysates and supernatants. Ourresults suggest that age is an important factor that affects composition of the conjunctival microbiome, and relative abundance of specific microorganism may vary according to the environment of the human host.

Recovery of High Quality Metagenome-Assembled Genomes From Full-Scale Activated Sludge Microbial Communities in a Tropical Climate Using Longitudinal Metagenome Sampling.

The analysis of metagenome data based on the recovery of draft genomes (so called metagenome-assembled genomes, or MAG) has assumed an increasingly central role in microbiome research in recent years. Microbial communities underpinning the operation of wastewater treatment plants are particularly challenging targets for MAG analysis due to their high ecological complexity, and remain important, albeit understudied, microbial communities that play ssa key role in mediating interactions between human and natural ecosystems. Here we consider strategies for recovery of MAG sequence from time series metagenome surveys of full-scale activated sludge microbial communities. We generate MAG catalogs from this set of data using several different strategies, including the use of multiple individual sample assemblies, two variations on multi-sample co-assembly and a recently published MAG recovery workflow using deep learning. We obtain a total of just under 9,100 draft genomes, which collapse to around 3,100 non-redundant genomic clusters. We examine the strengths and weaknesses of these approaches in relation to MAG yield and quality, showing that co-assembly may offer advantages over single-sample assembly in the case of metagenome data obtained from closely sampled longitudinal study designs. Around 1,000 MAGs were candidates for being considered high quality, based on single-copy marker gene occurrence statistics, however only 58 MAG formally meet the MIMAG criteria for being high quality draft genomes. These findings carry broader broader implications for performing genome-resolved metagenomics on highly complex communities, the design and implementation of genome recoverability strategies, MAG decontamination and the search for better binning methodology.

Short-range contributions of local sources to ambient air.

Recent developments in aerobiology have enabled the investigation of airborne biomass with high temporal and taxonomic resolution. In this study, we assess the contributions of local sources to ambient air within a 160,000 m2 tropical avian park (AP). We sequenced and analyzed 120 air samples from seven locations situated 160 to 400 m apart, representing distinct microhabitats. Each microhabitat contained a characteristic air microbiome, defined by the abundance and richness of its airborne microbial community members, supported by both, PCoA and Random Forest analysis. Each outdoor microhabitat contained 1% to 18.6% location-specific taxa, while a core microbiome of 27.1% of the total taxa was shared. To identify and assess local sources, we compared the AP dataset with a DVE reference dataset from a location 2 km away, collected during a year-round sampling campaign. Intersection of data from the two sites demonstrated 61.6% of airborne species originated from local sources of the AP, 34.5% from ambient air background, and only 3.9% of species were specific to the DVE reference site. In-depth taxonomic analysis demonstrated association of bacteria-dominated air microbiomes with indoor spaces, while fungi-dominated airborne microbial biomass was predominant in outdoor settings with ample vegetation. The approach presented here demonstrates an ability to identify local source contributions against an ambient air background, despite the prevailing mixing of air masses caused by atmospheric turbulences.