This study quantifies the influence of ventilation and indoor emissions on concentrations and particle sizes of airborne indoor allergenic fungal taxa and further examines geographical variability, each of which may affect personal exposures to allergenic fungi. Quantitative PCR and multiplexed DNA sequencing were employed to count and identify allergenic fungal aerosol particles indoors and outdoors in seven school classrooms in four different countries. Quantitative diversity analysis was combined with building characterization and mass balance modeling to apportion source contributions of indoor allergenic airborne fungal particles. Mass balance calculations indicate that 70% of indoor fungal aerosol particles and 80% of airborne allergenic fungal taxa were associated with indoor emissions; on average, 81% of allergenic fungi from indoor sources originated from occupant-generated emissions. Principal coordinate analysis revealed geographical variations in fungal communities among sites in China, Europe, and North America (p < 0.05, analysis of similarity), demonstrating that geography may also affect personal exposures to allergenic fungi. Indoor emissions including those released with occupancy contribute more substantially to allergenic fungal exposures in classrooms sampled than do outdoor contributions from ventilation. The results suggest that design and maintenance of buildings to control indoor emissions may enable reduced indoor inhalation exposures to fungal allergens.

Sewage sludge and biosolids production and management are a central component of water and sanitation engineering. The culmination of previous incremental technologies and regulations aimed at solving a current treatment problem, rather than developing the practice for the higher goals of sustainability have resulted in sludge becoming an economic and social liability. Sludge management practice must shift from treatment of a liability toward recovery of the embedded energy and chemical assets, while continuing to protect the environment and human health. This shift will require new research, treatment technologies and infrastructure and must be guided by the application of green engineering principles to ensure economic, social, and environmental sustainability.

The large diversity of viruses that exist in human populations are potentially excreted into sewage collection systems and concentrated in sewage sludge. In the U.S., the primary fate of processed sewage sludge (class B biosolids) is application to agricultural land as a soil amendment. To characterize and understand infectious risks associated with land application, and to describe the diversity of viruses in human populations, shotgun viral metagenomics was applied to 10 sewage sludge samples from 5 wastewater treatment plants throughout the continental U.S, each serving between 100 000 and 1 000 000 people. Nearly 330 million DNA sequences were produced and assembled, and annotation resulted in identifying 43 (26 DNA, 17 RNA) different types of human viruses in sewage sludge. Novel insights include the high abundance of newly emerging viruses (e.g., Coronavirus HKU1, Klassevirus, and Cosavirus) the strong representation of respiratory viruses, and the relatively minor abundance and occurrence of Enteroviruses. Viral metagenome sequence annotations were reproducible and independent PCR-based identification of selected viruses suggests that viral metagenomes were a conservative estimate of the true viral occurrence and diversity. These results represent the most complete description of human virus diversity in any wastewater sample to date, provide engineers and environmental scientists with critical information on important viral agents and routes of infection from exposure to wastewater and sewage sludge, and represent a significant leap forward in understanding the pathogen content of class B biosolids.

Human adenovirus diversity in sewage sludge was assessed by Ion Torrent sequencing and annotation of partial adenovirus hexon genes. The most abundant species identified were HAdV-C (average 78%) and -B (average 20%), which are associated with respiratory infections. These findings reinforce the necessity to consider aerosol exposure to sewage-derived pathogens.

Fungi are ubiquitous in outdoor air, and their concentration, aerodynamic diameters and taxonomic composition have potentially important implications for human health. Although exposure to fungal allergens is considered a strong risk factor for asthma prevalence and severity, limitations in tracking fungal diversity in air have thus far prevented a clear understanding of their human pathogenic properties. This study used a cascade impactor for sampling, and quantitative real-time PCR plus 454 pyrosequencing for analysis to investigate seasonal, size-resolved fungal communities in outdoor air in an urban setting in the northeastern United States. From the 20 libraries produced with an average of ~800 internal transcribed spacer (ITS) sequences (total 15 326 reads), 12 864 and 11 280 sequences were determined to the genus and species levels, respectively, and 558 different genera and 1172 different species were identified, including allergens and infectious pathogens. These analyses revealed strong relationships between fungal aerodynamic diameters and features of taxonomic compositions. The relative abundance of airborne allergenic fungi ranged from 2.8% to 10.7% of total airborne fungal taxa, peaked in the fall, and increased with increasing aerodynamic diameter. Fungi that can cause invasive fungal infections peaked in the spring, comprised 0.1–1.6% of fungal taxa and typically increased in relative abundance with decreasing aerodynamic diameter. Atmospheric fungal ecology is a strong function of aerodynamic diameter, whereby through physical processes, the size influences the diversity of airborne fungi that deposit in human airways and the efficiencies with which specific groups of fungi partition from outdoor air to indoor environments.

The inactivation of fecal coliforms in anaerobic batch reactors has been investigated at the thermophilic temperatures of 50, 55 and 60 °C. Throughout inactivation experiments at each temperature, individual colonies were isolated and identified by 16S rDNA gene sequencing to illustrate how the diversity of fecal coliforms is affected by thermophilic treatment. Results indicate that even though fecal coliforms in raw sewage sludge are comprised of several different bacterial species, each with variable temperature induced decay rates, the overall inactivation of fecal coliforms in raw sewage sludge was found to follow a first-order relationship. No tailing was observed across the range of fecal coliform concentrations measured. Fecal coliforms in raw sludge contained six different genera of bacteria and were 62% enriched in E. coli. Within 1.5 log removal of fecal coliform concentration by thermophilic treatment, the populations had shifted to, and remained at 100% E. coli. Subsequent inactivation rates measured in isolated fecal coliform strains confirmed that E. coli cells isolated post-treatment were more thermotolerant than E. coli and non-E coli bacteria isolated prior to thermal treatment. Overall, this study describes the potential enrichment of thermotolerant E. coli in biosolids fecal coliforms and demonstrates that while thermotolerant species are present at the end of treatment, pure first-order approximations are appropriate for estimating residence times to reduce fecal coliforms to levels promulgated in U.S. Class A biosolids standards.

The science linking processed sewage sludge (biosolids) land application with human health has improved in the last ten years. The goal of this review is to develop a consensus view on the human health impacts associated with land-applying biosolids. Pre-existing risk studies are integrated with recent advances in biosolids pathogen exposure science and risk analysis. Other than accidental direct ingestion, the highest public risks of infection from land application are associated with airborne exposure. Multiple, independent risk assessments for enteroviruses similarly estimate the yearly probabilities of infection near 10–4. However, the inclusion of other emerging pathogens, specifically norovirus, increases this yearly infectious risk by over 2 orders of magnitude. Quantitative microbial risk assessment for biosolids exposure more effectively operates as a tool for analyzing how exposure can be reduced rather than being used to assess “safety”. Such analysis demonstrates that the tradition of monitoring pathogen quality by Salmonella spp. and enterovirus content underestimates the infectious risk to the public, and that a rigorous biosolids pathogen treatment process, rather than extending community separation distances, is the most efficient method for reducing pathogen exposure and infectious risk.

This study investigated cytotoxicity and inflammation caused by human bronchial epithelial cells exposed to respirable aerosols produced during the land application of stabilized sewage sludges (biosolids). BEAS-2B cells were exposed to respirable aerosols (PM10) derived from soils, biosolids stabilized by mesophilic anaerobic digestion (MAD), temperature-phased anaerobic digestion (TPAD), and composting (COM) as well as animal manures stabilized by mesophilic anaerobic digestion (AMAD) and composting (ACOM). Anaerobically digested particles (MAD, TPAD, AMAD) induced the highest cytotoxicity with LD50 levels of 70 μg/cm2, 310 μg/cm2 for, and 375 μg/cm2 for MAD, AMAD, and TPAD, respectively. Conversely, there was no observed cytotoxicity for soils, composted biosolids, or composted manures at the in vitro doses tested. Inflammatory responses, measured by interleukin (IL)-6 and IL-8 release, were 2- to 15-fold greater in biosolids and manures than for equivalent doses in soils. Biosolids treatment rankings for human bronchial epithelial cell toxicity and inflammation were similar to the rankings found in recent biosolids pathogen content studies—from lowest pathogen content or toxicity to highest, rankings were as follows: COM < TPAD < MAD. Coupling in vitro responses with modeled tracheobronchial lung surface doses that may occur during a biosolids land application event suggests that an inflammatory aerosol exposure in the TB region could only occur under worst case scenarios (exercising human with reduced lung capacity at <65 m set backs), but examination of lower in vitro doses as well as consideration of the head and lower lung respiratory tract regions are needed to more definitively describe the links between biosolids aerosols and the potential for respiratory inflammation.

•Sewage sludge contains elevated concentrations of PBDEs.•Inhalation dose and PBDE body burdens due to sewage sludge spreading were estimated.•PBDEs aerosolized from sludge are insignificant contributors to human PBDE body burden.•Sewage sludge land application is the largest source of environmental PBDE’s.Elevated sewage sludge concentrations of polybrominated diphenyl ethers (PBDEs) are due to their broad utilization in textiles and polymers, their resistance to biological degradation, and also their hydrophobic nature—which drives partitioning into wastewater solids. This study estimated the total U.S. emissions of PBDE due to sewage sludge land application and then determined the human inhalation exposure to sludge-associated PBDEs as a function meteorological conditions and downwind distances from an application site. These aerosol exposures have also been incorporated into pharmacokinetic models to predict contributions to steady-state body burden. Our results suggest that while the amount of PBDEs aerosolized during the land application process is small compared to aerosol emissions associated with product use, the application of sludges onto U.S. soils constitutes a major source of PBDEs entering the outdoor environment. Regarding aerosol exposure to nearby residents, the maximum daily inhalation dosages from a common land application scenario occur immediately after sewage sludges are applied and were 137, 27, 1.9, and 81 pg/day for significant congeners PBDE-47, -99, -153 and 209 respectively. These doses are 1–2 orders of magnitude less than the standard daily inhalation exposure to the same PBDEs associated with home indoor air and are similar to doses from inhalation of urban and rural outdoor air. Under the worst-case atmospheric transport scenario, the dosages are reduced by approximately 1 order of magnitude when the setback distance between the sludge aerosolization source and human receptor is increased to 200 m. Though the health implications of low-level exposures are not well-understood, these sludge-derived PBDE dosages contribute less than a tenth of 1% to the estimated total body burden of PBDE produced from inhalation of indoor and outdoor air, exposure to house dust, and exposure to PBDE from food and water intake. Overall, the inhalation of PBDE aerosols from sludge-applied fields does not represent a significant contribution to human exposure compared to other common indoor exposures. However, land application is a major environmental source of PBDEs and sludge health impact analyses should focus on the practice's impacts on other exposures, such as biomagnification in aquatic and terrestrial food webs.

Waste-streams containing nitrogen (commonly as NH3/NH4+) have been promoted as a means to lower the energy burden and improve the overall sustainability of microalgae-based fuel and chemical production. However, beyond a concentration threshold, ammonia (NH3) is toxic to many microalgae. This study investigated the ammonia tolerances of oleaginous microalgae. Four microalgae that are often considered in biofuel production studies (Neochloris oleoabundans, Dunaliella tertiolecta, Chlorella sorokiniana and Nannochloropsis oculata) were grown in batch reactors maintained at 10 different NH4Cl concentrations at a constant pH = 8. Growth rates and lipid profiles were monitored. Ammonia acted as an inhibiting substrate for N. oleoabundans and D. tertiolecta at 2.3 and 3.3 mg L− 1 NH3, respectively. Growth rates for C. sorokiniana and N. oculata were largely unaffected by ammonia concentrations. D. tertiolecta demonstrated significant neutral lipid alterations during ammonia inhibition.

Favorable growth characteristics continue to generate interest in using triacylglycerides (TAGs) produced from microalgae for biodiesel feedstocks. In this opinion article, we suggest that due to the energy consumption associated with the production of external nitrogen fertilizers, the manner in which nitrogen is supplied to microalgae biorefineries will be an important driver of energy yields, sustainability, and commercial success. Schemes including the reuse of urban wastewater represent improvements on the overall energy balance, but will not allow for significant production of biofuels unless the nitrogen from the non-TAG portions of microalgae is recycled. Approaches to recycling nitrogen require an improved understanding of the tradeoffs between the different potential uses of the non-TAG microalgal portion (i.e., energy production via anaerobic digestion or thermal catalytic processes), and the development of nitrogen separation technologies.Highlights► The supply of external nitrogen to microalgae cultivation is energy intensive. ► Recycling the nitrogen contained in the non-triacylglyceride portion of microalgae may circumvent the negative energy impacts associated with external nitrogen supply. ► Decisions on how to reuse nitrogen must be made within the context of the overall energy balance, sustainability, and commercialization potential of the microalgae biorefinery.

BackgroundAllergic and nonallergic asthma severity in children can be affected by microbial exposures.ObjectiveWe sought to examine associations between exposures to household microbes and childhood asthma severity stratified by atopic status.MethodsParticipants (n = 196) were selected from a cohort of asthmatic children in Connecticut and Massachusetts. Children were grouped according to asthma severity (mild with no or minimal symptoms and medication or moderate to severe persistent) and atopic status (determined by serum IgE levels). Microbial community structure and concentrations in house dust were determined by using next-generation DNA sequencing and quantitative PCR. Logistic regression was used to explore associations between asthma severity and exposure metrics, including richness, taxa identification and quantification, community composition, and concentration of total fungi and bacteria.ResultsAmong all children, increased asthma severity was significantly associated with an increased concentration of summed allergenic fungal species, high total fungal concentrations, and high bacterial richness by using logistic regression in addition to microbial community composition by using the distance comparison t test. Asthma severity in atopic children was associated with fungal community composition (P = .001). By using logistic regression, asthma severity in nonatopic children was associated with total fungal concentration (odds ratio, 2.40; 95% CI, 1.06-5.44). The fungal genus Volutella was associated with increased asthma severity in atopic children (P = .0001, q = 0.04). The yeast genera Kondoa might be protective; Cryptococcus species might also affect asthma severity.ConclusionAsthma severity among this cohort of children was associated with microbial exposure, and associations differed based on atopic status.

Common indoor and outdoor environmental fungi such as Aspergillus fumigatus produce asexual spores containing a collection of proteins that can bind IgE antibodies and trigger allergic reactions. We characterized the impact of sporulation temperature on the IgE-binding capacity (allergenicity) of A. fumigatus and explored the links between variable allergenicity and temperature-dependant expression of genes encoding these allergenic proteins. A 12-fold increase in A. fumigatus allergenicity per spore was observed when sporulation temperatures were decreased from 32 °C to 17 °C. Per spore protein mass and Asp f 1 allergen mass also followed this trend. Functional gene expression analysis of A. fumigatus sporulating cultures by real-time reverse-transcription PCR and gene expression microarrays revealed that a greater number of genes encoding known, major allergens are more highly expressed at lower sporulation temperatures. The results of this study indicate that environmental conditions at growth significantly influence the allergenicity of this common mould through the differential production of allergenic proteins, and highlight the importance of in vivo or in vitro allergenicity measurements for understanding environmental exposure to airborne allergenic fungi.

•Study combined NGS and qPCR to evaluate the aerodynamic diameters of fungal taxa.•Aerodynamic diameters determined for >50 fungal genera in atmospheric bioaerosols.•Good agreement obtained between diameters estimated by NGS and taxon-specific qPCR.Aerodynamic diameter is an important determinant of the physical processes that act upon airborne fungi. Processes include gravitational settling, respiratory deposition, penetration into buildings, resuspension from surfaces into air, and long-range transport. This study combined next-generation DNA sequencing (NGS) with quantitative PCR (qPCR) to evaluate diverse, taxon-specific, fungal aerodynamic diameters from bioaerosol samples. The accuracy of the method was demonstrated by comparing geometric mean aerodynamic diameters of selected taxa produced by the NGS-based method to the diameters produced by taxon-specific qPCR (r=0.996). Geometric means (dg) and geometric standard deviations (σg) of aerodynamic diameters were characterized for more than 50 fungal taxa, spanning 55 genera, 9 classes, and 2 phyla. The results reported in this study demonstrate the robust nature of this method, provide novel insights into aerodynamic properties of diverse airborne fungal species, and potentially enable a better accounting of taxon-specific fungal fate and exposure both in indoor air and in the atmosphere.

Bacteria and fungi in buildings exert an influence on the human microbiome through aerosol deposition, surface contact, and human and animal interactions. As the identities and functions of beneficial human microbes emerge, the consequences of building design, operation, and function must be understood to maintain the health of occupants in buildings.

The inactivation of fecal coliforms in anaerobic batch reactors has been investigated at the thermophilic temperatures of 50, 55 and 60 °C. Throughout inactivation experiments at each temperature, individual colonies were isolated and identified by 16S rDNA gene sequencing to illustrate how the diversity of fecal coliforms is affected by thermophilic treatment. Results indicate that even though fecal coliforms in raw sewage sludge are comprised of several different bacterial species, each with variable temperature induced decay rates, the overall inactivation of fecal coliforms in raw sewage sludge was found to follow a first-order relationship. No tailing was observed across the range of fecal coliform concentrations measured. Fecal coliforms in raw sludge contained six different genera of bacteria and were 62% enriched in E. coli. Within 1.5 log removal of fecal coliform concentration by thermophilic treatment, the populations had shifted to, and remained at 100% E. coli. Subsequent inactivation rates measured in isolated fecal coliform strains confirmed that E. coli cells isolated post-treatment were more thermotolerant than E. coli and non-E coli bacteria isolated prior to thermal treatment. Overall, this study describes the potential enrichment of thermotolerant E. coli in biosolids fecal coliforms and demonstrates that while thermotolerant species are present at the end of treatment, pure first-order approximations are appropriate for estimating residence times to reduce fecal coliforms to levels promulgated in U.S. Class A biosolids standards.