Recycling urine for bioelectrochemical hydrogen production using a MoS2 nano carbon coated electrode in a microbial electrolysis cell.

In this study, a novel molybdenum disulfide (MoS2) nano-carbon (NC) coated cathode was developed for hydrogen production in a microbial electrolysis cell (MEC), while treating simulated urine with 2-6 times dilution (conductivity <20 mS cm-1). MoS2 nanoparticles were electrodeposited on the NC coated cathodes at -100, -150 and -200 µA cm-2 and their performances were evaluated in the MEC. The chronopotentiometry (CP) tests showed the improved catalytic activity of MoS2-NC cathodes with much lower cathode overpotential than non-MoS2 coated electrodes. The MoS2-NC200 cathode, electrodeposited at -200 µA cm-2, showed the maximum hydrogen production rate of 0.152 ± 0.002 m3 H2 m-2 d-1 at 0.9V of Eap, which is comparable to the previously reported Pt electrodes. It was found that high solution conductivity over 20 mS cm-1 (>600 mg L-1 NH3-N) can adversely affect the biofilm architecture and the bacterial activity at the anode of the MEC. Exoelectrogenic bacteria for this system at the anode were identified as Tissierella (Clostridia) and Bacteroidetes taxa. Maximum ammonia-nitrogen (NH3-N) and phosphorus (PO4 3--P) removal were 68.7 and 98.6%, respectively. This study showed that the newly fabricated MoS2-NC cathode can be a cost-effective alternative to the Pt cathode for renewable bioelectrochemical hydrogen production from urine.

Bacterial diversity and predicted enzymatic function in a multipurpose surface water system - from wastewater effluent discharges to drinking water production.

BACKGROUND: Rivers and lakes are used for multiple purposes such as for drinking water (DW) production, recreation, and as recipients of wastewater from various sources. The deterioration of surface water quality with wastewater is well-known, but less is known about the bacterial community dynamics in the affected surface waters. Understanding the bacterial community characteristics -from the source of contamination, through the watershed to the DW production process-may help safeguard human health and the environment. RESULTS: The spatial and seasonal dynamics of bacterial communities, their predicted functions, and potential health-related bacterial (PHRB) reads within the Kokemäenjoki River watershed in southwest Finland were analyzed with the 16S rRNA-gene amplicon sequencing method. Water samples were collected from various sampling points of the watershed, from its major pollution sources (sewage influent and effluent, industrial effluent, mine runoff) and different stages of the DW treatment process (pre-treatment, groundwater observation well, DW production well) by using the river water as raw water with an artificial groundwater recharge (AGR). The beta-diversity analysis revealed that bacterial communities were highly varied among sample groups (R = 0.92, p <  0.001, ANOSIM). The species richness and evenness indices were highest in surface water (Chao1; 920 ± 10) among sample groups and gradually decreased during the DW treatment process (DW production well; Chao1: 320 ± 20). Although the phylum Proteobacteria was omnipresent, its relative abundance was higher in sewage and industrial effluents (66-80%) than in surface water (55%). Phyla Firmicutes and Fusobacteria were only detected in sewage samples. Actinobacteria was more abundant in the surface water (≥13%) than in other groups (≤3%). Acidobacteria was more abundant in the DW treatment process (≥13%) than in others (≤2%). In total, the share of PHRB reads was higher in sewage and surface water than in the DW treatment samples. The seasonal effect in bacterial communities was observed only on surface water samples, with the lowest diversity during summer. CONCLUSIONS: The low bacterial diversity and absence of PHRB read in the DW samples indicate AGR can produce biologically stable and microbiologically safe drinking water. Furthermore, the significantly different bacterial communities at the pollution sources compared to surface water and DW samples highlight the importance of effective wastewater treatment for protecting the environment and human health.

A strategy for power generation from bilgewater using a photosynthetic microalgal fuel cell (MAFC).

Microbial fuel cells (MFCs) have recently been applied to generate electricity from oily wastewater. Although MFCs that utilize microalgae to provide a self-supporting oxygen (O2) supply at the cathode have been well discussed, those with microalgae at the anode as an active biomass for treating wastewater and producing electrons are still poorly studied and understood. Here, we demonstrated a bilgewater treatment using single- and double-chamber microalgal fuel cells (SMAFC and DMAFC) capable of generating energy with a novel microalgal strain (Chlorella sorokiniana) that was initially isolated from oily wastewater. Compared to previous MFC studies using green algae, relatively high voltage output (151.3-160.1 mV, 71.3-83.4 mV m-2 of power density) was observed in the SMAFC under O2 controlled systems (i.e., acetate addition or light/dark cycle). It was assumed that, under the O2 depletion, alternative electron acceptors such as bicarbonate may be utilized for power generation. A DMAFC showed better power density (up to 23.9%) compared to the SMAFC due to the separated cathode chamber which fully utilizes O2 as an electron acceptor. Both SMAFC and DMAFC removed 67.2-77.4% of soluble chemical oxygen demands (SCOD) from the synthetic bilgewater. This study demonstrates that the application of algae-based MFCs is a feasible strategy to treat oil-in-water emulsion while generating electricity.

High-throughput environmental DNA analysis informs a biological assessment of an urban stream.

There is growing interest in the use of DNA barcoding and metabarcoding approaches to aid biological assessments and monitoring of waterbodies. While biodiversity measured by morphology and by DNA often has been found correlated, few studies have compared DNA data to established measures of impairment such as multimetric pollution tolerance indices used by many bioassessment programs. We incorporated environmental DNA (eDNA) metabarcoding of seston into a rigorous watershed-scale biological assessment of an urban stream to examine the extent to which eDNA richness and diversity patterns were correlated with multimetric indices and ecological impairment status designations. We also evaluated different filtering approaches and taxonomic classifications to identify best practices for environmental assessments. Seston eDNA revealed a wide diversity of eukaryotic taxa but was dominated by diatoms (36%). Differentiation among sites in alpha and beta diversity was greater when operational taxonomic units (OTUs) were classified taxonomically, but coarse resolution taxonomy (kingdom) was more informative than finer resolution taxonomy (family, genus). Correlations of DNA richness and diversity with multimetric indices for fish and macroinvertebrates were generally weak, possibly because Metazoa were not highly represented in our DNA dataset. Nonetheless, sites could be differentiated based on ecological impairment status, with more impaired sites having lower eDNA diversity as measured by the Shannon index, but higher taxonomic richness. Significant environmental drivers of community structure, as inferred from constrained ordination analyses, differed among kingdoms within the eDNA dataset, as well as from fish and macrobenthos, suggesting that eDNA provides novel environmental information. These results suggest that even a simple seston eDNA filtering protocol can provide biodiversity information of value to stream bioassessment programs. The approach bears further investigation as a potentially useful rapid assessment protocol to supplement more intensive field sampling efforts.

Biodiversity and dynamics of cyanobacterial communities during blooms in temperate lake (Harsha Lake, Ohio, USA).

Cyanobacterial blooms are intensifying global ecological hazards. The fine structure and dynamics of bloom community are critical to understanding bloom development but little understood. Here, the questions whether dominant bloomers have high diversity and whether dominant OTUs (operational taxonomical units) compete with one another were addressed. 16S rRNA gene amplicons from an annual bloom at five locations in Harsha Lake (Ohio, USA) showed cyanobacteria were the dominant phylum, and co-existing major bacterial phyla included Proteobacteria, Bacteroidetes, Actinoacteria, and Verrucomicrobia. On the genus level, the initial dominance by Dolichospermum in June yielded to Planktothrix in July, which were replaced by Microcystis and Cylindrospermopsis in August throughout the bloom. Based on the number of verified unique OTUs (a within-genus biodiversity metric), dominant genera tended to have high within-genus diversity. For example, Dolichospermum had 57 unique OTUs, Planktothrix had 36, Microcystis had 12, and Cylindrospermopsis had 4 unique OTUs. Interestingly, these different OTUs showed different dynamics and association with other OTUs. First, no between-OTU competitions were observed during the bloom cycle, and dominant OTUs were abundant throughout the bloom. Such biodiversity of OTUs and their dynamics were verified in Microcystis aeruginosa with two microcystin synthetase genes (mcyA and mcyG): the relative abundance of both genes varied during the bloom based on quantitative PCR. Two Dolichospermum circinale OTUs and one P. rubescens OTU were most abundant and persistently present throughout the entire bloom. Second, these OTUs differed in the OTUs they were associated with. Third, these OTUs tended to have different levels of association with the environmental factors, even they belonged to the same genera. These findings suggest the structure and dynamics of a cyanobacterial bloom community is complex, with only few OTUs dominating the bloom. Thus, high-resolution molecular characterization will be necessary to understand bloom development.

Opportunistic Pathogens and Microbial Communities and Their Associations with Sediment Physical Parameters in Drinking Water Storage Tank Sediments.

The occurrence and densities of opportunistic pathogens (OPs), the microbial community structure, and their associations with sediment elements from eight water storage tanks in Ohio, West Virginia, and Texas were investigated. The elemental composition of sediments was measured through X-ray fluorescence (XRF) spectra. The occurrence and densities of OPs and amoeba hosts (i.e., Legionella spp. and L. pneumophila, Mycobacterium spp., P. aeruginosa, V. vermiformis, Acanthamoeba spp.) were determined using genus- or species-specific qPCR assays. Microbial community analysis was performed using next generation sequencing on the Illumina Miseq platform. Mycobacterium spp. were most frequently detected in the sediments and water samples (88% and 88%), followed by Legionella spp. (50% and 50%), Acanthamoeba spp. (63% and 13%), V. vermiformis (50% and 25%), and P. aeruginosa (0 and 50%) by qPCR method. Comamonadaceae (22.8%), Sphingomonadaceae (10.3%), and Oxalobacteraceae (10.1%) were the most dominant families by sequencing method. Microbial communities in water samples were mostly separated with those in sediment samples, suggesting differences of communities between two matrices even in the same location. There were associations of OPs with microbial communities. Both OPs and microbial community structures were positively associated with some elements (Al and K) in sediments mainly from pipe material corrosions. Opportunistic pathogens presented in both water and sediments, and the latter could act as a reservoir of microbial contamination. There appears to be an association between potential opportunistic pathogens and microbial community structures. These microbial communities may be influenced by constituents within storage tank sediments. The results imply that compositions of microbial community and elements may influence and indicate microbial water quality and pipeline corrosion, and that these constituents may be important for optimal storage tank management within a distribution system.

Comparison of two poultry litter qPCR assays targeting the 16S rRNA gene of Brevibacterium sp.

Chicken feces commonly contain human pathogens and are also important sources of fecal pollution in environmental waters. Consequently, methods that can detect chicken fecal pollution are needed in public health and environmental monitoring studies. In this study, we compared a previously developed SYBR green qPCR assay (LA35) to a novel TaqMan qPCR assay (CL) for the environmental detection of poultry-associated fecal pollution. We tested both assays against chicken litter (n = 40), chicken fecal samples (n = 186), non-chicken fecal sources (n = 484), and environmental water samples (n = 323). Most chicken litter samples (i.e., ≥ 98%) were positive for both assays with relatively high signal intensities, whereas only 23% and 12% of poultry fecal samples (n = 186) were positive with the LA35 and the CL assays, respectively. Data using fecal samples from non-target animal species showed that the assays are highly host-associated (≥ 95%). Bayesian statistical models showed that the two assays are associated with relatively low probability of false-positive and false-negative signals in water samples. The CL marker had a lower prevalence than the LA35 assay when tested against environmental water samples (i.e., 21% vs. 31% positive signals). However, by combining the results from the two assays the detection levels increased to 41%, suggesting that using multiple assays can improve the detection of chicken-fecal pollution in environmental waters.

Effects of stock use and backpackers on water quality in wilderness in Sequoia and Kings Canyon National Parks, USA.

During 2010-2011, a study was conducted in Sequoia and Kings Canyon National Parks (SEKI) to evaluate the influence of pack animals (stock) and backpackers on water quality in wilderness lakes and streams. The study had three main components: (1) a synoptic survey of water quality in wilderness areas of the parks, (2) paired water quality sampling above and below several areas with differing types and amounts of visitor use, and (3) intensive monitoring at six sites to document temporal variations in water quality. Data from the synoptic water quality survey indicated that wilderness lakes and streams are dilute and have low nutrient and Escherichia coli concentrations. The synoptic survey sites were categorized as minimal use, backpacker-use, or mixed use (stock and backpackers), depending on the most prevalent type of use upstream from the sampling locations. Sites with mixed use tended to have higher concentrations of most constituents (including E. coli) than those categorized as minimal-use (P ≤ 0.05); concentrations at backpacker-use sites were intermediate. Data from paired-site sampling indicated that E. coli, total coliform, and particulate phosphorus concentrations were greater in streams downstream from mixed-use areas than upstream from those areas (P ≤ 0.05). Paired-site data also indicated few statistically significant differences in nutrient, E. coli, or total coliform concentrations in streams upstream and downstream from backpacker-use areas. The intensive-monitoring data indicated that nutrient and E. coli concentrations normally were low, except during storms, when notable increases in concentrations of E. coli, nutrients, dissolved organic carbon, and turbidity occurred. In summary, results from this study indicate that water quality in SEKI wilderness generally is good, except during storms; and visitor use appears to have a small, but statistically significant influence on stream water quality.

Distribution and potential significance of a gull fecal marker in urban coastal and riverine areas of southern Ontario, Canada.

To better understand the distribution of gull fecal contamination in urban areas of southern Ontario, we used gull-specific PCR and qPCR assays against 1309 water samples collected from 15 urban coastal and riverine locations during 2007. Approximately, 58% of the water samples tested positive for the gull-assay. Locations observed to have higher numbers of gulls and their fecal droppings had a higher frequency of occurrence of the gull marker and a higher gull marker qPCR signal than areas observed to be less impacted by gulls. Lower gull marker occurrence and lower qPCR signals were associated with municipal wastewater (7.4%) and urban stormwater effluents (29.5%). Overall, there were no statistically significant differences in gull marker occurrence at beach sites for pore water, ankle, and chest-depth samples, although signals were generally higher in interstitial beach sand pore water and ankle-depth water than in chest-depth water samples. Overall, the results indicated that gull fecal pollution is widespread in urban coastal and riverine areas in southern Ontario and that it significantly contributes to fecal indicator bacterial loads.

Characterization of Enterococcus faecalis-infecting phages (enterophages) as markers of human fecal pollution in recreational waters.

Enterophages are a novel group of phages that specifically infect Enterococcus faecalis and have been recently isolated from environmental water samples. Although enterophages have not been conclusively linked to human fecal pollution, we are currently characterizing enterophages to propose them as viral indicators and possible surrogates of enteric viruses in recreational waters. Little is known about the morphological or genetic diversity which will have an impact on their potential as markers of human fecal contamination. In the present study we are determining if enterophages can be grouped by their ability to replicate at different temperatures, and if different groups are present in the feces of different animals. As one of the main objectives is to determine if these phages can be used as indicators of the presence of enteric viruses, the survival rate under different conditions was also determined as was their prevalence in sewage and a large watershed. Coliphages were used as a means of comparison in the prevalence and survival studies. Results indicated that the isolates are mainly DNA viruses. Their morphology as well as their ability to form viral plaques at different temperatures indicates that several groups of enterophages are present in the environment. Coliphage and enterophage concentrations throughout the watershed were lower than those of thermotolerant coliforms and enterococci. Enterophage concentrations were lower than coliphages at all sampling points. Enterophages showed diverse inactivation rates and T(90) values across different incubation temperatures in both fresh and marine waters and sand. Further molecular characterization of enterophages may allow us to develop probes for the real-time detection of these alternative indicators of human fecal pollution.

Turkey fecal microbial community structure and functional gene diversity revealed by 16S rRNA gene and metagenomic sequences.

The primary goal of this study was to better understand the microbial composition and functional genetic diversity associated with turkey fecal communities. To achieve this, 16S rRNA gene and metagenomic clone libraries were sequenced from turkey fecal samples. The analysis of 382 16S rRNA gene sequences showed that the most abundant bacteria were closely related to Lactobacillales (47%), Bacillales (31%), and Clostridiales (11%). Actinomycetales, Enterobacteriales, and Bacteroidales sequences were also identified, but represented a smaller part of the community. The analysis of 379 metagenomic sequences showed that most clones were similar to bacterial protein sequences (58%). Bacteriophage (10%) and avian viruses (3%) sequences were also represented. Of all metagenomic clones potentially encoding for bacterial proteins, most were similar to low G+C Gram-positive bacterial proteins, particularly from Lactobacillales (50%), Bacillales (11%), and Clostridiales (8%). Bioinformatic analyses suggested the presence of genes encoding for membrane proteins, lipoproteins, hydrolases, and functional genes associated with the metabolism of nitrogen and sulfur containing compounds. The results from this study further confirmed the predominance of Firmicutes in the avian gut and highlight the value of coupling 16S rRNA gene and metagenomic sequencing data analysis to study the microbial composition of avian fecal microbial communities.

Quantitative PCR for detection and enumeration of genetic markers of bovine fecal pollution.

Accurate assessment of health risks associated with bovine (cattle) fecal pollution requires a reliable host-specific genetic marker and a rapid quantification method. We report the development of quantitative PCR assays for the detection of two recently described bovine feces-specific genetic markers and a method for the enumeration of these markers using a Markov chain Monte Carlo approach. Both assays exhibited a range of quantification from 25 to 2 x 10(6) copies of target DNA, with a coefficient of variation of <2.1%. One of these assays can be multiplexed with an internal amplification control to simultaneously detect the bovine-specific genetic target and presence of amplification inhibitors. The assays detected only cattle fecal specimens when tested against 204 fecal DNA extracts from 16 different animal species and also demonstrated a broad distribution among individual bovine samples (98 to 100%) collected from five geographically distinct locations. The abundance of each bovine-specific genetic marker was measured in 48 individual samples and compared to quantitative PCR-enumerated quantities of rRNA gene sequences representing total Bacteroidetes, Bacteroides thetaiotaomicron, and enterococci in the same specimens. Acceptable assay performance combined with the prevalence of DNA targets across different cattle populations provides experimental evidence that these quantitative assays will be useful in monitoring bovine fecal pollution in ambient waters.