Response of microbial community and catabolic genes to simulated petroleum hydrocarbon spills in soils/sediments from different geographic locations.

AIMS: Study the response of microbial communities and selected petroleum hydrocarbon (PH)-degrading genes on simulated PH spills in soils/sediments from different geographic locations. METHODS AND RESULTS: A microcosm experiment was conducted by spiking mixtures of petroleum hydrocarbons (PHs) to soils/sediments collected from four different regions of China, including the Dagang Oilfield (DG), Sand of Bohai Sea (SS), Northeast China (NE) and Xiamen (XM). Changes in bacterial community and the abundance of PH-degrading genes (alkB, nah and phe) were analysed by denaturing gradient electrophoresis (DGGE) and qPCR, respectively. Degradation of alkanes and PAHs in SS and NE materials were greater (P < 0·05) than those in DG and XM. Clay content was negatively correlated with the degradation of total alkanes by 112 days and PAHs by 56 days, while total organic carbon content was negatively correlated with initial degradation of total alkanes as well as PAHs. Abundances of alkB, nah and phe genes increased 10- to 100-fold and varied by soil type over the incubation period. DGGE fingerprints identified the dominance of α-, ß- and γ-Proteobacteria (Gram -ve) and Actinobacteria (Gram +ve) bacteria associated with degradation of PHs in the materials studied. CONCLUSION: The geographic divergence resulting from the heterogeneity of physicochemical properties of soils/sediments appeared to influence the abundance of metabolic genes and community structure of microbes capable of degrading PHs. SIGNIFICANCE AND IMPACT OF THE STUDY: When developing practical in-situ bioremediation approaches for PHs contamination of soils/sediment, appropriate microbial community structures and the abundance of PH-degrading genes appear to be influenced by geographic location.

Bacteriophages as indicators of faecal pollution and enteric virus removal.

Bacteriophages are an attractive alternative to faecal indicator bacteria (FIB), particularly as surrogates of enteric virus fate and transport, due to their closer morphological and biological properties. Based on a review of published data, we summarize densities of coliphages (F+ and somatic), Bacteroides spp. and enterococci bacteriophages (phages) in individual human waste, raw wastewater, ambient fresh and marine waters and removal through wastewater treatment processes utilizing traditional treatments. We also provide comparisons with FIB and enteric viruses whenever possible. Lastly, we examine fate and transport characteristics in the aquatic environment and provide an overview of the environmental factors affecting their survival. In summary, concentrations of bacteriophages in various sources were consistently lower than FIB, but more reflective of infectious enteric virus levels. Overall, our investigation indicates that bacteriophages may be adequate viral surrogates, especially in built systems, such as wastewater treatment plants. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacteriophage are alternative fecal indicators that may be better surrogates for viral pathogens than fecal indicator bacteria (FIB). This report offers a summary of the existing literature concerning the utility of bacteriophage as indicators of viral presence (fecal sources and surface waters) and persistence (in built infrastructure and aquatic environments). Our findings indicate that bacteriophage levels in all matrices examined are consistently lower than FIB, but similar to viral pathogens. Furthermore, in built infrastructure (e.g. wastewater treatment systems) bacteriophage closely mimic viral pathogen persistence suggesting they may be adequate sentinels of enteric virus removal.

QMRA and water safety management: review of application in drinking water systems.

Quantitative microbial risk assessment (QMRA), the assessment of microbial risks when model inputs and estimated health impacts are explicitly quantified, is a valuable tool to support water safety plans (WSP). In this paper, research studies undertaken on the application of QMRA in drinking water systems were reviewed, highlighting their relevance for WSP. The important elements for practical implementation include: the data requirements to achieve sufficient certainty to support decision-making; level of expertise necessary to undertake the required analysis; and the accessibility of tools to support wider implementation, hence these aspects were the focus of the review. Recommendations to support the continued and growing application of QMRA to support risk management in the water sector are provided.

Changes in bacterial composition of biofilm in a metropolitan drinking water distribution system.

AIMS: This study examined the development of bacterial biofilms within a metropolitan distribution system. The distribution system is fed with different source water (i.e. groundwater, GW and surface water, SW) and undergoes different treatment processes in separate facilities. METHODS AND RESULTS: The biofilm community was characterized using 16S rRNA gene clone libraries and functional potential analysis, generated from total DNA extracted from coupons in biofilm annular reactors fed with onsite drinking water for up to 18 months. Differences in the bacterial community structure were observed between GW and SW. Representatives that explained the dissimilarity were associated with the classes Betaproteobacteria, Alphaproteobacteria, Actinobacteria, Gammaproteobacteria and Firmicutes. After 9 months the biofilm bacterial community from both GW and SW were dominated by Mycobacterium species. The distribution of the dominant operational taxonomic unit (OTU) (Mycobacterium) positively correlated with the drinking water distribution system (DWDS) temperature. CONCLUSIONS: In this study, the biofilm community structure observed between GW and SW were dissimilar, while communities from different locations receiving SW did not show significant differences. The results suggest that source water and/or the water quality shaped by their respective treatment processes may play an important role in shaping the bacterial communities in the distribution system. In addition, several bacterial groups were present in all samples, suggesting that they are an integral part of the core microbiota of this DWDS. SIGNIFICANCE AND IMPACT OF THE STUDY: These results provide an ecological insight into biofilm bacterial structure in chlorine-treated drinking water influenced by different water sources and their respective treatment processes.

Dose-response algorithms for water-borne Pseudomonas aeruginosa folliculitis.

We developed two dose-response algorithms for P. aeruginosa pool folliculitis using bacterial and lesion density estimates, associated with undetectable, significant, and almost certain folliculitis. Literature data were fitted to Furumoto & Mickey's equations, developed for plant epidermis-invading pathogens: N l = A ln(1 + BC) (log-linear model); P inf = 1-e(-r c C) (exponential model), where A and B are 2.51644 × 107 lesions/m2 and 2.28011 × 10-11 c.f.u./ml P. aeruginosa, respectively; C = pathogen density (c.f.u./ml), N l = folliculitis lesions/m2, P inf = probability of infection, and r C = 4·3 × 10-7 c.f.u./ml P. aeruginosa. Outbreak data indicates these algorithms apply to exposure durations of 41 ± 25 min. Typical water quality benchmarks (≈10-2 c.f.u./ml) appear conservative but still useful as the literature indicated repeated detection likely implies unstable control barriers and bacterial bloom potential. In future, culture-based outbreak testing should be supplemented with quantitative polymerase chain reaction and organic carbon assays, and quantification of folliculitis aetiology to better understand P. aeruginosa risks.

Impact of drinking water conditions and copper materials on downstream biofilm microbial communities and Legionella pneumophila colonization.

AIMS: This study examined the impact of pipe materials and introduced Legionella pneumophila on downstream Leg. pneumophila colonization and microbial community structures under conditions of low flow and low chlorine residual. METHODS AND RESULTS: CDC biofilm(™) reactors containing either unplasticized polyvinylchloride (uPVC) or copper (Cu) coupons were used to develop mature biofilms on Norprene(™) tubing effluent lines to simulate possible in-premise biofilm conditions. The microbial communities were characterized through 16S and 18S rRNA gene clone libraries and Leg. pneumophila colonization was determined via specific qPCR assays. The Cu significantly decreased downstream microbial diversity, approximately halved bacterial and eukaryotic abundance, with some groups only detected in uPVC-reactor tubing biofilms. However, some probable amoeba-resisting bacteria (ARB) like Mycobacterium spp. and Rhodobacteraceae were significantly more abundant in the Cu than uPVC-reactor tubing biofilms. In particular, Leg. pneumophila only persisted (postinoculation) within the Cu-reactor tubing biofilms, and the controlled low chlorine residue and water flow conditions led to a general high abundance of possible free-living protozoa in all tubing biofilms. The higher relative abundance of ARB-like sequences from Cu-coupons vs uPVC may have been promoted by amoebal selection and subsequent ARB protection from Cu inhibitory effects. CONCLUSIONS: Copper pipe and low flow conditions had significant impact on downstream biofilm microbial structures (on plastic pipe) and the ability for Leg. pneumophila colonization post an introduction event. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report that compares the effects of copper and uPVC materials on downstream biofilm communities grown on a third (Norprene(™)) surface material. The downstream biofilms contained a high abundance of free-living amoebae and ARB, which may have been driven by a lack of residual disinfectant and periodic stagnant conditions. Given the prevalence of Cu-piping in buildings, there may be increased risk from drinking water exposures to ARB following growth on pipe/fixture biofilms within premise drinking water systems.

Evaluation of Bacteroides fragilis GB-124 bacteriophages as novel human-associated faecal indicators in the United States.

UNLABELLED: Phages infecting human-associated Bacteroides fragilis (GB-124 phages) have been employed in the European Union (EU) to identify human faecal pollution, but their utility for the United States was unclear. Primary sewage samples were collected seasonally from seven wastewater treatment plants (WWTP) across the continental United States, and more time-intensive sampling was conducted at local WWTPs. All samples were assayed for plaque-forming units (PFU) of GB-124 phages, somatic and FRNA-specific coliphages, as well as adenoviruses (by quantitative PCR [qPCR]). Animal faecal samples (>250) from 14 different species were tested for the presence of the three phage groups. GB-124 phages were consistently detected in sewage (10-10(2) PFU ml(-1) ), but not in animal faeces. While density estimates of both coliphages in sewage were approximately one order of magnitude higher than GB-124 phages, they were both randomly detected in animal faecal samples (10(2) -10(5) g(-1) dry weight). Stability of all three phages was inversely proportional to temperature; persistence was greatest at 5°C compared to 20 and 35°C, where no phages were detectable after a week. In summary, GB-124 phages appear to be a feasible alternative indicator organism and benefit from being sewage associated, while providing an inexpensive detection technique for infectious virions. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacteroides fragilis GB-124 phages appear to be restricted to human sewage sources in the United States, being absent from 264 animal faecal samples from 14 different species and present in approx. 90% (34/38) of primary sewage effluent samples collected across the country. Although somatic and F-specific coliphages were present in sewage samples at higher densities, unlike GB-124 phages, both coliphage types were also detected in animal faecal samples. Hence, GB-124 phages may prove to be a useful novel indicator group for human faecal pollution in the continental United States.

Pseudomonas aeruginosa dose response and bathing water infection.

Pseudomonas aeruginosa is the opportunistic pathogen mostly implicated in folliculitis and acute otitis externa in pools and hot tubs. Nevertheless, infection risks remain poorly quantified. This paper reviews disease aetiologies and bacterial skin colonization science to advance dose-response theory development. Three model forms are identified for predicting disease likelihood from pathogen density. Two are based on Furumoto & Mickey's exponential 'single-hit' model and predict infection likelihood and severity (lesions/m2), respectively. 'Third-generation', mechanistic, dose-response algorithm development is additionally scoped. The proposed formulation integrates dispersion, epidermal interaction, and follicle invasion. The review also details uncertainties needing consideration which pertain to water quality, outbreaks, exposure time, infection sites, biofilms, cerumen, environmental factors (e.g. skin saturation, hydrodynamics), and whether P. aeruginosa is endogenous or exogenous. The review's findings are used to propose a conceptual infection model and identify research priorities including pool dose-response modelling, epidermis ecology and infection likelihood-based hygiene management.

Estimating the risk from sewage treatment plant effluent in the Sydney catchment area.

This study employed Quantitative Microbial Risk Assessment (QMRA) to estimate the gastrointestinal risks associated with Cryptosporidium and Giardia discharged from three STPs located within the Lake Burragorang catchment. The QMRA considered baseline and various hazardous event scenarios (e.g. plant failure and heavy rainfall). Under baseline conditions, the combined effect of constructed barriers, catchment barriers and dilution reduced pathogen numbers from the discharge of all three STPs by 10 to 14 orders of magnitude. This was sufficient for the risk to be well below currently mooted benchmarks of 'tolerable risk', even when relatively conservative assumptions were applied. For all hazardous event scenarios, the level of risk remained low, which illustrated the benefit of multiple barriers. Provisionally it appears that the STPs currently discharging into the waterways of the catchment do not pose an unacceptable or unmanageable risk to Sydney's drinking water consumers.

Differential growth of Legionella pneumophila strains within a range of amoebae at various temperatures associated with in-premise plumbing.

AIMS: The potential effect of in-premise plumbing temperatures (24, 32, 37 and 41°C) on the growth of five different Legionella pneumophila strains within free-living amoebae (Acanthamoeba polyphaga, Hartmannella vermiformis and Naegleria fowleri) was examined. METHODS AND RESULTS: Compared with controls that actively fed on Escherichia coli prey, when Leg. pneumophila was used as prey, strains Lp02 and Bloomington-2 increased in growth at 30, 32 and 37°C while strains Philadelphia-1 and Chicago 2 did not grow at any temperature within A. polyphaga. Strains Lp02, Bloomington-2 and Dallas 1E did not proliferate in the presence of H. vermiformis nor did strain Philadelphia-1 in the presence of N. fowleri. Yet, strain Bloomington-2 grew at all temperatures examined within N. fowleri, while strain Lp02 proliferated at all temperatures except 41°C. More intriguing, strain Chicago 2 only grew at 32°C within H. vermiformis and N. fowleri suggesting a limited temperature growth range for this strain. CONCLUSIONS: Identifying the presence of pathogenic legionellae may require the use of multiple host amoebae and incubation temperatures. SIGNIFICANCE AND IMPACT OF THE STUDY: Temperature conditions and species of amoeba host supported in drinking water appear to be important for the selection of human-pathogenic legionellae and point to future research required to better understand Legionella ecology.

Comparing probabilistic microbial risk assessments for drinking water against daily rather than annualised infection probability targets.

Some national drinking water guidelines provide guidance on how to define 'safe' drinking water. Regarding microbial water quality, a common position is that the chance of an individual becoming infected by some reference waterborne pathogen (e.g. Cryptsporidium) present in the drinking water should < 10(-4) in any year. However the instantaneous levels of risk to a water consumer vary over the course of a year, and waterborne disease outbreaks have been associated with shorter-duration periods of heightened risk. Performing probabilistic microbial risk assessments is becoming commonplace to capture the impacts of temporal variability on overall infection risk levels. A case is presented here for adoption of a shorter-duration reference period (i.e. daily) infection probability target over which to assess, report and benchmark such risks. A daily infection probability benchmark may provide added incentive and guidance for exercising control over short-term adverse risk fluctuation events and their causes. Management planning could involve outlining measures so that the daily target is met under a variety of pre-identified event scenarios. Other benefits of a daily target could include providing a platform for managers to design and assess management initiatives, as well as simplifying the technical components of the risk assessment process.

Quantitative Bayesian predictions of source water concentration for QMRA from presence/absence data for E. coli O157:H7.

A hierarchical Bayesian framework was applied for describing variability in pathogen concentration (with associated uncertainty) from presence/absence observations for E. coli O157:H7. Laboratory spiking experiments (method performance) and environmental sample assays were undertaken for a surface drinking water source in France. The concentration estimates were strongly dependent upon the assumed statistical model used (gamma, log-gamma or log-gamma constrained), highlighting the need for a solid theoretical basis for model choice. Bayesian methods facilitate the incorporation of additional data into the statistical analysis; this was illustrated using faecal indicator results of E. coli (Colilert) to reduce the posterior parameter uncertainty and improve model stability. While conceptually simple, application of these methods is still specialised, hence there is a need for the development of data analysis tools to make Bayesian simulation techniques more accessible for QMRA practitioners.

The role of biofilms and protozoa in Legionella pathogenesis: implications for drinking water.

Current models to study Legionella pathogenesis include the use of primary macrophages and monocyte cell lines, various free-living protozoan species and murine models of pneumonia. However, there are very few studies of Legionella spp. pathogenesis aimed at associating the role of biofilm colonization and parasitization of biofilm microbiota and release of virulent bacterial cell/vacuoles in drinking water distribution systems. Moreover, the implications of these environmental niches for drinking water exposure to pathogenic legionellae are poorly understood. This review summarizes the known mechanisms of Legionella spp. proliferation within Acanthamoeba and mammalian cells and advocates the use of the amoeba model to study Legionella pathogenicity because of their close association with Legionella spp. in the aquatic environment. The putative role of biofilms and amoebae in the proliferation, development and dissemination of potentially pathogenic Legionella spp. is also discussed. Elucidating the mechanisms of Legionella pathogenicity development in our drinking water systems will aid in elimination strategies and procedural designs for drinking water systems and in controlling exposure to Legionella spp. and similar pathogens.

Solar radiation disinfection of drinking water at temperate latitudes: inactivation rates for an optimised reactor configuration.

Solar radiation-driven inactivation of bacteria, virus and protozoan pathogen models was quantified in simulated drinking water at a temperate latitude (34 degrees S). The water was seeded with Enterococcus faecalis, Clostridium sporogenes spores, and P22 bacteriophage, each at ca 1x10(5) mL(-1), and exposed to natural sunlight in 30-L reaction vessels. Water temperature ranged from 17 to 39 degrees C during the experiments lasting up to 6h. Dark controls showed little inactivation and so it was concluded that the inactivation observed was primarily driven by non-thermal processes. The optimised reactor design achieved S90 values (cumulative exposure required for 90% reduction) for the test microorganisms in the range 0.63-1.82 MJ m(-2) of Global Solar Exposure (GSX) without the need for TiO2 as a catalyst. High turbidity (840-920 NTU) only reduced the S(90) value by <40%. Further, when all S90 means were compared this decrease was not statistically significant (prob.>0.05). However, inactivation was significantly reduced for E. faecalis and P22 when the transmittance of UV wavelengths was attenuated by water with high colour (140 PtCo units) or a suboptimally transparent reactor lid (prob.<0.05). S90 values were consistent with those measured by other researchers (ca 1-10 MJ m(-2)) for a range of waters and microorganisms. Although temperatures required for SODIS type pasteurization were not produced, non-thermal inactivation alone appeared to offer a viable means for reliably disinfecting low colour source waters by greater than 4 orders of magnitude on sunny days at 34 degrees S latitude.

Microbial challenge-testing of treatment processes for quantifying stormwater recycling risks and management.

Pathogenic microorganisms have been identified as the main human health risks associated with the reuse of treated urban stormwater (runoff from paved and unpaved urban areas). As part of the Smart Water initiative (Victorian Government, Australia), a collaborative evaluation of three existing integrated stormwater recycling systems, and the risks involved in non-potable reuse of treated urban stormwater is being undertaken. Three stormwater recycling systems were selected at urban locations to provide a range of barriers including biofiltration, storage tanks, UV disinfection, a constructed wetland, and retention ponds. Recycled water from each of the systems is used for open space irrigation. In order to adequately undertake exposure assessments, it was necessary to quantify the efficacy of key barriers in each exposure pathway. Given that none of the selected treatment systems had previously been evaluated for their treatment efficiency, experimental work was carried out comprising dry and wet weather monitoring of each system (for a period of 12 months), as well as challenging the barriers with model microbes (for viruses, bacteria and parasitic protozoa) to provide input data for use in Quantitative Microbial Risk Assessment.

Chemical contaminants in feedlot wastes: concentrations, effects and attenuation.

Commercial feedlots for beef cattle finishing are potential sources of a range of trace chemicals which have human health or environmental significance. To ensure adequate protection of human and environmental health from exposure to these chemicals, the application of effective manure and effluent management practices is warranted. The Australian meat and livestock industry has adopted a proactive approach to the identification of best management practices. Accordingly, this review was undertaken to identify key chemical species that may require consideration in the development of guidelines for feedlot manure and effluent management practices in Australia. Important classes of trace chemicals identified include steroidal hormones, antibiotics, ectoparasiticides, mycotoxins, heavy metals and dioxins. These are described in terms of their likely sources, expected concentrations and public health or environmental significance based on international data and research. Androgenic hormones such as testosterone and trenbolone are significantly active in feedlot wastes, but they are poorly understood in terms of fate and environmental implications. The careful management of residues of antibiotics including virginiamycin, tylosin and oxytetracycline appears prudent in terms of minimising the risk of potential public health impacts from resistant strains of bacteria. Good management of ectoparasiticides including synthetic pyrethroids, macrocyclic lactones, fluazuron, and amitraz is important for the prevention of potential ecological implications, particularly towards dung beetles. Very few of these individual chemical contaminants have been thoroughly investigated in terms of concentrations, effects and attenuation in Australian feedlot wastes.

A new approach to aid urban water management decision making using trade-off sacrifice modelled by fuzzy logic.

An approach to aid decision making for urban water management is presented that is based on the concept of trade-off sacrifice level in pairwise comparisons between criteria, modelled using fuzzy logic. This approach is illustrated by a case study - selection of alternative water supplies for a Sydney household. Four key decision making criteria covering health, economic, environment and technical aspects are selected: annual probability of infection, life cycle energy use, life cycle cost and reliability. The decision making problem is to select between cases with different volume and application of recycled greywater and rainwater in light of the four criteria. Decision maker's preference is expressed by five levels of trade-off sacrifice between pairs of criteria. The decision makers can assign their preferences for sacrifice level by linguistic assessment and the output trade-off weight (TOW). Measures of decision makers' perceived trade-off level are modelled by a rule-based fuzzy logic control system. The final analysis shows the performance for each sacrifice class for each case, to aid overall decision making with stakeholders.

Microbial exposure assessment of an urban recreational lake: a case study of the application of new risk-based guidelines.

New WHO and Australian guidelines promote a risk-management approach for minimising exposure to pathogens in recreational waters. Between 2003 and 2005, they were applied to Lake Parramatta (10 ha, 450 ML), a potential recreation site in Sydney, Australia. A three stage approach was developed involving (1) initial suitability assessment using historic data, (2) revised suitability assessment based on new data and (3) characterisation of hazardous (especially wet weather) events. Contrary to the stage 1 suitability classification, stage 2 baseline data indicated that during dry weather the lake had water quality sufficient for primary contact recreation (95th percentiles for enterococci = 19 MPN/100, n = 50) and the major pathogen source was wildfowl. Guideline principles provided a rationale for collecting microbiological and geographic data needed to understand local cycles of lake contamination/recovery. The concept of hazardous events was particularly useful. Studies of stormwater events led us to identify a transition point (> 10 mm rainfall in 24 h) where human-faecal pathogen risks increased and access needed to be controlled. Together baseline and event data yielded operational tools (i.e. event detection methods, action triggers, auditing criteria, remediation priorities) for minimising bather exposure.

Microbial risk and removal--a utility perspective.

In this paper the results of a sampling programme, undertaken as part of the EU MicroRisk project, are described. This project was undertaken to ascertain the occurrence of pathogens and indicators in the River Thames and their subsequent removal through a treatment works. Appropriate physico-chemical surrogates, as determined by statistical correlation are proposed for the microorganisms identified in the raw water. This study shows that under normal raw water conditions the treatment works is able to remove microbial contamination with a significant margin of safety.

Pathogen monitoring offers questionable protection against drinking-water risks: a QMRA (quantitative microbial risk analysis) approach to assess management strategies.

Risk mitigation provided by human monitoring and control over a water supply system has been consistently overlooked when estimating pathogen exposure to consumers. The Systems-Actions-Management (SAM) framework lends itself neatly to Quantitative Microbial Risk Assessment (QMRA) as one way to establish this link. The general premise is that an organisational protocol will influence how a human controller behaves, in turn influencing the system performance. For illustrative purposes, the framework was applied to a hypothetical water supply system to quantify the risk reduction offered by routine Cryptosporidium monitoring and the response to oocyst 'detects'. Our findings suggest that infrequent direct pathogen monitoring may provide a negligible risk barrier. The practice of sampling treated water to verify microbiological integrity is also dubious: oocyst densities were largely under-estimated, in part due to the spatial dispersion of oocysts in the waterbody, but predominantly from imperfect detection methods. The development of 'event-driven' monitoring schemes with barrier performance-based treatment verification methods, as promoted in new guidelines, is supported as a pressing issue to reduce the likelihood of undetected pathogen passage through a treatment plant.