Profiling bacterial survival through a water treatment process and subsequent distribution system

AIMS: To profile fractions of active bacteria and of bacteria culturable with routine heterotrophic plate count (HPC) methods through a typical water treatment process and subsequent distribution system. In doing so, investigate how water treatment affects both bacterial abundance and diversity, and reveal the identities of active bacteria not detected by traditional HPC culture. METHODS AND RESULTS: Profiling active fractions was performed by flow cytometric cell sorting of either membrane-intact (BacLight kit) or enzymatically active (carboxyfluorescein diacetate, CFDA) bacteria, followed by eubacterial 16S rDNA-directed PCR and denaturing gradient gel electrophoresis (DGGE). Water treatment significantly reduced active bacterial numbers detected by the BacLight kit and CFDA assay by 2.89 and 2.81 log respectively. Bacterial diversity was also reduced from > 20 DGGE bands in the active fractions of reservoir water to only two bands in the active fractions of finished water. These two bands represented Stenotrophomonas maltophila, initially culturable by HPC, and a Burkholderia-related species. Both species maintained measurable traits of physiological activity in distribution system bulk water but were undetected by HPC. CONCLUSIONS: Flow cytometric cell sorting with PCR-DGGE, to assess water treatment efficacy, identified active bacteria from a variety of major phylogenetic groups undetected by routine HPC. Following treatment S. maltophila and a Burkholderia-related species retained activity and entered distribution undetected by HPC. SIGNIFICANCE AND IMPACT OF THE STUDY: Methods used here demonstrate how water treatment operators can better monitor water treatment plant efficacy and assess distribution system instability by the detection and identification of active bacteria recalcitrant to routine HPC culture.     

Molecular analysis of microbial population transition associated with the start of denitrification in a wastewater treatment process

AIMS: The objective of this study is to determine the bacteria playing an important role in denitrification by monitoring the molecular dynamics accompanying the start of denitrification. METHODS AND RESULTS: cDNA reverse-transcribed from 16S rRNA was amplified with fluorescent labelled primer for terminal restriction fragment length polymorphism (T-RFLP) analysis and an unlabelled primer for cloning analysis. The terminal restriction fragments (T-RFs) that increased in association with the start of denitrification were determined. These T-RFs were identified by in silico analysis of 16S rRNA sequences obtained from cloning. As a result, it was clearly observed that the bacteria belonging to the genera Hydrogenophaga and Acidovorax increased in number after the start of denitrification. CONCLUSIONS: It was demonstrated that T-RFLP analysis targeting 16S rRNA is appropriate for the daily monitoring of a bacterial community to control wastewater treatment processes. Combination of the results of T-RFLP analysis and 16S rRNA clone library indicated that the bacteria belonging to the genera Hydrogenophaga and Acidovorax play an important role in denitrification. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of this study provide new insight to the 16S rRNA level of active denitrifying bacteria in wastewater treatment processes.     

Effect of the pH of growth on the survival of Lactobacillus delbrueckii subsp. bulgaricus to stress conditions during spray-drying

AIMS: The aim of this study was to optimize survival of Lactobacillus delbrueckii subsp. bulgaricus during spray-drying and subsequent storage through optimizing the pH of growth conditions. METHODS AND RESULTS: Cell concentrates previously grown without or with pH controlled were spray-dried and stored at 20 degrees C and heat treated at 57 degrees C. Cells grown under noncontrolled pH were more resistant to both drying and heating than cells grown under controlled pH but no significant differences were observed during storage. The intracellular proteins profile of cells grown under both conditions was studied by two-dimensional SDS-polyacrylamide gel electrophoresis. Eight proteins were identified using automated mass spectrometry (MS) and tandem mass spectrometry (MS/MS) data acquisition. Of the identified proteins, only cochaperonin GroES corresponded to a known heat shock protein (HSP). The other proteins identified are proteins involved in glycolysis. For cells grown under noncontrolled pH the expression of the Hsp70, GroES and GroEL, measured by Western blotting, was enhanced. CONCLUSIONS: The higher resistance of cells grown under noncontrolled pH correlates with the enhanced production of heat shock proteins. SIGNIFICANCE AND IMPACT OF THE STUDY: Growth of L. bulgaricus under controlled pH (commonly used by the starter cultures production industry) results in cells more sensitive to stresses frequently encountered by the cells during starter cultures preparation/storage/utilization.     

An estimation of the floristic richness of Bolivia’s Andean dry valleys

We estimate plant species richness of Bolivian Andean dry valleys applying the species/genus ratio of Compositae to all reported genera for those regions. The results suggest the existence of around 1500 species in the Bolivian dry valleys (12.6% more than the species recorded). A brief biogeographical analysis of our results suggests that the use of species/genus ratio of Compositae to estimate the size of the flora in Bolivian Andean dry valleys should be considered with caution, since the central Andes are geologically very young, and explosive speciation processes have taken place only in some taxa (mainly in Bromeliaceae and Cactaceae). We offer some evidence pointing to the possibility that the Andean dry valleys may possess many more species than suggested by the Compositae species/genus ratio. We discuss the implications of our estimation for conservation of the Andean dry valleys, which only represent 0.7% of the protected area of Bolivia.     

Expression of ABC-1 transporter is elevated in human glioma cells under irradiation and temozolomide treatment

Summary. Objective: Chemo-therapeutic treatment of glioma patients has minor success. Little is known about mechanisms of a pronounced resistance of gliomas towards actual therapies, yet. ABC-1 belongs to the group of transporters known to be involved in the export of hydrophobic substances and vascular regulation. This study investigates an effect of both temozolomide (TMZ) treatment and/or irradiation on the expression of the ABC-1 transporter in human U87-MG glioma cells.Material and methods: In parallel experiments U87-MG cells underwent either irradiation (RT), chemo-treatment (CT) using TMZ, and combined chemo/radiation-treatment (CT/RT). After each treatment the cells were incubated either 2 or 24 hours at 37°C and counted before protein analysis using Western-Blot technique.Results and conclusions: An exponential growth of cellular density was observed for both untreated and irradiated cells being, however, about 2-times slower in irradiated compared to untreated cells. In contrast the density increase of chemo-treated cells as well as that of cells, which underwent the combined CT/RT treatment was of linear nature. ABC-1 expression was detected in untreated as well as treated cells. Increasing cell density and all kinds of treatment resulted in a considerably enhanced ABC-1 expression. CT treatment resulted in highly up-regulated ABC-1 expression especially in non-confluent cultures compared to untreated cells. Irradiation had a comparable or even higher inducible effect on the ABC-1 expression rates depending, however, on cell density. The highest expression rates were observed in cultures with high cellular density 2 hours after application of the combined treatment. Strong up-regulation of ABC-1 expression under both irradiation and chemo-treatment might be a clue to multidrug and irradiation cross-resistance mechanisms of malignant glioma cells converting the ABC-1 transporter into an attractive pharmacological target for a clinical breakthrough in the therapy of malignant gliomas.     

Waste Treatment and Assessment of Long-Term Emissions (8pp)

Goal, Scope and Background The disposal phase of a product’s life cycle in LCA is often neglected or based on coarse indicators like ‘kilogram waste’. The goal of report No. 13 of the ecoinvent project (Doka 2003) is to create detailed Life Cycle Inventories of waste disposal processes. The purpose of this paper is to give an overview of the models behind the waste disposal inventories in ecoinvent, to present exemplary results and to discuss the assessment of long-term emissions. This paper does not present a particular LCA study. Inventories are compiled for many different materials and various disposal technologies. Considered disposal technologies are municipal incineration and different landfill types, including sanitary landfills, hazardous waste incineration, waste deposits in deep salt mines, surface spreading of sludges, municipal wastewater treatment, and building dismantling. The inventoried technologies are largely based on Swiss plants. Inventories can be used for assessment of the disposal of common, generic waste materials like paper, plastics, packaging etc. Inventories are also used within the ecoinvent database itself to inventory the disposal of specific wastes generated during the production phase. Inventories relate as far as possible to the specific chemical composition of the waste material (waste-specific burdens). Certain expenditures are not related to the waste composition and are inventoried with average values (process-specific burdens). Methods The disposal models are based on previous work, partly used in earlier versions of ecoinvent/ETH LCI data. Important improvements were the extension of the number of considered chemical elements to 41 throughout all disposal models and new landfill models based on field data. New inventories are compiled for waste deposits in deep salt mines and building material disposal. Along with the ecoinvent data and the reports, also Excel-based software tools were created, which allow ecoinvent members to calculate waste disposal inventories from arbitrary waste compositions. The modelling of long-term emissions from landfills is a crucial part in any waste disposal process. In ecoinvent long-term emissions are defined as emissions occurring 100 years after present. They are reported in separate emission categories. The landfill inventories include long-term emissions with a time horizon of 60’000 years after present. Results and Discussion As in earlier studies, the landfills prove to be generally relevant disposal processes, as also incineration and wastewater treatment processes produce landfilled wastes. Heavy metals tend to concentrate in landfills and are washed out to a varying degree over time. Long-term emissions usually represent an important burden from landfills. Comparisons between burdens from production of materials and the burdens from their disposal show that disposal has a certain relevance. Conclusion The disposal phase should by default be included in LCA studies. The use of a material not only necessitates its production, but also requires its disposal. The created inventories and user tools facilitate heeding the disposal phase with a similar level of detail as production processes. The risk of LCA-based decisions shifting burdens from the production or use phase to the disposal phase because of data gaps can therefore be diminished. Recommendation and Perspective Future improvements should include the modelling of metal ore refining waste (tailings) which is currently neglected in ecoinvent, but is likely to be relevant for metals production. The disposal technologies considered here are those of developed Western countries. Disposal in other parts of the World can differ distinctly, for logistic, climatic and economic reasons. The cross-examination of landfill models to LCIA soil fate models could be advantageous. Currently only chemical elements, like copper, zinc, nitrogen etc. are heeded by the disposal models. A possible extension could be the modelling of the behaviour of chemical compounds, like dioxins or other hydrocarbons.     

Influence of organic selenium on hsp70 response of heat-stressed and enteropathogenic Escherichia coli-challenged broiler chickens (Gallus gallus)

The effect of dietary selenium yeast, a source of organic selenium, on heat shock protein 70 (hsp70) responses, redox status, growth and feed utilization were evaluated either in enteropathogenic Escherichia coli-challenged (EPEC) or in heat-stressed (HS) male broiler chickens grown to 42 days of age. One day-old chicks in experiment 1 were challenged orally with EPEC (10(6) cfu/chicken on day 1 and boosted by water application on days 2, 3, and 4) and fed diets with or without selenium yeast. Body weight (BW), feed conversion ratio (FCR), and total mortality were determined at 42 days of age, and this was followed by collection of ileal tissue for the quantification of total glutathione (TGSH), reduced glutathione (GSH), oxidized glutathione (GSSG), and hsp70 in randomly selected chickens from each treatment. In experiment 2, male broiler chickens were fed diets with or without selenium yeast under a thermoneutral rearing condition. At four weeks of age, blood and hepatic tissue were collected from chickens maintained in the thermoneutral environment and from chickens subjected to HS (40 degrees C for 1 h) and analyzed for TGSH, GSH, GSSG, and hsp70. Selenium yeast improved BW, FCR, and decreased mortality in both control and EPEC-challenged chicks. Selenium yeast significantly attenuated hsp70 expression in EPEC-challenged chickens and in those subjected to HS. The EPEC challenge increased TGSH and GSSG levels and decreased GSH/GSSG ratio. However, GSSG level accumulated in chickens fed diets without selenium supplementation resulting in a lower GSH/GSSG ratio in the selenium yeast-fed group. Heat stress increased GSSG level and decreased GSH/GSSG ratio. Selenium yeast-fed groups maintained higher levels of GSSG before and after HS with a resultant lower GSH/GSSG ratio. The hsp70 response was significantly less in those chickens fed selenium yeast and challenged with either EPEC or HS than in those chickens given no supplemental selenium. The results of this study suggest that selenium yeast supplementation had imparted resistance to oxidative stress associated with enteric bacteria infection and to high temperature exposure. It is believed that the resistance to the stressors was due to an improved redox status of the selenium yeast-fed chickens.     

Occurrence and Fate of Estrone, 17β-estradiol and 17α-ethynylestradiol in STPs for Domestic Wastewater

Estrone (E1), 17β-estradiol (E2) and 17α-ethynylestradiol (EE2) discharged from sewage treatment plants (STPs) into surface waters, are seen as a threat effecting aquatic life by its estrogenic character. Therefore, much research is conducted on the fate and removal of these compounds. Since these compounds are present in influents and effluents in the ng/l range, methods for detection deserve special attention. Most important processes that play a role in the removal of estrogens are: adsorption, aerobic degradation, anaerobic degradation, anoxic biodegradation and photolytic degradation. Halflifes tend to vary and are remarkably shorter when low initial concentrations are applied. In general anaerobic conditions result in longer halflifes then aerobic conditions. EE2 shows far most persistence of the compounds, thereby also the estrogenic effect in vitro is about 2–3-fold higher compared to E2. The three compounds show a higher affinity to sorb to sludge compared to other tested adsorption materials like sediment. Aerobic degradation is far the most efficient in removing these compounds, but adsorption seems to play a significant role in retaining the estrogens inside full-scale STPs. Removal rates in full scale plants depend on the HRT, SRT and loading rates, but lack of information on the exact dependency so far prevents an optimal design able to fully eliminate estrogens from wastewater.     

Effects of pH amendment on metal working fluid wastewater biological treatment using a defined bacterial consortium

The aim of this study was to determine whether pH amendment of a highly alkaline metal working fluid (MWF) wastewater would improve biological treatment in a bioreactor system following introduction of a bacterial inoculum (comprised of the following strains: Agrobacterium radiobacter, Comamonas testosteroni, Methylobacterium mesophilicum, Microbacterium esteraromaticum, and Microbacterium saperdae). The pH values tested were 6, 7, 8, and 9. Three replicate batch mode bioreactors inoculated with the bacterial inoculum (plus an abiotic control bioreactor) were operated for each of the four pH conditions. After 14 days, the final mean chemical oxygen demand (COD) reduction at pH 9 was 50 +/- 1.4%; at pH 8, 58 +/- 1.4%; pH 7, 65 +/- 1.0%; and pH 6, 75 +/- 2.7% of the initial COD (approximately 10,000 mg L(-1)), respectively. Interestingly, within 5 days, the pH in all inoculated bioreactors progressed toward pH 8. However, all abiotic control bioreactors remained at the pH at which they were amended. The fate of the inoculum, determined by denaturing gradient gel electrophoresis (DGGE) and by cluster analysis of the resulting DGGE profiles, revealed that the inocula survived throughout operation of all pH-amended bioreactors. Length-heterogeneity polymerase chain reaction (PCR) was used to track the population dynamics of individual strains. After 7 days of operation, M. esteraromaticum was the most abundant population in all bioreactors, regardless of pH. From our findings, it appears necessary to adjust the MWF wastewater from pH 9 to between 6 and 7, to achieve optimal biological treatment rates.