The demonstration of a novel sulfur cycle-based wastewater treatment process: Sulfate reduction, autotrophic denitrification, and nitrification integrated (SANI®) biological nitrogen removal process

Saline water supply has been successfully practiced for toilet flushing in Hong Kong since 1950s, which saves 22% of freshwater in Hong Kong. In order to extend the benefits of saline water supply into saline sewage management, we have recently developed a novel biological organics and nitrogen removal process: the Sulfate reduction, Autotrophic denitrification, and Nitrification Integrated (SANI®) process. The key features of this novel process include elimination of oxygen demand in organic matter removal and production of minimal sludge. Following the success of a 500‐day lab‐scale trial, this study reports a pilot scale evaluation of this novel process treating 10 m³/day of 6‐mm screened saline sewage in Hong Kong. The SANI® pilot plant consisted of a sulfate reduction up‐flow sludge bed (SRUSB) reactor, an anoxic bioreactor for autotrophic denitrification and an aerobic bioreactor for nitrification. The plant was operated at a steady state for 225 days, during which the average removal efficiencies of both chemical oxygen demand (COD) and total suspended solids (TSS) at 87% and no excess sludge was purposefully withdrawn. Furthermore, a tracer test revealed 5% short circuit flow and a 34.6% dead zone in the SRUSB, indicating a good possibility to further optimize the treatment capacity of the process for full‐scale application. Compared with conventional biological nitrogen removal processes, the SANI® process reduces 90% of waste sludge, which saves 35% of the energy and reduces 36% of fossil CO₂ emission. The SANI® process not only eliminates the major odor sources originating from primary treatment and subsequent sludge treatment and disposal during secondary saline sewage treatment, but also promotes saline water supply as an economic and sustainable solution for water scarcity and sewage treatment in water‐scarce coastal areas. Biotechnol. Bioeng. 2012; 109: 2778–2789. © 2012 Wiley Periodicals, Inc.     

Detection and molecular identification of human adenoviruses and enteroviruses in wastewater from Morocco

Aims: Reclaimed wastewater is a considerable water resource in Morocco. Its agricultural reuse requires an assessment of viral contamination. The aim of this study was to detect both infectious and noninfectious human adenoviruses (HAdV) and enteroviruses (EV) in raw wastewater and treat effluent from wastewater treatment plants (WWTPs) and domestic sewage in Morocco. Methods and Results: A total of 22 samples were analysed. A polyethylene glycol (PEG) precipitation method was used, followed by integrated cell culture‐PCR (ICC‐PCR) using two cell lines: human rhabdomyosarcoma tumour tissue and laryngeal carcinoma cells (RD and Hep2 cells). Furthermore, viral genome amplification was confirmed by sequencing. HAdV were detected in 10 (45·5%) of the 22 samples involving two species: HAdV‐B and HAdV‐D. EV was detected in 5 (23%) samples belonging to Coxsackievirus B5 and poliovirus vaccine strain (Sabin 2). Conclusions: Human adenoviruses and EV were detected in the analysed samples from two WWTPs and HAdV in domestic sewage. Significance and Impact of the Study: This work is the first study in Morocco using cell culture, PCR and sequencing of enteric viruses from wastewater. The presence of infectious HAdV and EV in treated effluent emphasizes the need of wastewater treatment surveillance.     

Benthic resources are the key to Daphnia middendorffiana survival in a high arctic pond

1. Shallow arctic lakes and ponds have simple and short food webs, but large uncertainties remain about benthic–pelagic links in these systems. We tested whether organic matter of benthic origin supports zooplankton biomass in a pond in NE Greenland, using stable isotope analysis of carbon and nitrogen in the pond itself and in a ¹³C‐enrichment enclosure experiment. In the latter, we manipulated the carbon isotope signature of benthic algae to enhance its isotopic discrimination from other potential food sources for zooplankton. 2. The cladoceran Daphnia middendorffiana responded to the ¹³C‐enrichment of benthic mats with progressively increasing δ¹³C values, suggesting benthic feeding. Stable isotope analysis also pointed towards a negligible contribution of terrestrial carbon to the diet of D. middendorffiana. This agreed with the apparent dominance of autochthonous dissolved organic matter in the pond revealed by analysis of coloured dissolved organic matter. 3. Daily net production by phytoplankton in the pond (18 mg C m^?2 day^?1) could satisfy only up to half of the calculated minimum energy requirements of D. middendorffiana (35 mg C m^?2 day^?1), whereas benthic primary production alone (145 mg C m^?2 day^?1) was more than sufficient. 4. Our findings highlight benthic primary production as a major dietary source for D. middendorffiana in this system and suggest that benthic organic matter may play a key role in sustaining pelagic secondary production in such nutrient‐limited high arctic ponds.     

Establishment of reed canary grass with perennial legumes or barley and different fertilization treatments: effects on yield,botanical composition and nitrogen fixation

In two field experiments in northern Sweden, we investigated if intercropping reed canary grass (RCG; Phalaris arundinacea L.) with nitrogen‐fixing perennial legumes could reduce N‐fertilizer requirements and also if RCG ash or sewage sludge could be used as a supplement for mineral P and K. We compared biomass production, N uptake and N‐fixation of RCG in monoculture and mixtures of RCG with alsike clover (Trifolium hybridum L.), red clover (Trifolium pratense L.), goat's rue (Galega orientalis Lam.) and kura clover (Trifolium ambiguum M. Bieb.). In one experiment, RCG was also undersown in barley (Hordeum vulgare L.). Three fertilization treatments were applied: 100 kg N ha^?1, 50 kg N ha^?1 and 50 kg N ha^?1 + RCG ash/sewage sludge. We used a delayed harvest method: cutting the biomass in late autumn, leaving it on the field during the winter and harvesting in spring. The legume biomass of the mixtures at the inland experimental site was small and did not affect RCG growth negatively. At the coastal site, competition from higher amount of clover biomass affected RCG growth and spring yield negatively. N‐fixation in red clover and alsike clover mixtures in the first production year approximately covered half of recommended N‐fertilization rate. Goat's rue and kura clover did not establish well at the costal site, but at the inland site goat's rue formed a small but vital undergrowth. RCG undersown in barley gave lower yield, both in autumn and spring, than the other treatments. The high N treatment gave a higher spring yield at the inland site than the low N treatments, but there were no differences due to fertilization treatments at the coastal site. For spring harvest, there were no yield benefits of RCG/legume intercropping compared with RCG monoculture. However, intercropping might be more beneficial in a two‐harvest system.     

Grazing on toxic cyanobacterial blooms by tadpoles of edible frog Rana grylio

Tadpoles of Rana grylio were raised as edible frogs in fishponds of Guanqiao in Wuhan City, Hubei, China, during cyanobacterial blooms from June to October. The dominant cyanobacterial species was Microcystis, which was found to be lethally toxic by intraperitoneal (i.p.) mouse bioassay. Little is known about the effect of tadpoles on toxic cyanobacterial blooms. To evaluate the potential of the tadpoles to graze on cyanobacterial blooms, the tadpoles were fed on Microcystis collected from the field in the laboratory. The Microcystis cells decreased from 1.19 × 10⁷ cells mL^?1 to 3.23 × 10⁶ cells mL^?1, with a sharp reduction of 73% of the initial Microcystis population observed in the first 24 h after introduction of the tadpoles. The ponds containing tadpoles had a markedly lower density of Microcystis than those lacking tadpoles. Tadpoles exposed to either cultured Microcystis aeruginosa (NIES–90, 2.768 µg microcystins mg^–1 dw^–1) cells or lysed M. aeruginosa cells grew well, however, indicating that they were unaffected by Microcystis toxins. We found a significant increase in tadpole body weight after feeding on either field Microcystis or cultured M. aeruginosa. The mean increase in individual body weight was 20 mg day^?1 when fed on Microcystis from the pond, and 7 mg day^?1 when fed on M. aeruginosa from culture. Our study strongly suggested that there is a direct trophic relationship between R. grylio tadpoles and toxic Microcystis blooms and they possess the potential to graze on toxic Microcystis. The results imply that R. grylio tadpoles may play an important ecological role in reducing toxic cyanobacterial blooms caused by Microcystis.     

Effects of shoreline development on the nearshore environment in large deep oligotrophic lakes

1. In large deep oligotrophic lakes, the shallow nearshore waters may provide the most important habitat for animals to feed and breed, and it is this area of the lake where humans are most likely to have initial impacts as the shoreline is developed. Nutrients in fertilizers, sediments and sewage effluents are likely to be rapidly intercepted by nearshore algae at the lake edge, having heterogeneous effects nearshore before offshore effects are noted. 2. Here we examined the spatially explicit effects of residential development on nearshore periphyton communities in three large deep oligotrophic lakes that have all experienced modest residential development in the Pacific Northwest of the United States. We demonstrate that substantial nearshore changes in the basal food web are detectable even with low levels of shoreline development. These changes can potentially affect whole‐lake food web dynamics. 3. For our primary study site (Lake Crescent, Washington, USA), we found that algal biomass and accumulation of detritus were higher at developed sites. In addition, both macroinvertebrate and periphyton communities exhibited a shift in composition with more detritivores and filamentous green algae at developed sites. These differences were more pronounced during the spring than at other times of year. 4. A complementary investigation of field patterns in Priest Lake and Lake Pend Oreille (Idaho, USA) suggested that, although spatial and temporal patterns were idiosyncratic, indicators of productivity and the presence of filamentous green algae were generally higher at developed sites across lakes. 5. Stable isotope signatures and water column nutrients were not useful in distinguishing developed and undeveloped sites, increasing the potential usefulness of periphyton monitoring during early stages of lake development. 6. A laboratory investigation suggested that common macroinvertebrate grazers assimilated a much greater proportion of diatoms than the filamentous green algae that are associated with fertilization at developed sites. 7. These findings have at least two clear implications: (i) periphyton may be used to detect human impacts before disturbance is evident in offshore monitoring programmes and (ii) nearshore impacts in response to modest residential development have the potential to disrupt lake food web dynamics.     

The association of algicidal bacteria and raphidophyte blooms in South Carolina brackish detention ponds

Over the past 5 years, raphidophyte blooms have been frequently observed along the South Carolina coastal zone. During the 2002, 2003, and 2004 sampling seasons, we investigated temporal fluctuations of algicidal bacteria abundance against raphidophycean flagellates (Heterosigma akashiwo, Chattonella subsalsa, and Fibrocapsa japonica) using the microplate most probable number (MPN) method in three Kiawah Island brackish stormwater detention ponds (K1, K2, and K75). Local axenic isolates of H. akashiwo, C. subsalsa, and F. japonica were obtained and their susceptibility to algicidal bacteria tested. A total of 195 algicidal bacterial strains were isolated from raphidophyte blooms in the study ponds, and 6 of them were identified at the genus level, and the taxonomic specificity of their algicidal activity was tested against local (pond) and nonlocal isolates of raphidophytes (3 species, 10 total strains). In the ponds, a consistent association was found between raphidophyte bloom development and an increase in bacteria algicidal to the bloom species. In 12 of 15 cases, bloom decline followed the increase in algicidal bacteria to maximum abundances. Although variability was found in the taxonomic specificity of the algicidal bacteria effect (i.e. the number of raphidophyte species affected by a particular bacteria strain) and raphidophyte susceptibility (i.e. the number bacteria strains affecting a particular raphidophyte species), a toxic effect was always found when strains of a raphidophyte species were exposed to algicidal bacteria isolated from a bloom caused by that same species. The results suggest that algicidal bacteria may be an important limiting factor in raphidophyte bloom sustenance and can promote bloom decline in brackish lagoonal eutrophic estuaries.     

Growth kinetics of 1–2 mm and 3–4 mm colonies of Nostoc sphaeroides (Cyanophyta) in outdoor culture

Nostoc sphaeroides Kützing was cultivated in paddlewheel-driven raceway ponds and the growth kinetics of 1-2 mm and 3-4 mm colonies of N. sphaeroides was studied. The biomass productivities in 2.5 m(2) raceway ponds inoculated with 1-2 mm and 3-4 mm colonies were 5.2 and 0.25 g dry wt m(-2) d(-1), respectively. Furthermore, differently sized colonies showed different relative water content, total soluble carbohydrates, chlorophyll a content and density of filaments. This is the first report on mass culture of N. sphaeroides under outdoor conditions.     

Isolation of bacteria producing polyhydroxyalkanoates (PHA) from municipal sewage sludge

Bacterial isolates from sludge samples collected at a local municipal sewage treatment plant were screened for bacteria producing polyhydroxyalkanoates (PHA). Initially Sudan black B staining was performed to detect lipid cellular inclusions. Lipid-positive isolates were then grown in a nitrogen limitation E2 medium containing 2% (w/v) glucose to promote accumulation of PHA before the subsequent staining with Nile blue A. The positive isolates were quantified initially with a u.v. spectrophotometer, for a very large number of isolates (105) and among them high PHA-producing isolates (15) were selected and were confirmed by gas chromatographic analysis. The GC analysis showed the polymers produced by 13 of the selected isolates to be polyhydroxybutyrate (PHB), and the remaining two isolates produced polyhydroxybutyrate-co-hydroxyvalerate (PHB-co-HV) copolymer. The proportion of the PHA-positive bacterial isolates showed variability in the number of PHA accumulators during various months. The correlation of PHB production with the cell dry weight (CDW) was found to be statistically significant. The metabolism of PHB in these selected 15 isolates was studied using the Nile blue A staining, which showed an initial increase in the fluorescence followed by a decline, on further incubation. All the selected 15 isolates were classified to genus level by studying their morphological and biochemical characteristics. There were seven Bacillus species, three Pseudomonas species, two Alcaligenes species, two Aeromonas species, and one Chromobacterium species.     

Effects of Sewage Sludge on Plant Community Composition in Restored Limestone Quarries

The effects of sewage sludge, used to improve fertility of replaced soil, on vegetation were studied in limestone quarry restoration. Plant community growing in the first stages after sludge application was surveyed in six quarries of NE Spain. Areas with a mixture of sewage sludge and residual soil were compared to areas where the application consisted only of residual soil (a mixture of previous top soil and mine spoils). Sewage sludge was hypothesized to increase total biomass and cover, modifying species composition and delaying the early successional recover of the community. The results showed that both biomass and plant cover increased because of sewage sludge addition. The floristic composition was dominated by ruderal species that did not show any dependence on sewage sludge application. Convergence on similarity between sludge and control plots was not detected along a 5‐year period. Although species richness was significantly lower in sludge plots, diversity and equitability indexes did not show differences between treatments. The results did not show differences in the proportion of non‐native species. The proportion of legumes was lower in sludge plots. These results show that the plant communities resulting from the addition of sewage sludge to the soil used in limestone quarry restoration have more biomass and cover, but less number of species, and they do not show a clear trend to converge to those areas restored only with non‐amended soil.