A comparison of fluorescein dye and antibiotic-resistantEscherichia coli as indicators of pollution in groundwater

Field experiments were conducted to evaluate the performance of antibiotic-resistanceEscherichia coli and yellow fluorescein dye as tracers of subsurface water flow. These materials were released into three horizontal lines installed in the A, B, and C horizons of a western Oregon hillslope soil. Movement was evaluated by collecting groundwater samples from rows of modified piezometers (6 piezometers/row) placed at various depths and distances downslope from the injection lines. TheE. coli cells were transported rapidly through the experimental site and were recovered from piezometers 15 m dowslope only 1 h after inoculation while the rate by which the bacteria were moved was at least 1500 cm h^?1 in the B-horizon. The strains ofE. coli survived in large numbers for the duration of the 12 h sampling periods and appeared to be satisfactory as tracers of subsurface water flow. In contrast, the fluorescein dye was never visually detectable in any of the piezometers during any of the sampling periods although very small, residual amounts could be fluorometrically measured. The widespread use of dyes as water tracers and the potential for incorporating antibiotic-resistant fecal bacteria to this effect is discussed.     

Thirteen-year survival study of an environmentalEscherichia coli in field mini-plots

A multiple-antibiotic resistantE. coli was applied to rye-grass covered field mini-plots to simulate point-source contamination. Using three mini-plots for testing and a fourth as a control, the ability of the tracer bacterium to survive under field conditions was studied. Three test plots each received separately 10⁷, 10⁸, or 10¹⁰ cfu mL^?1 E. coli grown for 24 h. in 5 L one-third strength Tryptic soy broth. In Phase I of the study, it was determined that the tracer disappeared from leaf surfaces of rye-grass covering the plots after 41 days. In Phase II, determination of the presence of the tracer in the top 2″ (5 cm) of soil after two months elapsed time indicated that tracer cfu/g dry wt. of soil had declined five, three, and three-logs for test plots 1,2, and 3. In Phase III, subsurface soil sampling using a soil auger on the three test minei-plots indicated the tracer had penetrated through the top-soil and into the underlying B horizon (20 to 50 cm down). In Phase IV, detailed sampling by excavation of the subsurface soil Horizons of the third test mini-plot showed that the tracer had also penetrated through the hardpan (C Horizon) located 0.6 m below the surface to enter the groundwater (1.06 m deep) (Phase V).E. coli counts fell precipitously to 10³ cfu g^?1 in soil and then, in the groundwater at the groundwater-soil interface, persisted at a concentration of 10³ cfu 100 mL^?1 for 2 yr. As time past, tracer counts fell to 145 cfu/100 mL in 6 yr. rose to 820 cfu 100 mL^?1 in 1986 (8 yr elapsed time), and then fell to 25 cfu 100 mL^?1 in 1991 after 13 yr. Serotyping of 1986E. coli isolates indicated that 62% were of the original tracer serotype (0.128:B12) while only 43% of the 1991 isolates were of the same serotype. The penetration rate of the tracer down through the mini-plot soil into the groundwater was 0.02 m day^?1 while downslope dispersion occurred at an estimated rate of 1.0 m day^?1. The implications of the above findings are discussed.     

A quantitative evaluation and phylogenetic characterization of oligotrophic denitrifying bacteria harbored in subsurface upland soil using improved culturability

This is the first investigation to show that oligotrophic denitrifying bacteria are dominant denitrifiers in subsurface upland soil. We examined the vertical distribution of denitrifying bacterial populations in upland soil using two kinds of enumeration media. The number of denitrifying bacteria, enumerated in subsurface soil layers by a 100-fold diluted nutrient broth (DNB) medium with NO₃ ⁻, was two to three orders of magnitude greater than those enumerated by a conventional nutrient broth medium with NO₃ ⁻, suggesting the dominance of oligotrophic denitrifying bacteria. Seventy-four percent of the total denitrifying bacterial isolates were DNB organisms of the oligotrophic type, which did not show appreciable growth on a conventional nutrient broth medium. The isolates were heterogeneous and were categorized as alpha (35 strains) and beta (19 strains) subdivisions of proteobacteria and high G+C gram-positive bacteria (7 strains) by 16S rRNA gene sequence analysis. The 35-alpha subdivision of proteobacterial isolates was of oligotrophic type and widely distributed from the surface to subsurface soil layers. Phylogenetic analysis indicated that some isolates belonged to groups with few or no cultivated representatives, and that one isolate may be a member of a new genus. This isolation procedure, using diluted media, is valuable in detecting diverse and novel denitrifying bacteria in the subsurface soil.     

Soil water chemistry dependence on water pathways and turnover

Water flow and transit in the soil are important to water supply and transport of chemical compounds. Flows through forested till soils are by vertical percolation in the upslope areas, groundwater flows along the slope to discharge in downslope sites. The flow is divided into slow and fast parts both in the unsaturated percolation and the saturated groundwater. Soil development influences the flow and in coarse grained and well drained soils groundwater flow is stable but in finer grained physically stratified soils, variations in flow are considerable. Large flows occur in the upper soil layers and small flows in deep layers. Chemical composition of soil water varies in a similar way. In the upper layers, water is more acid and has a lower content of base cations compared to deeper layers. In downslope areas discharging deep groundwater contributes to less acidic conditions also in the upper soil layers. This discharge and a dominating lateral water flow in the upper layers partly protect deep layers from acidification which is emphasised by slow water turnover in these layers.     

Extraction of bacterial lipopolysaccharides from soil samples and their determination using the Limulus amebocyte lysate reaction

Lipopolysaccharides were extracted from soil using an aqueous phenol procedure and determined by using the Limulus amebocyte lysate coagulation reaction. When heat-killed Escherichia coli cells were added to soil at a concentration of organisms expected to occur in subsurface material, there was a 10% recovery of the added cellular lipopolysaccharides (LPS); because of the sensitivity of the Limulus amebocyte lysate (LAL) reaction this recovery is sufficient for the method to be a rapid (4 h) estimate of the Gram-negative bacteria present in soil samples. By using a kinetic assay LPS could be measured over a concentration range of five orders of magnitude, which is important with environmental samples.     

Vertical transport of a field-released genetically engineered microorganism through soil

The vertical movement of a genetically-engineered bacterium released into a sub-surface soil contained within a lysimeter was investigated. The strain used was Pseudomonas fluorescens HK44, a lux-based microbe capable of bioremediating specific polyaromatic hydrocarbons (PAHs). Strain HK44 was inoculated into a 4 m deep by 2.5 m diameter soil lysimeter outfitted with a hydraulic assembly to control groundwater levels. After 230 days, the lysimeter was subjected to a water table manipulation that resulted in significant transport of HK44 cells to depths 60 cm below the original inoculation zone within 20 days. A lysimeter not exposed to water level fluctuations exhibited no significant vertical transport. These results demonstrate the potential distribution of genetically engineered microorganisms in subsurface soil due to groundwater effects.     

Faecal Indicator Bacteria: Groundwater Dynamics and Transport Following Precipitation and River Water Infiltration

Faecal contamination of drinking water extracted from alluvial aquifers can lead to severe problems. River water infiltration can be a hazard for extraction wells located nearby, especially during high discharge events. The high dimensionality of river?Cgroundwater interaction and the many factors affecting bacterial survival and transport in groundwater make a simple assessment of actual water quality difficult. The identification of proxy indicators for river water infiltration and bacterial contamination is an important step in managing groundwater resources and hazard assessment. The time resolution of microbial monitoring studies is often too low to establish this relationship. A proxy-based approach in such highly dynamic systems requires in-depth knowledge of the relationship between the variable of interest, e.g. river water infiltration, and its proxy indicator. In this study, continuously recorded physico-chemical parameters (temperature, electrical conductivity, turbidity, spectral absorption coefficient, particle density) were compared to the counts for faecal indicator bacteria, Escherichia coli and Enterococcus sp. obtained from intermittent sampling. Sampling for faecal indicator bacteria was conducted on two temporal scales: (a) routine bi-weekly monitoring over a month and (b) intense (bi-hourly) event-based sampling over 3 days triggered by a high discharge event. Both sampling set-ups showed that the highest bacterial concentrations occurred in the river. E. coli and Enterococcus sp. concentrations decreased with time and length of flow path in the aquifer. The event-based sampling was able to demonstrate differences in bacterial removal between clusters of observation wells linked to aquifer composition. Although no individual proxy indicator for bacterial contamination could be established, it was shown that a combined approach based on time-series of physico-chemical parameters could be used to assess river water infiltration as a hazard for drinking water quality management.     

外包土工布暗管排盐条件下水盐运移规律

为揭示外包土工布暗管埋设在非饱和带时淋洗后水分和盐分的运移规律,该文设计了模拟暗管排水的室内试验,研究2种土壤初始状态下(非饱和状态和田持状态),排水初期暗管与地下水位的相对位置及其排水排盐情况,从开始淋洗至暗管停止排水全过程中地下水埋深及含盐量变化规律、暗管的排水排盐效果及土壤剖面的水盐动态运移规律。结果表明:在暗管周围包裹土工布的情况下,土壤初始状态无论是非饱和还是田持,当暗管开始排水时地下水均已完全淹没暗管,此时的排盐量最大,流量呈先增大后减小的变化趋势,且地下水位先升高后降低,地下水含盐量随着淋洗水量的增加由累积转变为脱盐。对比淋洗非饱和土壤(试验1)和淋洗田持土壤(试验2)的试验结果,试验2中暗管的排水、排盐效果优于试验1,在试验1中淋洗非饱和土壤时,土壤脱盐率在垂直方向上随土壤深度的增加逐渐降低,0~20 cm土层的脱盐率(>85%)最大,降至无盐水平,暗管周围土壤脱盐率相对较小(<60%),仍处于中度盐渍化水平;水平方向上,0~20 cm土层的脱盐率差异不大,20~40 cm土层中距暗管越远其脱盐率越小。试验2在试验1基础上进行,淋洗田持土壤时,0~20 cm土层盐分不再变化,30~40 cm土层的脱盐率增大(>60%)。此外,试验1中淋洗脱盐效果大于暗管排盐效果,暗管主要排出暗管以上土壤盐分;试验2中暗管排盐效果增强,暗管不仅排出暗管周围土壤盐分,而且排出暗管以下土层及地下水中盐分,随着淋洗水量的增加,土壤由脱盐型转变为排盐型。研究结果表明外包土工布暗管的应用效果受地下水与暗管相对位置的影响,合理提高淋洗水量可以增强暗管排水排盐效果及土壤脱盐效果,有效改善土壤盐渍化。研究结果可为西北内陆干旱地区不同地下水埋深条件下暗管排盐技术的推广和应用提供理论支撑和科学指导。     

Sub-surface colluviation: An example from West Sussex, UK

Reported here is an unusual type of colluvial deposit whose associated transport pathway involves a substantial subsurface component. The deposit represents the downslope transfer of nitrogenenriched agricultural soil into a seminatural woodland soil, creating significant localised eutrophication. The amount of material involved was c. 65 kg, and covered an area of c. 7.7 m². It appears that the deposition was rapid, and the outcome of a single transport event. The absence of more general observations prevent the calculation of an aerally-representative long-term rate of operation. The status of this process in the general modification of temperate hillslopes is still a matter for conjecture.     

Influence of Desiccation on the Transport of <Emphasis Type="Italic">Escherichia coli</Emphasis> through Saturated Sand Packs

Bacterial cells that enter the groundwater system commonly experience desiccation stresses (i.e., bacterial cells are directly exposed to air) when traveling through the unsaturated layer of soil. Little is known about the effects of desiccation on the transport of bacterial cells in the groundwater system. In this research, we investigated the transport of desiccated and non-desiccated Escherichia coli K12 (ATCC 10798) cells through saturated sand packs using laboratory column transport experiments. Cell desiccation was performed at 25°C under relative humidity (RH) levels of 22%, 53%, 75%, and 97%, respectively, and the desiccation duration was 22 h. Our results showed that desiccation reduced the viability of E. coli cells under all RH levels and increased the transport of E. coli cells under ≥75% RH levels. The increase in the transport of the desiccated E. coli cells was not related to changes in cell size or cell zeta potential. Desiccation under high (i.e., ≥75%) RH levels, however, led to lower cell hydrophobicity, which was found to be positively correlated with cell transport.     

Vadose Zone Microbial Transport Below At-Grade Distribution of Wastewater Effluent

The attenuation of Escherichia coli and total coliform from secondary treated wastewater effluent under two “at-grade” effluent distribution systems was evaluated in a sandy silt vadose zone in a cold climate. The two at-grade distribution lines had different designs and hydraulic loading rates. Effluent transport was examined using chloride as a tracer. Coliform fate was evaluated relative to the chloride using a combination of in situ pore water sampling and destructive soil sampling, combined with the observation of a dye tracer along excavation sidewalls. Although bacteria attenuation in the subsoil appeared to decrease during colder, winter temperatures (likely due to decreased viability and decreased predation), the subsoil provided about a four log reduction in E. coli over 90 cm of vertical transport. Horizontal transport of bacteria (up to 1.5 m from the line) was likely aided by flow on top of a microbial biomat observed at the soil surface. Both the subsurface dye patterns and the E. coli sampling suggested less preferential flow occurred below the lower loading rate design. At-grade distribution of secondary treated wastewater appears to be a viable alternative to conventional distribution fields at sites with similar climate and soils.     

Distribution with depth of protozoa,bacteria and fungi in soil profiles from three Danish forest sites

The numbers and biomass of protozoa, bacteria and fungi were measured at various depths (1.5–122.5 cm) in the unsaturated zone of three contrasting pristine Danish forest site profiles: a dry beech (Fagus silvatica) forest on mor, a wet peaty spruce (Picea abies)/birch (Betula pubescens) forest and a dry spruce (P. abies) forest on mor. All sites were situated on a Weichel moraine. Except for a bacterial peak at 42.5 cm in the peat profile, the general tendency was a decrease in biomass with increasing depth for all groups examined. Protozoa decreased more rapidly with increasing depth than the other two groups of organisms examined. An evaluation of the bacterial–protozoan relationship by a simple mathematical model indicated that the subsurface protozoan populations are active and not accidental percolated cysts. The low protozoan numbers found in shallow subsurface sites contrast markedly with the results from contaminated sites where much larger protozoan populations have been reported even at considerable depths. Consequently, the results suggest that protozoa are good indicators of organic pollution in subsurface soils; however, more work involving the comparison of polluted and unpolluted soils is needed to confirm this suggestion.     

Survival of Escherichia coli in soil with modified microbial community composition

Understanding the survival and persistence of Escherichia coli in soil with different microbial composition is essential for the accuracy of water quality assessment and microbial source tracking. This microcosm experiment was conducted to investigate the survival pattern of three E. coli strains (originated from soil, dog feces and human feces, separately) in soil with modified microbial community composition. Bile salt No. 3 (BS3) of progressively increased density (0.05%, 0.15%, 0.30% and 0.50%) was added into sandy loam soils and incubated for 90 days. Laboratory cultured E. coli were then inoculated into soil and incubated for another 150 days to monitor their survival pattern. Change of bacterial community diversity by BS3 was detected by both cultivation based and cultivation independent (PCR-Denaturing Gradient Gel Electrophoresis) methods. In general, progressively increased BS3 concentration resulted in decreased CFU counts both at 10 days and 90 days incubation. DGGE analysis indicated only a slight change in bacterial community composition at 10 days but a significant change at 90 days. Cluster analysis suggested that BS3 treatment grouped separately from controls. Survival of E. coli in soil was significantly influenced by the complexity of the microbial community, as die-off rate of E. coli progressively declined with the reduction of microbial community diversity. Differential survival of E. coli under different soil microbial stress highlights the importance of incorporating biotic factors in predictive models for water quality management and microbial source tracking study.     

Discharge of Escherichia Coli from Agricultural Surface and Subsurface Drainage Water: Tillage Effects

Drainage water from agricultural fields with applied manure can degrade the bacterial quality of surface and groundwater. The impact of conventional tillage (CT) and zero tillage (ZT) practices on Escherichia coli (E.coli) discharge through artificially drained soils is not well understood. Consequently, two field trials were conducted during 2002–2004. The first trial involved fall applications of beef manure while the second involved spring applications of dairy manure. Both surface and subsurface drainage water were monitored in the first trial while only subsurface drainage water was monitored in the second. Under fall applied beef manure (trial 1), no differences (p?>?0.05) were observed in E.coli concentrations (cfu/100 ml) in combined drainage water under both tillage systems. However, during 2003–2004, subsurface drainage water under ZT had higher E.coli concentrations and loads than drainage water under CT. When the combined (surface + subsurface) annual E.coli loads were considered, CT loads were greater than ZT during 2002–2003 with an opposite situation during 2003–2004. Overall, annual E.coli loads were similar under ZT (4.7?×?10¹⁰ cfu/ha) and CT (4.8?×?10¹⁰ cfu/ha). Spring dairy manure application (trial 2) produced significant (p?>?0.03) tillage effect on E.coli loads in subsurface drainage water only during the second year. During the study period, ZT plots (1.55?×?10¹⁰ cfu/ha) discharged 5× more E.coli than CT (0.23?×?10¹⁰ cfu/ha). A longer duration of ZT practices resulted in higher subsurface flow volumes and subsequently greater loads of E.coli discharge in both trials.     

Finger-Printing Biodegradation of Petroleum Contamination in Shallow Groundwater and Soil System Using Hydro-bio-geochemical Markers and Modelling Support

This study was conducted to determine the potential of in situ biodegradation and identify the geochemical and microbial processes of the petroleum-contaminated subsurface environment using integrated hydro-bio-geochemical markers so that the risk of contamination to subsurface environment can be better understood. The contamination process and corresponding bio-geo-chemistry were analysed in parallel with geochemical and multi-variant statistical modelling at a petroleum-contaminated site in the northeast China. The total petroleum hydrocarbon analysed in the monitoring wells and soil profile demonstrated heavy contamination with potential risk to human health and eco-environment. Further detailed analysis of petroleum fractions revealed a clear spatial variation of organic compositions in groundwater. It was evident that biodegradation and preferential biodegradability contributed considerably to the fraction distribution pattern, which can also be implicated by carbon and microbial respiration in the subsurface environment. The steady decrease in SO ₄ ^2- concentration, detection of S^2-, and increase in pH and alkalinity (HCO ₃ ^- ) in groundwater during the monitoring period demonstrated that sulphate reduction was the dominant biodegradation process in most contaminated zones. The results of statistical analysis further suggested that the hydro-geochemical environment was mainly controlled by the regional hydro-geochemical and sulphate reduction process associated closely with the total petroleum hydrocarbon. Knowledge from the comprehensive study provides useful insight on fate, transport and risk assessment of the petroleum contaminants in the shallow subsurface environment.     

Rhizobia nodulating African Acacia spp. and Sesbania sesban trees in southern Ethiopian soils are metabolically and genomically diverse

The diversity of 110 rhizobial strains isolated from Acacia abyssinica, A. seyal, A. tortilis, Faidherbia albida, Sesbania sesban, Phaseolus vulgaris, and Vigna unguiculata grown in soils across diverse agro-ecological zones in southern Ethiopia was assessed using the Biolog™ system and amplified fragment length polymorphism (AFLP) fingerprinting technique. By cluster analysis of the metabolic and genomic fingerprints, the test strains were grouped into 13 Biolog and 11 AFLP clusters. Twenty-two strains in the Biolog method and 15 strains in the AFLP analysis were linked to eight and four reference species, respectively, out of the 28 included in the study. Most of the test strains (more than 80% of 110) were not related to any of the reference species by both methods. Forty-six test strains (42% of 110) were grouped into seven corresponding Biolog and AFLP clusters, suggesting that these groups represented the same strains, or in some cases clonal descendants of the same organisms. In contrast to the strains from S. sesban, isolates from Acacia spp. were represented in several Biolog and AFLP clusters indicating the promiscuous nature of the latter and widespread occurrence of compatible rhizobia in most of the soil sampling locations. The results showed that indigenous rhizobia nodulating native woody species in Ethiopian soils constituted metabolically and genomically diverse groups that are not linked to reference species.     

重庆中梁山岩溶区耕作侵蚀影响因素研究

在重庆市中梁山岩溶区用示踪法对坡耕地进行耕作试验研究。结果表明:锄头和铁锹耕作引起的土壤位移距离、位移量与坡度呈正相关,而踩锹引起的土壤位移距离、位移量则与含水量具有一定的相关性。在顺坡耕作时铁锹引起的土壤位移距离、位移量与锄头相近,但它们远远大于踩锹;等高耕作时铁锹引起的土壤位移距离、位移量是锄头的1.3~3.8倍左右。铁锹顺坡耕作引起的土壤位移距离、位移量是它等高耕作的2~2.4倍;锄头顺坡耕作引起的土壤位移距离、位移量是它等高的3~8.8倍。引起土壤位移量的大小顺序依次是:锄头顺坡>铁锹顺坡>铁锹等高>锄头等高>踩锹顺坡。     

Studies on soil fumigation—II: Effects on bacteria

In a South Australian wheat-field soil the viable counts of “total” aerobic bacteria and of fluorescent pseudomonads were initially greatly depressed by fumigation with 220 kg·ha^?1 chloropicrin (CP) or with a combined application of 220 kg·ha^?1 of chloropicrin and 220 kg·ha^?1 methyl bromide (F). Bacterial numbers rose sharply within 10 days of the completion of fumigation. For a further 14 days the fluorescent pseudomonads formed the major part of the aerobic bacterial population counted and over 5 months later their numbers in F-treated soil remained about 10 times higher than in untreated soil. Numbers of aerobic spore-formers rose more slowly after CP or F treatment. but then remained significantly higher over the 159 days of the trial. Fumigation with 220 kg·ha^?1 of methyl bromide alone (MB) had little effect on soil bacterial numbers.A check of random isolates revealed a predominance of Gram-negative organisms in soil treated with CP, this dominance decreasing with time, whereas MB treatment did not result in any detectable change.Fluorescent pseudomonads from rhizospheres of wheat plants in soil fumigated with CP contained smaller proportions of strains antagonistic in vitro to Gaeumannomyces graminis var. tritici than isolates from MB-treated soil or from untreated soil.     

Evidence for the Accumulation and Steady-State Persistence of E. coli in Subtropical Drainage Basin Sediments

While the presence of fecal indicator bacteria such as Escherichia coli in urban stormwater has been widely documented, their occurrence and persistence in sediments are not as well understood. Recent investigations suggest that E. coli can accumulate in drainage basin sediments and act as a fecal bacterial reservoir within a watershed. We investigate the prevalence of E. coli populations in a tidal creek stormwater catchment and examine their interaction with overlying stormwater under wet and dry weather conditions. Two rain events are sampled more intensively with samples collected prior to, during, and after rainfall to profile bacteria in each matrix throughout a storm. Results of profile sampling and estimates of sediment resuspension provide evidence for E. coli accumulation during dry conditions and entrainment in overlying waters during storm conditions. Profile results suggest the occurrence of steady-state E. coli populations in drainage basin sediments.     

新疆玛纳斯河流域不同地貌类型土壤盐分累积变化

为详细分析干旱荒漠区土壤盐分长期累积变化的区域差异，以玛纳斯河流域为例，选择冲积洪积扇、冲积平原和干三角洲3种主要地貌类型，通过取样、典型调查等方法讨论其土壤盐分变化情况，结果表明：位于山前冲洪积扇缘以上部位是最稳定的地区，地下水位低，基本不存在盐碱化问题。相比之下，冲积洪积扇缘部分地区由于普遍较高的地下水位和地下水矿化度，短期内仍将处于持续积盐状态。冲积平原部位表土虽明显脱盐，但由于底土含盐量高，且部分地段地下潜水埋深上升至临界深度以下，仍处于脱盐不稳定或脱盐积盐反复型状态。干三角洲部位经过多年的开垦，地下水位出现上升，随着深层盐分上移，也将导致土壤盐渍化发生。分析认为冲洪积扇缘、冲积平原和干三角洲部位部分地区将来土壤盐渍化威胁依然很大。该研究将为区域土壤盐渍化治理提供依据。