RAPD marker and substrate utilization pattern applied to study microbial community diversity in the soil affected by agricultural chemicals

Present analyses of random amplified polymorphic DNA (RAPD) and Biolog GN substrate utilization pattern are combined to further study the diversity of microbial communities in four soils affected by agricultural chemicals. The results showed that the four soil microbial communities were apparently distinguishable in the diversity at RAPD level in terms of the richness and modified richness in the summer, which supports our previous report using the same soils in winter. A significant difference for the average well color development (AWCD) at 72 h incubation was found among the soils in winter using Biolog GN substrate utilization pattern, but this difference was not found among the soils in summer. However, Shannon-Weaver indices for microbial communities in the summer soils polluted by agricultural chemicals were significantly higher than those in winter at metabolic level; in contrast, no significant difference existed between the two seasons for microbial communities in the soil without chemical pollution. Present results suggest that the combined approach using RAPD and substrate utilization pattern could be used to effectively quantify microbial community diversity and its changes among the seasons in the soils affected by agricultural chemicals, simultaneously at molecular and physiological levels.     

Functional biodiversity of microbial communities in aerobic selector slaughterhouse wastewater.

The biodiversity of the microbial communities of an aerobic selector and a conventional system treating slaughterhouse wastewater were evaluated using the Biolog technique (Biolog Inc., Hayward, California). Principal components analysis of patterns and level of microbial activity indicate that microbial communities statistically differentiate between the selector and conventional system. Biolog data indicated that there was seasonal variation in the structure and function of the microbial community in conventional samples, which may be a useful indicator of wastewater community disturbance and unsteadiness, while, for the selector system, there were no recognizable statistical differences between winter and summer data communities, Biodiversity indices indicated that the selector system maintained a high functional diversity (Shannon-Weaver diversity index [H'] from 3.8 to 4.6) than the conventional system (H' from 1.8 to 2.8). The high values in the selector system were a reflection of the fact that most of the carbon sources were used, contributing to the very high Shannon indices. In addition, the high substrate equitability values (J) obtained for the selector samples indicated that the microbial communities between the summer and winter samples used the carbon sources in the same proportion. In contrast, differences in the equitability of the microbial communities within the conventional system were observed. This indicated a pattern representing unevenness.     

Spatial variability in the degradation rate of isoproturon in soil

Thirty samples of soil were taken at 50-m intersections on a grid pattern over an area of 250 x 200 m within a single field with nominally uniform soil characteristics. Incubations of isoproturon (3-(4-isopropylphenyl)-1,1-dimethylurea) under standard conditions (15 degrees C; -33 kPa soil water potential) indicated considerable variation in degradation rate of the herbicide, with the time to 50% loss (DT50) varying from 6.5 to 30 days. The kinetics of degradation also varied between the sub-samples of soil. In many of them, there was an exponential decline in isoproturon residues; in others, exponential loss was followed by more rapid rates of decline; in a few soil samples, rapid rates of loss began shortly after the start of the incubations. In more detailed studies with soils from a smaller number of sub-sites (20), measurements were again made of isoproturon degradation rate, and the soils were analysed for organic matter content, pH, and nutrient status (N, P, K). Measurements were also made of isoproturon adsorption by the soils and of soil microbial biomass. Patterns of microbial metabolism were assessed using 95 substrates in Biolog GN plates. Soils showing rapid biodegradation were generally of higher pH and contained more available potassium than those showing slower degradation rates. They also had a larger microbial biomass and greater microbial metabolic diversity as determined by substrate utilisation on Biolog GN plates. The implications of the results for the efficacy and environmental behaviour of isoproturon are discussed.     

Effects of lead and cadmium nitrate on biomass and substrate utilization pattern of soil microbial communities

A study was conducted to evaluate the effects of different concentrations of lead (Pb) and cadmium (Cd) applied as their nitrates on soil microbial biomass carbon (C(mic)) and nitrogen (N(mic)), and substrate utilization pattern of soil microbial communities. The C(mic) and N(mic) contents were determined at 0, 14, 28, 42 and 56 days after heavy metal application (DAA). The results showed a significant decline in the C(mic) for all Pb and Cd amended soils from the start to 28 DAA. From 28 to 56 DAA, C(mic) contents changed non-significantly for all other treatments except for 600 mgkg(-1) Pb and 100 mgkg(-1) Cd in which it declined significantly from 42 to 56 DAA. The N(mic) contents also decreased significantly from start to 28 DAA for all other Pb and Cd treatments except for 200 mgkg(-1) Pb which did not show significant difference from the control. Control and 200 mgkg(-1) Pb had significantly lower soil microbial biomass C:N ratio as compared with other Pb treatments from 14 to 42 DAA, however at 56 DAA, only 1000 mgkg(-1) Pb showed significantly higher C:N ratio compared with other treatments. No significant difference in C:N ratio for all Cd treated soils was seen from start to 28 DAA, however from 42 to 56 DAA, 100 mgkg(-1) Pb showed significantly higher C:N ratio compared with other treatments. On 56 DAA, substrate utilization pattern of soil microbial communities was determined by inoculating Biolog ECO plates. The results indicated that Pb and Cd addition inhibited the functional activity of soil microbial communities as indicated by the intensity of average well color development (AWCD) during 168 h of incubation. Multivariate analysis of sole carbon source utilization pattern demonstrated that higher levels of heavy metal application had significantly affected soil microbial community structure.     

Assessing microbial communities for a metabolic profile similar to activated sludge.

To search for reliable testing inocula alternatives to activated sludge cultures, several model microbial consortia were compared with activated sludge populations for their functional diversity. The evaluation of the metabolic potential of these mixed inocula was performed using the Biolog EcoPlates and GN and GP MicroPlates (Biolog, Inc., Hayward, California). The community-level physiological profiles (CLPPs) obtained for model communities and activated sludge samples were analyzed by principal component analysis and hierarchic clustering methods, to evaluate the ability of Biolog plates to distinguish among the different microbial communities. The effect of different inocula preparation methodologies on the community structure was also studied. The CLPPs obtained with EcoPlates and GN MicroPlates showed that EcoPlates are suitable to screen communities with a metabolic profile similar to activated sludge. New, well-defined, standardized, and safe inocula presenting the same metabolic community profile as activated sludge were selected and can be tested as surrogate cultures in activated-sludge-based bioassays.     

The veterinary antibiotic oxytetracycline and Cu influence functional diversity of the soil microbial community

There are increasing concerns over the effects of veterinary antibiotics and heavy metals in agricultural soils. The widely used veterinary antibiotic oxytetracycline (OTC), Cu and their combination on soil microbial community function were assessed with the Biolog method. The microbial community was extracted from the soil and exposed to a 0.85% sodium chloride solution containing OTC (0, 1, 5, 11, 43, 109 and 217 microM), or Cu (0, 10, 20, 100 and 300 microM), or combination of the two pollutants (OTC 0, 5, 11 microM and Cu 0, 20 microM). Functional diversity, evenness, average well color development (AWCD) and substrate utilization decreased significantly with increasing concentrations of OTC or Cu (p < 0.005). The critical concentrations were 11 microM for OTC and 20 microM for Cu. The combination of OTC and Cu significantly decreased Shannon's diversity, evenness and utilization of carbohydrates and carboxylic acids compared to individual one of the contaminants. The antibiotic OTC and Cu had significant negative effects on soil microbial community function, particularly when both pollutants were present.     

Microbial response to repeated treatments of manure containing sulfadiazine and chlortetracycline in soil

Substantive addition of antibiotic-contaminated manure to agricultural soil may lead to “persistent” residues of antibiotics and may affect soil health. Therefore, this study examines the effects of repeated manure treatments containing sulfadiazine (SDZ) and chlortetracycline (CTC) residues, both individually and combined, on the functional diversity and structure of soil microbial communities in the soils under laboratory conditions. The average well color development (AWCD), Simpson diversity index (1/D, dominant populations), Shannon-Wiener diversity index (H′, richness), and McIntosh diversity index (U, evenness) in the antibiotics-treated soils decreased in the first 60-day treatment and then gradually recovered or even exceeded the initial level in the unamended soils with increasing treatment frequency. A total of 11 specific bands in temperature gradient gel electrophoresis (TGGE) profiles were observed and sequence analyzed for five repeated treatments, and most of them belonged to the phyla Firmicutes, Actinobacteria, and Proteobacteria. These results indicate that repeated treatments of manure containing SDZ and CTC residues can alter soil microbial community structure, although they have a temporary suppression effect on soil microbial functional diversity.     

光合细菌改善Cd、Pb及呋喃丹污染土壤的微生物群落DNA序列多样性的研究

应用DNA随机扩增多态性(RAPD)分子标记技术研究了光合细菌(PSB)对Cd、Pb及呋喃丹污染土壤的微生物群落DNA序列多样性的影响.结果表明,Cd、Pb及呋喃丹单一污染或3者复合污染土壤的微生物群落DNA序列的丰富度相对对照土样(S0)都有不同程度的增加,受Cd、Pb或呋哺丹污染,可能会引起土壤微生物群落DNA序列...     

Influence of ciprofloxacin on the microbial catabolic diversity in soil

In this study, the effect of ciprofloxacin (CIP) on the catabolic diversity of soil microbial communities was evaluated. Soil samples were spiked with ciprofloxacin (0, 1, 5 and 50 mg?kg^?1) and were incubated for 1, 3, 9, 22 and 40 days. Untreated controls received only water. The functional diversity of the microbial community studied was characterized using a catabolic response profile (CRP). Six substrate groups were tested: carbohydrates, amino acids, carboxylic acids, aromatic chemicals, alcohols and polymers. After 40 days, the CIP concentrations in the soil samples ranged from 25% to 58% of the initial concentrations. Soil respiratory responses to the individual substrates D-glucose, lactose, D-mannose, L-glutamic, Na-citrate, malic acid and inosine were inhibited at the high CIP concentrations (5 and 50 mg·kg^?1) in the soils and were increased at the lowest CIP concentration (1 mg·kg^?1). Soil respiration was inhibited at all of the CIP concentrations after the addition of D-galactose and glycerol. The CIP concentration and incubation time explained 45.3% of the variance of the catabolic responses. The CRP analysis clearly discriminated among the different CIP concentrations. The results suggest that CIP strongly affects the catabolic diversities of soil microbial communities and that its effect is more significant than that of incubation time.     

Effect of green manure amendment on herbicide pendimethalin on soil

Manure amendment in agricultural practice can have a large effect on herbicide dissipation because the period of manure plowing is close to the period of herbicide application. In addition, manure amendment is among the frequently encountered options in ameliorating pesticide pollution. In this research, the dissipation of the herbicide pendimethalin was examined after amendment with two common green manures, Lupinus luteus (L) or Cosmos bipinnatus (C), for 110 days in pH 5.2 and 7.7 soils (Sankengtzu [Sk] and Erhlin [Eh] soil, respectively). The microbial activity and ecology changes were examined by using Biolog EcoPlate and denaturing gradient gel electrophoresis (DGGE). In Sk soil, the half-lives of pendimethalin with L, C, and blank treatment were 49.0, 54.9, and 62.2 days, respectively, whereas that in Eh soil they were 46.3, 52.6, and 34.8 days, respectively. Pendimethalin dissipated quickly in more neutral soil (Eh soil), but the addition of manure can only increase the dissipation rate in acidic soil (Sk soil), indicating that the amendment of manures exerted different effect in pendimethalin dissipation rates in different pH soils. The application of pendimethalin and/or manure altered the microbial community activity after 24 h of incubation. After 110 days, the microbial community activities in green manure–amended soil were more similar to that with blank than pendimethalin treatment in both types of soils. In comparison with treatment C, microbial communities were more similar between treatment L and blank, indicating the superior effect over pendimethalin on microbial communities when applying Lupinus luteus. The research showed that the application of herbicide pendimethalin changed soil microbial community, and the amendment of manures exerted different effect in pendimethalin dissipation rates in different pH soils. It is assumed that the change in dissipation rates was originated from the microbial community change after different manure amendment.     

Metabolic diversity of the heterotrophic microorganisms and potential link to pollution of the Rouge River

The heterotrophic microbial communities of the Rouge River were tracked using Biolog Ecoplates to understand the metabolic diversity at different temporal and spatial scales, and potential link to river pollution. Site less impacted by anthrophogenic sources (site 1), showed markedly lower metabolic diversity. The only substrates that were utilized in the water samples were carbohydrates. Sites more impacted by anthrophogenic sources (sites 8 and 9) showed higher metabolic diversity. Higher functional diversity was linked to the physico-chemical and biological properties of the water samples (i.e. higher concentrations of DO, DOC, chlorophyll, and bacterial density). Biolog analysis was found to be useful in differentiating metabolic diversity between microbial communities; in determining factors that most influence the separation of communities; and in identifying which substrates were most utilized by the communities. It can also be used as an effective ecological indicator of changes in river function attributable to urbanization and pollution.     

Effect of methamidophos and urea application on microbial communities in soils as determined by microbial biomass and community level physiological profiles

In this study, we evaluated the effect of the application by two agrochemicals, methamidophos (O,S-dimethyl phosphoroamidothioate) and urea, on microbial diversity in soil, using the combined approaches of soil microbial biomass analysis and community level physiological profiles (CLPPs). The results showed that both a low and a high level of methamidophos application (CS2 and CS3) and urea application (CS4) significantly decreased microbial biomass C (Cmic) by 41-83% compared with the control (CS1). The soil organic C (Corg) values of CS3 and CS4 were significantly higher and lower by 24% and 14%, respectively, than that of CS1. Similarly to Cmic, the values of Cmic/Corg of the three applied soils which decreased were lower by 31-84% than that of CS1. In contrast, the respiration activity of the three applied soils were significantly higher than the control. Agrochemical application also significantly increased the soil total of N and P (Ntol and Ptol) and decreased the Corg/Ntol and Corg/Ptol values. The CLPPs results showed that the AWCD (average well color development) of the three applied soils were significantly higher than that of CS1 during the incubation period. Substrate richness, Shannon and Simpson indices of microbial communities under chemical stresses, increased significantly. In addition, the CFU (colony-forming unit) numbers of methamidophos metabolized bacteria in CS2 and CS3 also increased significantly by 86.1% and 188.9% compared with that of CS1. The combined results suggest that agrochemicals reduce microbial biomass and enhance functional diversities of soil microbial communities; meanwhile, some species of bacteria may be enriched in soils under methamidophos stress.     

蚯蚓生态滤池对农村生活污水的深度净化效果

本实验利用多级蚯蚓生态滤池对农村生活污水进行处理研究,分别考察了不同季节蚯蚓生态滤池对COD、TN、NH4+-N和TP的去除效果,同时利用PCR-DGGE技术对不同季节的微生物群落进行初步分析。结果表明,夏季COD、TN、NH4+-N和TP的平均去除率分别达到86.05%、89.02%、98.48%和99.1%;出水浓度17.86、4.96、0.605和0.047 mg/L。冬季COD、TN、NH4+-N和TP的平均去除率分别达到83.29%、93.26%、96.96%和92.7%;出水浓度22.68、2.63、1.02和0.37 mg/L。不同季节出水水质均达到了《城镇污水处理厂污染物排放标准》(GB18918-2002)一级A类标准。蚯蚓生态滤池内的微生物多样性冬季的要比夏季的丰富,且冬季滤池内微生物种类从上至下逐渐增加,符合污染物去除效率的变化。     

Ni2＋对活性污泥活性及群落多样性的影响

通过检测活性污泥的电子传递体系活性以及生物多样性，研究Ni2＋对活性污泥微生物活性及群落多样性的影响。结果表明：与对照系统相比，5mg／L的Ni2＋对2，3，5-lriphenylteItrazoliumchloride（TTC．ETS）活性未产生显著的影响；但当Ni2＋的浓度进一步增大到10、20和40mg／L后，其对序批式反应器内活性污泥TTC—ETS活性的抑制率分别达到（36．794-11．14）％、（55．88±13．90）％和（70．97±6．78）％。低浓度Ni2＋．能增强活性污泥微生物对碳源的利用，但高于10mg／L的Ni2＋则显著抑制了活性污泥微生物对碳源的利用。各个SBR系统中微生物群落最常见的物种相近，物种丰富度和均一性则均有所不同，其中群落物种丰富度随着Ni2＋浓度的增加而逐渐减小。TTC—ETS活性、平均每孔颜色变化率、Shan—liOn指数和Simpson指数，与Ni2＋的胁迫浓度之间的显著相关性表明，它们均可有效地表征Ni2＋胁迫对活性污泥微生物活性及群落多样性的影响程度。     

Structural and functional diversity of microbial communities from a lake sediment contaminated with trenbolone, an endocrine-disrupting chemical

Effects of trenbolone (TBOH), a hormone used in cattle production, on the structure and function of microbial communities in a fresh water sediment from a lake in Southern Germany were studied in a microcosm experiment. The microbial community structure and the total gene pool of the sediment, assessed by 16S rRNA/rDNA and RAPD fingerprint analysis, respectively, were not significantly affected by TBOH. In contrast, the N-acetyl-glucosaminidase activity was almost 50% lower in TBOH treated samples (P<0.05). Also, the substrate utilization potential, measured using the BIOLOG system, was reduced after TBOH treatment. Interestingly, this potential did not recover at the end of the experiment, i.e. 19 days after the addition of the chemical. Repeated application of TBOH did not lead to an additional reduction in the substrate utilization potential. Overall results indicate that microbial community function was more sensitive to TBOH treatment than the community structure and the total gene pool.     

In situ fixation of metals in soils using bauxite residue: biological effects

Soils polluted with heavy metals can cause phytotoxicity and exhibit impared microbial activities. In this paper we evaluate the responses of different biological endpoints to in situ remediation processes. Three soil amendments (red mud, beringite and lime) were applied to two soils polluted by heavy metals. Oilseed rape, wheat, pea and lettuce were grown successively in pots on the untreated and amended soils and their yield and metal uptake were determined. A suite of microbial tests (lux-marked biosensors, Biolog and soil microbial biomass) were performed to determine the effect of the soil amendments on the functionality and size of the soil microbial community. In both soils all three amendments reduced phytotoxicity of heavy metals, enhanced plant yields and decreased the metal concentrations in plants. The red mud treatment also increased soil microbial biomass significantly. The microbial biosensors responded positively to the remediation treatments in the industrially-contaminated soil used in the experiment. Red mud applied at 2% of soil weight was as effective as beringite applied at 5%. The results also showed that since the biological systems tested respond differently to the alleviation of metal toxicity, a suite of biological assays should be used to assess soil remediation processes.     

In-field variation in 2,4-D mineralization in relation to sorption and soil microbial communities

This study was undertaken to assess 2,4-D mineralization in an undulating cultivated field, along a sloping transect (458 m to 442 m above sea level), as a function of soil type, soil microbial communities and the sorption of 2,4-D to soil. The 2,4-D soil sorption coefficient (Kd) ranged from 1.81 to 4.28 L kg(-1), the 2,4-D first-order mineralization rate constant (k) ranged from 0.04 to 0.13 day(-1) and the total amount of 2,4-D mineralized at 130 days (M(130)) ranged from 24 to 39%. Both k and M(130) were significantly negatively associated (or correlated) with soil organic carbon content (SOC) and Kd. Both k and M(130) were significantly associated with two fatty-acid methyl esters (FAME), i17:1 and a18, but not with twenty-two other individual FAME. Imperfectly drained soils (Gleyed Dark Grey Chernozems) in lower-slopes showed significantly lesser 2,4-D mineralization relative to well-drained soils (Orthic Dark Grey Chernozems) in mid- and upper-slopes. Well-drained soils had a greater potential for 2,4-D mineralization because of greater abundance and diversity of the microbial community in these soils. However, the reduced 2,4-D mineralization in imperfectly drained soils was predominantly because of their greater SOC and increased 2,4-D sorption, limiting the bioavailability of 2,4-D for degradation. The wide range of 2,4-D sorption and mineralization in this undulating cultivated field is comparable in magnitude and extent to the variability of 2,4-D sorption and mineralization observed at a regional scale in Manitoba. As such, in-field variations in SOC and the abundance and diversity of microbial communities are determining factors that require greater attention in assessing the risk of movement of 2,4-D by runoff, eroded soil and leaching.     

In-field variation in 2,4-D mineralization in relation to sorption and soil microbial communities

This study was undertaken to assess 2,4-D mineralization in an undulating cultivated field, along a sloping transect (458 m to 442 m above sea level), as a function of soil type, soil microbial communities and the sorption of 2,4-D to soil. The 2,4-D soil sorption coefficient (Kd) ranged from 1.81 to 4.28 L kg^?1, the 2,4-D first-order mineralization rate constant (k) ranged from 0.04 to 0.13 day^?1 and the total amount of 2,4-D mineralized at 130 days (M₁₃₀) ranged from 24 to 39%. Both k and M₁₃₀ were significantly negatively associated (or correlated) with soil organic carbon content (SOC) and Kd. Both k and M₁₃₀ were significantly associated with two fatty-acid methyl esters (FAME), i17:1 and a18, but not with twenty-two other individual FAME. Imperfectly drained soils (Gleyed Dark Grey Chernozems) in lower-slopes showed significantly lesser 2,4-D mineralization relative to well-drained soils (Orthic Dark Grey Chernozems) in mid- and upper-slopes. Well-drained soils had a greater potential for 2,4-D mineralization because of greater abundance and diversity of the microbial community in these soils. However, the reduced 2,4-D mineralization in imperfectly drained soils was predominantly because of their greater SOC and increased 2,4-D sorption, limiting the bioavailability of 2,4-D for degradation. The wide range of 2,4-D sorption and mineralization in this undulating cultivated field is comparable in magnitude and extent to the variability of 2,4-D sorption and mineralization observed at a regional scale in Manitoba. As such, in-field variations in SOC and the abundance and diversity of microbial communities are determining factors that require greater attention in assessing the risk of movement of 2,4-D by runoff, eroded soil and leaching.     

Response of denitrification genes <Emphasis Type="Italic">nirS</Emphasis>, <Emphasis Type="Italic">nirK</Emphasis>, and <Emphasis Type="Italic">nosZ</Emphasis> to irrigation water quality in a Chinese agricultural soil

Purpose  

Denitrification is an important biochemical process in global nitrogen cycle, with a potent greenhouse gas product N₂O. Wastewater irrigation can result in the changes of soil properties and microbial communities of agricultural soils. The purpose of this study was to examine how the soil denitrification genes responded to different irrigation regimes.     

2,4-Dichlorophenoxy Acetic Acid Mineralization in Amended Soil

The application of municipal biosolid or liquid hog manure to agricultural soils under laboratory conditions at 20°C influenced the fate of the herbicide 2,4-D [2,4-(dichlorophenoxy)acetic acid] in soil. When 2,4-D was added to soil at agronomic rates immediately after the addition of manure or biosolids to a coarse-textured soil, the percentage of 2,4-D mineralized at 100 days was about 47% for both treatments, compared to only 31% for control soils without amendments. The enhanced 2,4-D mineralization as a result of amendment addition was due to an increased heterotrophic microbial activity, with the greatest increases in soil respiration occurring for soils amended with biosolids. When additions of 2,4-D were delayed for one, two, or four weeks after the amendments were applied, the additions of amendments generally reduced 2,4-D mineralization in soil, particularly for manure, indicating that the effect of amendments on enhancing soil microbial activities diminished over time. In contrast, the mineralization of 2,4-D in control soils was less dependent on when 2,4-D was applied in relation to pre-incubations of soil for zero, one, two, or four weeks. The effect of manure on decreasing 2,4-D mineralization in specific soils was as large as the effect of soil texture on differences in 2,4-D mineralization across soils. Because manure was not found to impact 2,4-D sorption by soil, it is possible that 2,4-D mineralization decreased because 2,4-D transformation products were strongly sorbed onto organic carbon constituents in manure-amended soils and were therefore less accessible to microorganisms. Alternatively, microorganisms were less likely to metabolize the herbicide because they preferentially consumed the type of organic carbon in manure that is a weak sorbent for 2,4-D.