Influence of different temperatures on metal tolerance measurements and growth response in bacterial communities from unpolluted and polluted soils

The effects of temperature on the growth rate and metal toxicity in soil bacterial communities extracted from unpolluted and polluted soils were investigated using the thymidine and leucine incorporation techniques. An agricultural soil, which was contaminated in the laboratory with Cu, Cd, Zn, Ni or Pb, and an uncontaminated forest soil were used. Measurements were made at 0°C and 20°C. Leucine incorporation was found to be as sensitive to heavy metals as thymidine incorporation in the short-term trial used to indicate heavy metal tolerance. Similar IC₅₀ values (the log of the metal concentration that reduced incorporation to 50%) were also obtained at 0 and 20°C, independently of the technique used. Metal tolerance could thus be measured using both techniques at any temperature in the range 0–20°C. In the long-term experiment different temperature-growth relationships were obtained on the basis of the rate of thymidine or leucine incorporation into bacterial assemblages from unpolluted and polluted soils, as judged from the minimum temperature values. This could not be attributed to the metal addition alone since different patterns were observed when different metals were added to the soil. Thus, the minimum temperature for thymidine incorporation was similar in Cu-polluted and unpolluted soil, while in soils polluted with Cd and Zn the minimum temperature increased by 2°C, and Ni and Pb additions increased the minimum temperature by 4°C compared to the unpolluted soil. This suggested that heavy metal pollution led to bacterial communities showing different temperature characteristics to those in the corresponding unpolluted soil. Similar observations were deduced from the minimum temperatures required for leucine incorporation. Three groups of bacterial communities were distinguished according to the growth response to temperature in polluted soils, one group in Cu-polluted soil, a second group in soil polluted with Zn and Cd, and a third group in soils polluted with Ni and Pb.     

Distribution of different fractions of cadmium,zinc, lead,and copper in unpolluted and polluted soils

McLaren and Crawford's method for fractionating soil Cu was modified, and used to fractionate soil Cd, Zn, Pb, and Cu in 38 soil samples from 11 soil profiles from industrially polluted and nearby unpolluted areas. Pollutant metals, especially Cd and Zn, were more soluble than the native soil metals. On average, approximately 45% of Cd was present in the CaCl₂ soluble (CA) fraction, whereas corresponding values for the other metals were below 10%. The percentages of each metal in the CA fraction followed the order Cd > Zn > Pb > Cu. The same order was observed for the acetic acid soluble (AAC) fraction. Approximately 30% of total Pb and Cu were present in the pyrophosphate soluble (PYR) fraction, and only 10% of total Cd and Zn. Approximately 20% of total Zn or Pb and 10% of Cd or Cu were present in the free oxide (OX) fraction. Only 20% of Cd and between 40–50% of the other 3 metals were present in the residual (RES) fraction. The results show that Cd is more labile than the other 3 metals.     

Dynamics of C natural abundance in wood decomposing fungi and their ecophysiological implications

Factors that affect the δ¹³C values of fungi need to be analyzed for the progress of isotope-based studies of food-chain or organic matter dynamics in soils. To analyze the factors that control δ¹³C values of the fungal body, basidiomycete and ascomycete species were grown on a beechwood substrate (six species) and in glucose medium (nine species), and the δ¹³C value of produced fungal body was compared to that of the carbon source. The ¹³C enrichment (Δδ¹³C) in the fungal aggregates compared to the decomposed wood varied from 1.2 to 6.3‰ among six species. In the glucose substrate experiment, the degree of ¹³C enrichment in the hyphal mat was relatively small and varied from −0.1 to 2.8‰ among nine basidiomycetes species depending on their growth stage. Calculated δ¹³C values of the respired CO₂ were lower than those of the hyphal mat, organic metabolites and the glucose used. The degree of ¹³C enrichment was affected by fungal species, substrate and growth stage. Fungal internal metabolic processes are the plausible mechanism for the observed isotopic discrimination between fungal bodies and substrates. Especially, dark fixation of ambient CO₂ and kinetic isotope fractionation during assimilation and dissimilation reactions could well explain Δδ¹³C dynamics in our experiments. Through the analysis of field Δδ¹³C, we could know undisturbed fungal status about starvation, aeration and type of decomposition.     

Comparison of factors limiting bacterial growth in different soils

Lack of carbon has been assumed to be the most common limiting factor for bacterial growth in soil, although there are reports of limitation by other nutrients, e.g. nitrogen and phosphorus. We have studied which nutrient(s) limited instantaneous growth rates of bacteria in 28 Swedish soils using the thymidine or leucine incorporation technique to measure increased growth rate after adding different combinations of organic carbon (glucose), nitrogen and phosphorus. The soils ranged in pH between 3.1 and 8.9, in organic matter content between 1% and 91% and in soil C/N ratio between 10 and 28. We also tested the effect of adding different amounts of carbon on the bacterial change in growth rate for two soils with different organic matter content. We found that bacterial growth in most of the 28 soils was limited by a lack of carbon, indicated by an increased bacterial growth rate 48 h after adding glucose. In some soils, adding carbon together with nitrogen increased the bacterial growth rates even further. In three soils no effects were seen upon adding nutrients separately, but adding carbon and nitrogen together increased bacterial growth rates. Nitrogen addition tended to decrease bacterial growth rates, while phosphorus addition had little effect in most soils. No correlations were found between the soil C/N ratio, ammonium or nitrate content in soil and bacterial growth limitation, indicating that even soils with a C/N ratio of 28 could be carbon limited. Although the interpretation of the effects of a single limiting nutrient was in most cases straightforward, an interaction between the amount of carbon added and the organic matter content of the soil confounded the interpretation of the extent of a second limiting nutrient.     

The effects of different amendments on the mobility and toxicity of nickel and copper in polluted soils

The efficiency of different amendments for the remediation of acidic sandy soils contaminated with heavy metals was assessed in a model laboratory experiment with podzols from the Kola Peninsula. The addition of SLOVAKITE®, lime, and vermiculite leads to significant reduction of the acidity and mobility of nickel and copper in polluted soils, as well as an increase in the length and phytomass of shoots and roots of Festuca rubra. The application of superphosphate for the remediation of soils that were polluted with nickel and copper was less helpful, while the apatite and zeolite additives were inefficient.     

Molecular diversity of bacterial chitinases in arable soils and the effects of environmental factors on the chitinolytic bacterial community

The molecular diversity of bacterial chitinases in the bulk soils of arable land was investigated using culture-independent methods. The results demonstrate that bacterial chitinases in arable soils are highly diverse and comprise unique groups when their sequences were compared to those in public databases. The diversity of bacterial chitinases in arable soil was further evaluated using conventional phylogenetic analysis, the UniFrac analysis of the phylogenetic data, and the multidimensional scaling (MDS) analysis of T-RFLP profiles to elucidate the relationship between the diversity of bacterial chitinases and soil characteristics. These analyses indicate that environmental factors such as soil type and pH are responsible for shaping the composition of bacterial chitinases.     

Molecular community analysis of arbuscular mycorrhizal fungi associated with five selected plant species from heavy metal polluted soils

Arbuscular mycorrhizal fungi (AMF) are known to play an important role in plant tolerance to heavy metals (HMs) stress. This study aimed to understand the diversity of AMF communities associated with five selected plant species (Phytolacca americana, Rehmannia glutinosa, Perilla frutescens, Litsea cubeba and Dysphania ambrosioides) from severely HMs polluted soils in Dabaoshan Mine region, China, using molecular methods. Plant roots and rhizospheric soils were sampled from four sites, respectively. Targeting the fungal small subunit (SSU) rRNA gene, PCR-denaturing gradient gel electrophoresis (DGGE) analysis indicated that varied AMF communities colonized different plant species, and the AMF communities in rhizospheric soils were different from those in plant roots. Total six SSU rRNA gene clone libraries including four root samples and two rhizospheric soil samples were constructed. Screening clone libraries by DGGE and sequence analysis revealed that Glomus dominated all of the samples except for the roots of D. ambrosioides, while Kuklospora and Ambispora dominated the roots of D. ambrosioides and the rhizosphere of P. americana. This study indicates that diverse AMF are associated with these selected plants, and they are potentially useful to promote the phytoremediation of this HMs polluted area.     

植烟土壤提取物质对烟株生长及根际土壤细菌多样性的影响

对盆栽烟草外源添加不同浓度植烟土壤提取物质(T1:40μg·mL-1;T2:120μg·mL-1;CK:蒸馏水对照),探讨植烟土壤提取物质对烟草生长及土壤细菌多样性的影响。结果表明,植烟土壤提取物处理使烟株生长受抑制,且随处理浓度的增加受抑制程度显著提高,具体表现为烟株变矮,叶面积变小,光合作用能力降低,且烟草的保护酶系统受到破坏,丙二醛含量随处理浓度加大而增加,T2处理的丙二醛含量是对照的3.44倍。对外源添加物质处理后烟草根际土壤微生物T-RFs分析发现,在对照检测到17个门24个纲,T1处理有14个门21个纲,T2有10个门17个纲;丰富度指数的变化也和门纲的变化一致,随着处理浓度的增加而显著降低。可见外源添加物质处理后,根际土壤细菌群落减少,多样性水平下降。对各处理的根际土壤微生物T-RFs变化与烟株生长变化进行相关性分析表明,在外源添加物质处理的土壤中存在较多的负相关T-RFs片段,且这些片段中较多为病原菌;而正相关的T-RFs片段主要存在于对照土壤中,其中有较多与土壤营养元素循环相关的微生物。本研究结果显示,在外源添加植烟土壤提取物质处理下,烟草的生长受抑制,烟草根际土壤的微生态受到破坏,且随浓度的提升而加重。因此,连作土壤中自毒物质的富集是造成烟草连作障碍的主要原因。关键词烟草连作障碍根际细菌自毒作用T-RFLP     

Characterization of cultivable heterotrophic bacterial communities in Cr-polluted and unpolluted soils using Biolog and ARDRA approaches

The bacterial communities of two soils with different chromium levels were characterized by Biolog carbon substrate utilization patterns and amplified 16S ribosomal DNA restriction analysis (ARDRA). For each bacterial community sample, cell suspensions containing 10,000 or 100 colony-forming units (CFU) were inoculated in each well of Biolog-GN microplates. The number of carbon compounds utilized by the bacterial community consisting of 100 CFU from unpolluted soil was significantly lower than that detected for the bacterial community consisting of 10,000 CFU. The size of inoculum did not substantially influence the percentage of carbon sources utilized by the Cr-polluted soil bacterial community. ARDRA approach was applied to about 100 bacterial isolates for each soil sample. A similar number of clusters for Gram-negative bacteria were found in both soils, but there were differences in percentages of isolates belonging to each group and specific genomic groups were found in each soil. Pseudomonas was the dominant taxon in both soils. Comparing the ARDRA clusters obtained from Gram-positive isolates it was evident that the culturable bacterial communities of Cr-polluted and unpolluted soils were dominated by the genus Arthrobacter and the genus Bacillus, respectively.     

Fungal diversity in soils and historic wood from the Ross Sea Region of Antarctica

Microorganisms play a dominant role in Antarctic ecosystems, yet little is known about how fungal diversity differs at sites with considerable human activity as compared to those that are remote and relatively pristine. Ross Island, Antarctica is the site of three historic expedition huts left by early explorers to the South Pole, Robert F. Scott and Ernest Shackleton. The fungal diversity of these wooden structures and surrounding soils was investigated with traditional culturing methods as well as with molecular methodology including denaturing gradient gel electrophoresis (DGGE) using the internal transcribed spacer (ITS) regions of ribosomal DNA for identification. From historic wood and artifact samples and soils adjacent to the huts as well as soil samples obtained from the Lake Fryxell Basin, a remote Dry Valley location, and remote sites at Mt. Fleming and the Allan Hills, 71 fungal taxa were identified. The historic huts and associated artifacts have been colonized and degraded by fungi to various extents. The most frequently isolated fungal genera from the historic woods sampled include Cadophora, Cladosporium and Geomyces. Similar genera were found in soil samples collected near the huts. Sampling of soils from locations in the Transantarctic Mountains and Lake Fryxell Basin at considerable distances from the huts and with different soil conditions revealed Cryptococcus spp., Epicoccum nigrum and Cladosporium cladosporioides as the most common fungi present and Cadophora species less commonly isolated. DGGE revealed 28 taxa not detected by culturing including four taxa which possibly have not been previously described since they have less than 50% ITS sequence identity to any GenBank accessions. Fungi capable of causing degradation in the wood and artifacts associated with the expedition huts appear to be similar to those present in Antarctic soils, both near and at more remote locations. These species of fungi are likely indigenous to Antarctica and were apparently greatly influenced by the introduction of organic matter brought by early explorers. Considerable degradation has occurred in the wood and other materials by these fungi.     

Fungal and bacterial growth following the application of slurry and anaerobic digestate of livestock manure to temperate pasture soils

How land-application of digestate sourced from anaerobic digestion (AD) of animal waste influences the functioning of a mixed pasture agroecosystem is not well characterised, particularly with regard to the response of the actively growing microbial community. We studied the impact of the liquid AD digestate on the decomposer community in two different soils, seeded with two different common grassland crops; a mixture of either grass or grass/clover in a greenhouse experiment. We studied bacterial (leucine incorporation into bacteria) and fungal (acetate incorporation into ergosterol) growth responses to AD cattle slurry digestate, undigested cattle slurry, mineral fertiliser (NPK and N) added at a rate equivalent to 150?kg?N?ha^?1, and a no-fertiliser control treatment. Differences in fungal and bacterial growth were evident between the soil and sward types. However, the fertilisers consistently stimulated a higher bacterial growth than the no-fertiliser control, and liquid digestate resulted in a level of bacterial growth higher or equal to that of mineral fertiliser, whilst undigested slurry resulted in lower bacterial growth. These fertiliser effects on bacterial growth mirrored the effects on plant growth. In contrast, the fungal community responded only marginally to fertiliser treatments. We conclude that the application of digestate stimulates the bacterial decomposer community in a similar way to that of mineral fertilisers. Our results suggest that mineral fertiliser can be exchanged for liquid digestate with limited impact on the actively growing soil microbial community that, in turn, regulate important soil processes including nutrient cycling in agricultural soils.     

Cereal/legume rotation effects on rhizosphere bacterial community structure in west african soils

The increased use of cereal/legume crop rotation has been advocated as a strategy to increase cereal yields of subsistence farmers in West Africa, and is believed to promote changes in the rhizosphere that enhance early plant growth. In this study we investigated the microbial diversity of the rhizoplane from seedlings grown in two soils previously planted to cereal or legume from experimental plots in Gaya, Niger, and Kaboli, Togo. Soils from these legume rotation and continuous cereal plots were placed into containers and sown in a growth chamber with maize (Zea mays L.), millet (Pennisetum glaucum L.), sorghum (Sorghum bicolor L. Moench.), cowpea (Vigna unguiculata L.) or groundnut (Arachis hypogaea L.). At 7 and 14 days after sowing, 16S rDNA profiles of the eubacterial and ammonia-oxidizing communities from the rhizoplane and bulk soil were generated using denaturing gradient gel electrophoresis (DGGE). Community profiles were subjected to peak fitting analyses to quantify the DNA band position and intensities, after which these data were compared using correspondence and principal components analysis. The data showed that cropping system had a highly significant effect on community structure (p <0.005), irrespective of plant species or sampling time. Continuous cereal-soil grown plants had highly similar rhizoplane communities across crop species and sites, whereas communities from the rotation soil showed greater variability and clustered with respect to plant species. Analyses of the ammonia-oxidizing communities provided no evidence of any effects of plant species or management history on ammonia oxidizers in soil from Kaboli, but there were large shifts with respect to this group of bacteria in soils from Gaya. The results of these analyses show that crop rotation can cause significant shifts in rhizosphere bacterial communities.     

污染区千金子和酢浆草根际土壤中PAHs结合态残留的梯度分布

采集某污染区千金子(Euphorbia lathyris L.)和酢浆草(Oxalis corniculata L.)的离根表0~3、3~6、6~9 mm的根际土壤,分析了多环芳烃(PAHs)结合态残留中母体化合物(Parent compound of bound residue,PCBR)在根际土壤中的含量及梯度分布规律。供试土壤类型为黄棕壤。结果表明,在非根际和根际土壤中均可检出10种PAHs的PCBR,非根际土壤中PCBR总含量为3.31 mg kg-1,高于根际土壤(1.07~1.82mg kg-1)。根际土壤中PAHs的PCBR含量随离根表距离(0~9 mm)的增加而增大。可用根际效应(R)来衡量根际土壤中PAHs的PCBR含量与非根际土壤相比减少的比例;R值随离根表距离(0~9 mm)的增加而变小。3个连续根际区中,PAHs总PCBR的R值为45.15%~67.66%,其中2环PAH的R值最大(61.18%~93.50%),4环和5环PAHs的R值最小(2.39%~6.31%),低环PAHs的PCBR在根际土壤中更易转化。PAHs的PCBR在千金子根际土壤中R值大于酢浆草,表明前者有更利于PAHs结合态残留转化的根际环境。PAHs结合态残留的根际梯度分布与根系分泌物的梯度分布关系密切,而PAHs种类、植物根际环境对PAHs结合态残留的分布影响显著。     

Direct and indirect effects of enhanced UV-B radiation on the decomposing and competitive abilities of saprobic fungi

Increases in UV-B radiation have been shown to slow the rate of litter decomposition in ecosystems. However, it is unclear if this is a result of direct UV-B effects on saprobic microorganisms, or a result of UV-B-induced changes in litter quality that indirectly affect decay by saprobes. In this study, we evaluated the magnitude of direct and indirect effects on litter decomposition of Brassica napus by soil fungi, under growth chamber conditions. We found that, both, direct and indirect UV-B negatively influenced litter decomposition, however, direct effects were much more pronounced. We then tested whether UV-B radiation would have species-specific effects on fungal colonization and competitive ability, rather than influencing all fungal species equally. We predicted that darkly pigmented fungi would increase their relative competitive ability under high UV-B. The test fungi were all isolated from field soil under Brassica napus. Two fungi were hyaline (Aspergillus terreus, Trichoderma koningii), two were darkly-pigmented (Cladosporium sphaerospermum, Epicoccum purpurascens) and one had a hyaline mycelium but darkly-pigmented conidia (Aspergillus niger). Elevated UV-B radiation had differential direct and indirect effects on fungal growth, and caused shifts in the competitive balances between pigmented and non-pigmented fungi. However, in only two of six pair-wise challenges did the pigmented species increase their relative competitive ability under UV-B conditions. It is clear that UV-B profoundly influence fungal community structure in soil, but the direction of such effects remains unpredictable.     

不同强化调控措施对多氯联苯污染土壤的修复效应

通过微宇宙试验,以添加碳源、翻耕、覆膜为调控因子,研究生物强化措施对多氯联苯污染土壤的修复效果,并对修复前后土壤微生物数量以及功能多样性进行研究。结果表明,土壤翻耕处理是最有效的修复措施,修复效率达到29.3%;添加淀粉与覆膜处理,效果相近,并且均显著高于对照。添加淀粉和翻耕均显著增加了土壤中细菌、真菌和放线菌的数量,但降低了土壤微生物功能多样性。而覆膜处理则使土壤微生物数量及其活性和多样性均下降。可见,不同的调控措施不仅能够强化土壤中多氯联苯的降解,同时也会影响到土著微生物群落结构及其活性和功能。     

Fungal and bacterial recolonisation of acid and alkaline forest soils following artificial heat treatments

The direct response and the short-term recolonisation of soil by fungi and bacteria were studied after heat treatments of a humus soil with high carbon content and low pH, and a calcareous soil with lower carbon content and high pH. Heating was administered using a muffle furnace or an autoclave, with different temperatures and times of heat exposure, after which fresh soil (1%) was added as inoculum. Autoclaved soil showed more marked increases in bacterial growth during the recovery phase than oven-heated soil, and the bacterial growth response was more rapid in calcareous than in humus soil. Fungal growth recovered more rapid and reached values higher than the control in humus soil, while it remained low until the end of the study in calcareous soil. Respiration rate showed similar patterns in both soils. Fungal biomass (ergosterol and PLFA 18:2ω6,9) indicated that fungi benefited by autoclaving in humus soil, while they were disfavoured by this treatment in calcareous soil. The sum of bacterial PLFAs did not change due to heating, but some bacterial PLFAs (e.g. cy17:0) increased in both soils. We propose that the community assembly of the microbial communities after heating were mainly driven by pH, in that the high pH soil selected primarily for bacteria and the low pH soil for fungi.     

Influence of sodium polyacrylate on the water-holding capacity of three different soils and effects on growth of wheat

Abstract. The effectiveness of sodium polyacrylate to increase soil water retention and to enhance growth of wheat under water deficit was evaluated. Water-holding capacity of the soils was considerably increased only when the soil was amended with the polymer at a rate 3 g L⁻¹. The effect on plant-available water was greater at soil matric potentials up to −1000 hPa. The biomass and grain yield of plants without water deficit were increased by the polymer amendment, but decreased under severe water deficit stress. The polymer had no significant affect on plant N, grain N or grain Na content.     

Effects of indigenous and introduced arbuscular mycorrhizal fungi on the growth of Allium fistulosum under field conditions

ABSTRACT

Enhanced phosphorus (P) uptake from the soil and increased plant growth related to arbuscular mycorrhizal (AM) fungi in pot culture, using sterilized soil, are well-known phenomena. However, these enhancements are not widely observed under field conditions because field sterilization is difficult. The aim of this study was to investigate the effects of AM fungi on P uptake and the growth of Allium fistulosum in non-fumigated and fumigated fields, under different levels of P availability. Plants were inoculated with the AM fungus Glomus R-10 and grown in fumigated soil. For the uninoculated treatment, a sterilized inoculum was applied directly. The field was fumigated using dazomet. Superphosphate was applied to the field at the rates of 0 (P0) or 500 (P500) kg P₂O₅ ha^?1. The inoculated and uninoculated plants were transplanted into the fields and sampled three times to measure AM fungal colonization, shoot P concentration, and shoot dry weight of the plants. At the transplanting stage, AM fungal colonization was observed in the inoculated plants (>70%) but not in the uninoculated plants. At the third sampling, irrespective of P treatment, AM fungal colonization was observed both in the uninoculated and inoculated plants in the non-fumigated field, and there was no difference in shoot P content and shoot dry weight between the inoculated and uninoculated plants. AM fungal colonization in the fumigated field was higher in the inoculated than uninoculated plants, irrespective of P treatment; shoot P content and shoot dry weight were both higher in the inoculated plants than in the uninoculated plants with P0. These results suggest that the responses of A. fistulosum to AM fungal inoculation under the low-P and fumigated conditions are similar to those observed in sterilized pot culture conditions.     

接种溶磷真菌对玉米和大豆根际细菌群落结构多样性的影响

Application of phosphate-solubilizing microorganisms （PSMs） has been reported to increase P uptake and plant growth. However, no information is available regarding the ecological consequences of the inoculation with PSMs. The effect of inoculation with phosphate-solubilizing fungal （PSF） isolates Aspergillus niger P39 and Penicillium oxalicum P66 on the bacterial communities in the rhizospheres of maize （Zea mays L. ‘Haiyu 6＇） and soybean （Glycine max Merr. ‘Heinong 35＇） was examined using culture-dependent methods as well as a culture-independent method, polymerase chain reaction-denaturing gradient gel electrophoresis （PCR-DGGE）. Compared with the control, the number of culturable microbes for soybean was significantly greater with P39, whereas for maize, the same was significantly greater with P66. In addition, a greater number of microbes were found in the rhizosphere of maize compared with soybean. The fingerprint of DGGE for 16S rDNA indicated that inoculation with PSF also increased bacterial communities, with the P66 treatment having higher numbers of DGGE bands and a higher Shannon-Weaver diversity index compared with P39; the composition of the microbial community was also more complex with the P66 treatment. Overall, complex interactions between plant species and exotic PSMs affected the structure of the bacterial community in the rhizosphere, but plant species were more important in determining the bacterial community structure than the introduction of exotic microorganisms.