Influence of long-term conservation tillage on soil and rhizosphere microorganisms

 In long-term field experiments on sandy loam and loamy sand soils, the influence of conservation and conventional tillage on soil and rhizosphere microorganisms was studied. Conservation tillage stimulated rhizosphere bacteria on winter wheat, winter barley, winter rye and maize in different soil layers. Particularly the populations of Agrobacterium spp. and Pseudomonas spp. were increased. On the sandy loam, N₂ fixation and nodulation of pea plants were significantly increased. No influence of different soil tillage was determined on the colonization of the rhizosphere by mycorrhiza and saprophytic fungi. Stubble residues infected with Gaeumanomyces graminis were infectious for a longer time on the soil surface than after incorporation into the soil. Received: 10 March 1998     

Phosphate-solubilizing microorganisms associated with the rhizosphere of oil palm (Elaeis guineensis Jacq.) in Colombia

This study determined the cell density in the field and the biological activity of culturable phosphate-solubilizing microorganisms (PSMs) present in the Elaeis guineensis Jacq. rhizosphere at two locations in a commercial plantation. Promising isolates found in two soils under different agronomic management conditions were selected. We first calculated the cell density of cultivable PSMs grown in SRS medium (Sundara-Rao and Sinha, 1963) supplemented with the insoluble phosphate sources Ca₃(PO₄)₂, AlPO₄, and FePO₄. Twenty-two bacteria, ten filamentous fungi, and eight yeast isolates were found. The 16 isolates with the clearest P solubilization halo in Petri dishes were selected to estimate their P solubilization potential in SRS medium with Ca₃(PO₄)₂. No solubilization activity was registered using AlPO₄ or FePO₄ as the P sources. Ten of the isolates presented solubilization efficiencies between 20 and 82%. Some of these isolates showed high percentages of identity with the 16S and ITS rDNA sequences of the genera Aspergillus, Penicillium, Klebsiella, Burkholderia, and Staphylococcus according to the NCBI and EzTaxon-e databases. The solubilization activity of the isolates was associated with a decrease in the pH and the release of organic acids, such as gluconic, citric, succinic, and acetic acids. Gluconic acid was mainly released by the genera Aspergillus and Penicillium, and these isolates also showed the highest solubilization activities (82 and 80%, respectively). Therefore, these isolates were selected as the most promising isolates present in the oil palm rhizosphere for phosphate solubilization.     

Nitrification and denitrification in the rhizosphere of rice: the detection of processes by a new multi-channel electrode

 N turnover in flooded rice soils is characterized by a tight coupling between nitrification and denitrification. Nitrification is restricted to the millimetre-thin oxic surface layer while denitrification occurs in the adjacent anoxic soil. However, in planted rice soil O₂ released from the rice roots may also support nitrification within the otherwise anoxic bulk soil. To locate root-associated nitrification and denitrification we constructed a new multi-channel microelectrode that measures NH₄ ⁺, NO₂ ^–, and NO₃ ^– at the same point. Unfertilized, unplanted rice microcosms developed an oxic-anoxic interface with nitrification taking place above and denitrification below ca. 1 mm depth. In unfertilized microcosms with rice plants, NH₄ ⁺, NO₂ ^– and NO₃ ^– could not be detected in the rhizosphere. Assimilation by the rice roots reduced the available N to a level where nitrification and denitrification virtually could not occur. However, a few hours after injecting (NH₄)₂HPO₄ or urea, a high nitrification activity could be detected in the surface layer of planted microcosms and in a depth of 20–30 mm in the rooted soil. O₂ concentrations of up to 150 μM were measured at the same depth, indicating O₂ release from the rice roots. Nitrification occurred at a distance of 0–2 mm from the surface around individual roots, and denitrification occurred at a distance of 1.5–5.0 mm. Addition of urea to the floodwater of planted rice microcosms stimulated nitrification. Transpiration of the rice plants caused percolation of water resulting in a mass flow of NH₄ ⁺ towards the roots, thus supporting nitrification. Received: 23 July 1999     

Microsensor analysis of oxygen and pH in the rice rhizosphere under field and laboratory conditions

 O₂ and pH microsensors were used to analyse the microdistribution of O₂ and pH inside and outside roots of lowland rice (Oryza sativa L.). The roots of 3-week-old transplants had O₂ concentrations of about 20% air saturation at the surface, but due to a high rate of O₂ consumption in the rhizosphere, the oxic region only extended about 0.4 mm into the surrounding soil. Also the fine lateral roots created an oxic zone extending about 0.15 mm into the soil. The O₂ concentration within the roots approached air saturation close to the base, but only about 40–60% of air saturation in a region about 8 cm below the base where lateral rootlets were present. A shift from air to N₂ around the leaves led to a drop of 50% in the O₂ concentration after 12 min at a distance of 8.5 cm from the base. Flowering plants did not export O₂ to the soil from the majority of their roots, but high microbial activity was present in a very thin biofilm covering the root surface. A brown colour around the thin lateral roots indicated some O₂ export from these also during flowering. No oxidized zone was present around the roots at later stages of crop growth. The roots caused only minor minima in pH (<0.2 pH units) in the rhizosphere as compared to the bulk soil. Illumination of the plants had no effect on rhizosphere pH. Received: 28 April 1998     

Effect of organic nitrogen fertilizers on microbial populations associated with bermudagrass putting greens

 Four natural organic fertilizers, alone or in combination with the synthetic organic fertilizer isobutylidene diurea (IBDU), were compared with IBDU alone for their effect on soil/root microbial populations associated with bermudagrass grown on a golf course putting green in southern Florida, USA. Populations of total fungi, total bacteria, fluorescent pseudomonads, Stenotrophomonas maltophilia, actinomycetes and heat-tolerant bacteria were monitored every 3 months during the 2-year study. On only one sampling date and for only one bacterial population (S. maltophilia) was a significant difference in microbial populations obtained among the fertilizer treatments. However, the S. maltophilia populations associated with the natural organic fertilizer treatments were not significantly different from the synthetic organic IBDU fertilizer treatment. Received: 4 April 1997     

Antagonistic effect of Pseudomonas spp. on pathogenic fungi and enhancement of growth of green gram (Vigna radiata)

species were isolated from the rhizosphere of green gram [Vigna radiata (L.) Wilczek] and some of the rhizobacterial isolates were found to have a wide range of antifungal activity inhibiting growth of the phytopathogenic fungi Aspergillus sp., Curvularia sp., Fusarium oxysporum and Rhizoctonia solani in culture. These isolates also showed slight inhibition of the growth of a Bradyrhizobium strain (Vigna) in a spot test which was mainly a result of nutrient competition as culture supernatants of the Pseudomonas isolates did not inhibit the growth of bradyrhizobia but inhibited the growth of fungi. The rhizobacterial isolates produced siderophores in Fe-deficient succinate medium. However, the inhibition of fungal growth by different Pseudomonas isolates in Luria Bertani and King's medium B which were not limiting in Fe³⁺ ions suggested that, besides siderophores, other antifungal compounds (antibiotics) produced by these rhizobacteria were involved in antagonism. On coinoculation of green gram with Pseudomonas strains MRS13 and MRS16 and Bradyrhizobium sp. (Vigna) strain S24, there was a significant increase in nodule weight, plant dry weight and total plant N as compared to inoculation with Bradyrhizobium strain S24 alone, suggesting that the nodule-promoting effects of Pseudomonas sp. lead to an increase in symbiotic N fixation and plant growth. Received: 27 October 1997     

Survival and cell culturability of biocontrol Pseudomonas fluorescens CHA0 in the rhizosphere of cucumber grown in two soils of contrasting fertility status

 The effect of cucumber roots on survival patterns of the biocontrol soil inoculant Pseudomonas fluorescens CHA0-Rif was assessed for 22 days in two non-sterile soils, using a combination of total immunofluorescence cell counts, Kogure's direct viable counts and colony counts on plates containing rifampicin. In Eschikon soil (high fertility status for cucumber), CHA0-Rif persisted as culturable cells in bulk soil and in the rhizosphere, but colony counts were lower than viable counts and total cell counts inside root tissues. The occurrence of viable but non-culturable (VBNC) cells inside root tissues (5 log cells g^–1 root) was unlikely to have resulted from the hydrogen peroxide treatment used to disinfect the root surface, as hydrogen peroxide caused the death of CHA0-Rif cells in vitro. In Siglistorf soil (low fertility status for cucumber), the inoculant was found mostly as non-culturable cells. Colony counts and viable counts of CHA0-Rif were similar, both in bulk soil and inside root tissues, whereas in the rhizosphere viable counts exceeded colony counts at the last two samplings (giving about 7 log VBNC cells g^–1). In conclusion, soil type had a significant influence on the occurrence of VBNC cells of CHA0-Rif, although these cells were found in root-associated habitats (i.e. rhizosphere and root tissues) and not in bulk soil. Received: 12 November 1999     

The number of bacteria in the rhizosphere during plant development: relating colony-forming units to different reference units

 The number of bacteria was determined during the growth of chrysanthemum plants on young (tip) and old (base) root parts. We assessed if the same conclusions could be drawn on the dynamics of bacterial populations during plant development when different reference units were used to express the bacterial counts. The results indicated that the total number of bacteria on the base decreased significantly during plant development, when expressed per root length, per root fresh weight or per root surface. The number of bacteria on the tip only decreased significantly when expressed per root length. Using the unit of dry weight of adhering soil, contradictory results were obtained for both base and tip; in general, the number of bacteria increased significantly during plant development. Thus, different reference units may lead to different conclusions. Root surface seemed to be the best unit to use, but the use of this unit requires time-consuming measurements. Regression analyses indicated that the reference unit "root surface" was highly correlated with root fresh weight (R ²=93%). Thus, once this relation is determined, the less time-consuming unit can be measured in the experimental work. To analyse the data, the colony-forming units should be expressed per root surface. Besides bacterial numbers during plant development, we assessed whether the bacterial populations collected showed different growth rates on agar plates. The growth rates of bacteria from the tip and base and different development stages of the plants showed differences, indicating differences in the metabolic state of the collected populations. Received: 9 December 1997     

皖南烤烟根际微生物与焦甜香特色风格形成的关系

2006-2007 年对皖南烟区 4 种主要质地土壤冲积粉沙土、河滩沙壤土、冲积沙壤土和水稻土烤烟根际微生物进行了系统研究,发现不同生长期烤烟根际微生物的数量团棵期最少,至旺长期最多,成熟期和采收期逐渐减少.与之相似,根际微生物生物量C(MBC)以团棵期最低,旺长期和成熟期较高,采收期有所下降.在皖南选取具代表性的 19 个土样,包括 7 种不同质地土壤,种植 4 个烤烟品种,分析发现根际 MBC 与有机质和碱解N含量呈极显著的正相关,据此可将土样分为 3 大类型,分别是低 MBC 的相对贫瘠的土壤、中等 MBC 的中等肥力的土壤和高 MBC 的肥沃土壤.焦甜香气较好的烤烟多集中于MBC在 350～380 mg/kg干土的中等肥力的土壤上.表明烤烟旺长期根际 MBC 与烤烟品种无关,而仅与土壤性质有关,焦甜香特色风格的形成与根际 MBC 有密切的关系.     

Effects of marigold (Tagetes sp.) roots on soil microorganisms

 Marigolds (genus Tagetes) suppress populations of soil endopathogenic nematodes such as Pratylenchus penetrans and Meloidogyne species. Nematode suppression by marigolds is thought to be due to thiophenes, heterocyclic sulfur-containing molecules abundant in this plant. When activated, thiophenes such as α-terthienyl produce oxygen radicals. If marigold roots release such a powerful biocidal agent and it is activated in soil, microbial populations in the marigold rhizosphere should be substantially perturbed. We made various measurements of microbial population size and activity in soils that had been cropped to marigolds (Crackerjack, Creole) in the field and in the greenhouse, and compared these with bare soil and soil cropped to rye (Secale cereale L.). Total extractable microbial biomass (measured by the fumigation extraction method), total bacteria (measured by epifluorescence microscopy on 5-(4,6-dichlorotriazine-2-γl) aminofluorescein-stained preparations), heterotrophic bacteria (measured by plate count on various media), and nitrite-oxidizing bacteria (measured by the most-probable-number method) were not significantly different in any of the treatments. Residues of ¹⁴C-labelled rye were mineralized slightly more rapidly in rye-cropped soil than in the other treatments, which were comparable. The rates of die-back of introduced cells of the bacteria Escherichia coli and Rhodococcus TE1 were similar in marigold-cropped and control soils, suggesting that there was not a noteworthy accumulation of biocidal agents in soils cropped to marigolds. We conclude that marigolds do not cause a general depression in the numbers of microorganisms in soils, and that nematode control by this plant may not be due to the release of a biocidal agent into the soil. Received: 4 September 1997     

Plasmid transfer between introduced and indigenous bacteria in leaf litter, soil and vermicompost as affected by soil invertebrates

 Plasmid transfer between introduced strains of Pseudomonas stutzeri JM302 (pLV1013), Pseudomonas putida PAW340 (pLV1017), Pseudomonas aeruginosa PAO5 (RP4), and Enterobacter cloacae MF10 (RP4), all of them harbouring genetically modified or naturally occurring plasmids and bacteria indigenous to oak leaf litter, soddy-podzolic soil or vermicompost was monitored using non-sterile laboratory microcosms inhabited by the millipede Pachyiulus flavipes, the woodlouse Armadillidium vulgare, or the earthworms Aporrectodea caliginosa and Eisenia fetida, respectively. Plasmid transfer from genetically modified Streptococcus faecalis JH2-2 (pAMβ₁) to Bacillus thuringiensis var. israelensis 1-5 under similar conditions was also followed. A recombinant conjugative plasmid (pLV1017) encoding resistance to antibiotics and expressing catechol 2,3-dioxygenase was transferred into two facultatively anaerobic gram-negative bacteria, isolated from the excrement of A. vulgare. Presumed transconjugants of the facultatively anerobic gram-negative bacteria given above were isolated from the leaves and soil (four strains) and the excrement of the invertebrates (11 strains). They carried the same plasmids and exhibited the same enzymatic activity as the respective inoculant strains; for most of them, however, DNA fingerprints were slightly different. Transfer from S. faecalis JH2-2 to B. thuringiensis var. israelensis of a naturally occurring plasmid, pAMb₁, encoding resistance to erythromycin was observed in vermicompost. Received: 18 November 1997     

Nodulation and nitrogen fixation of Lupinus species with Bradyrhizobium (lupin) strains in iron-deficient soil

 The effect of six Bradyrhizobium sp. (lupin) strains (WPBS 3201D, WPBS 3211D, USDA 3040, USDA 3041, USDA 3042 and CB 2272) and Fe supply on nodulation, N₂-fixation and growth of three lupin species (Lupinus termis, L. albus and L. triticale) grown under Fe deficiency in an alkaline soil, were examined in sterilized and non-sterilized pot experiments. When inoculated with USDA 3040, 3041, 3042 and CB2272 without Fe addition, the three lupin species had a very low nodule number and mass, low shoot and root dry matter accumulation and lower N yield. However, inoculation with WPBS 3201D and 3211D without Fe treatments increased all these parameters substantially. The ability of WPBS 3201D and 3211D to form nodules on the three lupin species under conditions of Fe stress could be attributed to their ability to scavenge Fe from Fe-deficient environments through their siderophore production. Addition of Fe to the other four strains significantly increased nodulation and N₂-fixation of the three lupin species, indicating that the poorer nodulation and N₂-fixation of these strains in the absence of Fe, resulted from a low ability to obtain Fe from alkaline soils. Bradyrhizobium strains WPBS 3201D and 3211D were superior to the other four strains in terms of promoting greater nodulation, N₂-fixation, plant growth and N accumulation of L. termis and L. albus. However, the other four strains were more efficient in symbiotic association with L. triticale. The greater variations in nodule efficiencies (specific nitrogenase activity) under different levels of Fe supply could be attributed to the quantities of bacteroid protein and leghaemoglobin in the nodules. The results suggested that Bradyrhizobium (lupin) strains differ greatly in their ability to obtain Fe from alkaline soils, and that the selection of bradyrhizobial strains which are tolerant of Fe deficient soils could complement plant breeding for the selection of legume crops for Fe-deficient soils. Received: 5 January 1998     

Response of nitrogen-transforming microorganisms to arbuscular mycorrhizal fungi

 We studied fluctuations in the numbers of autotrophic ammonium oxidizers, ammonifying microorganisms and denitrifying microorganisms in pot cultures of mycorrhizal and non-mycorrhizal maize. The populations were enumerated after 0, 15, 30, 45 and 60 days of plant growth. Two arbuscular mycorrhizal (AM) fungi belonging to different Glomus species were investigated. Pot cultures with AM-infected maize had significant quantitative and qualitative changes in the root-associated population of N-transforming bacteria compared with the non-mycorrhizal controls. The occurrence of autotrophic ammonium oxidizers in pot cultures of the AM fungi Glomus mosseae and G. fasciculatum was significantly higher than in non-mycorrhizal cultures throughout maize growth. The occurrence of these bacteria was delayed by 15 days in non-mycorrhizal as opposed to Glomus-colonized soil. Ammonifying and denitrifying bacterial populations were significantly decreased in the pot cultures of AM plants compared with the control. The distribution patterns of the physiological groups of bacteria tested were similar for both AM treatments but different from that of the non-mycorrhizal controls. Activity measurements expressed on a per cell basis showed changes with respect to the form of N in the mycorrhizal soil. G. fasciculatum was more active than G. mosseae during the earlier stages of plant growth. Received: 8 July 1997     

Inoculation with phosphate-solubilizing microorganisms and a vesicular-arbuscular mycorrhizal fungus improves dry matter yield and nutrient uptake by wheat grown in a sandy soil

 The effect of inoculating wheat (Triticum aestivum L.) with the PO₄ ^3–-solubilizing microorganisms (PSM) Bacillus circulans and Cladosporium herbarum and the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus sp. 88 with or without Mussoorie rock phosphate (MRP) amendment in a nutrient-deficient natural sandy soil was studied. In the sandy soil of low fertility root colonization by VAM fungi was low. Inoculation with Glomus sp. 88 improved root colonization. At maturity, grain and straw yields as well as N and P uptake improved significantly following inoculation with PSM or the VAM fungus. These increases were higher on combined inoculation of PSM and the VAM fungus with MRP amendment. In general, a larger population of PSM was maintained in the rhizosphere of wheat in treatments with VAM fungal inoculation and MRP amendment. The results suggest that combined inoculation with PSM and a VAM fungus along with MRP amendment can improve crop yields in nutrient-deficient soils. Received: 4 September 1997     

Phosphate solubilizing microorganisms isolated from rhizosphere of maize cultivated in an oxisol of the Brazilian Cerrado Biome

Many soil microorganisms are able to transform insoluble forms of phosphorus to an accessible soluble form, contributing to plant nutrition as plant growth-promoting microorganisms (PGPM). The objective of this work was to isolate, screen and evaluate the phosphate solubilization activity of microorganisms in maize rhizosphere soil to manage soil microbial communities and to select potential microbial inoculants. Forty-five of the best isolates from 371 colonies were isolated from rhizosphere soil of maize grown in an oxisol of the Cerrado Biome with P deficiency. These microorganisms were selected based on the solubilization efficiency of inorganic and organic phosphate sources in a modified Pikovskaya's liquid medium culture containing sodium phytate (phytic acid), soybean lecithin, aluminum phosphate (AlPO₄), and tricalcium phosphate (Ca₃(PO₄)₂). The isolates were identified based on nucleotide sequence data from the 16S ribosomal DNA (rDNA) for bacteria and actinobacteria and internal transcribed spacer (ITS) rDNA for fungi. Bacteria produced the greatest solubilization in medium containing tricalcium phosphate. Strains B17 and B5, identified as Bacillus sp. and Burkholderia sp., respectively, were the most effective, mobilizing 67% and 58.5% of the total P (Ca₃(PO₄)₂) after 10 days, and were isolated from the rhizosphere of the P efficient L3 maize genotype, under P stress. The fungal population was the most effective in solubilizing P sources of aluminum, phytate, and lecithin. A greater diversity of P-solubilizing microorganisms was observed in the rhizosphere of the P efficient maize genotypes suggesting that the P efficiency in these cultivars may be related to the potential to enhance microbial interactions of P-solubilizing microorganisms.     

施氮肥与隔根对间作大豆农艺性状和根际微生物数量及酶活性的影响

采用盆栽试验及种间根系分隔技术研究了施氮肥和不施氮肥两种条件下根系互作在大豆玉米间作中所发挥的优势作用。结果表明,无论在施氮肥和不施氮肥条件下相对于尼龙网分隔和完全分隔根系,未分隔处理的大豆玉米间作能增加大豆各个生育时期的叶绿素含量。施氮肥在改善大豆株高、主茎节数、有效分枝数、单株荚数和百粒重农艺性状方面的效果要大于间作,间作中根系互作只在改善大豆单株荚数和百粒重方面起到显著(p<0.05)作用。通过地下部隔根处理同样证明了大豆玉米间作由于根系的互作能提高大豆的经济产量和生物产量,但效果没有施氮肥明显。施氮肥和间作作物根系的互作不仅可以增加土壤中的细菌、真菌、放线菌和固氮菌的数量,而且可以提高脲酶、磷酸酶、转化酶和蛋白酶的活性。经相关分析得出,该试验中细菌、真菌、放线菌和固氮菌数量和脲酶、磷酸酶、转化酶和蛋白酶活性与大豆的叶绿素含量及大豆单株的经济产量和生物产量之间皆存在正相关关系,且多数之间存在显著(p<0.05)或极显著(p<0.01)正相关。本研究认为间作系统中根系的互作在改善土壤微生态环境,增加土壤微生物数量和酶活性,提高作物经济产量和生物产量方面发挥着重要作用。     

Contribution of termites to the breakdown of straw under Sahelian conditions

 The effect of termites on the breakdown of Pennisetum pedicellatum straw under Sahelian conditions was studied in northern Burkina Faso during the dry and wet seasons of 1995. Litterbags were laid on plots treated with drieldrin at a rate of 500 g a.i. (active ingredient) ha^–1 to exclude termite activity and on untreated plots. Termites were the only macroscopically visible consumers which were observed in the litterbags during the dry and wet season. In litterbags with no termites present the rate of breakdown of straw was lower during both the dry or wet seasons than in litterbags with termites. In the litterbags with termites, much of the mulch was taken off and replaced by termite-made sheeting. From April to September over 60% of the added material disappeared from termite litterbags, while only 18% of the added straw disappeared from litterbags without termites. From October to December, 28% of the straw disappeared from termite-infested litterbags versus 8% from litterbags without termites. The contribution of termites to the breakdown of straw was estimated to be over 70% for each of the two experiments. Under Sahelian conditions, the breakdown of organic residues is strongly influenced by termites. Received: 6 May 1998     

Phosphate-solubilizing rhizobacteria enhance the growth and yield but not phosphorus uptake of canola (Brassica napus L.)

The ability of phosphate-solubilizing rhizobacteria to enhance the growth and phosphorus uptake of canola (Brassica napus L., cv. Legend) was studied in potted soil experiments in the growth chamber. One hundred and eleven bacteria isolated from the rhizosphere of field-grown plants, and a collection of nine bacteria known to be effective plant growth-promoting rhizobacteria (PGPR), were screened for P-solubilization in vitro. All rhizobacteria were identified using whole-cell fatty acids methyl ester (FAME) profiles. The best P-solubilizing isolates were two Bacillus brevis strains, B. megaterium, B. polymyxa, B. sphaericus, B. thuringiensis, and Xanthomonas maltophilia (PGPR strain R85). The P-solubilizers were tested for their effects on growth and P-uptake of canola plants in a P-deficient soil amended with rock phosphate. Although some of the P-solubilizing rhizobacteria significantly increased plant height or pod yield, none increased P-uptake. The most effective inoculant was a B. thuringiensis isolate which significantly increased the number and weight of pods and seed yield without rock phosphate. Xanthomonas maltophilia increased plant height, whereas the other bacilli increased the number on weight of pods. These results demonstrate the potential use of these P-solubilizing rhizobacteria as inoculants for canola, but indicate that P-solubilization was not the main mechanism responsible for positive growth response. Received: 8 February 1996     

Inoculation of rice plants with the endophytic diazotrophs Herbaspirillum seropedicae and Burkholderia spp.

 Four experiments were performed under gnotobiotic conditions to select strains of the endophytic diazotrophs Herbaspirillum seropedicae and Burkholderia spp. as inocula of rice plants. Eighty strains of H. seropedicae originally isolated from rice, sorghum and maize plants, were tested in test tube cultures with N-free agar as the substrate. Rice plants showed medium and high increases in their fresh weight in response to inoculation with nineteen strains. These strains were tested again, and six strains were then selected to evaluate their contribution to the N of the plant via biological N₂ fixation (BNF) using an agar growth medium containing 5 mg N l^–1of ¹⁵N-labelled (NH₄)₂SO₄. The contribution of the strains to plant N via BNF varied from 54% when rice plants were inoculated with strain ZAE94, to 31% when strain ZAE67 was used. These results were confirmed in the fourth gnotobiotic experiment, which also included strains of the new N-fixing bacteria belonging to the genus Burkholderia, isolated from rice, as well as a strain of Burkholderia vietnamiensis, isolated from rice rhizosphere. Burkholderia spp. strains showed similar effects to those observed for H. seropedicae strains, while B. vietnamiensis fixed only 19% of plant total N. The best four strains were tested in a pot experiment where pre-germinated, inoculated rice seedlings were grown in soil labelled with ¹⁵N. The results confirmed the gnotobiotic experiments, although the levels of N in the rice plants derived from BNF of the selected H. seropedicae and Burkholderia spp. strains were lower. Nevertheless, there was an increase in N content in grains of inoculated plants, and the results showed that the method used for strain selection is very useful and can be applied to other strains of N₂-fixing bacteria and plants. Received: 4 May 1999     

Influence of microorganisms on Cu speciation in the rhizosphere of forest soils

Our study is one of the first attempts to document the copper (Cu) speciation in the rhizosphere of forest soils and to determine the importance and the influence of the microbial community on Cu speciation. In order to do this, bulk and rhizosphere samples were collected from field sites located close to industrial facilities. The rhizosphere materials were sampled under trembling aspen (Populus tremuloides Michx.) and separated from the bulk soils in the field. A characterization of the microbial populations was made by measuring microbial biomass C and N, urease and dehydrogenase activities. In soil water extracts, chemical properties were also measured, as well as total water-soluble Cu concentration (Cu_WS) and free-ion Cu activity (Cu²⁺). The residual Cu speciation was obtained by modelling, using MINEQL+ 4.5. In all cases, the Cu speciation was dominated by organic forms of Cu, the proportion of which increases with increasing pH. The reverse pH effect was observed for Cu²⁺. Moreover, almost systematically higher concentrations for all Cu variables were reported in the rhizosphere as compared to the bulk soils. The results also showed that microbial variables explained 22% of the distribution of Cu_WS and Cu²⁺ in bulk samples, a proportion that reached 61.5% in rhizospheric samples. In the rhizosphere, relationships between pH, microbial biomass N and Cu²⁺ indicated that microorganisms influenced Cu by modifying the pH of the solution through nitrogen assimilation. Furthermore, links found between urease activity, biomass variables, solid- and liquid-phase organic carbon and Cu_WS suggested that microbial mineralization could partly supply Cu to the solution fraction of the rhizosphere through root decay. This study reveals that microorganisms have a strong influence on Cu speciation in the rhizosphere of forest soils and suggests that a realistic understanding and representation of Cu dynamic in the rhizosphere must take microbial activity into account. Further investigations are needed to identify and establish precisely how microbial processes impact on Cu speciation.