Zhongmin Dong

Managing soil microorganisms to improve productivity of agro-ecosystems

Historically, agricultural production has relied on practices designed to manage nutrients, water, weeds, and crop diseases. Precision agriculture and integrated pest management programs have gone one step further by recognizing the need to target inputs where they are required in the field. The major objective of these programs has been to minimize adverse environmental impacts of intensive agriculture practices and reduce per unit production costs. This review surveys the literature, examining the manipulation of microbial (primarily bacterial) populations as linked to agricultural production, and discusses new approaches that involve the precision management of microorganisms in the agro-ecosystem. It is proposed that our understanding of plant–soil interactions can be greatly refined through the development of “smart” field technology, where real-time, computer-controlled electronic diagnostic devices can be used to monitor rhizosphere and plant health. We submit that “smart field” generated information could be used to develop a prescription for timely and low-level production interventions that will avoid the traditional inundative approaches to crop maintenance and soil husbandry. Consequently, a lesser impact on the agricultural soil environment is envisioned. The maximization of production efficiencies will also involve the development of crop cultivars that are bred specifically to capitalize on beneficial plant–microbial associations.

A Nitrogen-Fixing Endophyte of Sugarcane Stems (A New Role for the Apoplast)

The intercellular spaces of sugarcane (Saccharum officinarum L.) stem parenchyma are filled with solution (determined by cryoscanning microscopy), which can be removed aseptically by centrifugation. It contained 12% sucrose (Suc; pH 5.5.) and yielded pure cultures of an acid-producing bacterium (approximately 104 bacteria/mL extracted fluid) on N-poor medium containing 10% Suc (pH 5.5). This bacterium was identical with the type culture of Acetobacter diazotrophicus, a recently discovered N2-fixing bacterium specific to sugarcane, with respect to nine biochemical and morphological characteristics, including acetylene reduction in air. Similar bacteria were observed in situ in the intercellular spaces. This demonstrates the presence of an N2-fixing endophyte living in apoplastic fluid of plant tissue and also that the fluid approximates the composition of the endophytess optimal culture medium. The apoplastic fluid occupied 3% of the stem volume; this approximates 3 tons of fluid/ha of the crop. This endogenous culture broth consisting of substrate and N2-fixing bacteria may be enough volume to account for earlier reports that some cultivars of sugarcane are independent of N fertilizers. It is suggested that genetic manipulation of apoplastic fluid composition may facilitate the establishment of similar symbioses with endophytic bacteria in other crop plants.

A putative new endophytic nitrogen-fixing bacterium Pantoea sp. from sugarcane.

Aims:  To isolate and identify endophytic nitrogen‐fixing bacteria in sugarcane growing in Cuba without chemical fertilizers.

H2 oxidation, O2 uptake and CO2 fixation in hydrogen treated soils

In many legume nodules, the H2 produced as a byproduct of N2 fixation diffuses out of the nodule and is consumed by the soil. To study the fate of this H2 in soil, a H2 treatment system was developed that provided a 300 cm3 sample of a soil:silica sand (2:1) mixture with a H2 exposure rate (147 nmol H2 cm−3hr−1) similar to that calculated exist in soils located within 1–4 cm of nodules (30–254 nmol H2 cm−3hr−1). After 3 weeks of H2 pretreatment, the treated soils had a Km and Vmax for H2 uptake (1028 ppm and 836 nmol cm−3 hr−1, respectively) much greater than that of control, air-treated soil (40.2 ppm and 4.35 nmol cm−3 hr−1, respectively). In the H2 treated soils, O2, CO2 and H2 exchange rates were measured simultaneously in the presence of various pH2. With increasing pH2, a 5-fold increase was observed in O2 uptake, and CO2 evolution declined such that net CO2 fixation was observed in treatments of 680 ppm H2 or more. At the H2 exposure rate used to pretreat the soil, 60% of the electrons from H2 were passed to O2, and 40% were used to support CO2 fixation. The effect of H2 on the energy and C metabolism of soil may account for the well-known effect of legumes in promoting soil C deposition.

RAPD polymorphisms in spring wheat cultivars and lines with different level of Fusarium resistance

Abstract. Random amplified polymorphic DNA (RAPD) markers have been used to characterize the genetic diversity among 35 spring wheat cultivars and lines with different levels of Fusarium resistance. The objectives of this study were to determine RAPD-based genetic similarity between accessions and to derive associations between Fusarium head blight (FHB) and RAPD markers. Two bulked DNA from either highly resistant lines or susceptible lines were used to screen polymorphic primers. Out of 160 screened primers, 17 primers generated reproducible and polymorphic fragments. Genetic similarity calculated from the RAPD data ranged from 0.64 to 0.98. A dendrogram was prepared on the basis of a similarity matrix using the UPGMA algorithm, which corresponded well with the results of principal component analysis and separated the 35 genotypes into two groups. Association analysis between RAPD markers and the FHB index detected three RAPD markers, H191000, F2500 and B12400, significantly associated with FHB-resistant genotypes. These results suggest that a collection of unrelated genotypes can be used to identify markers linked to an agronomically important quantitative trait like FHB. These markers will be useful for marker-assistant breeding and can be used as candidate markers for further gene mapping and cloning.

Microbial nature of the hydrogen-oxidizing agent in hydrogen-treated soil

Abstract. Experiments have shown that legume soil can promote non-legume plant growth. Although generally attributed to N fertilization of the soil, there are other factors that are additionally responsible. Work has been done in an attempt to characterize this response; fungal agents have been implicated. Other studies have shown H2 to be responsible for plant growth promotion. This work attempts to understand the nature of the H2 uptake agent in soil. The H2 uptake ability of H2-treated soil disappeared when the soil was autoclaved or when glucose was added, suggesting the biotic nature of the H2 uptake agent. Physical disturbance reduced the H2 uptake ability of the soil, indicating the special colonial or long filamentous structure of the H2 uptake agent in soil. The fact that the addition of fungicides did not, in most cases, significantly affect the H2 uptake ability of the soil excluded the fungi as the H2 uptake microorganisms in soil. Addition of antibiotics significantly affected the H2 uptake ability of the soil suggesting the H2 uptake agent in soil is bacterial. The H2 uptake ability of soil was greatly reduced by extremely high concentrations of benomyl and a low concentration of penicillin, indicating that actinomycetes might be the active H2 uptake bacteria in H2-treated soils.

Effect of hydrogen on soil bacterial community structure in two soils as determined by terminal restriction fragment length polymorphism

The metabolism of hydrogen evolved from HUP− legume nodules can alter bacterial community structures in the rhizosphere. Our earlier experiments demonstrated increased hydrogen uptake and appearance of white spots within bacterial colonies in H2-treated soil. We were also able to isolate hydrogen-oxidizing bacteria from soil samples exposed to hydrogen, but not from samples exposed to air. To further understand the effect of hydrogen metabolism on soil microbial communities, in this study 16S rRNA terminal restriction fragment (TRF) profiles of different soil samples exposed to hydrogen gas under laboratory, greenhouse, and field conditions were analyzed. Relationships between soil bacterial community structures from hydrogen-treated soil samples and controls, illustrated by UPGMA (unpaired group mathematical averages) dendrograms, indicated a significant contribution of hydrogen metabolism to the variation in bacterial community. The intensity variation of TRF peaks includes both hydrogen-utilizing bacteria, whose growth were stimulated by hydrogen exposure, and other bacterial species whose growth was inhibited. Comparison of TRF profiles between laboratory and greenhouse samples showed that T-RFLP is a useful technique in the detection of root-related effects on soil bacterial community structure.

Soybean nodule hydrogen metabolism affects soil hydrogen uptake and growth of rotation crops

To test the beneficial effect on the following crop of hydrogen released by Hup− soybean nodules, soybean was inoculated with either a Hup− (JH47) or a Hup+ (JH) strain of Bradyrhizobium japonicum. These isogenic strains differ only in that JH47 has a Tn5 inserted in the gene coding for the small hydrogenase subunit which eliminates hydrogenase activity; thus when present in soybean nodules, hydrogen is released into the rhizosphere. Inoculated alfalfa plants were used as the positive control as no hydrogenase activity has ever been found in alfalfa nodules. Soil adjacent to hydrogen releasing (Hup−strain) legume nodules had a significantly higher hydrogen uptake rate than that around the nodules containing the Hup+ strain. Barley grown following soybean inoculated with the Hup− strain exhibited an increased grain yield under field conditions. Key words: Soil, hydrogen oxidization, rotation benefit

Retention of vacuole contents of plant cells during fixation

Changes in the semi‐permeability of tonoplast during fixation were studied using beetroot tissue. Using vacuole betacyanin as the indicator, the permeability of the tonoplast was assessed by the leakage of this pigment as determined by changes in the optical density of the solution bathing the tissue. Cryo‐analytical scanning electron microscopy was used to monitor the changes in ion concentration in the cells during fixation. Fixatives were 3% glutaraldehyde or 4% formaldehyde in 0·025 m phosphate buffer at room temperature or on ice. Results showed that glutaraldehyde, especially at low temperature, takes as long as 30 h to disrupt the semi‐permeability of the tonoplast of beetroot cells, while in formaldehyde on ice, these cells begin to lose their selective permeability in 15 min. This study confirms that the semi‐permeability of the tonoplast may not be lost until long after the cytoplasm has been fixed and suggests that this explains why cold fixation in 3% glutaraldehyde for about 12 h has become the most reliable standard procedure for the successful preservation of vacuolated plant cells.