Gregory E. Welbaum

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.

BIOPHYSICAL, PHYSIOLOGICAL AND BIOCHEMICAL PROCESSES REGULATING SEED GERMINATION

In the physiological sense, germination begins with seed water uptake and ends with the initiation of elongation by the embryonic axis, usually the radicle. The driving forces and constraints on expansion by the embryo are examined, particularly for seeds in which the embryo is surrounded by endosperm and testa tissues that restrict growth. Models have been developed to predict germination based on thermal time, hydrotime and combined hydrothermal time. These population-based models indicate that the timing of germination is closely tied to physiologically determined temperature and water potential thresholds for radicle emergence which vary among individual seeds in a population. The restraint imposed by tissues surrounding the radicle is a major determinant of the threshold water potential. Enzymatic weakening of these tissues is a key event regulating the timing of radicle emergence. Considerable evidence suggests that endo-β-mannanase is involved in this process in a number of species, although it is doubtful that it is the sole determinant of when radicle emergence occurs. Molecular and biochemical studies are revealing the complexity of events occurring in endosperm and embryo cells associated with the completion of germination. Unique permeability properties and the presence of enzymes associated with pathogen resistance suggest additional functional roles for the tissues enclosing the embryo. The insights gained from physiology and modelling are being extended by the application of molecular techniques to identify and determine the function of genes expressed in association with germination. Single-seed assay methods, in vivo reporters, specific modification of gene expression and mutagenesis will be critical technologies for advancing our understanding of germination

Changes in spinach phylloepiphytic bacteria communities following minimal processing and refrigerated storage described using pyrosequencing of 16S rRNA amplicons

Aims:  To investigate the changes in bacterial diversity on fresh spinach phyllosphere associated with storage at refrigeration temperatures.

A gel diffusion assay for visualization and quantification of chitinase activity

Higher plants, bacteria, fungi, insects, and crustaceans all produce chitinases. Chitinase genes in many organisms are currently under investigation. Chitinase activity is usually assayed with radiolabeled or fluorogenic substrates. We developed a simple, inexpensive, nonradioactive gel-diffusion assay for chitinase that can be used to screen large numbers of samples. In this assay, chitinase diffuses from a small circular well cut in an agarose or agar gel containing the substrate glycol chitin, a soluble, modified form of chitin. Chitinase catalyzes the cleavage of glycol chitin as it diffuses through the gel, leaving a dark, unstained circular zone around the well, because the fluorescent dye calcofluor binds only to undigested chitin. Sample activities can be determined from linear regression of logstandard enzyme concentration versus the zone diameter of internal standards on each Petri dish used for a diffusion assay.

Seed ultrastructure and water absorption pathway of the root-parasitic plant Phelipanche aegyptiaca (Orobanchaceae).

BACKGROUND AND AIMS Obligate root parasitic plants of the Orobanchaceae do not germinate unless they chemically detect a host plant nearby. Members of this family, like Orobanche, Phelipanche and Striga, are noxious weeds that cause heavy damage to agriculture. In spite of their economic impact, only a few light microscopical studies of their minute seeds have been published, and there is no knowledge of their ultrastructure and of the role each tissue plays during the steps preceding germination. This paper describes the ultrastructure of Phelipanche seeds and contributes to our understanding of seed tissue function. METHODS Seeds of P. aegyptiaca were examined under light, scanning electron, transmission electron and fluorescence microscopy following various fixations and staining protocols. The results were interpreted with physiological data regarding mode of water absorption and germination stimulation. KEY RESULTS AND CONCLUSIONS The endothelium, which is the inner layer of the testa, rapidly absorbs water. Its interconnected cells are filled with mucilage and contain labyrinthine walls, facilitating water accumulation for germination that starts after receiving germination stimuli. Swelling of the endothelium leads to opening of the micropyle. The perisperm cells underneath this opening mediate between the rhizosphere and the embryo and are likely to be the location for the receptors of germination stimuli. The other perisperm cells are loaded with lipids and protein bodies, as are the endosperm and parts of the embryo. In the endosperm, the oil bodies fuse with each other while they are intact in the embryo and perisperm. Plasmodesmata connect the perisperm cells to each other, and the cells near the micropyle tightly surround the emerging seedling. These perisperm cells, and also the proximal embryo cells, have dense cytoplasmic contents, and they seem to represent the two seed components that are actively involved in transfer of reserve nutrients to the developing seedling during germination.

Characterization of chitinase activity and gene expression in muskmelon seeds

Chitinase is often produced in higher plants as a general defence response after wounding or pathogenic attack. Since germinating seeds are exposed to soil pathogens, the activity and expression of chitinase in muskmelon (Cucumis melo L.) seeds was investigated. One acidic and three basic chitinase isoforms were detected, beginning 40 d after anthesis in developing and fully mature seeds. Both acidic and basic chitinase isoforms were found in endosperm tissue during imbibition and after radicle emergence. Basic chitinase isoforms, but not acidic isoforms, were detected in the embryonic axes of imbibed seeds and in seeds before germination, indicating that chitinases are developmentally regulated in specific seed tissues. Two complete cDNAs, Cmchi1 and Cmchi2, were cloned from germinated muskmelon seeds and are predicted to encode chitinases that show 95% identity to a class III chitinase from cucumber (Cucumis sativus L.) and 61% identity to a class II chitinase from soybean (Glycine max L.), respectively. Southern blotting indicated that Cmchi2 was present only once in the muskmelon genome, while Cmchi1 may be present in one or two copies. Cmchi1 and Cmchi2 mRNAs were only detected in radicles of germinating seeds and in roots of mature plants, so additional genes other than Cmchi1 and Cmchi2 must be responsible for the chitinase activity in developing seeds. Salicylic acid and benzothiadiazole stimulated the expression of Cmchi1, but not Cmchi2, after radicle emergence. A putative role for chitinase in muskmelon seeds is defence against fungal pathogens.

Shelf Life Determination of Fresh Blueberries (Vaccinium corymbosum) Stored under Controlled Atmosphere and Ozone

Fresh blueberries are commonly stored and transported by refrigeration in controlled atmospheres to protect shelf life for long periods of storage. Ozone is an antimicrobial gas that can extend shelf life and protect fruit from microbial contamination. Shelf life of fresh highbush blueberries was determined over 10-day storage in isolated cabinets at 4°C or 12°C under different atmosphere conditions, including air (control); 5% O2 : 15% CO2 : 80% N2 (controlled atmosphere storage (CAS)); and ozone gas (O3) 4 ppm at 4°C or 2.5 ppm at 12°C, at high relative humidity (90–95%). Samples were evaluated for yeast and molds growth, weight loss, and firmness. CAS and O3 did not delay or inhibit yeast and molds growth in blueberries after 10 days at both temperatures. Fruit stored at 4°C showed lower weight loss values compared with 12°C. Blueberries stored under O3 atmosphere showed reduced weight loss at 12°C by day 10 and loss of firmness when compared to the other treatments. Low concentrations of ozone gas together with proper refrigeration temperature can help protect fresh blueberries quality during storage.

Survival of Listeria monocytogenes on fresh blueberries (Vaccinium corymbosum) stored under controlled atmosphere and ozone.

Listeria monocytogenes is a foodborne pathogen that represents a high risk for consumers because it can grow under refrigeration conditions and can also develop acid tolerance. Fresh blueberries are hand-picked, packed, and transported under refrigeration without receiving a microbial inactivation treatment. The aim of this work was to study the survival of L. monocytogenes in fresh highbush blueberries stored at 4 or 12 °C under different controlled atmosphere conditions, including air (control); 5% O2, 15% CO2, 80% N2 (controlled atmosphere storage [CAS]); or ozone gas (O3), 4 ppm at 4 °C or 2.5 ppm at 12 °C, at high relative humidity (90 to 95%) for a total of 10 days. Fresh blueberries inside a plastic clamshell were spot inoculated with the bacteria and were stored at 4 or 12 °C in isolated cabinets under air, CAS, and O3 atmospheric conditions. Samples were evaluated on days 0, 1, 4, 7, and 10 for microbial growth using modified Oxford agar. CAS did not delay or inhibit L. monocytogenes growth in fresh blueberries after 10 days. O3 achieved 3- and 2-log reductions when compared with air treatment at 4 and 12 °C, respectively. Low concentrations of O3 together with proper refrigeration temperature can ensure product safety throughout transportation. O3 is a strong antimicrobial that safely decomposes to oxygen and water without leaving residues and can be used as an alternative method to prevent bacterial growth during a long transport period.

Organic broccoli production on transition soils: comparing cover crops, tillage and sidedress N.

Little information is available about how farmers in transition to organic practices should manage short- and long-term N fertility. The objectives of this research were (1) to evaluate the leguminous cover crops lablab ( Dolichos lablab L.), soybean ( Glycine max L.), sunn hemp ( Crotalaria juncea L.) and a mixture of sunn hemp and cowpea ( Vigna sinensis Endl.) as N sources; (2) to compare N availability and broccoli yield when cover crops were incorporated with conventional tillage (CT) or retained as a surface mulch using no-tillage (NT) practices; and (3) to quantify the amount of supplemental sidedress nitrogen required to maximize the yield of organic broccoli ( Brassica oleracea Group Italica) on transition soils. Broccoli was grown during the first year after organic transition in the spring and fall of 2006 at the Kentland Agricultural Research Farm near Blacksburg, VA. Spring ( P P −1 , and showed a quadratic correlation with leaf N ( P = R 2 =0.80 and P = R 2 =0.38, respectively). There was no difference in spring broccoli yield between CT and NT; however, CT produced the highest yield in the fall crop. At low sidedress N rates, leaf N was highest in CT plots, but tillage had no effect on N uptake at high N rates. This indicates that early season and perhaps total plant-available mineralized N was greater in CT than NT; however, potential N deficiency in NT soil may be compensated by sidedress N. Broccoli yield was not affected by leguminous cover crop, even though the quantity of cover crop biomass and N contribution was different among species. This suggests that N availability from leguminous cover crops may be impacted by other ecological processes such as soil microbial activity. This study shows that organic CT and NT growers can maximize broccoli yield in transition soils low in N availability, by using leguminous cover crops in combination with moderate amounts of sidedress N.

Heat Stress Related Physiological and Metabolic Traits in Peanut Seedlings

ABSTRACT To maintain high yields under an increasingly hotter climate, high temperature resilient peanut cultivars would have to be developed. Therefore, the mechanisms of plant response to heat need to be understood. The objective of this study was to explore the physiological and metabolic mechanisms developed by virginia-type peanut at early growth stages in response to high temperature stress. Peanut seedlings were exposed to 40/35 C (heat) and 30/25 C (optimum temperature) in a growth chamber. Membrane injury (MI), the Fv/Fm ratio, and several metabolites were evaluated in eight genotypes at four time-points (day 1, 2, 4, and 7) after the heat stress treatment initiation. Even though we were able to highlight some metabolites, e.g., hydroxyproline, galactinol, and unsaturated fatty acid, explaining specific differential physiological (MI) responses in peanut seedlings, overall our data suggested general stress responses rather than adaptive mechanisms to heat. Rather than individual metabolites, a co...