The challenge of improving nitrogen use efficiency in crop plants: towards a more central role for genetic variability and quantitative genetics within integrated approaches

In this review, recent developments and future prospects of obtaining a better understanding of the regulation of nitrogen use efficiency in the main crop species cultivated in the world are presented. In these crops, an increased knowledge of the regulatory mechanisms controlling plant nitrogen economy is vital for improving nitrogen use efficiency and for reducing excessive input of fertilizers, while maintaining an acceptable yield. Using plants grown under agronomic conditions at low and high nitrogen fertilization regimes, it is now possible to develop whole-plant physiological studies combined with gene, protein, and metabolite profiling to build up a comprehensive picture depicting the different steps of nitrogen uptake, assimilation, and recycling to the final deposition in the seed. A critical overview is provided on how understanding of the physiological and molecular controls of N assimilation under varying environmental conditions in crops has been improved through the use of combined approaches, mainly based on whole-plant physiology, quantitative genetics, and forward and reverse genetics approaches. Current knowledge and prospects for future agronomic development and application for breeding crops adapted to lower fertilizer input are explored, taking into account the world economic and environmental constraints in the next century.     

Effect of Substrate and Cell Surface Hydrophobicity on Phosphate Utilization in Bacteria

We measured the rates of utilization of hydrophobic and hydrophilic phosphate compounds in gram-negative bacteria with different surface hydrophobicities, isolated from wetland habitats. Three hydrophobic and two hydrophilic bacterial species were selected for study by measuring cell adherence to hydrocarbons. The bacteria were grown under phosphorus-limited conditions with P(infi), hydrophilic (beta)-glycerophosphate, or hydrophobic phosphatidic acid as the phosphate source. Hydrophilic bacteria grew most rapidly on P(infi), followed by (beta)-glycerophosphate. Phosphatidic acid did not support growth or did so at a much later time (40 h) than did the other phosphate treatments. Although all hydrophobic species grew well on these substrates, the rate of growth of two Acinetobacter baumannii isolates on phosphatidic acid exceeded the rate of growth on phosphate or (beta)-glycerophosphate. A membrane phospholipid and lipopolysaccharide were used as a source of phosphorus by hydrophobic species, whereas hydrophilic species could not use the membrane phospholipids and used lipopolysaccharide to a lesser extent. Besides hydrophobic interaction between cells and substrate, phosphatase activity, which was cell bound in hydrophilic species but 30 to 50% unbound in hydrophobic species, affected cell growth. Dialyzed culture supernatant containing phosphatase from hydrophobic species increased the phosphate availability to hydrophilic species. Additionally, cellular extracts from a hydrophilic species, when added to hydrophilic cells, permitted growth on hydrophobic phosphate sources. Naturally occurring amphiphilic humic acids affected the utilization of P(infi) and (beta)-glycerophosphate in bacteria with hydrophilic surfaces but did not affect hydrophobic bacteria. Our results indicate that hydrophobic phosphate sources can be used by bacteria isolated from aquatic environments as the sole phosphorus source for growth. This utilization, in part, appears to be related to cell surface hydrophobicity and extracellular enzyme production.     

Fluorometric analysis of iodinated aflatoxin in minicultures ofAspergillus flavus andAspergillus parasiticus

Summary A convenient miniassay for aflatoxin has been developed for cultures ofAspergillus flavus andA. parasiticus grown for 3–10 days in 10 ml of a coconut extract medium. The sensitivity of the assay, as measured by photofluorometry (365 nm maximum excitation; 445 nm maximum emission), is of the order of 0.01 M (3.12 ng/ml) for aflatoxin B₁ dissolved in aqueous iodine (0.26 mM). High performance liquid chromatography, monitored by fluorometric analysis of both an aflatoxin B₁ standard and selected culture filtrates, confirmed the sensitivity of the assay and indicated specificity for iodine-enhanced fluorescence of aflatoxin in the coconut extract medium. Thin layer chromatography further confirmed the aflatoxin titers and the specificity for enhancement of aflatoxins B₁ and G₁ in culture filtrates.Alabama Agricultural Experiment Station Journal No. 6-871297.     

A sight and video record of the oilbird, Steatornis caripensis , in Peruvian lowland Amazonia

This paper reports on the observation of an oilbird, Steatornis caripensis, in Peruvian lowland Amazonia in September 2006, and discusses this sighting in relation to other lowland reports of oilbirds. Taken together, available information indicates that oilbirds seem to occur in lowland Amazonia more regularly than previously known.     

Isolation and analysis of the gene encoding peripheral myelin protein zero

We have isolated the gene encoding the Schwann cell glycoprotein P0, the major structural protein of the peripheral myelin sheath. In rats and mice, this gene is split into six exons distributed over 7 kb of DNA. The segregation of these exons is consistent with the functional segregation of the P0 protein into extracellular, membrane-spanning, and cytoplasmic domains. We find that the P0 extracellular domain is similar in structure to a single immunoglobulin variable region domain. In contrast to prototypical immunoglobulin domains, however, this P0 domain is encoded by two exons, the partitioning of which provides genetic evidence for the evolution of immunoglobulin-related domains from an ancestral half-domain. We also describe procedures for transfection of cultures of nontransformed rat Schwann cells and use these procedures to show that the Schwann cell-specific expression of the P0 gene is controlled by cis-acting elements localized upstream of exon I.     

Pupae survival following fire in the frosted elfin (Callophrys irus)

1. The frosted elfin (Callophrys irus) butterfly inhabits landscapes that may be subject to frequent fire to be sustained. Frosted elfins pupate primarily in leaf litter, at the soil surface, or just below it, and may suffer high mortality rates when fires occur. Gathering better information on this source of mortality is critical to planning prescribed fire operations in a manner conducive to the long-term survival of the species.
2. We buried lab-reared frosted elfin pupae (n = 61) at 0.75 cm (n = 31) or 1.75 cm (n = 30) below the ground and conducted two experimental fires that mimicked typical prescribed fires.
3. Eighteen of 30 (60%) buried at 1.75 cm survived 4 weeks postburn; no pupae buried at 0.75 cm survived. Most (n = 17) of the pupae that survived successfully enclosed the following year. Surviving pupae encountered lower maximum temperatures and were exposed to shorter durations of above-lethal temperatures compared to those that died.
4. Our data demonstrate that high mortality rates can be expected due to fire, yet fire remains a critical tool for maintaining the habitat. Fire practitioners should mitigate losses by using ignition patterns and suboptimal burn conditions to reduce fire intensity, or burn in a mosaic pattern across the landscape to ensure enough survival to perpetuate frosted elfin populations.

     

Online deployment of an O-PLS model for dielectric spectroscopy-based inline monitoring of viable cell concentrations in Chinese hamster ovary cell perfusion cultivations

Viable cell concentration (VCC) is an essential parameter that is required to support the efficient cultivation of mammalian cells. Although commonly determined using at-line or off-line analytics, in-line capacitance measurements represent a suitable alternative method for the determination of VCC. In addition, these latter efforts are complimentary with the Food and Drug Administration's initiative for process analytical technologies (PATs). However, current applications for online determination of the VCC often rely on single frequency measurements and corresponding linear regression models. It has been reported that this may be insufficient for application at all stages of a mammalian cell culture processes due to changes in multiple cell parameters over time. Alternatively, dielectric spectroscopy, measuring capacitance at multiple frequencies, in combination with multivariate mathematical models, has proven to be more robust. However, this has only been applied for retrospective data analysis. Here, we present the implementation of an O-PLS model for the online processing of multifrequency capacitance signals and the on-the-fly integration of the models’ VCC results into a supervisory control and data acquisition (SCADA) system commonly used for cultivation observation and control. This system was evaluated using a Chinese hamster ovary (CHO) cell perfusion process.     

High resolution multicolor-banding: a new technique for refined FISH analysis of human chromosomes.

A new multicolor-banding technique has been developed which allows the differentiation of chromosome region specific areas at the band level. This technique is based on the use of differently labeled overlapping microdissection libraries. The changing fluorescence intensity ratios along the chromosomes are used to assign different pseudo-colors to specific chromosome regions. The multicolor banding of human chromosome 5 is presented as an example.     

Industrial mycology and the new genetics

Summary The genetic investigation of fungi has been extended substantially by DNA-mediated transformation, providing a supplement to more conventional genetic approaches based upon sexual and parasexual processes. Initial transformation studies with the yeastSaccharomyces cerevisiae provided the model for transformation systems in other fungi with regard to methodology, vector construction and selection strategies. There are, however, certain differences betweenS. cerevisiae and filamentous fungi with regard to type of genomic insertion and the availability of shuttle vectors. Single-site linked insertions are common in yeast due to the high level of homology required for recombination between vectored and genomic sequences, whereas mycelial fungi often show a high frequency of heterologous and unlinked insertions, often in the form of random and multiple-site integrations. While extrachromosomally-maintained or replicative vectors are readily available for use with yeasts, such vectors have been difficult to construct for use with filamentous fungi. The development of vectors for replicative transformation with these fungi awaits further study. It is proposed that replicative vectors may be inherently less efficient for use with mycelial fungi relative to yeasts, since the mycelium, as an extended and semicontinuous network of cells, may delimit an adequate diffusion of the vector carrying the selectable gene, thus leading to a high frequency of abortive or unstable transformants.Based on the Charles Thom Award Lecture presented to the Society for Industrial Microbiology on August 4, 1994.