Extreme resistance to cucumber mosaic virus (CMV) in transgenic tomato expressing one or two viral coat proteins

For the production of broad commercial resistance to cucumber mosaic virus (CMV) infection, tomato plants were transformed with a combination of two coat protein (CP) genes, representing both subgroups of CMV. The CP genes were cloned from the CMV-D strain and Italian CMV isolates (CMV-22 of subgroup I and CMV-PG of subgroup II) which have been shown to produce severe disease symptoms. Four plant transformation vectors were constructed: pMON18774 and pMON18775 (CMV-D CP), pMON18831 (CMV-PG CP) and pMON18833 (CMV-22 CP and CMV-PG CP). Transformed R0 plants were produced and lines were selected based on the combination of three traits: CMV CP expression at the R0 stage, resistance to CMV (subgroup I and/or II) infection in growth chamber tests in R1 expressing plants, and single transgene copy, based on R1 segregation. The results indicate that all four vector constructs generated plants with extremely high resistant to CMV infection. The single and double gene vector construct produced plants with broad resistance against strains of CMV from both subgroups I and II at high frequency. The engineered resistance is of practical value and will be applied for major Italian tomato varieties. This revised version was published online in June 2006 with corrections to the Cover Date.     

Cephalexin synthesis by immobilised penicillin G acylase under non-isothermal conditions: reduction of diffusion limitation

The effect of thermodialysis on the enzymatic kinetic synthesis of the antibiotic cephalexin was investigated. As reference points, two existing models for an immobilised enzyme (Assemblase^®) and for the free enzyme were used. For Assemblase^®, it is known that diffusion limitation occurs and that therefore considerably more of the undesired side-product phenylglycine is formed.

The enzyme was immobilised on a membrane, and under isothermal conditions (293 K) the course of the reaction resembled that of the Assemblase^® enzyme. However, if a temperature gradient was applied across the membrane, with an average temperature of 293 K for the enzyme, than the course of the reaction changed. For large temperature gradients (30° and more), the course of the reaction resembled that of free enzyme. Thermodialysis enhances mass transfer across the membrane and therewith reduces diffusion limitations in the immobilised enzyme on the membrane.

The stability of the immobilised enzyme is such that the reactor can be re-used repeatedly. This, together with the positive effect of the temperature gradient on the course of the reaction, makes thermodialysis an interesting new technique that has potential to be applied on a larger scale if the membrane surface area per volume of reactor can be improved.     

Inter-strain competition and dispersal in aphids: evidence from a greenhouse study

1. Crops are often colonised by aphids having different life-history traits, and the aphids and their offspring compete for the shared resource. The intraspecific competition and dispersal characteristics of two strains (A and B) of the cotton aphid Aphis gossypii on cucumber were examined in a greenhouse study. Strain A normally feeds on cucumber, whereas strain B originated from melon but develops on cucumber. Both strains reproduce exclusively via parthenogenesis and can be discriminated using molecular markers. 2. The strains were released at the same time on different plants in a greenhouse. Total density and their dispersion within and between plants were recorded through time, and the frequency of each strain was assessed using DNA fingerprinting. The Lotka–Volterra competition model was used to test for inter-strain competition and to estimate its intensity. 3. Strain A had the highest rate of increase and carrying capacity, and became the most frequent strain, nearly displacing strain B. Strain B was the most common only on the plants it infested first. The competition was unbalanced as strain B was affected strongly by strain A, but strain A was only affected minimally by strain B. This asymmetrical relationship for strain B was not due only to competition for the shared resource. 4. Such competition may in part explain the reduction of genetic polymorphism observed by others in the field where colonising strains coming from different hosts compete and some are eliminated. Polymorphism in A. gossypii populations in nature is thought to be maintained by heterogeneous patches of hosts differentially favourable for different clones.     

Resistance of transgenic tomato to cucumber mosaic cucumovirus under field conditions

Since the summer of 1993, transgenic tomato plants expressing the coat protein (CP) genes of cucumber mosaic cucumovirus have been tested under field conditions to assess the level of resistance and agronomic performance. Trials were performed in different areas in Italy and the target virus in the majority of tests was spread naturally by the indigenous aphid populations. Twenty-three homozygous lines of variety UC82B, transformed to contain four different CP genes of CMV, were evaluated. The lines were preselected for CP expression, single gene copy, and virus resistance in growth chamber experiments. In general, CMV resistance was confirmed under field conditions though resistance in the field was less effective than what was observed in growth chamber experiments. The resistance observed in multi-year and multi-location experiments is of commercial value for several of the most resistant lines. Engineered resistance upon transfer to Italian varieties by breeding or direct transformation will be used in tomato production in Italy or elsewhere. This revised version was published online in June 2006 with corrections to the Cover Date.     

Effects of the Extraction of Internal Organs of the Holothurian Cucumaria japonica on the Indices of Nonspecific Resistance

The experiments performed on livestock showed that a tissue preparation from the internal organs of the Japanese sea cucumber stimulates bactericide, lysozyme, beta-lysine, and phagocytic activities of the blood in adult cows and newborn calves.     

Structural Analysis of A-Protein of Cucumber Green Mottle Mosaic Virus and Tobacco Mosaic Virus by Synchrotron Small-Angle X-Ray Scattering

The size and shape of A-protein of tobacco mosaic virus coat protein (TMVP) and cucumber green mottle mosaic virus coat protein (CGMMVP) were evaluated by means of small-angle X-ray scattering (SAXS) using a synchrotron radiation source, complemeted by electron microscopic observations. The results imply that TMV and CGMMV A-proteins are composed of three and two subunits, respectively, stacked in the shape of an isosceles triangular prism at lower ionic strength. Considering the difference of the A-protein structure at higher and lower ionic strength, the globular core structure was proposed as a subunit which might be modeled as a thin isosceles triangular prism composed of four globular cores joined by rather flexible segments. These cores correspond probably to four helical regions in a subunit, and rearrange their relative positions according to the external conditions. A slight rearrangement of core positions in a subunit may result in the formation of A-proteins of various shapes.     

Physiological and cytochemical studies on activator calcium in contraction by smooth muscle of a sea cucumber,Isostichopus badionotus

Summary The physiological properties of mechanical responses and the intracellular localization and translocation of calcium as a pyroantimonate precipitate were studied in the longitudinal retractor muscle (LRM) of a Bermuda sea cucumber. Acetylcholine (ACh)-induced contraction was reduced by lowering the external Ca concentration, and suppressed completely by prolonged soaking in Ca-free solution. The magnitude of ACh-induced contraction was decreased by Mn and La ions. Furthermore, procaine reduced the ACh-induced contraction. The complete removal of Ca and Mg ions from the external medium induced a socalled Ca · Mg-removal contraction. Electron microscopically, numerous subsarcolemmal vesicles were observed in the LRM fibers. In the resting fibers, pyroantimonate precipitates were localized in the subsarcolemmal vesicles and along the inner surface of plasma membrane. While, in the fiber fixed during mechanical activity, the pyroantimonate precipitates were decreased remarkably in the subsarcolemmal vesicles and at the plasma membrane, and diffusely distributed in the myoplasm. Electronprobe X-ray microanalysis showed that the precipitate contains Ca in a significant amount. These results indicate that the contraction of the LRM fibers is caused not only by Ca-influx but also by Ca-release from the intracellular storage sites, such as the subsarcolemmal vesicles and the inner surface of plasma membrane.     

Heterorhabditis sp. (Nematoda: Heterorhabditidae): A Nematode Parasite Isolated from the Banded Cucumber Beetle Diabrotica balteata

A nematode identified as Heterorhabditis sp. was discovered in June 1982 in larval cadavers of the banded cucumber beetle, Diabrotica balteata, in soil on wooded land. Effective beetle control (over 95%) was obtained when larvae were exposed to potted soil containing infective stage nematode juveniles or infected larval cadavers. The nematode was propagated in vivo on larvae of D. balteata, Diaphania nitidalis (the pickleworm), and Galleria mellonella (the greater wax moth). This Heterorhabditis sp. has promising potential as a biocontrol agent for the banded cucumber beetle.