Electrophoretic Investigations on Nematode Resistant Sugar Beets

Hybridizations were made between beta vulgaris and three wild species of the patellares section being resistant to the best cyst nematode {heterodera schachtii). Monosomic addition lines (2 n = 19) with full nematode resistance were investigated together with wild beets by means of electrophoretical techniques. One alkaline esterase band and a complex of several acidic esterase bands were localized on the resistance-carrying B. procumbens chromosome. The alkaline esterase marker also appeared in B. patellaris addition lines. An aconitase double band was visible in two of four B. webbiana addition lines. One resistant monotelosomic addition line with a small B. procumbens fragment had lost the esterase gene. Evidence is given that more than one chromosome is carrying genes for nematode resistance. The use of electrophoretic screening together with a nematode testing program is discussed.     

Using engineered scaffold interactions to reshape MAP kinase pathway signaling dynamics

Scaffold proteins link signaling molecules into linear pathways by physically assembling them into complexes. Scaffolds may also have a higher-order role as signal-processing hubs, serving as the target of feedback loops that optimize signaling amplitude and timing. We demonstrate that the Ste5 scaffold protein can be used as a platform to systematically reshape output of the yeast mating MAP kinase pathway. We constructed synthetic positive- and negative-feedback loops by dynamically regulating recruitment of pathway modulators to an artificial binding site on Ste5. These engineered circuits yielded diverse behaviors: ultrasensitive dose response, accelerated or delayed response times, and tunable adaptation. Protein scaffolds provide a flexible platform for reprogramming cellular responses and could be exploited to engineer cells with novel therapeutic and biotechnological functions.     

Intragastric gavage with denatonium benzoate acutely induces neuronal activation in the solitary tract nucleus via the vagal afferent pathway

Natural toxic substances have a bitter taste and their ingestion sends signals to the brain leading to aversive oral sensations. In the present study, we investigated chronological changes in c-Fos immunoreactivity in the nucleus tractus solitarius (NTS) to study the bitter taste reaction time of neurons in the NTS. Equal volumes (0.5 mL) of denatonium benzoate (DB), a bitter tastant, or its vehicle (distilled water) were administered to rats intragastrically. The rats were sacrificed at 0, 0.5, 1, 2, 4, 8, or 16 h after treatment. In the vehicle-treated group, the number of c-Fos-positive nuclei started to increase 0.5 h after treatment and peaked 2 h after gavage. In contrast, the number of c-Fos-positive nuclei in the DB-treated group significantly increased 1 h after gavage. Thereafter, the number of c-Fos immunoreactive nuclei decreased over time. The number of c-Fos immunoreactive nuclei in the NTS was also increased in a dose-dependent manner 1 h after gavage. Subdiaphragmatic vagotomy significantly decreased DB-induced neuronal activation in the NTS. These results suggest that intragastric DB increases neuronal c-Fos expression in the NTS 1 h after gavage and this effect is mediated by vagal afferent fibers.     

Sterol dynamics of heterotrophic Tetraselmis suecica and its nutritional implication in the bivalve aquaculture

Employment of heterotrophic culture of microalgal foods in the hatchery‐based seed production of oysters is still controversial because some algae produced using the method appear to loose sterols, a key nutritional factor for bivalve growth. We traced the changes in sterol content of Tetraselmis suecica growing under photoautotrophic and heterotrophic conditions with the aid of gas chromatography (GC) and GC–mass spectrometry. The photoautotrophic T. suecica at the mid‐logarithmic growth phase contained six major (cholesta‐5,22‐dien‐3β‐ol, ergost‐5‐en‐3β‐ol, cholest‐5‐en‐3β‐ol, 24‐methylcholesta‐5,22‐dien‐3β‐ol, 24‐methylcholesta‐5,24‐dien‐3β‐ol, and 24‐ethylchlolesta‐5,24‐dien‐3β‐ol) and two minor sterols (24‐methylcholesta‐5‐en‐3β‐ol and 24‐ethylchlolesta‐5en‐3β‐ol). In the comparison of algal growth and sterol level, photoautotrophic alga appeared to need higher amounts of major sterols for cell growth over heterotrophic alga. These findings, that heterotrophic alga needed less amount of sterols for growth, may have significant implications in the introduction of the method in bivalve hatcheries. We also reviewed the sterolic properties of the alga obtained from heterotrophic method with respect to bivalve aquaculture.     

The production and characterization of monoclonal antibodies against Photobacterium damselae ssp. piscicida and initial observations using immunohistochemistry

Five monoclonal antibodies (MAbs: F2B1, 1E4, 13B10, 4D4 and F3G12) were produced against lysed Photobacterium damselae ssp. piscicida ( Ph. d . ssp. piscicida ). The MAbs recognized three antigens of differing molecular weight on the Western blot of Ph. d . ssp. piscicida . They also cross-reacted with five different species of Vibrio . An enzyme linked immunosorbent assay (ELISA) with MAbs, F3G12 and 4D4 demonstrated differences between Ph. d . ssp. piscicida and three Ph. d . ssp. damselae type strains, indicating differences in the surface antigenicity between these two groups of bacteria. Antigen retrieval in conjunction with immunohistochemistry (IHC) using MAb 13B10, revealed colonies of bacteria in the kidney, spleen and liver of sea bass, Dicentrarchus labrax , infected with pasteurellosis. A number of positive colonies were observed around the mucosal layers of the intestinal tissue, especially within the lamina propria. In addition, a number of bacterial colonies were associated with red blood cells and blood vessels of the organs examined.     

DNA fingerprinting: application to conservation of the CITES-listed dragon fish, Scleropages formosus (Osteoglossidae)

Since 1975,CITES has listed the dragon fish, Scleropages formosus, as anendangered species. In 1995, a captive-bred population was set upby a commercial fish farm with assistance from the PrimaryProduction Department in Singapore. Other farms in Indonesia andMalaysia followed suit. These populations have contributed to animmediate conservation of the species. Due to very high demandfor this ornamental fish, these venues may be its last sanctuary.DNA fingerprints of the dragon fish were obtained by different methods from the green, red and gold varieties grown in a Singapore fish farm to determine which method was most suitable in providing information on genetic variability. Because a DNA fingerprint is a pattern made up of DNA fragments that are resolved by electrophoresis, each individual has its own unique fingerprint due to a genetic make-up different from another individual. Thus, genetic variability was best studied by developing DNA fingerprints.Firstly, restriction fragment length polymorphisms (RFLPs) were obtained. DNA fragments formed by cleavage with nine restriction endonucleases used singly were hybridized individually to four non-radioactively labelled probes to give RFLPs. The RFLPs for each variety were similar and genomic DNA from each variety had many binding sites to the probes. This made differentiating RFLPs specific to individual varieties difficult. Secondly, random amplified polymorphic DNA (RAPD) fingerprints were developed. DNA fragments that were resolved on a denaturing polyacrylamide gel were hybridized to seven arbitrary primers used singly. RAPD fingerprints for each variety were different for each primer tested. The similarity index indicated low genetic variability between varieties. Lastly, DNA was screened for microsatellite loci which refer to short tandem repeats of two or three bases. The occurrence of other microsatellite loci, their chromosome location and frequency is being investigated while primers have been designed to detect more loci by the polymerase chain reaction. As this method provides undisputed and reproducible evidence of relatedness and stock identification, and can be applied for long-term management of domesticated populations through pedigree construction and evaluation of heterozygosity, it is the preferred choice to determine genetic variability     

Effect of natural free gossypol and gossypol‐acetic acid on growth performance and resistance of channel catfish (Ictalurus punctatus) to Edwardsiella ictaluri challenge

A study was conducted to evaluate the effect of free gossypol from glanded‐cottonseed meal (G‐CSM) (natural free gossypol) or gossypol‐acetic acid on growth performance, body composition, haematology, immune response and resistance of channel catfish (Ictalurus punctatus) to Edwardsiella ictaluri challenge. Soya bean meal‐based diets supplemented with 0, 100, 200, 400, and 800 mg kg^?1 free gossypol from G‐CSM or gossypol‐acetic acid were fed to juvenile channel catfish in triplicate aquaria to apparent satiation twice daily for 12 weeks. Neither sources nor levels of dietary gossypol significantly influenced the final weight gain, feed intake, feed efficiency and survival of channel catfish. Similarly, whole‐body proximate composition, haematological parameters (red blood cell, white blood cell counts, haemoglobin and haematocrit), serum protein concentration, macrophage chemotaxis ratio, phagocytic activity and antibody production against E. ictaluri 21‐day postinfection were not significantly affected at either dietary sources or levels of gossypol. Gossypol concentrations of liver were linearly related to dietary level of gossypol but the retention rate varied dependent on sources of the dietary gossypol. At dietary gossypol levels of 400 or 800 mg kg^?1, total gossypol concentrations in liver of fish fed dietary gossypol from G‐CSM were significantly higher than those of fish fed the corresponding levels of gossypol from gossypol‐acetic acid. The (+)‐isomer of gossypol was predominantly retained in liver regardless of dietary sources of gossypol. The ratio of (+) to (?) gossypol isomers in liver decreased with increasing dietary concentrations of gossypol. Serum lysozyme activity of fish fed dietary gossypol levels of 200 mg kg^?1 or higher, either from G‐CSM or gossypol‐acetic acid, was significantly higher than that of the control. At a level of 800 mg kg^?1 diet, gossypol from G‐CSM stimulated significantly higher lysozyme activity than gossypol from gossypol‐acetic acid. Fish fed diets containing 400 mg kg^?1 gossypol or higher from G‐CSM or 800 mg kg^?1 gossypol from gossypol‐acetic acid had significantly increased superoxide anion (O) production. However, neither the sources nor the levels of dietary free gossypol influenced the resistance of juvenile channel catfish to E. ictaluri challenge.