Molecular characterization of a stable antisense chalcone synthase phenotype in strawberry (Fragaria x ananassa)

An octaploid (Fragaria x ananassa cv. Calypso) genotype of strawberry was transformed with an antisense chalcone synthase (CHS) gene construct using a ripening related CHS cDNA from Fragaria x ananassa cv. Elsanta under the control of the constitutive CaMV 35S promoter via Agrobacterium tumefaciens. Out of 25 transgenic lines, nine lines showed a reduction in CHS mRNA accumulation of more than 50% as compared to the untransformed cv. Calypso control. The antisense CHS construct was found to be integrated into the genome, with a copy number ranging from one to four. The pigmentation of the fruit was only affected when less than 5% of the control CHS expression level was detected. A stable antisense phenotype over a period of 4 years was obtained in the primary transgenic lines at a rate of 1:20. As a consequence of the reduced activity of CHS, the levels of anthocyanins, flavonols, and proanthocyanidins were downregulated and precursors of the flavonoid pathway were shunted to the phenylpropanoid pathway leading to highly increased levels of cinnamoyl glucose (520% of control), caffeoyl glucose (816% of control), and feruloyl glucose (1092% of control) as well as p-coumaryl alcohol (363% of control) and p-coumaryl-1-acetate (1079% of control), which occur only as trace components in untransformed control fruits. These results demonstrate that the introduction of an antisense CHS construct in strawberry results in an unpredictable biochemical phenotype, thereby confirming that CHS function is an important regulatory point of substrate flow between the flavonoid and the phenylpropanoid pathways.     

Dynamics of microcrack propagation in hardwood during heat treatment investigated by synchrotron-based X-ray tomographic microscopy

The process of crack propagation in wood during pyrolysis is strongly linked to heterogeneities in its hierarchical porous structure. Fundamental understanding of this process is necessary for the analysis of the behavior of wood structural elements during fire exposure. Synchrotron-based X-ray tomographic microscopy combined with a recently developed laser-based furnace at the TOMCAT beamline of the Swiss Light Source provides a unique opportunity to study the heat-induced propagation of microcracks in hardwood in situ with high spatial and temporal resolutions. In this study, attention was focused on the 3D microstructure of beech and the interconnectivity between morphology and cracking patterns. It is shown that thermal cracks initiate mainly along the ray cells in hardwood and in the junction of seasonal growth layers. There is a clear indication of increased total porosity of the wood due to charring.     

Analysis of genetic diversity and population structure of confectionery sunflower (<Emphasis Type="Italic">Helianthus annuus</Emphasis> L.) native to Iran

Genetic diversity within and among 50 populations of confectionery sunflower (Helianthus annuus L.) collected from different geographical areas of Iran was evaluated by using microsatellite and retrotransposon markers. The number of alleles (Na) in SSR loci ranged from 2 to 3 with an average of 2.1. The polymorphic bands in retrotransposon markers ranged from 7 in locus CR-UR1 to 15 in locus CR-816 with a mean value of 11.33. Herarchical clustering of individuals (50 × 5 = 250) by neighbor joining method in DARwin5 software subdivided them into three groups. Using Bayesian method in the software pakage of Structure, the studied individuals were subdivided into two sub-populations. Principal coordinate analysis revelaed that the two first components explaine 7.86 and 6.16% of the total variance, respectively. Analysis of molecular variance revealed a higher level of genetic variation within (70%) than between (30%) populations. High molecular variation among individuals within population possibly is due to high allogamy nature of the sunflower plant. Low genetic variation observed between populations could be considered as a consequence of genetic equilibrium that has occurred over the long period of cultivation of confectionery sunflower in this area as well as seed exchange among regions. The traditional assumption that selecting genotypes of different geographical origin will maximize the diversity available to a breeding project does not hold in confectionery sunflower.     

Improvement of Tissue Survival of Skin Flaps by 5α-Reductase Inhibitors: Possible Involvement of Nitric Oxide and Inducible Nitric Oxide Synthase

Background:

Skin flap grafting is a popular approach for reconstruction of critical skin and underlying soft tissue injuries. In a previous study, we demonstrated the beneficial effects of two 5α-reductase inhibitors, azelaic acid and finasteride, on tissue survival in a rat model of skin flap grafting. In the current study, we investigated the involvement of nitric oxide and inducible nitric oxide synthase (iNOS) in graft survival mediated by these agents.

Methods:

A number of 42 male rats were randomly allocated into six groups: 1, normal saline topical application; 2, azelaic acid (100 mg/flap); 3, finasteride (1 mg/flap); 4, injection of L-N^G-nitroarginine methyl ester (L-NAME) (i.p., 20 mg/kg); 5, L-NAME (20 mg/kg, i.p.) + azelaic acid (100 mg/flap, topical); 6, L-NAME (20 mg/kg, i.p.) + finasteride (1 mg/flap, topical). Tissue survival, level of nitric oxide, and iNOS expression in groups were measured.

Results:

Our data revealed that azelaic acid and finasteride significantly increased the expression of iNOS protein and nitric oxide (NO) levels in graft tissue (P < 0.05). These increases in iNOS expression and NO level were associated with higher survival of the graft tissue.

Conclusion:

It appears that alterations of the NO metabolism are implicated in the azelaic acid- and finasteride-mediated survival of the skin flaps.Key Words: Finasteride, Azelaic acid, Surgical flaps, Nitric oxide, Nitric oxide synthase Type II     

Effects of cover crops on soil mineral nitrogen and on the yield and nitrogen content of maize

Current agricultural practice favours winter cover crops, which can not only optimize N management in field crop rotation; but also affect subsequent crops. Three field experiments were carried out in Eastern Slovenia to examine the effects of Italian ryegrass (Lolium multiflorum Lam.), winter rape (Brassica napus ssp.oleifera (Metzg.) Sinsk), subclover (Trifolium subterraneum L.), and crimson clover (Trifolium incarnatum L.) as winter cover crops on the mineral N (N_min) content of soil and on the yield and N content of subsequent maize (Zea mays L.), fertilized with 120 kg N ha⁻¹. Italian ryegrass and winter rape decreased soil N_min contents before winter and in spring more than both clovers. In contrast, clovers accumulated significantly higher amounts of N in organic matter and had lower C/N ratios than winter rape and especially Italian ryegrass. In comparison to the control (bare fallow without cover crop), clovers increased the whole above ground maize dry matter yield, maize grain yield and N contents in whole above ground plants and in grain. The yields and N contents of maize following winter rape were on the same level as the control, while yields and N contents of maize following Italian ryegrass were, in two of the experiments, at the same level as the control. The effects of Italian ryegrass on the maize as subsequent crop in the third experiment were markedly negative. Maize in the control treatment exploited N much more efficiently than in treatments with cover crops. Therefore, cover crop N management should be improved, especially with a view to optimizing the timing of net N mineralization in accordance with the N demands of the subsequent crop.     

Comparison of the pathogensis of two isolates of <Emphasis Type="Italic">Besnoitia caprae</Emphasis> in inbred BALB/c mice

This study was performed to evaluate the infectivity of bradyzoites of two Besnoitia caprae isolates, BC-1 and BC-2, to inbred BALB/c mice. Each group of inbred BALB/c mice was inoculated intraperitoneally with 1 × 10³, 1 × 10⁴, 1 × 10⁵, 5 × 10⁵ and 1 × 10⁶ of one of the two isolates of B. caprae bradyzoites. The mice were monitored daily for a period of 40 days for survival. After death of each mice, several passages from its peritoneal washing and tissues were analyzed using ribosomal DNA-specific PCR assay. Marked differences in pathogenicity between the isolates were seen. All the inbred BALB/c mice infected with BC-2 survived but all the mice that were administered with 1 × 0⁵, 5 × 10⁵ and 1 × 10⁶ BC-1 bradyzoites were died within 4–9 days post-infection (DPI). Histopathological examination of the tissues of the dead mice revealed hyperemia and necrosis with presence of mononuclear and polymorphonuclear cell infiltration in myocardium, spleen and intestines together with interstitial pneumonia and peritonitis. All inbred BALB/c mice in the 1 × 10³ and 1 × 10⁴ groups of BC-1 inoculated mice survived and they were euthanized after 40 DPI. Chronic inflammation with infiltration of mononuclear cells was evident in myocardium, spleen, alveolar septa of the lungs of most of the examined tissues with hemorrhagic enteritis in the mice infected with 1x10⁶ bradyzoites. The mice infected with different doses of BC-2 were euthanized after 40 DPI and no lesion was seen in histopathological sections of their organs. All peritoneal washings and examined tissues were PCR positive in BC-1 group. This experiment is the first report to show inbred BALB/c mice as a relevant model for B. caprae and demonstrates that this strain of inbred BALB/c mice is a suitable animal model for biological studies and examination of pathogenesis for this species of Besnoitia. The present findings also provide evidence for significant differences between the two isolates of B. caprae.     

Effects of artemisinin in broiler chickens following chronic oral intake

Artemisinin has been used for centuries to treat malaria, intestinal tract helminthosis, diarrhea, and used as an antipyretic and sedative agent, but the usage in veterinary medicine is a new field. Recently, it has been used successfully to control experimental poultry coccidiosis. The present study aimed to determine the effects of different doses of artemisinin in broiler chickens with chronic usage. Sixty birds divided into one control and four treatment groups that fed rations mixed with artemisinin at doses of 17, 34, 68, and 136 ppm for 36 days. During the experiment, birds showed no clinical signs except anemia. In microscopic examinations, heart, lung, and spleen had no lesion, but liver, kidney, and brain showed various lesions. Degenerative lesions like intracytoplasmic eosinophilic inclusions were seen in both kidney and liver but fatty change was seen only in liver. There was no relationship between severity of the liver lesions and drug dosage. Central chromatolysis, scattered neuronal necrosis, and mild spongy changes were observed in five regions of the brain that were chosen for sectioning (motor cortex, cerebellar nuclei, midbrain nuclei, and hindbrain nuclei at two separate levels). Severity of lesions in brain was dose-dependent, and cerebral cortex was the most vulnerable area. Haematologic tests showed lower values for hematocrit and red blood cell count dose-dependently. In conclusion, artemisinin is a promising drug for prevention and control of coccidiosis in broiler chickens and its side effects are not too much serious especially at therapeutic doses.     

Assessment of chronic gastritis in pet dogs and its relation with helicobacter-like organisms

The aim of this study was to evaluate the prevalence of chronic gastritis in pet dogs, to determine the histopathologic changes of gastric mucosa and, to determine its relationship with canine gastric Helicobacter infection. Sixty percent (n = 18), 27% (n = 8) and 13% (n = 4) of the examined stomachs showed normal, congested and erosive gastric mucosa respectively. Histopathologic examination was confirmed the presence of chronic gastritis in 40% of dogs (n = 12). Lymphocytic-plasmacytic gastritis was the most common type of chronic gastritis. Gastric Helicobacter was detected in cytological examination of 26 out of 30 dogs (86.6%) but in the PCR analysis, 93% of gastric samples were positive for GHLO. There was no significant relation between the presence of Helicobacters and chronic gastritis (p>0.05). Follicular gastritis was detected in 12 cases (40%) and there was also no significant correlation between its presence and GHLO's infection (p>0.05). In conclusion, chronic gastritis can be considered as a prevalent disease especially in dogs. Nutritional and environmental factors as well as individual immune response may have role in induction of chronic gastritis, but the clinical significance of these histopathologic changes should be evaluated.     

MR relaxivity measurement of iron oxide nano-particles for MR lymphography applications

The aim of this study was to assess the T1, T2 and T2* relaxivity of Ultrasmall Super Paramagnetic Iron Oxide (USPIO) nano-particles in vitro and in vivo in rat models with magnetic resonance imaging at 1.5T. First, relaxation properties of USPIO nano-particles at different doses were measured using related SE and GRE MR imaging protocols. The relation between dose and relaxation were observed which is linear; Higher dose of the nano-particles means higher relaxivity. Based on this relation, an optimum protocol can be proposed for obtaining the best image contrast at each situation. Then detection ability of MRI protocols was studied for USPIO nano-particles with injection of the particles in the rat. The optimum MR protocols were used to observe the signal change of lymph nodes in rat.