Nitric oxide effectively curtails neck bending and mitigates senescence in isolated flowers of Calendula officinalis L.

In recent years, there has been a considerable and renewed upsurge in research to ascertain the physiological and biochemical role of Nitric oxide (NO) in plants. The present investigation is focused to study the role of NO on neck bending associated with senescence and postharvest performance in isolated flowers of Calendula officinalis. The flower buds harvested at one day before anthesis stage were supplied with sodium nitroprusside (SNP) as a source of NO at different concentrations viz., 50, 100, 150 and 200 µM. A distinct set of flowers held in distilled water designated the control. The investigation revealed that SNP delayed the senescence in flowers of C. officinalis significantly manifested by prolonged longevity. The maximum longevity of 12 days was recorded in flowers supplemented with 100 µM SNP. The flowers held in distilled water (control) displayed early senescence symptoms and lasted for 6 days only. Our research suggested that improved flower longevity by SNP was commensurate with delayed neck bending, inhibition of bacterial growth in the vase, increased solution uptake, high membrane stability, besides an up-regulated activities of antioxidant enzymes in the tissue samples. In addition, the treated flowers exhibited increased content of  sugar fractions, total phenols and soluble proteins in the petal tissues compared to control. Further, 100 µM SNP was observed as most effective treatment and increased the longevity of flowers by 6 days. The concentration above 150 µM provoked early senescence compared to control, whereas concentration lower than 100 µM was less efficacious in improving the postharvest life and longevity of cut Calendula flowers.     

Organic biopolymers of venus clams: Collagen-related matrix in the bivalve shells with crossed-lamellar ultrastructure

BackgroundBiochemical studies and spectroscopic techniques have shown that chitin-silk fibroins are common in nacroprismatic bivalve shells. However, the nature of organic biopolymers in the less well studied shell architectures, such as crossed lamellar shells, remain unknown. Here, two venus shells, Callista disrupta and Callista kingii, with crossed lamellar ultrastructure have been studied.MethodsWe employed thermal gravimetric analysis, optical-, confocal- and scanning electron-microscopes, gel-sodium dodecyl sulfate (gel-SDS), FTIR, ultra-performance liquid chromatography and high-performance anion-exchange chromatography system with pulsed amperometric detection to analyse organic macromolecules in the shells.ResultsThermal analysis showed a low concentration of organic macromolecules in C. disrupta (1.38 wt%) and in C. kingii (1.71 wt%). A combination of biochemical protocols, including Calcofluor White staining and FTIR spectroscopic assessment, indicate that amino-polysaccharide chitin together with proteins, are present in the organic scaffolding of the shells. Scanning electron microscope of insoluble acid biopolymer extracts as well as FTIR technique show that the hierarchical structural organizations of organic biopolymers consist collagen-related matrix. Our histochemical fixing and staining techniques reveal many discrete proteins and glycoproteins from soluble organic macromolecules on the gel-SDS. We show here ‘singlet’ and ‘doublet’ glycosaminoglycan bands that are far above 260 kDa.General significance/conclusionsThe presence of collagen matrix in Callista shells shows promise for the new source of biomaterials. Most importantly, the structural organization of the proteinaceous motif is predominantly helical structures and not silk-fibroin unlike in nacreous bivalve shells.     

Redox potentials, laccase oxidation, and antilarval activities of substituted phenols

Laccases are copper-containing oxidases that are involved in sclerotization of the cuticle of mosquitoes and other insects. Oxidation of exogenous compounds by insect laccases may have the potential to produce reactive species toxic to insects. We investigated two classes of substituted phenolic compounds, halogenated di- and trihydroxybenzenes and substituted di-tert-butylphenols, on redox potential, oxidation by laccase and effects on mosquito larval growth. An inverse correlation between the oxidation potentials and laccase activity of halogenated hydroxybenzenes was found. Substituted di-tert-butylphenols however were found to impact mosquito larval growth and survival. In particular, 2,4-di-tert-butyl-6-(3-methyl-2-butenyl)phenol (15) caused greater than 98% mortality of Anophelesgambiae larvae in a concentration of 180nM, whereas 2-(3,5-di-tert-butyl-4-hydroxyphenyl)-2-methylpropanal oxime (13) and 6,8-di-tert-butyl-2,2-dimethyl-3,4-dihydro-2H-chromene (33) caused 93% and 92% mortalities in concentrations of 3.4 and 3.7μM, respectively. Larvae treated with di-tert-butylphenolic compounds died just before pupation.     

Protective effect of selenium on protein-undernutrition-induced brain damage in rats

The effect of ad libitum ingestion of selenium (Se) in drinking water (0.15 mg SeO₂/L) for 3 wk on the brain weight, total brain protein, glutathione (GSH) level, catalase activity, and lipid peroxidation in the brain of protein-undernourished (PU) rats was investigated, in an attempt to determine whether antioxidants alone can reverse some of the neuropathological changes associated with protein undernutrition in rats. Feeding on a normal diet (16% casein) by well-fed rats or a low-protein diet (5% casein) by PU rats and Se-treated PU rats lasted 14 wk. Setreated PU rats were given Se in drinking water during the last 3 wk of the experiment. Results show that protein undernutrition induced significant reductions (p<0.001) in brain weight, total brain protein, and catalase activity (p<0.05) while it induced a significant increase (p<0.05) in lipid peroxidation when compared with well-nourished rats; but no significant effect was observed for the GSH level. However, the ingestion of Se in drinking water by PU rats for 3 wk resulted in significant increases (p<0.05) in brain weight, catalase activity, and total brain protein but induced a significant reduction (p<0.05) in lipid peroxidation when compared with PU rats given water. The values obtained for Setreated PU rats are comparable with those obtained for well-nourished rats. The GSH level was, however, not affected by Se ingestion. We suggest that Se, by inducing increases in the concentration of certain proteins, including catalase, in the brain, abolished some of the pathological changes associated with protein undermutrition in the brain, and appears as a promising antioxidant in the prevention and management of pro-oxidant-induced brain damage.     

An improved protocol for coupling synthetic peptides to carrier proteins for antibody production using DMF to solubilize peptides.

We present an improved protocol for coupling synthetic peptides to carrier proteins. In this protocol, dimethyl-formamide is used as the solvent to solubilize peptides instead of phosphate-buffered saline (PBS) or 6 M guanidine-HCl/0.01 M phosphate buffer (pH 7). Additionally, the last desalting or dialyzing step to remove uncoupled peptides as in the traditional method is eliminated. Finally, 3 ml of 0.1 M ammonium bicarbonate is added to the carrier protein conjugated peptide solution to help the lyophilization process. Coupling of Cys-containing synthetic peptides to keyhole limpet hemocyanin or bovine serum albumin using m-maleimidobenzoyl-N-hydroxysuccinimide ester are used as the test cases. This method produces high-quality antipeptide antibodies. Also, compared to the traditional method, this procedure is simpler and useful for peptides with solubility problems in PBS or 6 M guanidine-HCl.     

Activin-A, but not inhibin, regulates 17β-hydroxysteroid dehydrogenase type 1 activity and expression in cultured rat granulosa cells

17β-Hydroxysteroid dehydrogenase type 1 (17HSD type 1) catalyzes the reduction of estrone (E₁) to biologically more active estradiol (E₂). In the present study, the effect of activin, inhibin, and follistatin on 17HSD activity and 17HSD type 1 expression in cultured, unluteinized rat granulosa cells was examined. Furthermore, the effects of these hormones on 17HSD type 1 expression were compared with the expression of P450 aromatase (P450arom). Rat granulosa cells were pre-incubated in serum-free media for 3 days, followed by a 2-day treatment with activin, inhibin, follistatin and 8-Br-cAMP. Activin in increasing concentrations appeared to effect a dose-dependent increase in 17HSD activity. In addition, increasing concentrations of activin also increased 17HSD type 1 mRNA expression. Addition of 8-Br-cAMP at concentrations of 0.25 and 1.5 mmol/l together with activin significantly augmented the stimulatory effects of activin alone in the cultured cells. Neither inhibin, nor follistatin, either alone or in combination with 8-Br-cAMP, had any notable effects on 17HSD activity and 17HSD type 1 expression. Preincubation of activin with increasing concentrations of follistatin significantly diminished the stimulatory effect of activin. In the presence of follistatin, activin did not significantly increase the 8-Br-cAMP-induced 17HSD activity and 17HSD type 1 expression. The culturing of granulosa cells in the presence or the absence of inhibin or follistatin with or without 8-Br-cAMP did not alter the effect of these peptides on P450arom expression in rat granulosa cells as judged by Northern blot analysis of total RNA. However, cAMP-induced P450arom expression was enhanced by activin treatment, except when follistatin was present. This is in line with the suggested role of follistatin as an activin-binding protein, which limits the bioavailability of activin to its membrane receptors. Thus, the results support the notion of a paracrine/autocrine role of activin in follicular steroidogenesis of growing follicles.     

Black flies and Onchocerciasis: Knowledge,attitude and practices among inhabitants of Alabameta,Osun State,Southwestern, Nigeria

Background and objectivesThis study reports knowledge of residents of Alabameta community, Osun State, Nigeria on the bioecology and socio-economic burden of black flies and onchocerciasis.MethodsUsing structured questionnaires and Focus Group Discussion (FGD), a total of 150 community respondents participated in the study.ResultsThe knowledge of the residents on the existence of black flies in the community was significant (p<0.05) as all the 150 respondents confirmed the presence of black flies with the local name ‘Amukuru’ i.e causing itching. However, their lack of knowledge of the flies breeding site (104) (69%), prevention (134) (89%), cause (132) (88%), and treatment (133) (89%) of onchocerciasis was profound. Majority 147(98%) of the respondents reported that flies bite more in the wet season as against dry season 3(2%) and have a higher affinity (124) (82%) for biting the leg than any other part of the body. A larger percentage (89%) of the respondents are unaware of any medication for the treatment of onchocerciasis while 11% are aware. There had been no sensitization on onchocerciasis according to 89% of the respondents.ConclusionDue to lack of resident’s knowledge on black flies bioecology which may continuously expose them to the bite of the flies and ultimately infection, it is paramount that the Osun State government and the NTD implementing partner map out new public health education strategies during routine Mass Administration of Medicines with Ivermectin with a view to preventing onchocerciasis infection as well as man-vector contact.     

The chemokine‐binding protein encoded by the poxvirus orf virus inhibits recruitment of dendritic cells to sites of skin inflammation and migration to peripheral lymph nodes

Orf virus (ORFV) is a zoonotic parapoxvirus that induces acute pustular skin lesions in sheep and humans. ORFV can reinfect its host and the discovery of several secreted immune modulatory factors that include a chemokine‐binding protein (CBP) may explain this phenomenon. Dendritic cells (DC) are professional antigen presenting cells that induce adaptive immunity and their recruitment to sites of infection in skin and migration to peripheral lymph nodes is critically dependent on inflammatory and constitutive chemokine gradients respectively. Here we examined whether ORFV‐CBP could disable these gradients using mouse models. Previously we established that ORFV‐CBP bound murine inflammatory chemokines with high affinity and here we show that this binding spectrum extends to constitutive chemokines CCL19 and CCL21. Using cell‐based chemotaxis assays, ORFV‐CBP inhibited the movement of both immature and mature DC in response to these inflammatory and constitutive chemokines respectively. Moreover in C57BL/6 mice, intradermally injected CBP potently inhibited the recruitment of blood‐derived DC to lipopolysaccharide‐induced sites of skin inflammation and inhibited the migration of ex vivo CpG‐activated DC to inguinal lymph nodes. Finally we showed that ORFV‐CBP completely inhibited T responsiveness in the inguinal lymph nodes using intradermally injected DC pulsed with ovalbumin peptide and transfused transgenic T cells.     

Cairo-East Bank effluent re-use study 3 — Effect of field crop irrigation with secondary treated wastewater on biological and chemical properties of soil and groundwater

Large scale field trials were conducted in fertile soil and desert (virgin) soil to evaluate the effect of irrigation with secondary treated wastewater from two wastewater treatment plants in Cairo on biological and chemical properties of soil and groundwater. Soil samples were taken for physical and chemical analysis after crop harvest. Groundwater monitoring wells were installed in and around the experimental soil sites. Considerable amounts of macronutrients (NPK) were applied to the grown crops through the treated wastewater irrigation: N (19–79%), P (23–181%) and K (85–357%) of the recommended fertilizer rates according to the crop and the experimental site. Soil physical and chemical analysis showed that both soils are widely variable in water holding capacity, organic matter, pH value, CaCO₃, salinity, cation exchange capacity, and soil bulk density in the topsoil (0–30 cm). Groundwater samples which were examined for the presence of pathogenic bacteria (salmonella), faecal coliform bacteria and helminth ova. At the virgin soil site, 10–57% of the samples from each well contained salmonella, whereas at the fertile soil, salmonella was not detected in five wells, and the occurrence in the other four wells was 10–20% of the samples. The numbers of faecal coliforms were similar at both sites, in the range 10²–10³ MPN per 100 mL. Small numbers of parasite ova were also found in the majority of wells, with a greater number occurring at the virgin soil. The groundwater in both sites was similar and of poor quality, and would be unsuitable for potable or irrigation purposes.