Phenotypic and genotypic variation in Iranian sour and duke cherries

Phenotypic and genotypic variation and structure of 29 sour cherry (P. cerasus) and duke cherry (P. x gondouinii) genotypes from different regions of Iran were identified using random amplified polymorphic DNA (RAPD) markers and morphological characters. Furthermore, one Prunus mahaleb genotype was used as an outgroup for molecular analysis. For morphological analysis, 23 variables were recorded to detect similarities between and among studied sour and duke cherries. Most studied characteristics were showing a high degree of variability. Principal component analysis showed that the first three components explained a total of 73.87 % of the whole phenotypic variability. Based on the morphological cluster analysis, studied sour and duke cherry genotypes were placed into three main clusters. The first main cluster included 16 sour cherry genotypes. The second main cluster contained all duke cherry genotypes and eight sour cherry genotypes, while, only one sour cherry genotype was placed in third main cluster. For RAPD analysis, 17 primers generated a total of 233 discernible and reproducible bands across genotypes analyzed, out of which 214 (91.51 %) were polymorphic with varied band size from 300 to 3000 bp. According to the similarity matrix, the lowest similarity was obtained between P. mahaleb, as an outgroup, and sour cherry. Dendrogram based on molecular data separated genotypes according to their species and geographic origin. Low correlation was observed between the similarity matrices obtained based on morphological and RAPD data. The information obtained here could be valuable for devising strategies for conservation of Iranian sour and duke cherries.     

Amperometric sulfide detection using Coprinus cinereus peroxidase immobilized on screen printed electrode in an enzyme inhibition based biosensor

In the present work, an amperometric inhibition biosensor for the determination of sulfide has been fabricated by immobilizing Coprinus cinereus peroxidase (CIP) on the surface of screen printed electrode (SPE). Chitosan/acrylamide was applied for immobilization of peroxidase on the working electrode. The amperometric measurement was performed at an applied potential of -150 mV versus Ag/AgCl with a scan rate of 100 mV in the presence of hydroquinone as electron mediator and 0.1M phosphate buffer solution of pH 6.5. The variables influencing the performance of sensor including the amount of substrate, mediator concentration and electrolyte pH were optimized. The determination of sulfide can be achieved in a linear range of 1.09-16.3 μM with a detection limit of 0.3 μM. Developed sensor showed quicker response to sulfide compared to the previous developed sulfide biosensors. Common anions and cations in environmental water did not interfere with sulfide detection by the developed biosensor. Cyanide interference on the enzyme inhibition caused 43.25% error in the calibration assay which is less than the amounts reported by previous studies. Because of high sensitivity and the low-cost of SPE, this inhibition biosensor can be successfully used for analysis of environmental water samples.     

Endocrine profiles of somatic nuclear transfer-derived pregnancies in dairy cattle

In cattle, several hormones and proteins are necessary for maintenance of a normal pregnancy that will result in a viable calf. Deviation from the normal cascade or expected profile of reproductive hormones and proteins may be associated with impairment of somatic nuclear transfer-derived pregnancies and the high rate of fetal loss. The objectives of this study were to characterize maternal plasma concentrations of pregnancy-specific protein B (PSPB), progesterone (P4), estrone sulphate (E₁S), and estradiol (E2) during the last two-thirds of pregnancy (cloned calves), and to determine associations with gestational abnormalities. Cows with cloned fetuses, produced by either commercial (N = 16) or zona-free (N = 4) cloning techniques, were compared with pregnant animals derived from traditional embryo transfer (N = 6) or AI (N = 6), at various stages of gestation (Days 80, 120, 150, 180, 210, and 240; Day 0 = estrus). Fetal well-being was monitored with ultrasonography throughout gestation. At Day 80, progesterone concentration was lower (P < 0.0001) in nuclear transfer (NT) recipients than in control groups. Mean estrone sulphate concentrations did not vary significantly between NT and control groups. At Day 150, pregnancy-specific protein B concentrations were elevated (P < 0.002) in NT cows. Estradiol concentration was higher in NT recipients than control cows throughout the study period.     

Ultrasonographic and histological characterization of the placenta of somatic nuclear transfer-derived pregnancies in dairy cattle

The high incidence of pregnancy loss and prenatal morbidity and mortality in cloned animals may be due to placental insufficiency, thereby compromising fetal survival. Our objective was to characterize morphological changes in fetal membranes of cloned bovine pregnancies. Two groups of cows with cloned fetuses, produced by two cloning techniques, a commercial group (n=16) and a hand-made group (n=4), and control fetuses derived from traditional embryo transfer (n=6) or AI (n=6), were compared at various stages of gestation (Days 80, 120, 150, 180, 210, and 240; Day 0=estrus). Thickness and shape of the amniotic membrane, placentome shape and length, umbilical cord shape and diameter, and fetal fluid echodensities were assessed by ultrasonography, and the placenta was evaluated histologically. Only eight (40%) of cloned pregnancies reached term and seven calves (35%) were alive at birth. Both placentome length and umbilical cord diameter were larger (P<0.05) in clones than in normal fetuses at all stages of gestation. Amniotic membrane abnormalities (Day 120) including focal edema and the presence of a series of nodules were detected in 38% of the clones and were always accompanied by hyper-echodense spikes or irregularities (detected ultrasonographically) around the umbilical cord. Histopathology revealed degenerate inflammatory cells, edematous chorioallantoic membranes, and decreased epithelial thickness. We inferred that these morphological anomalies of placentomes compromised fetal development, and we concluded that ultrasonographic monitoring of pregnancies enabled characterization of changes in the placentae and may be useful to assess fetal well-being.     

Role of Jasmonic and Salicylic Acid on Enzymatic Changes in the Root of Two Alyssum inflatum Náyr. Populations Exposed to Nickel Toxicity

Phytohormones, including salicylic acid (SA) and jasmonic acid (JA) have the potential to ameliorate plant development and tolerance to deleterious effects of toxic metals like nickel (Ni). Therefore, the current study was carried out to evaluate SA and JA's interactive effect on the root antioxidative response of two Alyssum inflatum Nyár. populations against Ni-toxicity. Two A. inflatum species under different Ni concentrations (0, 100, 200, and 400 μM) were exposed to alone or combined levels of SA (0, 50, and 200 μM) and JA (0, 5, and 10 μM) treatments. Results showed that high Ni concentration (400 μM) reduced roots fresh weight in both populations than in control. However, external application of individual SA and JA or combined SA?+?JA in higher doses had ameliorated roots biomass by mitigating Ni-toxicity, especially in the NM population, in comparison to 400 μM Ni. Under Ni toxicity, SA and JA, especially their combination, induced high Ni accumulation in plants' roots. Moreover, the application of SA and JA alone, as well as combined SA?+?JA, was found to be effective in the scavenging of hydrogen peroxide by improving the activity of superoxide dismutase, peroxidase, catalase, and ascorbate peroxidase in both populations under Ni-toxicity. Overall, our results manifest that SA and JA's external use, especially combined SA?+?JA treatments, ameliorate root biomass and plant tolerance by restricting Ni translocation to the shoot, accumulating in roots, and enhancing antioxidant defense systems.

     

An improved cage system for the bioassay of Metarhizium anisopliae on Thrips tabaci (Thysanoptera: Thripidae)

The ability of thrips and other minute insects to escape from conventional assay cups led to development of a successful assay chamber. In the present study, we evaluated three bioassay systems for thrips; Tashiro cage, microtube assay and leaf sandwich assay and introduced an improved thrips entomopathogenic bioassay system (TEBS). Our objective was to investigate the effect of Metarhizium anisopliae on Thrips tabaci using an easy-to-use and inexpensive holding chamber that would produce bioassay results with acceptable levels of accuracy and precision. In our assays, we found that escape of second-instar larvae from TEBS was significantly less than those of other methods which we tested. Our results indicated that our assay system was more suitable than the other conventional assays for the bioassay of entomopathogenic fungi on T. tabaci.     

Bioelectricity generation enhancement in a dual chamber microbial fuel cell under cathodic enzyme catalyzed dye decolorization

Enzymatic decolorization of reactive blue 221 (RB221) using laccase was investigated in a dual-chamber microbial fuel cell (MFC). Suspended laccase was used in the cathode chamber in the absence of any mediators in order to decolorize RB221 and also improve oxygen reduction reaction in the cathode. Molasses was utilized as low cost and high strength energy source in the anode chamber. The capability of MFC for simultaneous molasses and dye removal was investigated. A decolorization efficiency of 87% was achieved in the cathode chamber and 84% COD removal for molasses was observed in the anode chamber. Laccase could catalyze the removal of RB221 and had positive effect on MFC performance as well. Maximum power density increased about 30% when enzymatic decolorization was performed in the cathode chamber.     

Immunochemical properties of a 60 kDa cell surface-associated heat shock-protein (Hsp60) from Helicobacter pylori

Abstract Western blot analysis (immunoblotting) of cell surface-associated proteins from Helicobacter pylori confirmed our previous findings that binding of human IgG is a common property (among H. pylori strains). Purification of the IgG-binding proteins (IGBP) was achieved by two purification steps, affinity chromatography on IgG-Sepharose and nickel chelate affinity chromatography. SDS-PAGE and immunoblotting analysis revealed a 60 kDa protein with affinity for peroxidase labeled human IgG. Solid phase binding assays showed that IgG binds to an immobilized protein (IGBP). The 60 kDa IGBP binds human IgG₁, IgG₃ and IgM. Binding could be inhibited by the kappa chain of the human IgG, but not with its Fc fragment, nor with IgA or IgM. In addition, rabbit polyclonal antibodies raised against the 60 kDa IGBP blocked IgG binding. Monoclonal antibodies, specific to the Hsp60 heat shock protein of H. pylori recognized the 60 kDa IGBP as revealed by immunoblotting analysis, both in crude preparations and in the purified fractions.     

Cryptic domains of a 60 kDa heat shock protein of Helicobacter pylori bound to bovine lactoferrin

Abstract Bovine lactoferrin binds to a 60 kDa heat shock protein of Helicobacter pylori . Binding ability was related to human immunoglobulin G because bovine lactoferrin binding proteins were isolated by extraction of cell surface associated proteins with distilled water, applied on IgG-Sepharose and nickel sulphate chelate affinity chromatography. Binding was demonstrated by Western blot after purified protein was digested with α-chymotrypsin and incubated with peroxidase-labeled bovine lactoferrin. Binding was inhibited by bovine lactoferrin, lactose, rhamnose, galactose, and two iron-containing proteins, ferritin and haptoglobin. Helicobacter pylori binds ferritin and haptoglobin via charge or hydrophobic interactions because this binding was not inhibited by specific and various glycoproteins or carbohydrates. Carbohydrate moieties of bovine lactoferrin molecules seem to be involved in binding because glycoproteins with similar carbohydrate structures strongly inhibited binding. Scatchard plot analysis of the binding of peroxidase-labeled bovine lactoferrin to H. pylori cells yielded a k _d 2.88 × 10⁻⁶ M. In addition, binding of H. pylori cells to bovine lactoferrin was enhanced when bacteria treated with pepsin or α-chymotrypsin after isolation from iron-restricted and iron-containing media.