Optimizing injection time of GFP plasmid into sheep zygote

Microinjection of exogenous DNA into the cytoplasm of matured oocytes or zygotes is a promising technique to generate transgenic animals. However, the data about the microinjection time and procedure in sheep are limited and have not treated in detail. To obtain more in-depth information, the Sarda sheep oocytes from abattoir-derived ovaries were subjected to IVM and IVF. Then, the GFP plasmid as a reporter gene was injected into the cytoplasm of MII oocytes (n: 95) and zygotes at different post-insemination intervals (6–8 hpi, n: 120; 8–10 hpi, n: 122; 10–12 hpi, n: 110 and 12–14 hpi, n: 96). There were no significant differences in the cleavage rates between the groups. However, blastocyst rate of injected zygotes at all-time intervals was significantly lower than injected MII oocytes and control group (p < 0.05). Interestingly, the proportion of GFP-positive embryos was higher at 8–10 hpi compared with other injected groups (4 % versus 0 %, p  < 0.01). Among these, the proportion of mosaic embryos was high and two of those embryos developed to the blastocyst stage. In conclusion, we settled on the cytoplasmic microinjection of GFP plasmid at 8–10 hpi as an optimized time point for the production of transgenic sheep and subsequent experiments.     

Effect of Bacillus thuringiensis var. kurstaki on survival and mortality of immature and mature stages of Diadegma insulare parasitizing Plutella xylostella

Effects of Bacillus thuringiensis (Bt) on immature stages of Diadegma insulare (Cresson) (Hym., Ichneumonidae) within larvae of diamondback moth (DBM), Plutella xylostella (L.) (Lep., Plutellidae), and on the adult stage as a Bt-honey solution were evaluated at 25?±?1?°C, 65?±?10% r.h. and 16:8?h (L:D) photoperiod. P. xylostella and D. insulare pupae were collected from cabbage fields in Karaj, Iran. A commercial wettable powder formulation of Bt (var. kurstaki, serotype H-3a3b, strain Z-52) was used in these experiments. The value of LC₅₀ for third instar larvae of P. xylostella was 210?ppm. Parasitoid??s adult mortality at field rate of Bt was not significantly different from that of control. Formation of parasitoid??s pupae in Bt-parasitoid combined treatments was significantly lower than parasitoid alone. Formation of parasitoid??s pupae in larvae treated with LC₁₀, LC₃₀ and LC₅₀ of Bt and then exposed to parasitoids after 48?h was 5%, 3% and 2%, respectively, which was significantly lower than that of the parasitoid alone (41%). Mean percentage mortality of larvae of DBM in LC₃₀ and LC₅₀ concentrations of Bt combined with the parasitoid was approximately 100%. Our results showed that B. thuringiensis kills D. insulare larvae indirectly by killing susceptible hosts in which they are developing. Successful use of multiple biological control agents will depend on close monitoring of their compatibility in the field. Our results could be useful in this purpose.     

Uptake and remobilization of selenium in Brassica napus L. plants supplied with selenate or selenium‐enriched plant residues

Selenium (Se) biofortification via crops is one of the best strategies to elevate the daily Se intake in areas where soil Se levels are low. However, Se fertilizer recovery (SeFR) is low and most of the Se taken up accumulates in non‐harvested plant parts and returns to the soil with plant residues. A pot experiment with soil was undertaken to study the efficiency of inorganic Se (Na₂SeO₄) and Se‐enriched plant residues for biofortification, as well as to identify the bottlenecks in Se utilization by Brassica napus L. The soil was fertilized with Na₂SeO₄ (0 and 7 µg Se kg^?1) or with Se in stem or leaf residues (0 and 7 µg Se kg^?1). A treatment with autoclaved soil was included (0 and 7 µg kg^?1 as Na₂SeO₄) to unravel the impact of microbial activity on Se uptake. The Se‐enriched plant residues produced a lower Se uptake efficiency (SeUPE) and SeFR than did inorganic Se, and soil autoclaving enhanced Se accumulation in the plants. The time required for decomposition seems to preclude crop residues as an alternative source of Se. Furthermore, B. napus had a limited capacity to accumulate Se in seeds. The study shows that the bottlenecks in Se biofortification appear to be its low bioavailability in soil and poor loading from the silique walls to seeds. Thus, improved Se translocation to seeds would be a useful breeding goal in B. napus to increase SeFR.     

A Bayesian Generalized Linear Model for Crimean–Congo Hemorrhagic Fever Incidents

Global spread of the Crimean–Congo hemorrhagic fever (CCHF) is a fatal viral infection disease found in parts of Africa, Asia, Eastern Europe and Middle East, with a fatality rate of up to 30%. A timely prediction of the prevalence of CCHF incidents is highly desirable, while CCHF incidents often exhibit nonlinearity in both temporal and spatial features. However, the modeling of discrete incidents is not trivial. Moreover, the CCHF incidents are monthly observed in a long period and take a nonlinear pattern over a region at each time point. Hence, the estimation and the data assimilation for incidents require extensive computations. In this paper, using the data augmentation with latent variables, we propose to utilize a dynamically weighted particle filter to take advantage of its population controlling feature in data assimilation. We apply our approach in an analysis of monthly CCHF incidents data collected in Turkey between 2004 and 2012. The results indicate that CCHF incidents are higher at Northern Central Turkey during summer and that some beforehand interventions to stop the propagation are recommendable. Supplementary materials accompanying this paper appear on-line.     

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.     

Larval feeding behaviour of angel fish Pterophyllum scalare (Cichlidae) fed copepod Eucyclops serrulatus and cladoceran Ceriodaphnia quadrangula

Three different live diets, Eucyclops serrulatus, Ceriodaphnia quadrangula and equal combination of E. serrulatus copepodid and C. quadrangula, were offered to angelfish (Pterophyllum scalare) larvae viz 1‐week, 2‐week and 3‐week old at prey densities of 2, 5 and 10 individuals mL^?1. Results showed that 1‐week‐old P. scalare larvae consumed E. serrulatus copepodid at a rate of 31.3–56.7 ind. h^?1, C. quadrangula at 8.0–12.0 ind. h^?1, and mixture of E. serrulatus and C. quadrangula at 20.7–40.7 ind. h^?1. For 2‐week‐ and 3‐week‐old larvae, consumption rate increased accordingly. The electivity indices (E) of P. scalare (1‐week‐old larvae) for E. serrulatus copepodid were +0.18, +0.23 and +0.22 at prey densities of 2, 5 and 10 ind. mL^?1 respectively. Tendency towards E. serrulatus copepodid consumption reduced by aging P. scalare as indicated by the E values for 2‐ and 3‐week‐old larvae. However, growth and survival of P. scalare larvae was greatest when fed on combination of copepod E. serrulatus and C. quadrangula.     

Study of infection with an Iranian field-isolated H9N2 avian influenza virus in vaccinated and unvaccinated Japanese quail

In the present study, we examined the mortality rate, egg production, and clinical signs of quail experimentally infected with a field isolate of A/Chicken/Iran/339/02 (H9N2) avian influenza virus obtained from an infected commercial layer farm with severe morbidity and mortality. A total of 120 quail at 14 days old were randomly divided into four groups of vaccinated (B and C) and unvaccinated (A and D) birds. Vaccination was done on days 20 and 32, and viral inoculation of birds in groups C and D was then carried out on day 43. For evaluation of viral transmission, at 24 hr postinoculation additional unvaccinated birds were placed in direct contact with challenged birds. All the birds were evaluated for clinical signs, egg production, antibody production, viral titration in lung homogenates, and viral transmission following inoculation. All unvaccinated-challenged birds were infected and showed clinical signs, whereas the infection rate along with clinical signs of vaccinated-challenged birds reached 30%-40%. Although vaccination induced high antibody titers, reduction in food and water consumption was evident in this vaccinated-challenged group compared with the unchallenged control group. These results could indicate that inactivated vaccine did not fully prevent the infection, although it was capable of protecting birds against clinical signs and significantly decreased viral titers in lungs after intranasal challenge.     

Pathological and Hormonal Changes in Freshwater Fishes Due to Exposure to Heavy Metals Pollutants

The concentrations of major heavy metals in organs of two cyprinid fish and in water collected from three sections of the Kor River, Iran, were determined using the induction coupled plasma method. The concentrations of heavy metals in tissue of fish from the middle sampling zone were significantly higher (p?<?0.05) than those from the other two sampling zones, whereas no significant differences (p?>?0.05) were detected between the two sexes and species. Estradiol in females and progesterone and testosterone in males from the middle study site were significantly (p?<?0.05) lower than values from the other two sites. Pathological changes in blood cells, liver, and kidneys of fishes were significantly higher in highly polluted areas (middle sampling zone). So heavy metals exposure can effectively decreases estrogenic and androgenic secretion in fish. These results show that industrial activities have polluted the river and that heavy metals exposure can induce pathological changes in fish organs.     

Cryptic diversity,multilocus phylogeny,and pathogenicity of cercosporoid fungi associated with common bean and cowpea

In recent years, common bean (Phaseolus vulgaris) and cowpea (Vigna unguiculata) plants in the north of Iran have exhibited symptoms resembling Cercospora leaf spot (CLS) disease. This study was initiated to elucidate the taxonomy and pathogenicity of cercosporoid taxa associated with leaf spot diseases of these two legume crops in Iran. A total of 138 samples with CLS symptoms were collected from cultivated common bean and cowpea species in northern Iran and subjected to microscopic examination, resulting in identification of 98 Cercospora and 59 Pseudocercospora samples. A six-locus phylogenetic analysis (ITS, actA, tef1, gapdh, his3, and cmdA) coupled with examination of the morphology of 42 representative isolates from these samples confirmed that several cercosporoid fungi occur on common bean and cowpea in Iran. Five Cercospora species (C. iranica, C. cf. flagellaris, Cercospora sp. G, Cercospora sp. T, and C. vignigena) and two Pseudocercospora species (P. griseola f. griseola and P. cf. cruenta) were found; of these, C. cf. flagellaris was the dominant species, occurring on both common bean and cowpea. Pathogenicity tests confirmed that all seven species could infect leaves of common bean and/or cowpea. This is the first report of C. iranica, Cercospora sp. G, and Cercospora sp. T associated with common bean and/or cowpea in the world. In addition, C. vignigena was recorded for the first time in Iran. Results achieved in this study will assist strategies for the management of CLS disease of common bean and cowpea.