Quantitative study of regression and regeneration of the thymic cell population after X-irradiation in the newtPleurodeles waltlii Michah

Summary The changes in cell numbers of different thymic cell populations and the conditions governing the regeneration of these populations and the thymus itself were examined after X-irradiation (700 rads) of different parts of the body. The general effects of the irradiation were studied in each experimental group in terms of mortality and growth rate. The particular effects on each thymic cell population were studied by the measurement of mitotic activity and of evaluation of the changes in numbers among these populations in the thymus itself, and were compared with the effects in the granulopoietic layer of the liver and in the spleen. The great reduction in the number of lymphocytes after irradiation demonstrates that they are more radiosensitive than other cell types; this reduction can be compensated for by the arrival of new lymphoid cells originating from other lymphoid organs (if they have been protected from irradiation) and by allowing thymic regeneration. Thus, irradiation has indirect effects on non-irradiated areas, and demonstrates that the lymphoid cell population has a high potential for multidirectional migration.     

Protection of Recombinant Mammalian Antibodies from Development-Dependent Proteolysis in Leaves of Nicotiana benthamiana

The expression of clinically useful proteins in plants has been bolstered by the development of high-yielding systems for transient protein expression using agroinfiltration. There is a need now to know more about how host plant development and metabolism influence the quantity and quality of recombinant proteins. Endogenous proteolysis is a key determinant of the stability and yield of recombinant proteins in plants. Here we characterised cysteine (C1A) and aspartate (A1) protease profiles in leaves of the widely used expression host Nicotiana benthamiana, in relation with the production of a murine IgG, C5-1, targeted to the cell secretory pathway. Agroinfiltration significantly altered the distribution of C1A and A1 proteases along the leaf age gradient, with a correlation between leaf age and the level of proteolysis in whole-cell and apoplast protein extracts. The co-expression of tomato cystatin SlCYS8, an inhibitor of C1A proteases, alongside C5-1 increased antibody yield by nearly 40% after the usual 6-days incubation period, up to ∼3 mg per plant. No positive effect of SlCYS8 was observed in oldest leaves, in line with an increased level of C1A protease activity and a very low expression rate of the inhibitor. By contrast, C5-1 yield was greater by an additional 40% following 8- to 10-days incubations in younger leaves, where high SlCYS8 expression was maintained. These findings confirm that the co-expression of recombinant protease inhibitors is a promising strategy for increasing recombinant protein yields in plants, but that further opportunity exists to improve this approach by addressing the influence of leaf age and proteases of other classes.     

Sodium Acetate,Sodium Acid Pyrophosphate,and Citric Acid Impacts on Isolated Peripheral Lymphocyte Viability,Proliferation, and DNA Damage

The present study examined the impacts of sodium acetate (SA), sodium acid pyrophosphate (SAPP), and citric acid (CA) on the viability, proliferation, and DNA damage of isolated lymphocytes in vitro. 3‐(4,5‐Dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) release assays were adopted to evaluate cell viability, while comet assay was employed to assess the genotoxic effects. The cells were incubated with different levels of SA (50, 100, and 200 mM), SAPP (25, 50, and 100 mM/L), or CA (100, 200, and 300 μg/mL). The lymphocytes treated with the tested food additives showed concentration‐dependent decreases in both cell viability and proliferation. A concentration‐dependent increase in LDH release was also observed. The comet assay results indicated that SA, SAPP, and CA increased DNA damage percentage, tail DNA percentage, tail length, and tail moment in a concentration‐dependent manner. The current results showed that SA, SAPP, and CA are cytotoxic and genotoxic to isolated lymphocytes in vitro.     

Nematicidal activity of silver nanoparticles of botanical products against root-knot nematode,Meloidogyne incognita

Current study investigated the nematicidal activity of leaf extracts of Conyza dioscoridis, Melia azedarach, and Moringa oleifera that were prepared as silver nanoparticles (Ag-NP). The characterisation and size confirmation of the Ag-NP were done by UV–vis spectrophotometry and the scanning electron microscopy (SEM). The phytochemical contents of crude extracts and the nano formulations were analysed using gas chromatography-mass spectroscopy (GC-MS). Results revealed that silver nanoparticles of C. dioscoridis extractives had great nematicidal activity against the 2^nd stage juvenile (J2) and eggs of Meloidogyne incognita. Also, the Ag-NP showed similar nematicidal effect to the reference nematicide; rugby. The GC-MS analysis revealed the increase of certain metabolites due to the formulation of the Ag-NPs. Aromadendrene, 1-hydroxy-1,7-dimethyl-4-isopropyl-2,7-cyclodecdiene, 6-epi-shyobunol, 4-hexylacetophenone, β-isocomene, caryophyllene, β- and α-selinene, α-cadinol, berkheyaradulen, and bis-(2-ethylhexyl)phthalate were increased more than 2.5-folds in the Ag-NP compared the extract. Therefore, the green synthesis of metal nanoparticles might be a safe, effective and affordable nematicide alternatives.     

Immunization of Chickens with Newcastle Disease Virus Expressing H5 Hemagglutinin Protects against Highly Pathogenic H5N1 Avian Influenza Viruses

Background

Highly-pathogenic avian influenza virus (HPAIV) and Newcastle disease virus (NDV) are the two most important poultry viruses in the world. Natural low-virulence NDV strains have been used as vaccines over the past 70 years with proven track records. We have previously developed a reverse genetics system to produce low-virulent NDV vaccine strain LaSota from cloned cDNA. This system allows us to use NDV as a vaccine vector for other avian pathogens.

Methodology/Principal Finding

Here, we constructed two recombinant NDVs (rNDVs) each of which expresses the hemagglutinin (HA) gene of HPAIV H5N1strain A/Vietnam/1203/2004 from an added gene. In one, rNDV (rNDV-HA), the open reading frame (ORF) of HA gene was expressed without modification. In the second, rNDV (rNDV-HAF), the ORF was modified so that the transmembrane and cytoplasmic domains of the encoded HA gene were replaced with those of the NDV F protein. The insertion of either version of the HA ORF did not increase the virulence of the rNDV vector. The HA protein was found to be incorporated into the envelopes of both rNDV-HA and rNDV-HAF. However, there was an enhanced incorporation of the HA protein in rNDV-HAF. Chickens immunized with a single dose of either rNDV-HA or rNDV-HAF induced a high titer of HPAIV H5-specific antibodies and were completely protected against challenge with NDV as well as lethal challenges of both homologous and heterologous HPAIV H5N1.

Conclusion and Significance

Our results suggest that these chimeric viruses have potential as safe and effective bivalent vaccines against NDV and. HPAIV. These vaccines will be convenient and affordable, which will be highly beneficial to the poultry industry. Furthermore, immunization with these vaccines will permit serological differentiation of vaccinated and avian influenza field virus infected animals.     

Origin of Saxitoxin Biosynthetic Genes in Cyanobacteria

Background

Paralytic shellfish poisoning (PSP) is a potentially fatal syndrome associated with the consumption of shellfish that have accumulated saxitoxin (STX). STX is produced by microscopic marine dinoflagellate algae. Little is known about the origin and spread of saxitoxin genes in these under-studied eukaryotes. Fortuitously, some freshwater cyanobacteria also produce STX, providing an ideal model for studying its biosynthesis. Here we focus on saxitoxin-producing cyanobacteria and their non-toxic sisters to elucidate the origin of genes involved in the putative STX biosynthetic pathway.

Methodology/Principal Findings

We generated a draft genome assembly of the saxitoxin-producing (STX+) cyanobacterium Anabaena circinalis ACBU02 and searched for 26 candidate saxitoxin­genes (named sxtA to sxtZ) that were recently identified in the toxic strain Cylindrospermopsis raciborskii T3. We also generated a draft assembly of the non-toxic (STX−) sister Anabaena circinalis ACFR02 to aid the identification of saxitoxin-specific genes. Comparative phylogenomic analyses revealed that nine putative STX genes were horizontally transferred from non-cyanobacterial sources, whereas one key gene (sxtA) originated in STX+ cyanobacteria via two independent horizontal transfers followed by fusion. In total, of the 26 candidate saxitoxin-genes, 13 are of cyanobacterial provenance and are monophyletic among the STX+ taxa, four are shared amongst STX+ and STX-cyanobacteria, and the remaining nine genes are specific to STX+ cyanobacteria.

Conclusions/Significance

Our results provide evidence that the assembly of STX genes in ACBU02 involved multiple HGT events from different sources followed presumably by coordination of the expression of foreign and native genes in the common ancestor of STX+ cyanobacteria. The ability to produce saxitoxin was subsequently lost multiple independent times resulting in a nested relationship of STX+ and STX− strains among Anabaena circinalis strains.     

Tubers from potato lines expressing a tomato Kunitz protease inhibitor are substantially equivalent to parental and transgenic controls

Recombinant protease inhibitors represent useful tools for the development of insect‐resistant transgenic crops, but questions have been raised in recent years about the impact of these proteins on endogenous proteases and chemical composition of derived food products. In this study, we performed a detailed compositional analysis of tubers from potato lines expressing the broad‐spectrum inhibitor of Ser and Asp proteases, tomato cathepsin D inhibitor (SlCDI), to detect possible unintended effects on tuber composition. A compositional analysis of key nutrients and toxic chemicals was carried out with tubers of SlCDI‐expressing and control (comparator) lines, followed by a two‐dimensional gel electrophoresis (2‐DE) proteomic profiling of total and allergenic proteins to detect eventual effects at the proteome level. No significant differences were observed among control and SlCDI‐expressing lines for most chemicals assayed, in line with the very low abundance of SlCDI in tubers. Likewise, proteins detected after 2‐DE showed no quantitative variation among the lines, except for a few proteins in some control and test lines, independent of slcdi transgene expression. Components of the patatin storage protein complex and Kunitz protease inhibitors immunodetected after 2‐DE showed unaltered deposition patterns in SlCDI‐expressing lines, clearly suggesting a null impact of slcdi on the intrinsic allergenic potential of potato tubers. These data suggest, overall, a null impact of slcdi expression on tuber composition and substantial equivalence between comparator and SlCDI‐expressing tubers despite reported effects on leaf protein catabolism. They also illustrate the usefulness of proteomics as a tool to assess the authenticity of foods derived from novel‐generation transgenic plants.     

Efficiency of diagnostic biomarkers among colonic schistosomiasis Egyptian patients

The schistosomal parasite plays a critical role in the development of malignant lesions in different organs. The pathogenesis of cancer is currently under intense investigation to identify reliable prognostic indices for disease detection. The objective of this paper is to evaluate certain biochemical parameters as diagnostic tools to efficiently differentiate between colonic carcinoma and colonic carcinoma associated with schistosomal infection among Egyptian patients. The parameters under investigation are interleukin 2 (IL-2), tumour necrosis factor alpha (TNF-α), carcinoembryonic antigen (CEA) levels, tissue telomerase, pyruvate kinase (PK), glucose-6-phosphate dehydrogenase (G-6-PD) and lactate dehydrogenase (LDH) enzyme activities. The results revealed a significant elevation in the level of the tumour markers IL-2, TNF-α and CEA as well as the activities of LDH, telomerase and G-6-PD among non-bilharzial and bilharzial colonic cancer groups, with a more potent effect in bilharzial infection-associated colonic cancer. A significant inhibition in PK activity was recorded in the same manner as compared to normal tissues. The efficacy of this biomarker was also evaluated through detecting sensitivity, specificity, negative and positive predictive values. In conclusion, schistosomal colonic carcinoma patients displayed more drastic changes in all parameters under investigation. The combination of the selected parameters succeeded in serving as biomarkers to differentiate between the two malignant types.     

Effect of site-specific bronchial radon progeny deposition on the spatial and temporal distributions of cellular responses

Inhaled short-lived radon progenies may deposit in bronchial airways and interact with the epithelium by the emission of alpha particles. Simulation of the related radiobiological effects requires the knowledge of space and time distributions of alpha particle hits and biological endpoints. Present modelling efforts include simulation of radioaerosol deposition patterns in a central bronchial airway bifurcation, modelling of human bronchial epithelium, generation of alpha particle tracks, and computation of spatio-temporal distributions of cell nucleus hits, cell killing and cell transformation events. Simulation results indicate that the preferential radionuclide deposition at carinal ridges plays an important role in the space and time evolution of the biological events. While multiple hits are generally rare for low cumulative exposures, their probability may be quite high at the carinal ridges of the airway bifurcations. Likewise, cell killing and transformation events also occur with higher probability in this area. In the case of uniform surface activities, successive hits as well as cell killing and transformation events within a restricted area (say 0.5 mm²) are well separated in time. However, in the case of realistic inhomogeneous deposition, they occur more frequently within the mean cycle time of cells located at the carinal ridge even at low cumulative doses. The site-specificity of radionuclide deposition impacts not only on direct, but also on non-targeted radiobiological effects due to intercellular communication. Incorporation of present results into mechanistic models of carcinogenesis may provide useful information concerning the dose–effect relationship in the low-dose range.