Predictive inference on cytoplasmic and mitochondrial thioredoxin peroxidases in the highly radioresistant Lepidopteran insect Spodoptera frugiperda

Lepidopteran insects show remarkable resistance to radiation and chemical stress than insects of other orders. Despite this, the antioxidant machinery of insects of this order is poorly understood. Recently we demonstrated the significance of cytoplasmic NOS and a stronger mitochondrial antioxidant enzyme system in the stress-resistance of Lepidopteran insects. In the present study, we hypothesize two thioredoxin peroxidase orthologues (Sf-TPx1 and Sf-TPx2) in Lepidopteran insect Spodoptera frugiperda and demonstrate their structural/functional features important for cellular antioxidant activity and stress resistance. Results show a higher mitochondrial localization score (WoLFPSORT) of Sf-TPx2 (mitochondria-18.0, cytoplasm-7.0, nucleus-4.0) than its Drosophila orthologue Jafrac2 (secretory-30.0; mitochondria/nucleus/cytoplasm-no signal), which is important for antioxidant activity, and a higher cytoplasmic localization score of Sf-TPx1 (mitochondria-no signal; cytoplasm-22.0; nucleus-3.5) than the Drosophila Jafrac1 (mitochondria-17; nucleus- 11; cytoplasm-no signal). Structural modeling data show certain motifs present in Jafrac1 and Jafrac2 that affect active site conformation and separate cysteine residues at distances not suitable for disulphide bridge formation (5.21Å; 5.73Å). These motifs are absent in Sf-TPx1 and Sf-TPx2, yielding shorter distance (2.01Å; 2.05Å) between the cysteine residues suitable for disulphide bridge formation. Taken together, the disulphide bridge as well as mitochondrial and cytoplasmic localization are crucial for peroxidatic activity of TPx''s. Therefore,we hypothesize that the Spodoptera TPx''s offer potentially stronger anti-oxidant activity than that of Drosophila orthologues, and may contribute in the high radioresistance of Lepidopteran insects.     

A molecular and evolutionary study of the beta-globin gene family of the Australian marsupial Sminthopsis crassicaudata

Beta-globin gene families in eutherians (placental mammals) consist of a set of four or more developmentally regulated genes which are closely linked and, in general, arranged in the order 5'-embryonic/fetal genes- adult genes-3'. This cluster of genes is proposed to have arisen by tandem duplication of ancestral beta-globin genes, with the first duplication occurring 200 to 155 MYBP just prior to a period in mammalian evolution when eutherians and marsupials diverged from a common ancestor. In this paper we trace the evolutionary history of the beta-globin gene family back to the origins of these mammals by molecular characterization of the beta-globin gene family of the Australian marsupial Sminthopsis crassicaudata. Using Southern and restriction analysis of total genomic DNA and bacteriophage clones of beta-like globin genes, we provide evidence that just two functional beta-like globin genes exist in this marsupial, including one embryonic- expressed gene (S.c-epsilon) and one adult-expressed gene (S.c-beta), linked in the order 5'-epsilon-beta-3'. The entire DNA sequence of the adult beta-globin gene is reported and shown to be orthologous to the adult beta-globin genes of the North American marsupial Didelphis virginiana and eutherian mammals. These results, together with results from a phylogenetic analysis of mammalian beta-like globin genes, confirm the hypothesis that a two-gene cluster, containing an embryonic- and an adult-expressed beta-like globin gene, existed in the most recent common ancester of marsupials and eutherians. Northern analysis of total RNA isolated from embryos and neonatals indicates that a switch from embryonic to adult gene expression occurs at the time of birth, coinciding with the transfer of the marsupial from a uterus to a pouch environment.      

Alleviation of water and osmotic stress-induced changes in nitrogen metabolizing enzymes in <Emphasis Type="Italic">Triticum aestivum</Emphasis> L. cultivars by potassium

Present communication reports laboratory and pot experiments conducted to study the influence of water and osmotic stress on nitrogen uptake and metabolism in two wheat (Triticum aestivum L) cultivars with and without potassium supplementation. Polyethylene glycol 6000-induced osmotic stress/restricted irrigation caused a considerable decline in the activity of nitrate reductase, glutamate synthase, alanine and aspartate aminotransferases, and glutamate dehydrogenase. Potassium considerably improved nitrogen metabolism under normal water supply conditions and also resulted in amelioration of the negative impact of water and osmotic stresses indicating that potassium supplementation can be used as a potential tool for enhancing the nitrogen use efficiency in wheat for exploiting its genetic potential.     

Large scale production of phage antibody libraries using a bioreactor

One of the limitations of the use of phage antibody libraries in high throughput selections is the production of sufficient phage antibody library at the appropriate quality. Here, we successfully adapt a bioreactor-based protocol for the production of phage peptide libraries to the production of phage antibody libraries. The titers obtained in the stirred-tank bioreactor are 4 to 5 times higher than in a standard shake flask procedure, and the quality of the phage antibody library produced is indistinguishable to that produced using standard procedures as assessed by Western blotting and functional selections. Availability of this protocol will facilitate the use of phage antibody libraries in high-throughput scale selections.     

The Yo-Yo intermittent recovery test level 1 is reliable in young high-level soccer players

The aim of the study was to investigate test reliability of the Yo-Yo intermittent recovery test level 1 (YYIR1) in 36 high-level youth soccer players, aged between 13 and 18 years. Players were divided into three age groups (U15, U17 and U19) and completed three YYIR1 in three consecutive weeks. Pairwise comparisons were used to investigate test reliability (for distances and heart rate responses) using technical error (TE), coefficient of variation (CV), intra-class correlation (ICC) and limits of agreement (LOA) with Bland-Altman plots. The mean YYIR1 distances for the U15, U17 and U19 groups were 2024 ± 470 m, 2404 ± 347 m and 2547 ± 337 m, respectively. The results revealed that the TEs varied between 74 and 172 m, CVs between 3.0 and 7.5%, and ICCs between 0.87 and 0.95 across all age groups for the YYIR1 distance. For heart rate responses, the TEs varied between 1 and 6 bpm, CVs between 0.7 and 4.8%, and ICCs between 0.73 and 0.97. The small ratio LOA revealed that any two YYIR1 performances in one week will not differ by more than 9 to 28% due to measurement error. In summary, the YYIR1 performance and the physiological responses have proven to be highly reliable in a sample of Belgian high-level youth soccer players, aged between 13 and 18 years. The demonstrated high level of intermittent endurance capacity in all age groups may be used for comparison of other prospective young soccer players.