Purification of recombinant nucleocapsid protein of Newcastle disease virus from unclarified feedstock using expanded bed adsorption chromatography

In the present work, a single-step purification of recombinant nucleocapsid protein (NP) of the Newcastle disease virus (NDV) directly from unclarified feedstock using an expanded bed adsorption chromatography (EBAC) was developed. Streamline 25 column (ID = 25 mm) was used as a contactor and Streamline chelating adsorbent immobilized with Ni2+ ion was used as affinity adsorbent. The dynamic binding capacity of Ni2+ -loaded Streamline chelating adsorbent for the NP protein in unclarified feedstock was found to be 2.94 mg ml(-1) adsorbent at a superficial velocity of 200 cm h(-1). The direct purification of NP protein from unclarified feedstock using expanded bed adsorption has resulted in a 31% adsorption and 9.6% recovery of NP protein. The purity of the NP protein recovered was about 70% and the volume of processing fluid was reduced by a factor of 10. The results of the present study show that the IMA-EBAC developed could be used to combine the clarification, concentration and initial purification steps into a single-step operation.     

Genome sequence of the Mycobacterium abscessus strain M93

Mycobacterium abscessus is a rapid-growing species of nontuberculous mycobacteria that is frequently associated with opportunistic infections in humans. We report herein the draft genome sequence of M. abscessus strain M93.     

In a subgroup of high-risk Asians, telmisartan was non-inferior to ramipril and better tolerated in the prevention of cardiovascular events

Background and Objectives

Results of the recently published ONTARGET study (The Ongoing Telmisartan Alone and in Combination with Ramipril Global Endpoint Trial) showed that telmisartan (80 mg/day) was non-inferior to ramipril (10 mg/day) in reducing cardiovascular events. Clinicians in Asia doubt tolerability of these doses for their patients. We therefore analyzed data from this study and a parallel study TRANSCEND (Telmisartan Randomized Assessment Study in ACE Intolerant Subjects with Cardiovascular Disease). Our objectives were to compare Asians and non-Asians with respect to the following:

1. 1) Effectiveness of telmisartan vs. ramipril in reducing cardiovascular events;
2. 2) Proportions who reached the full dose of telmisartan, ramipril or placebo; and
3. 3) Proportions of overall discontinuations, and discontinuations due to adverse effects.

Method

The ONTARGET study randomized 25,620 patients at risk of cardiovascular events to ramipril, telmisartan, or their combination. The primary composite endpoint was death caused by cardiovascular disease, acute MI, stroke, and hospitalization because of congestive heart failure. TRANSCEND randomized 5926 high-risk patients with a history of intolerance to ACE-inhibitors to telmisartan or placebo. The primary outcome was the same. In this substudy, we compared Asians and non-Asians as to how well they tolerated telmisartan (given in both studies) and ramipril (given in ONTARGET).

Results

1) Telmisartan was non-inferior to ramipril in lowering the primary endpoint among Asians (RR = 0.92; 95% CI: 0.74, 1.13); 2) more Asians achieved the full dose of either drug; 3) less withdrew (overall); and 4) less withdrew for adverse effects. Furthermore, telmisartan was better tolerated than ramipril. This advantage was greater among Asians.

Conclusion and Significance

Although Asians had lower BMI than non-Asians, Asians tolerated both drugs better. Regulatory agencies require reporting of safety and effectiveness data by ethnicity, but few comply with this requirement. This study shows that safety data in ethnic subgroups can help assess applicability of results to specific populations.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00153101","term_id":"NCT00153101"}}NCT00153101     

SOME BIOLOGICAL ASPECTS AND A UNIQUE HABITAT CHOICE OF DAMSELFLY INDOCNEMIS ORANG FÖRSTER(ODONATA: PLATYCNEMIDIDAE)FROM MALAYSIA.

Abstract The hemimetabolous preliminary studies on the life cycle of Indocnemis orang is here reported for the first time, being completed in 6 instars for a minimum duration of 69 days. It can be classified as a stenotopic species being sensitive to biotope requirements but having tolerance towards fluctuating water qualities during its developmental process. The need for caution in the interpretation of bioindicator species for habitat assessment and biomonitoring is here both indicated and discussed.     

Inorganic and Hybrid Interfacial Materials for Organic and Perovskite Solar Cells

As organic solar cells (OSCs) and perovskite solar cells (PVSCs) move closer to commercialization, further efforts toward optimizing both cell efficiency and stability are needed. As interfaces strongly affect device performance and degradation processes, interfacial engineering by employing various materials as hole transport layers (HTLs) and electron transport layers (ETLs) has been a very active field of research in OSCs and PVSCs. Among them, inorganic materials exhibit significant advantages in promoting device performance due to their excellent charge transporting properties and intrinsic thermal and chemical robustness. In this review, an extensive overview is provided of inorganic semiconductors such as copper‐based ones with emphasis on copper iodide and copper thiocyanate, transition metal chalcogenides, nitrides and carbides as well as hybrid materials based on these inorganic compounds that have been recently employed as HTLs and ETLs in OSCs and PVSCs. Following a short discussion of the main optoelectronic and physical properties that interfacial materials used as HTLs and ETLs should possess, the functionalities of the aforementioned materials as interfacial, charge transport, layers in OSCs and PVSCs are discussed in depth. It is concluded by providing guidelines for further developments that could significantly extend the implementation of these materials in solar cells.     

Expression, purification and characterization of flavin reductase from Citrobacter freundii A1

Flavin reductase plays an important biological role in catalyzing the reduction of flavin by NAD(P)H oxidation. The gene that codes for flavin reductase from Citrobacter freundii A1 was cloned and expressed in Escherichia coli BL21(DE3)pLysS. In this study, we aimed to characterize the purified recombinant flavin reductase of C. freundii A1. The recombinant enzyme was purified to homogeneity and the biochemical profiles, including the effect of pH, temperature, metal ions and anions on flavin reductase activity and stability, were determined. This enzyme exhibited optimum activity at 45 °C in a 10-min reaction at pH 7.5 and was stable at temperatures up to 30 °C. At 0.1 mM concentration of metal ions, flavin reductase activity was stimulated by divalent cations including Mn²⁺, Sr²⁺, Ni²⁺, Sn²⁺, Ba²⁺, Co²⁺, Mg²⁺, Ca²⁺ and Pb²⁺. Ag⁺ was noticeably the strongest inhibitor of recombinant flavin reductase of C. freundii A1. This enzyme should not be defined as a standard flavoprotein. This is the first attempt to characterize flavin reductase of C. freundii origin.     

Dengue Virus Type 2 (DENV2)-Induced Oxidative Responses in Monocytes from Glucose-6-Phosphate Dehydrogenase (G6PD)-Deficient and G6PD Normal Subjects

Background

Dengue virus is endemic in peninsular Malaysia. The clinical manifestations vary depending on the incubation period of the virus as well as the immunity of the patients. Glucose-6-phosphate dehydrogenase (G6PD) deficiency is prevalent in Malaysia where the incidence is 3.2%. It has been noted that some G6PD-deficient individuals suffer from more severe clinical presentation of dengue infection. In this study, we aim to investigate the oxidative responses of DENV2-infected monocytes from G6PD-deficient individuals.

Methodology

Monocytes from G6PD-deficient individuals were infected with DENV2 and infection rate, levels of oxidative species, nitric oxide (NO), superoxide anions (O₂ ⁻), and oxidative stress were determined and compared with normal controls.

Principal Findings

Monocytes from G6PD-deficient individuals exhibited significantly higher infection rates compared to normal controls. In an effort to explain the reason for this enhanced susceptibility, we investigated the production of NO and O₂ ⁻ in the monocytes of individuals with G6PD deficiency compared with normal controls. We found that levels of NO and O₂ ⁻ were significantly lower in the DENV-infected monocytes from G6PD-deficient individuals compared with normal controls. Furthermore, the overall oxidative stress in DENV-infected monocytes from G6PD-deficient individuals was significantly higher when compared to normal controls. Correlation studies between DENV-infected cells and oxidative state of monocytes further confirmed these findings.

Conclusions/Significance

Altered redox state of DENV-infected monocytes from G6PD-deficient individuals appears to augment viral replication in these cells. DENV-infected G6PD-deficient individuals may contain higher viral titers, which may be significant in enhanced virus transmission. Furthermore, granulocyte dysfunction and higher viral loads in G6PD-deificient individuals may result in severe form of dengue infection.     

Chemical characterization and antimicrobial activity of rhizome essential oils of very closely allied Zingiberaceae species endemic to Borneo: Alpinia ligulata K. Schum. and Alpinia nieuwenhuizii Val

Two poorly studied, morphologically allied Alpinia species endemic to Borneo, viz., A. ligulata and A. nieuwenhuizii, were investigated here for their rhizome essential oil. The oil compositions and antimicrobial activities were compared with those of A. galanga, a better known plant. A fair number of compounds were identified in the oils by GC‐FID and GC/MS analyses, with large differences in the oil composition between the three species. The rhizome oil of A. galanga was rich in 1,8‐cineole (29.8%), while those of A. ligulata and A. nieuwenhuizii were both found to be extremely rich in (E)‐methyl cinnamate (36.4 and 67.8%, resp.). The three oils were screened for their antimicrobial activity against three Gram‐positive and three Gram‐negative bacteria and two fungal species. The efficiency of growth inhibition of Staphylococcus aureus var. aureus was found to decline in the order of A. nieuwenhuizii>A. ligulata ～ A. galanga, while that of Escherichia coli decreased in the order of A. galanga>A. nieuwenhuzii ～ A. ligulata. Only the A. galanga oil inhibited the other bacteria and the fungi tested.     

NDV-induced apoptosis in absence of Bax; evidence of involvement of apoptotic proteins upstream of mitochondria

ABSTRACT: BACKGROUND: Recently it was shown that following infection of HeLa cells with Newcastle disease virus (NDV), the matrix (M) protein binds to Bax and subsequently the intrinsic pathway of apoptosis is activated. Moreover, there was very little alteration on mRNA and protein levels of Bax and Bcl-2 after infection with NDV.FindingIn order to further investigate the role of members of the Bcl-2 family, Bax-knockout and wild-type HCT116 cells were infected with NDV strain AF2240. Although both cells underwent apoptosis through the activation of the intrinsic pathway and the release of cytochrome c from mitochondria, the percentage of dead Bax-knockout cells was significantly lower than wt cells (more than 10 % at 48 h post-infection). In a parallel experiment, the effect of NDV on HT29 cells, that are originally Bcl-2-free, was studied. Apoptosis in HT29 cells was associated with Bax redistribution from cytoplasm to mitochondria, similar to that of HeLa and wt HCT116 cells. CONCLUSION: Although the presence of Bax during NDV-induced apoptosis contributes to a faster cell death, it was concluded that other apoptotic protein(s) upstream of mitochondria are also involved since cancer cells die whether in the presence or absence of Bax. Therefore, the classic Bax/Bcl-2 ratio may not be a major determinant in NDV-induced apoptosis.     

Overexpression of human mitochondrial valyl tRNA synthetase can partially restore levels of cognate mt-tRNAVal carrying the pathogenic C25U mutation

Phenotypic diversity associated with pathogenic mutations of the human mitochondrial genome (mtDNA) has often been explained by unequal segregation of the mutated and wild-type genomes (heteroplasmy). However, this simple hypothesis cannot explain the tissue specificity of disorders caused by homoplasmic mtDNA mutations. We have previously associated a homoplasmic point mutation (1624C>T) in MTTV with a profound metabolic disorder that resulted in the neonatal deaths of numerous siblings. Affected tissues harboured a marked biochemical defect in components of the mitochondrial respiratory chain, presumably due to the extremely low (<1%) steady-state levels of mt-tRNA^Val. In primary myoblasts and transmitochondrial cybrids established from the proband (index case) and offspring, the marked respiratory deficiency was lost and steady-state levels of the mutated mt-tRNA^Val were greater than in the biopsy material, but were still an order of magnitude lower than in control myoblasts. We present evidence that the generalized decrease in steady-state mt-tRNA^Val observed in the homoplasmic 1624C>T-cell lines is caused by a rapid degradation of the deacylated form of the abnormal mt-tRNA^Val. By both establishing the identity of the human mitochondrial valyl-tRNA synthetase then inducing its overexpression in transmitochondrial cell lines, we have been able to partially restore steady-state levels of the mutated mt-tRNA^Val, consistent with an increased stability of the charged mt-tRNA. These data indicate that variations in the levels of VARS2L between tissue types and patients could underlie the difference in clinical presentation between individuals homoplasmic for the 1624C>T mutation.