Acute sarcocystosis in a captive white-tailed deer in Virginia

A 19-mo-old female captive white-tailed deer in a public wild animal park in Richmond (Virginia, USA) was necropsied and evaluated histologically following spontaneous death after a 1 wk period of lethargy in a captive herd of 22 deer. An acute necrotizing pneumonia was associated with intraendothelial protozoal schizonts that were identified immunohistochemically as Sarcocystis spp. This is the first confirmed report of acute sarcocystosis in a wild ruminant.     

Strain typing of Toxoplasma gondii: comparison of antigen-coding and housekeeping genes

Molecular characterization of Toxoplasma gondii isolates is central for understanding differences in disease transmission and manifestations. Only 3 subgroups (lineages) have been discerned with subtle within-lineage variation, permitting low-resolution classification of isolates. Because proteins, coding sequences, and especially antigen-coding genes have been used extensively in previous studies, we focused on sequence variation in introns of housekeeping genes, which may be more informative for phylogenetic analysis because they evolve under lower selection. We compared sequence variation in introns of 5 housekeeping genes with 2 antigen-coding genes. Introns of housekeeping genes were slightly more polymorphic than coding and noncoding regions of antigen-coding genes and only the former showed intralineage variation. Intragenic linkage disequilibrium was complete, but intergenic linkage, although highly significant, was incomplete, suggesting that genes are partially uncoupled. Six of 7 substitutions found within the region coding for the tachyzoite surface antigen, SAG2, were nonsynonymous, indicating that diversifying selection acts on this locus. Typing isolates on the basis of housekeeping and antigen-coding genes was consistent, but the phylogenetic relationships among the resulting groups was inconsistent. A cougar isolate typed as lineage II using a restriction fragment length polymorphism assay possessed multiple unique polymorphisms, suggesting that it represents a new lineage. We concluded that introns of housekeeping genes are preferred markers for phylogenetic study, and that multilocus genotyping is preferred for typing parasites, especially from feral or unstudied environments.     

Ontogeny of solasodine-containing mucilage layer inSolanum viarum Dunal, ploidy types

Berries of steroid-bearingSolanum viarum Dunal are exploited commercially in India as raw material by steroid industries for solasodine, a glycoalkaloid, present in the mucilaginous exotesta of the seed. Comparative ontogeny of exotesta studied through histochemical studies in diploid, autotetraploid and trisomic plants indicated similarity in the histochemical changes occurring during ontogeny of the outermost seed coat layer which culminated in the transformation of this layer into the mucilage layer. The increased cell size in this layer in the autotetraploid plants probably accounts for the higher steroid content reported. Corroborative evidences for histochemical changes observed in the mucilage layer were obtained from studies of ultrastructure using transmission electron microscopy.     

Comparative Pharmacokinetic Study of Mangiferin After Oral Administration of Pure Mangiferin and US Patented Polyherbal Formulation to Rats

The US patented polyherbal formulation for the prevention and management of type II diabetes and its vascular complications was used for the present study. The xanthone glycoside mangiferin is one of the major effector constituents in the Salacia species with potential anti-diabetic activity. The pharmacokinetic differences of mangiferin following oral administration of pure mangiferin and polyherbal formulation containing Salacia species were studied with approximately the same dose 30 mg/kg mangiferin and its distribution among the major tissue in Wistar rats. Plasma samples were collected at different time points (15, 30, 60, 120, 180, 240, 360, 480, 600, 1,440, 2,160, and 2880 min) and subsequently analyzed using a validated simple and rapid LC-MS method. Plasma concentration versus time profiles were explored by non-compartmental analysis. Mangiferin plasma exposure was significantly increased when administered from formulation compared to the standard mangiferin. Mangiferin resided significantly longer in the body (last mean residence time (MRT_last)) when given in the form of the formulation (3.65 h). C_max values of formulation (44.16 μg/mL) administration were elevated when compared to equivalent dose of the pure mangiferin (15.23 μg/mL). Tissue distribution study of mangiferin from polyherbal formulation was also studied. In conclusion, the exposure of mangiferin is enhanced after formulation and administration and could result in superior efficacy of polyherbal formulation when compared to an equivalent dose of mangiferin. The results indicate that the reason which delays the elimination of mangiferin and enhances its bioavailability might the interactions of the some other constituents present in the polyherbal formulation. Distribution study results indicate that mangiferin was extensively bound to the various tissues like the small intestine, heart, kidney, spleen, and liver except brain tissue.

Electronic supplementary material

The online version of this article (doi:10.1208/s12249-014-0206-8) contains supplementary material, which is available to authorized users.KEY WORDS: bioavailability, mangiferin, pharmacokinetics, polyherbal formulation, tissue distribution     

R54C Mutation of NOTCH3 Gene in the First Rungus Family with CADASIL

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a rare hereditary stroke caused by mutations in NOTCH3 gene. We report the first case of CADASIL in an indigenous Rungus (Kadazan-Dusun) family in Kudat, Sabah, Malaysia confirmed by a R54C (c.160C>T, p.Arg54Cys) mutation in the NOTCH3. This mutation was previously reported in a Caucasian and two Korean cases of CADASIL. We recruited two generations of the affected Rungus family (n = 9) and found a missense mutation (c.160C>T) in exon 2 of NOTCH3 in three siblings. Two of the three siblings had severe white matter abnormalities in their brain MRI (Scheltens score 33 and 50 respectively), one of whom had a young stroke at the age of 38. The remaining sibling, however, did not show any clinical features of CADASIL and had only minimal changes in her brain MRI (Scheltens score 17). This further emphasized the phenotype variability among family members with the same mutation in CADASIL. This is the first reported family with CADASIL in Rungus subtribe of Kadazan-Dusun ethnicity with a known mutation at exon 2 of NOTCH3. The penetrance of this mutation was not complete during the course of this study.     

Functional Impairment of Myeloid Dendritic Cells during Advanced Stage of HIV-1 Infection: Role of Factors Regulating Cytokine Signaling

Introduction

Severely immunocompromised state during advanced stage of HIV-1 infection has been linked to functionally defective antigen presentation by dendritic cells (DCs). The molecular mechanisms behind DC impairment are still obscure. We investigated changes in DC function and association of key regulators of cytokine signaling during different stages of HIV-1 infection and following antiretroviral therapy (ART).

Methods

Phenotypic and functional characteristics of circulating myeloid DCs (mDCs) in 56 ART-naive patients (23 in early and 33 in advanced stage of disease), 36 on ART and 24 healthy controls were evaluated. Sixteen patients were studied longitudinally prior-to and 6 months after the start of ART. For functional studies, monocyte-derived DCs (Mo-DCs) were evaluated for endocytosis, allo-stimulation and cytokine secretion. The expression of suppressor of cytokine signaling (SOCS)-1 and other regulators of cytokine signaling was evaluated by real-time RT-PCR.

Results

The ability to respond to an antigenic stimulation was severely impaired in patients in advanced HIV-1 disease which showed partial recovery in the treated group. Mo-DCs from patients with advanced HIV-disease remained immature with low allo-stimulation and reduced cytokine secretion even after TLR-4 mediated stimulation ex-vivo. The cells had an increased expression of negative regulatory factors like SOCS-1, SOCS-3, SH2-containing phosphatase(SHP)-1 and a reduced expression of positive regulators like Janus kinase(JAK)2 and Nuclear factor kappa-light-chain-enhancer of activated B cells(NF-κB)1. A functional recovery after siRNA mediated silencing of SOCS-1 in these mo-DCs confirms the role of negative regulatory factors in functional impairment of these cells.

Conclusions

Functionally defective DCs in advanced stage of HIV-1 infection seems to be due to imbalanced state of negative and positive regulatory gene expression. Whether this is a cause or effect of increased viral replication at this stage of disease, needs further investigation. The information may be useful in design of novel therapeutic targets for better management of disease.     

Endo-β-N-acetylglucosamidases (ENGases) in the fungus Trichoderma atroviride: Possible involvement of the filamentous fungi-specific cytosolic ENGase in the ERAD process

N-Glycosylation is an important post-translational modification of proteins, which mainly occurs in the endoplasmic reticulum (ER). Glycoproteins that are unable to fold properly are exported to the cytosol for degradation by a cellular system called ER-associated degradation (ERAD). Once misfolded glycoproteins are exported to the cytosol, they are subjected to deglycosylation by peptide:N-glycanase (PNGase) to facilitate the efficient degradation of misfolded proteins by the proteasome. Interestingly, the ortholog of PNGase in some filamentous fungi was found to be an inactive deglycosylating enzyme. On the other hand, it has been shown that in filamentous fungi genomes, usually two different fungi-specific endo-β-N-acetylglucosamidases (ENGases) can be found; one is predicted to be localized in the cytosol and the other to have a signal sequence, while the functional importance of these enzymes remains to be clarified. In this study the ENGases of the filamentous fungus Trichoderma atroviride was characterized. By heterologous expression of the ENGases Eng18A and Eng18B in Saccharomyces cerevisiae, it was found that both ENGases are active deglycosylating enzymes. Interestingly, only Eng18B was able to enhance the efficient degradation of the RTL protein, a PNGase-dependent ERAD substrate, implying the involvement of this enzyme in the ERAD process. These results indicate that T. atroviride Eng18B may deglycosylate misfolded glycoproteins, substituting the function of the cytoplasmic PNGase in the ERAD process.     

Polyamine catabolism influences antioxidative defense mechanism in shoots and roots of five wheat genotypes under high temperature stress

Effect of high temperature stress on polyamine catabolism and antioxidant enzyme activity in relation to glutathione, ascorbate and proline accumulation was studied in five wheat (Triticum aestivum L.) genotypes (differently susceptible to temperature stress). High temperature significantly increased the activities of superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (GPX), ascorbate peroxidase (APX), dehydroascorbate reductase (DHAR), glutathione reductase (GR) and glutathione S-transferase (GST) in shoots of all genotypes. Higher activities of GPX in C 306, C 273 and APX in PBW 550, PBW 343 and PBW 534 demonstrate their important role in scavenging H₂O₂. Conversely, high temperature stress led to a significant decline in SOD, CAT, APX and GPX activities of roots with a subsequent increase in diamine oxidase (DAO) and polyamine oxidase (PAO) activities especially in PBW 550 and PBW 343. The concentration of ascorbic acid declined with the imposition of heat stress, however, polyamines responded to high temperature stress by increasing spermidine and spermine levels and decreasing putrescine levels. After exposure to high temperature, proline accumulation was significantly decreased in roots and increased in shoots though maximum concentration was achieved in C 306 genotype. Apparently, the wheat seedlings respond to high temperature mediated increase in reactive oxygen species (ROS) production by altering antioxidative defense mechanism and polyamine catabolism though differentially in five wheat genotypes. Among five genotypes studied, C 306 and C 273 seem to be better protected against temperature stress. The results suggested that shoots were more resistant against the destructive effects of ROS as is indicated by low levels of thiobarbituric acid reactive substances under high temperature stress.