Identification and characterisation of differentially expressed leaf proteins among Vitis species

Grapes are commercially grown worldwide for fresh fruit and wine. They are mainly classified into bunch and muscadine grapes. These species differ in their sugar content and composition, photosynthetic efficiency and tolerance to abiotic and biotic stresses. Grape berry relies on carbohydrates produced during photosynthesis to support its development and composition. In view of the unique physiology and genetic make‐up of muscadine grape, a proteomics study was performed to increase our knowledge of Vitis leaf proteome in order to improve enological and disease tolerance characteristics of grape species. A high throughput two‐dimensional gel electrophoresis (2‐DE) was conducted on muscadine, bunch and hybrid bunch leaf proteins. The differentially expressed proteins were excised from 2‐DE gels, subjected to in‐gel trypsin digestion, and analysed in MALDI/TOF mass spectrometer. The mass spectra were collected and protein identification was performed by searching against Viridiplantae database using Matrix Science algorithm. Proteins were mapped to universal protein resource to study gene ontology. We have discovered >255 proteins with pIs between 3.5 and 8.0 and molecular weight between 12 and 100 kDa among the Vitis species. Comparative analysis of leaf proteome showed that 54 polypeptides varied qualitatively and quantitatively among the three Vitis species studied. Of these, seven proteins were unique to muscadine, two proteins were present in both muscadine and bunch, while 28 proteins were common to all the three species. Bioinformatic analysis of these proteins showed that they are involved in signal transduction pathway, transport of metabolites, energy metabolism, protein trafficking, photosynthesis and defence. We have also identified proteins unique to muscadine grape that are involved in defence and stress tolerance. In addition, photosynthesis‐related proteins were found to be more abundant in Vitis vinifera grape compared to other Vitis species.     

Blastic plasmacytoid dendritic cell neoplasm: cytopathologic findings

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare, aggressive hematopoietic neoplasm, which in the past was also known variously as blastic NK cell lymphoma, agranular CD4+ natural killer cell leukemia, and CD4+/CD56+ hematodermic neoplasm. BPDCN is now believed to arise from plasmacytoid dendritic cells, but its exact etiology is still unknown. We report here on the cerebrospinal fluid (CSF) cytology of a BPDCN, a hypercellular specimen comprised of malignant, singly dispersed cells with scant to moderate amounts of pale blue, agranular cytoplasm, and uniform round to oval nuclei, fine chromatin, prominent nucleoli, occasional cytoplasmic microvacuoles, and pseudopodia. Neither mitoses nor karyorrhexis were identified. Flow cytometry of the CSF demonstrated that the malignant cells expressed bright CD45, HLA-DR and CD33, dim CD4, heterogeneous CD56, and partial CD123. The importance of clinical, histopathological, and phenotypic correlation is emphasized. Clinical and histopathological correlation and a literature review are also presented. The poor clinical outcome makes it important to accurately report this rare tumor in a CSF specimen.     

Age-dependent renal cortical microvascular loss in female mice

Renal function and blood flow decline during aging in association with a decrease in the number of intrarenal vessels, but if loss of estrogen contributes to this microvascular, rarefaction remains unclear. We tested the hypothesis that the decreased renal microvascular density with age is aggravated by loss of estrogen. Six-month-old female C57/BL6 mice underwent ovariectomy (Ovx) or sham operation and then were allowed to age to 18-22 mo. Another comparable group was replenished with estrogen after Ovx (Ovx+E), while a 6-mo-old group served as young controls. Kidneys were then dissected for evaluation of microvascular density (by micro-computed tomography) and angiogenic and fibrogenic factors. Cortical density of small microvessels (20-200 μm) was decreased in all aged groups compared with young controls (30.3 ± 5.8 vessels/mm2, P < 0.05), but tended to be lower in sham compared with Ovx and Ovx+E (9.9 ± 1.7 vs. 17.2 ± 4.2 and 18 ± 3.0 vessels/mm2, P = 0.08 and P = 0.02, respectively). Cortical density of larger microvessels (200-500 μm) decreased only in aged sham (P = 0.04 vs. young control), and proangiogenic signaling was attenuated. On the other hand, renal fibrogenic mechanisms were aggravated in aged Ovx compared with aged sham, but blunted in Ovx+E, in association with downregulated transforming growth factor-β signaling and decreased oxidative stress in the kidney. Therefore, aging induced in female mice renal cortical microvascular loss, which was likely not mediated by loss of endogenous estrogen. However, estrogen may play a role in protecting the kidney by decreasing oxidative stress and attenuating mechanisms linked to renal interstitial fibrosis.     

A spectrum of novel NPHS1 and NPHS2 gene mutations in pediatric nephrotic syndrome patients from Pakistan

Background

Mutations in the NPHS1 and NPHS2 genes are among the main causes of early-onset and familial steroid resistant nephrotic syndrome respectively. This study was carried out to assess the frequencies of mutations in these two genes in a cohort of Pakistani pediatric NS patients.

Methods

Mutation analysis was carried out by direct sequencing of the NPHS1 and NPHS2 genes in 145 nephrotic syndrome (NS) patients. This cohort included 36 samples of congenital or infantile onset NS cases and 39 samples of familial cases obtained from 30 families.

Results

A total of 7 homozygous (6 novel) mutations were found in the NPHS1 gene and 4 homozygous mutations in the NPHS2 gene. All mutations in the NPHS1 gene were found in the early onset cases. Of these, one patient has a family history of NS. Homozygous p.R229Q mutation in the NPHS2 gene was found in two children with childhood-onset NS.

Conclusions

Our results show a low prevalence of disease causing mutations in the NPHS1 (22% early onset, 5.5% overall) and NPHS2 (3.3% early onset and 3.4% overall) genes in the Pakistani NS children as compared to the European populations. In contrast to the high frequency of the NPHS2 gene mutations reported for familial SRNS in Europe, no mutation was found in the familial Pakistani cases. To our knowledge, this is the first comprehensive screening of the NPHS1 and NPHS2 gene mutations in sporadic and familial NS cases from South Asia.     

Association of the ACE-II genotype with the risk of nephrotic syndrome in Pakistani children

Nephrotic syndrome is a common pediatric glomerular disease associated with heavy proteinuria. Since, the angiotensin converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism is a putative genetic risk factor for NS, in this study, ACE (I/D) polymorphism was analyzed in 268 NS and 223 control samples by a PCR-based method. The genotypic and allelic frequencies were determined and the association between ACE I/D polymorphism and NS was evaluated. The frequency distribution of the II, ID and DD genotypes was 82 (30.6%), 128 (47.8%) and 58 (21.6%) in the NS patients and 9 (4.0%), 171 (76.7%) and 43 (19.3%) in the control samples respectively. In the Pakistani pediatric NS population, the II genotypic and allelic frequencies were found to be significantly associated with the disease (OR = 6.755; C.I = 3-14.9). No significant association was found between this polymorphism and the response to standard steroid therapy. Thus, in contrast to reports from other parts of the world, the II genotype was found to be significantly associated with NS in the Indian and Malay populations and in the Pakistani population described here. To our knowledge, this is the first report from Pakistan describing the association of the ACE I/D polymorphism with pediatric NS. On the basis of these results, it is suggested that analysis of the ACE (I/D) polymorphism should be performed for the early diagnosis in the high risk NS patients in South Asia.     

Additions to the cytologically investigated species of <Emphasis Type="Italic">Potentilla</Emphasis> L. (Rosaceae) from India

At present 14 species of Potentilla L. have been cytologically worked out from different geographical areas of Kashmir and Himachal Pradesh in the Western Himalayas. New chromosome numbers in nine species—Potentilla argyrophylla (n = 14), P. atrosanguinea (n = 7, 14), P. desertorum (n = 7), P. gerardiana (n = 14), P. indica (n = 14), P. micropetala (n = 14), P. nepalensis (n = 14), P. sibbaldia (n = 14) and P. thomsonii (n = 7)—have been reported on a worldwide basis for the first time. Additional chromosomal races of polyploid cytotypes for P. argyrophylla (n = 28) and P. desertorum (n = 14) along with a diploid cytotype for P. micropetala (n = 7) plus diploid cytotypes for the five species as P. fulgens (n = 7), P. gelida (n = 7), P. kleiniana (n = 7), P. sibbaldia (n = 7) and P. sundaica (n = 7) as well as a tetraploid cytotype for P. fruticosa (n = 14) all have been cytologically worked out from India for the first time. The course of meiosis varies from normal to abnormal in different populations of the majority of the species, such as P. argyrophylla, P. atrosanguinea, P. desertorum, P. fruticosa, P. fulgens, P. gelida, P. indica, P. nepalensis, P. sibbaldia and P. sundaica, except for normal meiosis observed in P. gerardiana, P. kleiniana, P. micropetala and P. thomsonii. The anomalous taxa are marked with meiotic abnormalities in the form of cytomixis, chromosomal stickiness, unoriented bivalents, formation of laggards and bridges resulting in abnormal microsporogenesis, and production of heterogenous-sized fertile pollen grains along with reduced pollen fertility. All the taxa with normal meiotic courses show nearly one hundred percent pollen fertility.     

Criteria for selection of surrogates used to study the fate and control of pathogens in the environment

This article defines the term surrogate as an organism, particle, or substance used to study the fate of a pathogen in a specific environment. Pathogenic organisms, nonpathogenic organisms, and innocuous particles have been used as surrogates for a variety of purposes, including studies on survival and transport as well as for method development and as "indicators" of certain conditions. This article develops a qualitative surrogate attribute prioritization process and allows investigators to select a surrogate by systematically detailing the experimental process and prioritizing attributes. The results are described through the use of case studies of various laboratories that have used this process. This article also discusses the history of surrogate and microbial indicator use and outlines the method by which surrogates can be used when conducting a quantitative microbial risk assessment. The ultimate goal of selecting a sufficiently representative surrogate is to improve public health through a health-based risk assessment framework. Under- or overestimating the resistance, inactivation, or movement may negatively impact risk assessments that, in turn, will impact health assessments and estimated safety levels. Reducing uncertainty in a risk assessment is one of the objectives of using surrogates and the ultimate motive for any experiment investigating potential exposure of a pathogen.     

Revisiting the conformation of xanthan and the effect of industrially relevant treatments

The structure and conformation of xanthan in aqueous solution following various processing treatments typically encountered in its application were investigated in this study. Treatments such as heating, autoclaving, high pressure homogenisation and irradiation were subjected to the same sample. Parameters such as weight average molecular weight (M(w)), polydispersity index, root mean square radius of gyration, intrinsic viscosity and Huggins constant were used to monitor the effect of these treatments. Additionally, we have quantified the mass recovery of samples examined by gel permeation chromatography and light scattering to properly account for all fractions present in xanthan solutions. Atomic force microscopy (AFM) images together with height measurements confirmed that xanthan conformation is double helical ordered renatured state (pre-heat treated by the manufacturer) in dilute solution conditions and random coil conformation in very dilute solution. The ordered (renatured) conformation is shown to have partially molten double helix, with more flexibility than the perfectly ordered native double helix. Heat treatment for 2h at 85°C reduces the M(w) of xanthan to half its initial value, and mass recovery measurements indicate that it completely overcomes its associative nature. Thermally treated xanthan solution in the dilute region leads to an order-disorder transition, as determined by contour length per unit mass. Similarly, irradiation of xanthan solution results in an order-disorder transition together with the production of single strand low molecular weight molecules. Autoclaving and high pressure homogenisation treatments cause degradation of xanthan. The results from treated xanthan solutions following high pressure homogenisation and irradiation confirm that xanthan does not reassociate. A revised summary of xanthan conformation in solution together with schematic models following the various treatments are proposed.