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Monkeypox is a viral zoonotic disease, often transmitted to humans from animals. While the whole world is haggling with the COVID-19 pandemic, the emergence of the monkeypox virus (MPXV) arose as a new challenge to mankind. Till date, numerous cases related to the MPXV have been reported in several countries across the globe, but, its momentary distribution in the current time has left everyone in fright with increasing mortality and limited clinically approved treatments. Therefore, it is of immense importance to develop a potent and highly effective vaccine capable of inducing desired immunogenic responses against the highly contagious MPXV. Herein, using various immunoinformatic and computational biology tools, we made an attempt to develop a multi-epitope vaccine construct against the MPXV which is antigenic, non-allergen and non-toxic in nature and capable of exhibiting immunogenic behavior. The sequence of vaccine construct was designed using the proposed 4 MHC-I, 3 MHC-II and 4 B-cell epitopes linked with suitable adjuvant and linkers. The modeled structure of the vaccine construct was used to assess its interaction with the Toll-like Receptor 4 (TLR4) using ClusPro and HADDOCK. All-atoms molecular dynamics simulation of the MPXV vaccine construct-TLR4 complex followed by a high level of gene expression of the construct within the bacterial system affirmed its stability along with induction of immunogenic response within the host cell. Altogether, our immunoinformatic approach aid in the development of a stable chimeric vaccine construct against MPXV and needs further experimental validation for its immunological relevance and usefulness as a vaccine candidate.Communicated by Ramaswamy H. Sarma.

While NMR-based metabolomics is only about 20 years old, NMR has been a key part of metabolic and metabolism studies for >40 years. Historically, metabolic researchers used NMR because of its high level of reproducibility, superb instrument stability, facile sample preparation protocols, inherently quantitative character, non-destructive nature, and amenability to automation. In this chapter, we provide a short history of NMR-based metabolomics. We then provide a detailed description of some of the practical aspects of performing NMR-based metabolomics studies including sample preparation, pulse sequence selection, and spectral acquisition and processing. The two different approaches to metabolomics data analysis, targeted vs. untargeted, are briefly outlined. We also describe several software packages to help users process NMR spectra obtained via these two different approaches. We then give several examples of useful or interesting applications of NMR-based metabolomics, ranging from applications to drug toxicology, to identifying inborn errors of metabolism to analyzing the contents of biofluids from dairy cattle. Throughout this chapter, we will highlight the strengths and limitations of NMR-based metabolomics. Additionally, we will conclude with descriptions of recent advances in NMR hardware, methodology, and software and speculate about where NMR-based metabolomics is going in the next 5-10 years., (© 2022. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Foot-and-mouth disease (FMD) is a major disease of livestock in India and causes huge economic losses. The formal FMD control program started in 2003-04 in selected districts and was gradually expanded. The present study provides a descriptive review of the FMD outbreaks, prevalent serotypes, and genetic and antigenic features of the FMD virus (FMDV) that circulated in the country between 2011 and 2020. FMD outbreaks were regularly reported in cloven-hoofed domestic livestock and wildlife, with three serotypes including O, A, and Asia1. During the study period, a total of 2226 FMD outbreaks were documented and serotypes confirmed. FMDV serotype O dominated the outbreak scenario, accounting for about 92% of all outbreaks, followed by Asia1 (5% of all outbreaks) and A (3% of all outbreaks). Two major epidemics of FMD on an unprecedented scale during the years 2013 and 2018 by serotype O were recorded. The spatial distribution of FMD was characterized by a larger number of outbreaks in the southern region of the country. In an annual-scale analysis, 2020 was the year with the lowest outbreaks, and 2013 was the year with the highest. The month-scale analysis showed that outbreaks were reported throughout the year, with the highest numbers between October and March. The emergence of three major lineages (O/ME-SA/Ind2001d, O/ME-SA/Ind2001e, and O/ME-SA/Ind2018) of serotype O was observed during the period. In the cases of serotype A and Asia1, the appearance of at least one novel lineage/genetic group, including A/G-18/non-deletion/2019 and Asia1/Group-IX, was documented. While serotype A showed the advent of antigenic variants, serotypes O and Asia1 did not show any antigenic diversity. It was noticed during the course of an outbreak that animal movement contributes significantly to disease transmission. Except for 2018, when numerous FMD outbreaks were recorded, the number of annual outbreaks reported after 2016 has been lower than in the first half of the decade, probably due to mass vaccination and COVID-19 pandemic-linked movement restrictions. Even during outbreaks, disease symptoms in ruminant populations, including cattle, were found to be less severe. Regular six-monthly immunization certainly has a positive impact on the reduction of disease burden and should be followed without fail and delay, along with intensive disease surveillance., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Purpose: Pseudoexfoliation is an age-related fibrillopathy characterized by the deposition of fibrillar material in the eye with an increased risk of complications during cataract surgery. Aim was to study visual outcomes and complications in patients with pseudoexfoliation undergoing manual small-incision cataract surgery (MSICS)., Methods: Prospective observational study was performed on 152 patients with pseudoexfoliation above 50 years undergoing MSICS in a tertiary hospital from December 2016 to November 2017. Intraoperative and postoperative complications were documented with follow-up on postoperative day 1, 1 st week, 1 st month, and 3 rd month., Results: Preoperative small pupil was noted in 49 eyes (32.2%), and 19 (12.5%) required intraoperative measures. Intraocular complications noted were zonular dialysis in five (3.3%), posterior capsular rupture in one (0.7%), and iridodialysis in one (0.7%). On postoperative day 1, the most common complication was corneal edema in 134 patients but clinically significant in only 23 (15.1%). Postoperative complications at 3 months were irregular pupil in 17 cases and decentered IOL in three cases. Intraocular pressure decreased with each visit [preoperative mean: 14.39 (±3.4) and 13.37 (±2.0) 12.53 (±1.4) mm Hg at 1 and 3 months, respectively]. There was a significant improvement in vision from the first day mean pinhole vision of 0.26 (±0.24) to mean best corrected visual acuity (BCVA) of 0.09 (±0.22) and 0.07 (±0.22) at 1 and 3 months, respectively. Mean endothelial cell loss was 193.16 (7.79%) and 266.01 (10.68%) at 1 and 3 months, respectively., Conclusion: Pseudoexfoliation has an increased risk of complications during cataract surgery. MSICS gives good outcomes in terms of visual recovery and postoperative outcomes., Competing Interests: None

Foot-and-mouth disease (FMD) is endemic in India, where circulation of serotypes O, A and Asia1 is frequent. Here, we provide an epidemiological assessment of the ongoing mass vaccination programs in regard to post-vaccination monitoring and outbreak occurrence. The objective of this study was assessing the contribution of mass vaccination campaigns in reducing the risk of FMD in India from 2008 to 2016 by evaluating sero-monitoring data and modelling the spatiotemporal dynamics of reported outbreaks. Through analyzing antibody titre data from >1 million animals sampled as part of pre- and post-vaccination monitoring, we show that the percent of animals with inferred immunological protection (based on ELISA) was highly variable across states but generally increased through time. In addition, the number of outbreaks in a state was negatively correlated with the percent of animals with inferred protection. We then analyzed the distribution of reported FMD outbreaks across states using a Bayesian space-time model. This approach provides better acuity to disentangle the effect of mass vaccination programs on outbreak occurrence, while accounting for other factors that contribute to spatiotemporal variability in outbreak counts, notably proximity to international borders and inherent spatiotemporal correlations in incidence. This model demonstrated a ∼50% reduction in the risk of outbreaks in states that were part of the vaccination program. In addition, after controlling for spatial autocorrelation in the data, states that had international borders experienced heightened risk of FMD outbreaks. These findings help inform risk-based control strategies for India as the country progresses towards reducing reported clinical disease., (© 2022 The Authors. Transboundary and Emerging Diseases published by Wiley-VCH GmbH.)

An emerging area of interest in understanding disease phenotypes is systems genomics. Complex diseases such as diabetes have played an important role towards understanding the susceptible genes and mutations. A wide number of methods have been employed and strategies such as polygenic risk score and allele frequencies have been useful, but understanding the candidate genes harboring those mutations is an unmet goal. In this perspective, using systems genomic approaches, we highlight the application of phenome-interactome networks in diabetes and provide deep insights. LINC01128, which we previously described as candidate for diabetes, is shown as an example to discuss the approach., Competing Interests: Conflict-of-interest statement: The authors declare no conflict of interest for this article., (©The Author(s) 2022. Published by Baishideng Publishing Group Inc. All rights reserved.)

The Natural Products Magnetic Resonance Database (NP-MRD) is a comprehensive, freely available electronic resource for the deposition, distribution, searching and retrieval of nuclear magnetic resonance (NMR) data on natural products, metabolites and other biologically derived chemicals. NMR spectroscopy has long been viewed as the 'gold standard' for the structure determination of novel natural products and novel metabolites. NMR is also widely used in natural product dereplication and the characterization of biofluid mixtures (metabolomics). All of these NMR applications require large collections of high quality, well-annotated, referential NMR spectra of pure compounds. Unfortunately, referential NMR spectral collections for natural products are quite limited. It is because of the critical need for dedicated, open access natural product NMR resources that the NP-MRD was funded by the National Institute of Health (NIH). Since its launch in 2020, the NP-MRD has grown quickly to become the world's largest repository for NMR data on natural products and other biological substances. It currently contains both structural and NMR data for nearly 41,000 natural product compounds from >7400 different living species. All structural, spectroscopic and descriptive data in the NP-MRD is interactively viewable, searchable and fully downloadable in multiple formats. Extensive hyperlinks to other databases of relevance are also provided. The NP-MRD also supports community deposition of NMR assignments and NMR spectra (1D and 2D) of natural products and related meta-data. The deposition system performs extensive data enrichment, automated data format conversion and spectral/assignment evaluation. Details of these database features, how they are implemented and plans for future upgrades are also provided. The NP-MRD is available at https://np-mrd.org., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: Hypertriglyceridemia is as an independent risk factor for cardiovascular disease (CVD). Apolipoprotein C-III (ApoC-III) is known to regulate triglyceride (TG) metabolism. However, the causal association between ApoC-III and CVD development is unclear. The objectives were to examine the impact of ApoC-III concentration on TG and lipoproteins and investigate the role of known rare loss-of-function APOC3 variants for modulating ApoC-III, TG concentrations and CVD risk in different ethnic groups., Methods: Plasma ApoC-III levels were measured in a multiethnic sample of 518 individuals comprising 271 Asian Indians (Sikhs), 87 Caucasians, 80 African Americans, and 80 Hispanics., Results: ApoC-III levels showed a robust association with TG in Asian Indians (r = 0.5, p = 1.1 × 10 -23 ), Caucasians (r = 0.4, p = 7.2 × 10 -4 ), and Hispanics (r = 0.9, p = 2.7x × 10 -28 ). African Americans had lowest ApoC-III and TG concentrations and highest (44%) prevalence of coronary artery disease (CAD). ApoC-III levels correlated with fasting blood glucose (r = 0.25, p = 6.1 × 10 -5 ) in Asian Indians and central adiposity in Hispanics (waist: r = 0.22, p = 0.05; waist-hip ratio: r = 0.24, p = 0.04). The carriers of rare variants IVS1-2G-A (rs373975305); A43T (rs147210663) and IVS3 + 1G-T (rs140621530) showed high TG but not low ApoC-III levels in Asian Indians and Caucasians., Conclusion: These results highlight the challenges of generalizing antisense ApoC-III inhibition for treating atherosclerotic disease in dyslipidemia that may benefit only specific sub-populations. The observed ethnic differences in ApoC-III concentrations and CAD risk factors, emphasize in-depth genetic and metabolomics evaluations on diverse ancestries.

The study of differential gene expression in persister cells is compounded by ceasure of conventional cellular metabolic pathways during persistence. There is, hence, a requirement to identify and validate suitable reference genes whose expression remains stable during persistence. We evaluated the suitability of five genes viz. dnaJ, groEL, rpoB, kp751, kp4432 as references to study gene expression using real-time polymerase chain reaction (qPCR) during persister cell formation in Klebsiella pneumoniae. Results obtained showed that while dnaJ and groEL suffered from unstable expression; rpoB, kp751 and kp4432 showed stable expression. Further, it was observed that data normalization using either of the stable genes viz. rpoB, kp751, kp4432 alone, resulted in either too low expression levels or too high variation among replicates. Our study indicates the concurrent use of kp4432 and rpoB as reference genes to be the most suitable for reliable analysis of differential gene expression during antibiotic induced persister formation in K. pneumoniae. kp4432 and rpoB encode NAD-dependant phenylacetaldehyde dehydrogenase and DNA-directed RNA polymerase beta subunit respectively. The outcome of this study will increase the utility of qPCR in studying the temporal changes in gene expression during persistence. The study will also aid in understanding mechanisms underlying persister cell formation particularly in K. pneumoniae., (Copyright © 2021 Elsevier B.V. All rights reserved.)

DZIP3/hRUL138 is a poorly characterized RNA-binding RING E3-ubiquitin ligase with functions in embryonic development. Here we demonstrate that DZIP3 is a crucial driver of cancer cell growth, migration, and invasion. In mice and zebrafish cancer models, DZIP3 promoted tumor growth and metastasis. In line with these results, DZIP3 was frequently overexpressed in several cancer types. Depletion of DZIP3 from cells resulted in reduced expression of Cyclin D1 and a subsequent G 1 arrest and defect in cell growth. Mechanistically, DZIP3 utilized its two different domains to interact and stabilize Cyclin D1 both at mRNA and protein levels. Using an RNA-binding lysine-rich region, DZIP3 interacted with the AU-rich region in 3' untranslated region of Cyclin D1 mRNA and stabilized it. Using a RING E3-ligase domain, DZIP3 interacted and increased K63-linked ubiquitination of Cyclin D1 protein to stabilize it. Remarkably, DZIP3 interacted with, ubiquitinated, and stabilized Cyclin D1 predominantly in the G 1 phase of the cell cycle, where it is needed for cell-cycle progression. In agreement with this, a strong positive correlation of mRNA expression between DZIP3 and Cyclin D1 in different cancer types was observed. Additionally, DZIP3 regulated several cell cycle proteins by modulating the Cyclin D1-E2F axes. Taken together, this study demonstrates for the first time that DZIP3 uses a unique two-pronged mechanism in its stabilization of Cyclin D1 to drive cell-cycle and cancer progression. SIGNIFICANCE: These findings show that DZIP3 is a novel driver of cell-cycle and cancer progression via its control of Cyclin D1 mRNA and protein stability in a cell-cycle phase-dependent manner. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/2/315/F1.large.jpg., (©2020 American Association for Cancer Research.)

Mutton and lamb sales continue to grow globally at a rate of 5% per year. However, sheep farming struggles with low profit margins due to high feed costs and modest carcass yields. Selecting those sheep expected to convert feed efficiently and have high carcass merit, as early as possible in their life cycle, could significantly improve the profitability of sheep farming. Unfortunately, direct measurement of feed conversion efficiency (via residual feed intake [RFI]) and carcass merit is a labor-intensive and expensive procedure. Thus, indirect, marker-assisted evaluation of these traits has been explored as a means of reducing the cost of its direct measurement. One promising and potentially inexpensive route to discover biomarkers of RFI and/or carcass merit is metabolomics. Using quantitative metabolomics, we profiled the blood serum metabolome (i.e., the sum of all measurable metabolites) associated with sheep RFI and carcass merit and identified candidate biomarkers of these traits. The study included 165 crossbred ram-lambs that underwent direct measurement of feed consumption to determine their RFI classification (i.e., low vs. high) using the GrowSafe System over a period 40 d. Carcass merit was evaluated after slaughter using standardized methods. Prior to being sent to slaughter, one blood sample was drawn from each animal, and serum prepared and frozen at -80 °C to limit metabolite degradation. A subset of the serum samples was selected based on divergent RFI and carcass quality for further metabolomic analyses. The analyses were conducted using three analytical methods (nuclear magnetic resonance spectroscopy, liquid chromatography mass spectrometry, and inductively coupled mass spectrometry), which permitted the identification and quantification of 161 unique metabolites. Biomarker analyses identified three significant (P < 0.05) candidate biomarkers of sheep RFI (AUC = 0.80), seven candidate biomarkers of carcass yield grade (AUC = 0.77), and one candidate biomarker of carcass muscle-to-bone ratio (AUC = 0.74). The identified biomarkers appear to have roles in regulating energy metabolism and protein synthesis. These results suggest that serum metabolites could be used to categorize and predict sheep for their RFI and carcass merit. Further validation using a larger (3×) and more diverse cohort of sheep is required to confirm these findings., (© The Author(s) 2020. Published by Oxford University Press on behalf of the American Society of Animal Science. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Even among households that have access to improved sanitation, children's faeces often do not end up in a latrine, the international criterion for safe disposal of child faeces., Methods: We collected data on possible determinants of safe child faeces disposal in a cross-sectional study of 851 children <5 y of age from 694 households in 42 slums in two cities in Odisha, India. Caregivers were asked about defecation and faeces disposal practices for all the children <5 y of age in the household., Results: Only a quarter (25.5%) of the 851 children's faeces were reported to be disposed of in a latrine. Even fewer (22.3%) of the 694 households reported that the faeces of all children <5 y of age in the home ended up in the latrine the last time the child defecated. In multivariate analysis, factors associated with being a safe disposal household were education and religion of the primary caregiver, number of children <5 y of age in the household, wealth, type and location of the latrine used by the household, household members >5 y of age using the latrine for defecation and mobility of children <5 y of age in the household., Conclusions: Few households reported disposing of all of their children's faeces in a latrine. Improving latrine access and specific behaviour change interventions may improve this practice., (© The Author(s) 2019. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Background: Recent cross-sectional studies have suggested that chronic inflammation resulting due to gingival and periodontal diseases may have potential reversible risk factors for dementia. Recently, cognition, memory deficit, and along with the behavioral defect have been considered as characteristics of dementia. Age is one of the main factors which is also known to be associated with dementia. The aim of the present study was to establish the correlation between tooth loss and diagnosis of cognitive impairment/dementia in elderly patients., Materials and Methods: A cross-sectional study, comprising of 300 patients (Age: 50-80 years), was conducted. For the assessment of cognitive impairment/dementia, Mini-Mental Status Examination (MMSE) was conducted. All the results were analyzed using the Chi-square and Kruskal-Wallis ANOVA test., Results: Number of teeth was found to be directly associated with MMSE. Less number of teeth was associated with lower MMSE score, irrespective of gender. The level of statistical significance was found to be P < 0.001. Factors, such as socioeconomic status, school education, and marital status, were found to have a statistically significant impact on the dementia scale., Conclusion: The current study found a significant association between tooth loss and cognitive impairment. Along with age factors such as socioeconomic status, education, and marital status were also found to have a direct or indirect impact on dementia. More prospective studies and clinical trials are required to be conducted to confirm the observations of this study., Competing Interests: There are no conflicts of interest.

Mycoplasma pneumoniae is a substantial respiratory pathogen that develops not only pneumonia but also other respiratory diseases, which mimic viral respiratory syndromes. Nevertheless, vaccine development for this pathogen delays behind as immunity correlated with protection is now predominantly unknown. In the present study, an immunoinformatics pipeline is utilized for epitope-based peptide vaccine design, which can trigger a critical immune response against M. pneumoniae. A total of 105 T-cell epitopes from 12 membrane associated proteins and 7 T-cell epitopes from 5 cytadherence proteins of M. pneumoniae were obtained and validated. Thus, 18 peptides with 9-mer core sequence were identified as best T-cell epitopes by considering the number of residues with > 75% in favored region. Further, the crucial screening studies predicted three peptides with good binding affinity towards HLA molecules as best T-cell and B-cell epitopes. Based on this result, visualization, and dynamic simulation for the three epitopes (WIHGLILLF, VILLFLLLF, and LLAWMLVLF) were assessed. The predicted epitopes needs to be further validated for their adept use as vaccine. Collectively, the study opens up a new horizon with extensive therapeutic application against M. pneumoniae and its associated diseases.

Discerning the relationship between molecules involved in diseases based on their underlying biological mechanisms is one of the greatest challenges in therapeutic development today. Gestational diabetes mellitus (GDM) is one of the most common complications during pregnancy, which adversely affects both mothers and offspring during and after pregnancy. We have constructed two datasets of (GDM associated genes from affected mother and placenta to systematically analyze and evaluate their interactions like gene-gene, gene-protein, gene-microRNA (miRNA), gene-transcription factors, and gene-associated diseases to enhance our current knowledge, which may lead to further advancements in disease diagnosis, prognosis, and treatment. The results identify the key genes with respect to maternal dataset as insulin receptor, insulin (INS), leptin (LEP), glucokinase, and hepatocyte nuclear factor 1 alpha, whereas from placenta include insulin-like growth factor 1, growth hormone receptor, and breast cancer anti-estrogen resistance protein 1, which are found to be highly enriched in pancreas, ovary, adipocyte, heart, and placental tissues. The key transcription factors include Sp1 transcription factor, pancreatic and duodenal homeobox 1, and hepatocyte nuclear factor 4 alpha, whereas miRNA includes has-miR-5699-5p and has-miR-3158-3p. The study also reveals that GDM has associations with diseases like type I and II diabetes mellitus, obesity, and preeclampsia. More significantly, we could trace out a significant connection between the key molecules like LEP and placental growth hormone from mother and placental dataset, which plays a critical role in INS secretion, INS signaling, and β-cell dysfunction pathways., (© 2018 Wiley Periodicals, Inc.)

'National foot-and-mouth disease (FMD) control programme' is being implemented in India and therefore predicting vaccine match is a key surveillance task. Recently, a considerable proportion of field viruses (75.6%) showed antigenic drift from the existing serotype A vaccine strain A IND 40/2000 necessitating search for an alternate strain. Here, antigenic relationship ('r 1 ' value) of 87 field viruses with each of the 8 candidate strains was estimated by virus neutralization test. A IND 27/2011 strain emerged to be the one with the widest spectrum of antigenic coverage showing 'r 1 ' value of more than 0.3 with 81.6% of field strains. It achieved a reasonably high titre of log 10 7.5 TCID 50 /ml in BHK-21 suspension cell which was accompanied by positive charge gaining substitutions (E 82 -K and E 131 -K in VP2) thought to have adaptive significance. However, potency trial remains to be conducted before A IND 27/2011 finds a place in the vaccine formulation., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Aim: The study was undertaken to assess the prevalence of antibodies to Capripoxviruses among small ruminants of Odisha, India., Materials and Methods: A total of 500 random serum samples collected from 214 sheep and 286 goats across 10 agro-climatic zones of Odisha, were screened using whole virus antigen-based indirect ELISA for antibodies against Capripoxviruses. Results were analyzed by suitable statistical methods., Results: Screening of 500 serum samples showed seropositivity of 8.88% and 31.47% in sheep and goats, respectively, for Capripoxviruses. The prevalence rate according to agro-climatic zone ranged from 0% (North Eastern coastal plain zone) to 48.57% (North central plateau zone) for goat pox, and 0% (Western undulating zone and North central plateau) to 22.22% (South Eastern ghat zone) for sheep pox. The difference in prevalence rates among the various agro-climatic zones was statistically significant (p<0.05) for goats, but not for sheep. Antibody prevalence rates among various districts were recorded to be the highest in Jagatsinghpur (30%) for sheep pox and Dhenkanal (80%) for goat pox., Conclusions: The study revealed serological evidence of Capripoxvirus infection in sheep and goat populations in the study area, in the absence of vaccination. Systematic investigation, monitoring, and reporting of outbreaks are necessary to devise control strategies.

Background: Research suggests that the lived experience of inadequate sanitation may contribute to poor health outcomes above and beyond pathogen exposure, particularly among women. The goal of this research was to understand women's lived experiences of sanitation by documenting their urination-related, defecation-related and menstruation-related concerns, to use findings to develop a definition of sanitation insecurity among women in low-income settings and to develop a conceptual model to explain the factors that contribute to their experiences, including potential behavioural and health consequences., Methods: We conducted 69 Free-List Interviews and eight focus group discussions in a rural population in Odisha, India to identify women's sanitation concerns and to build an understanding of sanitation insecurity., Findings: We found that women at different life stages in rural Odisha, India have a multitude of unaddressed urination, defecation and menstruation concerns. Concerns fell into four domains: the sociocultural context, the physical environment, the social environment and personal constraints. These varied by season, time of day, life stage and toilet ownership, and were linked with an array of adaptations (ie, suppression, withholding food and water) and consequences (ie, scolding, shame, fear). Our derived definition and conceptual model of sanitation insecurity reflect these four domains., Discussion: To sincerely address women's sanitation needs, our findings indicate that more is needed than facilities that change the physical environment alone. Efforts to enable urinating, defecating and managing menstruation independently, comfortably, safely, hygienically, privately, healthily, with dignity and as needed require transformative approaches that also address the gendered, sociocultural and social environments that impact women despite facility access. This research lays the groundwork for future sanitation studies to validate or refine the proposed definition and to assess women's sanitation insecurity, even among those who have latrines, to determine what may be needed to improve women's sanitation circumstances., Competing Interests: Competing interests: None declared.

Child feces represent a particular health risk to children due to increased prevalence of enteric agents and a higher risk of exposure owing to exploratory behaviors of young children. The safe management of such feces presents a significant challenge, not only for the 2.4 billion who lack access to improved sanitation, but also due to unhygienic feces collection and disposal and poor subsequent handwashing practices. We assessed potential sources of fecal exposure by documenting child feces management practices in a cross-sectional study of 851 children < 5 years of age from 694 households in 42 slums in two cities in Odisha, India. No preambulatory children and only 27.4% of ambulatory children defecated directly in the latrine. Children that did not defecate in a latrine mainly defecated on the ground, whether they were preambulatory or ambulatory. Use of diapers (1.2%) or potties (2.8%) was low. If the feces were removed from the ground, the defecation area was usually cleaned, if at all, only with water. Most children's feces were disposed of in surrounding environment, with only 6.5% deposited into any kind of latrine, including unimproved. Handwashing with soap of the caregiver after child feces disposal and child anal cleaning with soap after defecation was also uncommon. While proper disposal of child feces in an improved latrine still represents a major challenge, control of the risks presented requires attention to the full range of exposures associated to the management of child feces, and not simply the place of disposal.

Nonstructural proteins of hepatitis C virus had drawn much attention for the scientific fraternity in drug discovery due to its important role in the disease. 3D structure of the protein was predicted using molecular modelling protocol. Docking studies of 10 medicinal plant compounds and three drugs available in the market (control) with NS2 protease were employed by using rigid docking approach of AutoDock 4.2. Among the molecules tested for docking study, naringenin and quercetin revealed minimum binding energy of - 7.97 and - 7.95 kcal/mol with NS2 protease. All the ligands were docked deeply within the binding pocket region of the protein. The docking study results showed that these compounds are potential inhibitors of the target; and also all these docked compounds have good inhibition constant, vdW+Hbond+desolv energy with best RMSD value.

Foot-and-mouth disease (FMD) is a highly contagious and economically devastating disease of livestock, primarily affecting cattle, buffalo and pigs. FMD virus serotypes O, A and Asia1 are prevalent in India and systematic efforts are on to control and eventually eradicate the disease from the country. FMD epidemiology is complex due to factors like co-circulation, extinction, emergence and re-emergence of genotypes/lineages within the three serotypes, animal movement, diverse farm practices and large number of susceptible livestock in the country. Systematic vaccination, prompt diagnosis, strict biosecurity measures, and regular monitoring of vaccinal immunity and surveillance of virus circulation are indispensible features for the effective implementation of the control measures. Availability of suitable companion diagnostic tests is very important in this endeavour. In this review, the diagnostic assays developed and validated in India and their contribution in FMD control programme is presented.

Foot-and-mouth disease (FMD) virus serotype O Ind2001 lineage within the Middle East-South Asia topotype is the major cause of recent FMD incidences in India. A sub-lineage of Ind2001 caused severe outbreaks in the southern region of the country during 2013 and also reported for the first time from Libya. In this study, we conducted a detailed evolutionary analysis of Ind2001 lineage. Phylogenetic analysis of Ind2001 lineage based on maximum likelihood method revealed two major splits and three sub-lineages. The mean nucleotide substitution rate for this lineage was calculated to be 6.338×10(-3)substitutions/site/year (s/s/y), which is similar to those of PanAsian sub-lineages. Evolutionary time scale analysis indicated that the Ind2001 lineage might have originated in 1989. The sub-lineage Ind2001d that caused 2013 outbreaks seems to be relatively more divergent genetically from other Ind2001 sub-lineages. Seven codons in the VP1 region of Ind2001 were found to be under positive selection. Four out of 24 recent Ind2001 strains tested in 2D-MNT had antigenic relationship value of <0.3 with the serotype O vaccine strain indicating intra-epidemic antigenic diversity. Amino acid substitutions found in these minor variants with reference to antigenic diversity have been discussed. The dominance of antigenically homologous strains indicates absence of vaccine immunity in the majority of the affected hosts. Taken together, the evolution of Ind2001 lineage deviates from the strict molecular clock and a typical lineage evolutionary dynamics characterized by periodic emergence and re-emergence of Ind2001 and PanAsia lineage have been observed in respect of serotype O., (Copyright © 2015 Elsevier B.V. All rights reserved.)

In this study we describe the adaptive changes fixed on the capsid of several foot-and-mouth disease virus serotype A strains during propagation in cell monolayers. Viruses passaged extensively in three cell lines (BHK-21, LFBK and IB-RS-2) consistently gained positively charged amino acids in the putative heparin-sulfate-binding pocket (VP2 βE-βF loop, VP1 C-terminus and VP3 β-B knob) surrounding the fivefold symmetry axis (VP1 βF-βG loop) and at other discrete sites on the capsid (VP3 βG-βH loop, VP1 C-terminus, VP2 βC strand and VP1 βG-βH loop). A lysine insertion in the VP1 βF-βG loop of two of the BHK-21-adapted viruses supports the biological advantage of positively charged residues acquired in cell culture. The charge transitions occurred irrespective of cell line, suggesting their possible role in ionic interaction with ubiquitous negatively charged cell-surface molecules such as glycosaminoglycans (GAG). This was supported by the ability of the cell-culture-adapted variants to replicate in the integrin-deficient, GAG-positive CHO-K1 cells and their superior fitness in competition assays compared with the lower passage viruses with WT genotypes. Substitutions fixed in the VP1 βG-βH loop (-3, -2 and +2 'RGD' positions) or in the structural element known to be juxtaposed against that loop (VP1 βB-βC loop) suggest their possible role in modulating the efficiency and specificity of interaction of the 'RGD' motif with αv-integrin receptors. The nature and location of the substitutions described in this study could be applied in the rapid cell culture adaptation of viral strains for vaccine production., (© 2015 The Authors.)

The formation of metallic particulates from erosion was investigated by running a series of transducers at various frequencies in water. Two low frequency transducer sonotrodes were run for 7.5h at 18kHz and 20kHz. Three high frequency plates operating at megasonic frequencies of 0.4MHz, 1MHz, and 2MHz were run over a 7days period. Electrical conductivity and pH of the solution were measured before and after each run. A portion of the non-sonicated and treated water was partially evaporated to achieve an 80-fold concentration of particles and then sieved through nano-filters of 0.1μm, 0.05μm, and 0.01μm. An aliquot of the evaporated liquid was also completely dried on strips of carbon tape to determine the presence of finer particles post sieving. An aliquot was analyzed for detection of 11 trace elements by Inductively Coupled Plasma Mass Spectroscopy (ICPMS). The filters and carbon tapes were analyzed by FE-SEM imaging to track the presence of metals by EDS (Energy Dispersive Spectroscopy) and measure the particle size and approximate composition of individual particles detected. Light microscopy visualization was used to calculate the area occupied by the particles present in each filter and high resolution photography was used for visualization of sonotrode surfaces. The roughness of all transducers before and after sonication was tested through profilometry. No evidence of formation of nano-particles was found at any tested frequency. High amounts of metallic micron-sized particles at 18kHz and 20kHz formed within a day, while after 7day runs only a few metallic micro particles were detected above 0.4MHz. Erosion was corroborated by an increase in roughness in the 20kHz tip after ultrasound. The elemental analysis showed that metal leach occurred but values remained below accepted drinking water limits, even after excessively long exposure to ultrasound. With the proviso that the particles measured here were only characterized in two dimensions and could be nanoparticulate in terms of the third dimension, this research suggests that there are no serious health implications resulting from the formation of nanoparticles under the evaluation conditions. Therefore, high frequency transducer plates can be safely operated in direct contact with foods. However, due to significant production of metallic micro-particulates, redesign of lower frequency sonotrodes and reaction chambers is advised to enable operation in various food processing direct-contact applications., (Copyright © 2014 Elsevier B.V. All rights reserved.)

In this study, an RNA transfection was used to rescue infectious foot-and-mouth disease (FMD) virus from clinical samples in BHK-21 cell line for diagnosis of FMD. Tissue samples (n=190) were subjected to FMD virus isolation by conventional cell culture and also by RNA transfection. FMD virus was isolated from 62% of the clinical samples by RNA transfection, whereas virus was isolated only from 16% of the clinical samples in conventional cell culture method, suggesting better performance of the RNA transfection. Virus was rescued from 67% and 10% of ELISA negative but multiplex PCR positive samples by RNA transfection and conventional cell culture, respectively. The efficiency of transfection was studied on clinical samples subjected to temperature as high as 37°C and varying pH (pH 4-9). Except up to 1 week of storage at 4°C at pH 7.5, virus isolation was not possible by cell culture. Virus was rescued by transfection from samples stored at 4°C for any of the applied pH up to 4 weeks, and when stored at 37°C virus could be rescued up to 4 weeks at pH 7.5 suggesting the fitness of transfection to isolate virus from clinical samples stored under inappropriate conditions. The sequence data and antigenic relationships with the vaccine strains, between virus rescued by transfection and conventional cell culture, were comparable. The RNA transfection will help to increase the efficiency of virus isolation, diagnosis and molecular epidemiological studies., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Serological investigation to detect foot-and-mouth disease (FMD) virus circulation in the domestic small ruminant population of India was conducted. A total of 4407 and 4035 serum samples from sheep and goats, respectively were collected at random covering majority of the states across the country during 2010-2012. These samples were analyzed for antibodies against the non-structural proteins (NSP) of FMD virus in an indirect 3AB NSP ELISA and against the structural proteins (SP) in a liquid phase blocking (LPB) ELISA. A total of 20.35% sheep and 13.60% goats were found to be positive for 3AB NSP antibodies providing a serological evidence of extensive viral activity. In LPB ELISA, only 4.54% sheep and 6.27% goats were found to have protective antibody (log10 titre of ≥1.8) against all three serotype strains in the vaccine, which correlates with "no or sparse vaccination" scenario in these species in the country. Hence, to check silent amplification and dissemination of virus in a mixed farming set up, small ruminants may be brought under the ambit of routine vaccination and surveillance programmes., (Copyright © 2013 Elsevier B.V. All rights reserved.)

In India, emergence of Ind2001 lineage of foot-and-mouth disease virus (FMDV) serotype O was recorded in the year 2001. After causing sporadic incidences, the Ind2001 lineage that re-surged in 2008 out-competed PanAsia from the field during 2009 and continued its dominance during 2010 and 2011 as well. The lineage has diversified in due course of time, leading to two sub-lineages (Ind2001a and Ind2001b). The sub-lineage Ind2001a include isolates collected during 2001-2002 and sub-lineage Ind2001b is constituted largely by isolates collected during 2008-2012. The nucleotide substitution rate of sub-lineage Ind2001b was estimated at 6.58×10⁻³ substitutions/site/year. The most stable PanAsia lineage is restricted only to few outbreaks. During 2011, emergence of a new genetic group with >9% nucleotide divergence from rest of the lineages circulating in the country was detected and named as lineage Ind2011. Two specific amino acid substitutions at positions VP1-36 (F) and VP2-133 (T) were observed in the Ind2011 lineage. The new lineage at present is restricted only to southern states of the country. It is uncertain whether the emergence was triggered by immune pressure or due to a bottleneck in transmission or selected for higher fitness value. Six sites (4, 68, 83, 135, 138 and 209) in VP1 protein were identified to undergo episodic diversifying selection in serotype O field isolates. Both emerging and re-emerging lineages had appropriate antigenic match with currently used vaccine strain, INDR2/1975. Irrespective of genetic variability, the field isolates showed remarkable conservation at antigenically critical residues that might contribute to the observed antigenic stability. With the emergence of a new genetic group after a span of 10 years, the overall epidemiological scenario in the region is expected to change in the coming years., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Recent research has shown that high frequency ultrasound (0.4-3 MHz), can enhance milkfat separation in small scale systems able to treat only a few milliliters of sample. In this work, the effect of ultrasonic standing waves on milkfat creaming was studied in a 6L reactor and the influence of different frequencies and transducer configurations in direct contact with the fluid was investigated. A recombined coarse milk emulsion with fat globules stained with oil-red-O dye was selected for the separation trials. Runs were performed with one or two transducers placed in vertical (parallel or perpendicular) and horizontal positions (at the reactor base) at 0.4, 1 and/or 2 MHz (specific energy 8.5 ± 0.6 kJ/kg per transducer). Creaming behavior was assessed by measuring the thickness of the separated cream layer. Other methods supporting this assessment included the measurement of fat content, backscattering, particle size distribution, and microscopy of samples taken at the bottom and top of the reactor. Most efficient creaming was found after treatment at 0.4 MHz in single and double vertical transducer configurations. Among these configurations, a higher separation rate was obtained when sonicating at 0.4 MHz in a vertical perpendicular double transducer setup. The horizontal transducer configuration promoted creaming at 2 MHz only. Fat globule size increase was observed when creaming occurred. This research highlights the potential for enhanced separation of milkfat in larger scale systems from selected transducer configurations in contact with a dairy emulsion, or emulsion splitting in general., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The central dogma of molecular biology - DNA makes RNA makes proteins - is a flow of information that in eukaryotes encounters a physical barrier: the nuclear envelope, which encapsulates, organizes and protects the genome. Nuclear-pore complexes, embedded in the nuclear envelope, regulate the passage of molecules to and from the nucleus, including the poorly understood process of the export of RNAs from the nucleus. Recent imaging approaches focusing on single molecules have provided unexpected insight into this crucial step in the information flow. This review addresses the latest studies of RNA export and presents some models for how this complex process may work.

India is endemic for foot-and-mouth disease (FMD), and goats constitute the second largest susceptible population of domestic livestock. FMD surveillance and control strategies in the country largely ignore small ruminants, known to be critical in the epidemiology of the disease. Here, serological investigations were carried out to generate estimates of antibody prevalence in goats of Orissa state to both non-structural (NSP-Ab) and structural proteins (SP-Ab) of FMD. The apparent overall NSP-Ab and SP-Ab seroprevalences were 38% and 20.7%, respectively, which signifies a very high level of FMD virus circulation in the goat population despite the lack of clinical signs in this species. The apparent prevalence of NSP-Ab and SP-Ab was positively correlated in the sampling areas. Interestingly, the values found for NSP-Ab prevalence were almost consistently higher than those found for SP-Ab prevalence. This could have been attributable to either issues related to sensitivity and specificity of the test systems employed or differences in the post-infection kinetics of NSP- and SP-Ab. The pattern that emerged from SP-Ab analysis indicated goats being infected with all three prevalent serotypes (O, A and Asia 1) and reinforces the concept that non-vaccinated goats can be exploited as tracer animals for detecting serotypes involved in outbreaks. The results underscore the requirement to bring caprine species under comprehensive surveillance and vaccination campaigns to check silent amplification, excretion and transmission of the virus., (© 2010 Blackwell Verlag GmbH.)

The nuclear pore complex (NPC) mediates all transport between the nucleus and cytoplasm. Passage through the NPC is highly selective, yet the same channel must allow rapid specific transport of a wide range of cargoes. This chapter focuses mainly on the phenylalanine-glycine (FG) nucleoporins (nups), proteins carrying natively unfolded regions that are thought to form the selectively permeable barrier within the NPC. The physical properties of the FG nup barrier remain unclear. The high selectivity and rapidity of transport observed in vivo may be explained, in part, by competition for binding and space between transport factors and nontransported proteins. Future studies of FG nups will therefore also examine their interactions between FG nups and other proteins in their surroundings.

To investigate the prevalence of neurocysticercosis among free ranging pigs and to study the type of pathomorphological lesions in affected brains, a total of 200 brains were collected from pigs slaughtered at a local abattoir, between August, 2005 to March, 2006. Gross and histopathological examination revealed 3% (6/200) occurrence of neurocysticercosis in pigs. Taenia solium cysticercosis is an under-rated zoonosis and is a leading cause of epilepsy due to neurocysticercosis in human population of India. The prevailing situation warrants immediate implementation of effective control measures for this dreaded disease.

Objective: The objective of the study was to determine which vehicle factors are significantly related to pelvic injury in side impact collisions. Identification of relevant parameters could aid in the reduction of these injuries., Method: Side impact crashes from the CIREN database were separated into those in which the occupant sustained a pelvic fracture and those in which no pelvic fracture occurred, although all occupants had serious injuries. A multibody MADYMO model was created of a USDOT SINCAP (U.S. Department of Transportation Side Impact New Car Assessment Program) test of a vehicle with a large center console., Results: From a study of 113 side impact crashes in the ciren database, nearside occupants with pelvic fractures (n = 78) had (i) more door intrusion (mean, 37 vs. 32 cm, p = 0.02) than those who had serious injuries, but not pelvic fractures (ii) a greater likelihood that the lower border of the door intruded more than the upper part (40% vs. 18%, p < 0.025); and (iii) a greater likelihood that their vehicle had a center console (47 vs. 17%, p < 0.005). Other parameters such as occupant age, weight, gender, vehicle weight, and struck vehicle speed change were not significantly different. MADYMO modeling showed that with a center console, an initial positive pelvic acceleration occurred at about 30 msec, followed at about 45 msec by a second acceleration peak in the opposite direction. Reducing console stiffness reduced the second acceleration but not the initial peak. Allowing the seat to translate laterally when contacted by the door reduced the initial pelvic acceleration by 50% and eliminated the second acceleration peak., Conclusions: Redesigning the center console using less stiff materials and allowing some lateral translation of the seat could aid in reducing pelvic injuries in side impact collisions.

The present work was broadly divided into two parts: first, the efficiency of synthetically prepared ferrihydrite and Al-ferrihydrite to adsorb dissolved organic matter (DOM) was tested as a function of time, pH and ionic strength and its effects on specific surface area was found. In the second part, the effect of DOM concentration and solution pH was studied to elucidate the influence of this two parameters on As(V) leachability from As(V) loaded ferrihydrite and Al-ferrihydrite. It was found that both pH and ionic strength had significant influence on DOM adsorption. The behavior and magnitude of adsorption was significantly different for both the materials. In general, the efficiency of Al-ferrihydrite to adsorb DOM was better compared to ferrihydrite. Results indicated that ligand exchange was the dominant interaction mechanism for DOM adsorption on ferrihydrite and Al-ferrihydrite. The study also suggested a strong potential for DOM to mobilize As(V) from the studied materials. At pH 5.0, DOM concentrations of 2.5 and 5.0 mg C/L had negligible effect on As(V) leachability. With increasing DOM concentration from 10 to 30 mg C/L, increased the As(V) leachability into the solution. Net release from ferrihydrite was up to 1.07 mg/g (5.75%) of the total adsorbed arsenic. The corresponding figure for Al-ferrihydrite was 1.47 mg/g (6.30%). As the primary mechanisms for the arsenic release from solid phases we identified two different types of mechanisms i.e., dissolution of the solid phase and competition between arsenic and organic anions for sorption sites.

Protozoan parasites of the order Kinetoplastida are responsible for a significant proportion of global morbidity and economic hardship. These organisms also represent extremely distal points within the Eukarya, and one such organism, Trypanosoma brucei, has emerged as a major system for the study of evolutionary cell biology. Significant technical challenges have hampered the full exploitation of this organism, but advances in genomics and proteomics provide a novel approach to acquiring rapid functional data. However, the vast evolutionary distance between trypanosomes and the higher eukaryotes presents significant problems with functional assignment based on sequence similarity, and frequently homologues cannot be identified with sufficient confidence to be informative. Direct identification of proteins in isolated organelles has the potential of providing robust functional insight and is a powerful approach for initial assignment. We have selected the nucleus of T. brucei as a first target for protozoan organellar proteomics. Our purification methodology was able to reliably provide both nuclear and subnuclear fractions. Analysis by gel electrophoresis, electron microscopy, and immunoblotting against trypanosome subcellular markers indicated that the preparations are of high yield and purity, maintain native morphology, and are well resolved from other organelles. Minor developmental differences were observed in the nuclear proteome for the bloodstream and procyclic stages, whereas significant morphological alterations were visible. We demonstrate by direct sequencing that the NUP-1 nuclear envelope antigen is a coiled coil protein, containing approximately 20 near-perfect copies of a 144-amino acid sequence. Immunoelectron microscopy localized NUP-1 to the inner face of the nuclear envelope, suggesting that it is a major filamentous component of the trypanosome nuclear lamina.

The pathway and complete collection of factors that orchestrate ribosome assembly are not clear. To address these problems, we affinity purified yeast preribosomal particles containing the nucleolar protein Nop7p and developed means to separate their components. Nop7p is associated primarily with 66S preribosomes containing either 27SB or 25.5S plus 7S pre-rRNAs. Copurifying proteins identified by mass spectrometry include ribosomal proteins, nonribosomal proteins previously implicated in 60S ribosome biogenesis, and proteins not known to be involved in ribosome production. Analysis of strains mutant for eight of these proteins not previously implicated in ribosome biogenesis showed that they do participate in this pathway. These results demonstrate that proteomic approaches in concert with genetic tools provide powerful means to purify and characterize ribosome assembly intermediates.

The conformation of oat globulin dispersions (10% in D2O) under the influence of pH, chaotropic salts, protein structure perturbants, and heating conditions was studied by Fourier transform infrared (FTIR) spectroscopy. The FTIR spectrum of oat globulin showed major bands from 1670 to 1634 cm(-1), corresponding to the four major types of secondary structures, that is, beta-turns, beta-sheets, alpha-helices, and random coils. At extreme acidic and alkaline pH conditions, there were changes in intensity in the bands attributed to beta-sheet structures (1626, 1634, and 1682 cm(-1)), and shifts of the bands to higher or lower wavenumbers, indicating changes in conformation. In the presence of some chaotropic salts, the 1626 and 1634 cm(-1) bands were shifted upward, with a marked decrease in the intensity of the 1634 cm(-1) peak. The addition of several protein structure perturbants led to a slight shift in the alpha-helix/random coil bands and a marked reduction in the beta-sheet peaks, suggesting protein unfolding. Heating under aggregating conditions led to slight shifts in all of the major bands and progressive changes in the intensity of the alpha-helix, beta-sheet, and beta-turn peaks, suggesting protein denaturation. This was accompanied by marked increases in intensity of the two intermolecular beta-sheet bands (1682 and 1624-1626 cm(-1)) associated with the formation of aggregated strands. The IR spectra of soluble and insoluble aggregates showed a redistribution of native and extensively denatured proteins in the two fractions.

Nucleocytoplasmic transport is mediated by the interplay between soluble transport factors and nucleoporins resident within the nuclear pore complex (NPC). Understanding this process demands knowledge of components of both the soluble and stationary phases and the interface between them. Here, we provide evidence that Nup2p, previously considered to be a typical yeast nucleoporin that binds import- and export-bound karyopherins, dynamically associates with the NPC in a Ran-facilitated manner. When bound to the NPC, Nup2p associates with regions corresponding to the nuclear basket and cytoplasmic fibrils. On the nucleoplasmic face, where the Ran--GTP levels are predicted to be high, Nup2p binds to Nup60p. Deletion of NUP60 renders Nup2p nucleoplasmic and compromises Nup2p-mediated recycling of Kap60p/Srp1p. Depletion of Ran--GTP by metabolic poisoning, disruption of the Ran cycle, or in vitro by cell lysis, results in a shift of Nup2p from the nucleoplasm to the cytoplasmic face of the NPC. This mobility of Nup2p was also detected using heterokaryons where, unlike nucleoporins, Nup2p was observed to move from one nucleus to the other. Together, our data support a model in which Nup2p movement facilitates the transition between the import and export phases of nucleocytoplasmic transport.

Medico- etnobotanical exploration carried out in Dhenkanal district of Orissa during 1996-98 reveal that, people use 21 plant species belonging o 20 genera and 17 families on 10 different combinations for the treatment of diarrehoeal diseases. The method of preparation of medicine and details of application care recorded.

Analysis of Raman spectra of oat globulin showed that extreme pH values caused an increase in the amide and C-H stretching band intensity, indicating changes in the secondary structures of the protein due to denaturation. Similar changes were observed when oat globulin was treated with chaotropic salts and several protein perturbants. Sodium dodecyl sulfate, beta-mercaptoethanol, and ethylene glycol also caused a shift in the amide III' band, suggesting a transition from beta-sheet to a random coil conformation. Heating at temperatures near the denaturation temperature of oat globulin led to increases in the amide and C-H band intensity, indicating unfolding of the protein. The data indicate that FT-Raman spectroscopy is suitable for studying the secondary structure of plant proteins such as oat globulin.

An understanding of how the nuclear pore complex (NPC) mediates nucleocytoplasmic exchange requires a comprehensive inventory of the molecular components of the NPC and a knowledge of how each component contributes to the overall structure of this large molecular translocation machine. Therefore, we have taken a comprehensive approach to classify all components of the yeast NPC (nucleoporins). This involved identifying all the proteins present in a highly enriched NPC fraction, determining which of these proteins were nucleoporins, and localizing each nucleoporin within the NPC. Using these data, we present a map of the molecular architecture of the yeast NPC and provide evidence for a Brownian affinity gating mechanism for nucleocytoplasmic transport.

NPCs are the sole sites of exchange between the nucleus and cytoplasm. A large family of transport factors carry cargo between the nucleus and cytoplasm through the NPC. The NPC is a huge symmetric octagonal structure comprised of dozens of NUPs. NUPs make many contacts with surrounding structures, including the NE, the cytoplasm and nuclear interior. A subset of NUPs contain repeated peptide motifs that serve as docking sites for transport factors. The directionality of transport is determined by the transport factor, and its interactions with the small GTPase Ran and NUPs. Very little is known about how the NPC mediates transport, NPC assembly and the NPC's role in regulating transport, but these areas of research are beginning to emerge.

The 26 kD basic subunit of 280 kD buckwheat grain legumin has been partially characterized by measurement of its fluorescence and CD spectra. The protein has 22% alpha-helix, 36% beta-sheet, 12% beta-turn and 30% random coil secondary structure. In comparison with the basic subunits of other legumin-type proteins, the buckwheat legumin subunit has a high content of lysine and methionine. The protein also has higher ratios of lysine to arginine and methionine to arginine.

While much has been learned in recent years about the movement of soluble transport factors across the nuclear pore complex (NPC), comparatively little is known about intranuclear trafficking. We isolated the previously identified Saccharomyces protein Mlp1p (myosin-like protein) by an assay designed to find nuclear envelope (NE) associated proteins that are not nucleoporins. We localized both Mlp1p and a closely related protein that we termed Mlp2p to filamentous structures stretching from the nucleoplasmic face of the NE into the nucleoplasm, similar to the homologous vertebrate and Drosophila Tpr proteins. Mlp1p can be imported into the nucleus by virtue of a nuclear localization sequence (NLS) within its COOH-terminal domain. Overexpression experiments indicate that Mlp1p can form large structures within the nucleus which exclude chromatin but appear highly permeable to proteins. Remarkably, cells harboring a double deletion of MLP1 and MLP2 were viable, although they showed a slower net rate of active nuclear import and faster passive efflux of a reporter protein. Our data indicate that the Tpr homologues are not merely NPC-associated proteins but that they can be part of NPC-independent, peripheral intranuclear structures. In addition, we suggest that the Tpr filaments could provide chromatin-free conduits or tracks to guide the efficient translocation of macromolecules between the nucleoplasm and the NPC.

We report a novel connection between nuclear pore complexes (NPCs) and spindle pole bodies (SPBs) revealed by our studies of the Saccharomyces cerevisiae NDC1 gene. Although both NPCs and SPBs are embedded in the nuclear envelope (NE) in yeast, their known functions are quite distinct. Previous work demonstrated that NDC1 function is required for proper SPB duplication (Winey, M., M.A. Hoyt, C. Chan, L. Goetsch, D. Botstein, and B. Byers. 1993. J. Cell Biol. 122:743-751). Here, we show that Ndc1p is a membrane protein of the NE that localizes to both NPCs and SPBs. Indirect immunofluorescence microscopy shows that Ndc1p displays punctate, nuclear peripheral localization that colocalizes with a known NPC component, Nup49p. Additionally, distinct spots of Ndc1p localization colocalize with a known SPB component, Spc42p. Immunoelectron microscopy shows that Ndc1p localizes to the regions of NPCs and SPBs that interact with the NE. The NPCs in ndc1-1 mutant cells appear to function normally at the nonpermissive temperature. Finally, we have found that a deletion of POM152, which encodes an abundant but nonessential nucleoporin, suppresses the SPB duplication defect associated with a mutation in the NDC1 gene. We show that Ndc1p is a shared component of NPCs and SPBs and propose a shared function in the assembly of these organelles into the NE.

In eukaryotic cells, a regulated flux of molecules between the cytoplasm and the nucleus maintains two very different environments while allowing the controlled exchange of macromolecules necessary for their individual functions. Molecules entering or leaving the nucleus use nuclear localization signals or nuclear export signals to pass through selective channels in the nuclear envelope formed by nuclear pore complexes. The recognition of signal-bearing cargo, its interaction with the nuclear pore complex and its translocation through the pore complex are mediated by soluble transport factors. Recently, the list of potential transport factors has grown rapidly, suggesting a previously unanticipated level of complexity for nuclear transport.