Identification of a Novel Variant c.163delG in HBB Gene Resulting in a Beta Null Phenotype in a Proband with Thalassemia Intermedia.

A 21-year-old patient presented with a previous medical history of pallor, mild icterus, increased fatigue, low hemoglobin, and abnormal hemoglobin variant analysis with more than 70 transfusions. He was referred for genetic analysis to identify the pathogenic variations in the ß-globin gene. Sanger's sequencing of the proband and his family revealed the presence of a novel frame shift variant HBB:c.163delG in a compound heterozygous state with hemoglobin E (HbE) (HBB:c.79G > A) variant. The father and the sibling of the patient were found to be normal for the HBB gene. Mother was found to be heterozygous for HbE (HBB:c.79G > A) variant. In silico analysis by Mutalyzer predicted that c.163delG variant generated a premature stop codon after seven codons, leading to a truncated protein. FoldX protein stability analysis showed a positive ΔΔG value of 45.27 kcal/mol suggesting a decrease in protein stability. HBB:c.79G > A is a known variant coding for HbE variant, which results in the reduced synthesis of ß-globin chain and shows mild thalassemia. Combined effect of HBB:c.163delG and HBB:c.79G > A variants in the proband might have led to the reduced synthesis of ß-globin chains resulting in a thalassemia intermedia type of clinical manifestation.

Spatial distribution and potential biological risk of some metals in relation to granulometric content in core sediments from Chilika Lake, India.

The article presents first systematic report on the concentration of selected major elements [iron (Fe) and manganese (Mn)] and minor elements [zinc (Zn), copper (Cu), chromium (Cr), lead (Pb), nickel (Ni), and cobalt (Co)] from the core sediment of Chilika Lake, India. The analyzed samples revealed higher content of Pb than the background levels in the entire study area. The extent of contamination from minor and major elements is expressed by assessing (i) the metal enrichments in the sediment through the calculations of anthropogenic factor (AF), pollution load index (PLI), Enrichment factor (EF), and geoaccumulation index (Igeo) and (ii) potential biological risks by the use of sediment quality guidelines like effect range median (ERM) and effect range low (ERL) benchmarks. The estimated indices indicated that sediment is enriched with Pb, Ni, Cr, Cu and Co. The enrichment of these elements seems to be due to the fine granulometric characteristics of the sediment with Fe and Mn oxyhydroxides being the main metal carriers and fishing boats using low grade paints, fuel, and fishing technology using lead beads fixed to fishing nets. Trace element input to the Chilika lake needs to be monitored with due emphasis on Cr and Pb contaminations since the ERM and ERL benchmarks indicated potential biological risk with these metals.

Optimal Levothyroxine Replacement Adequately Improves Symptoms of Hypothyroidism; Residual Symptoms Need Further Evaluation for Other than Hypothyroidism Causation.

OBJECTIVES: Many patients with hypothyroidism complain of persistent residual symptoms, despite optimal treatment, although the similar prevalence is seen in patients with documented absence of thyroid disorder in primary health-care setup. We aimed to investigate symptomatic relief in new cases of primary hypothyroidism and compare with controls with other chronic conditions. METHODS: This prospective case-control follow-up study enrolled patients from July 2014 to May 2015 in an endocrine outpatient clinic of a tertiary hospital. Controls were age- and gender-matched ambulatory individuals with well-controlled other chronic diseases and no major comorbidity. Thyroid symptom questionnaire (TSQ) was administered at pretreatment to all the cases and then they were started on levothyroxine (LT). At euthyroidism, TSQ was readministered. For controls, TSQ was administered only once. TSQ was measured on Likert scale 1-4 for lack of energy, dry skin, constipation, aches and pains, cold intolerance, poor memory, depression, weight gain, tiredness after walking, and difficulty in getting up (DGU). P < 0.05 was considered statistically significant. RESULTS: A total of 194 cases (147 females and 47 males) and 259 controls (187 females and 72 males) were analyzed. A significant difference in the symptoms prevalence was seen between controls and pretreatment cases, except for DGU, and between subclinical and overt hypothyroidism. Pretreatment serum thyroid-stimulating hormone in cases correlated significantly with all their pretreatment symptoms score. All symptoms prevalence decreased significantly posttreatment. At euthyroidism, the mean symptoms score in posttreatment cases was similar or lower than the controls. CONCLUSION: LT effectively improves the symptoms of hypothyroidism in newly diagnosed cases of primary hypothyroidism. The residual symptoms need an alternation explanation other than hypothyroidism.

Sources and Variability of Petroleum Hydrocarbon Residues in Sediments of Chilika Lagoon, East Coast of India.

The spatio-temporal distribution and the controlling factors of petroleum hydrocarbons (PHCs) in sediments of Chilika lagoon was investigated. Samples were collected during three seasons and quantified using UV-fluorescence spectroscopy. Concentrations of PHCs in surface sediments varies from 0.18 to 12.13 ppm (mean 3.71 ± 3.94 ppm). Compared to the lagoon, the monitoring stations adjacent to jetties with high boating activities tend to have higher PHC concentrations, suggesting that the contribution is likely to be from fossil fuel combustion and accidental seepage. The sediment organic matter (OM) of Chilika ranges from 0.26% to 6.23%. PHC maintains a positive correlation with OM (p < 0.05; f = 0.334), indicating the long term deposition of PHC as sediment OM. However, there is no significant relation between PHC and sediment texture, indicating its negligible control over PHC. The recorded PHC concentrations are below the threshold limit (70 ppm) as classified by United States (US) National Academy of Sciences (NAS) and also lower than those reported from similar ecosystems in India and overseas. Since the long term deposition and the bioaccumulation of PHC cannot be avoided, it is essential to monitor these parameters periodically.

Spatiotemporal distribution and composition of phytoplankton assemblages in a coastal tropical lagoon: Chilika, India.

The Asia's largest lagoon, Chilika, is a shallow water estuary and a designated "Ramsar" site located in the east coast of India. The spatiotemporal diversity of phytoplankton based on the monthly sampling between July 2011 and June 2012 was investigated in relation to physicochemical variables of the surface water column from 13 stations. The salinity was minimum (average 9) during the monsoon which was primarily due to riverine discharge. As the season progressed towards post-monsoon, average salinity of the whole lagoon reached to 10 which further increased to 20 during pre-monsoon season. A total of 259 species of phytoplankton, mostly dominated by the Bacillariophyta (138 species) followed by Dinophyta (38 species), Chlorophyta (32 species), Cyanophyta (29 species), Euglenophyta (18 species), and Chrysophyta (4 species), were recorded in this study. Different ecological sectors of the lagoon (except the northern sector) were dominated by diatoms, while the northern sector due to its freshwater regime supported large population of euglenoids. Based on the multivariate ordination analysis, salinity regime and light availability played important role in determining the distribution, diversity, and composition of phytoplankton communities. Overall, this study documented a very high diversity of phytoplankton and highlighted the importance of taking extensive sampling in getting a clearer understanding of phytoplankton community structure in less-studied environments such as Chilika lagoon.

Alpha-1 giardin is an annexin with highly unusual calcium-regulated mechanisms.

Alpha-giardins constitute the annexin proteome (group E annexins) in the intestinal protozoan parasite Giardia and, as such, represent the evolutionary oldest eukaryotic annexins. The dominance of alpha-giardins in the cytoskeleton of Giardia with its greatly reduced actin content emphasises the importance of the alpha-giardins for the structural integrity of the parasite, which is particularly critical in the transformation stage between cyst and trophozoite. In this study, we report the crystal structures of the apo- and calcium-bound forms of α1-giardin, a protein localised to the plasma membrane of Giardia trophozoites that has recently been identified as a vaccine target. The calcium-bound crystal structure of α1-giardin revealed the presence of a type III site in the first repeat as known from other annexin structures, as well as a novel calcium binding site situated between repeats I and IV. By means of comparison, the crystal structures of three different alpha-giardins known to date indicate that these proteins engage different calcium coordination schemes, among each other, as well as compared to annexins of groups A-D. Evaluation of the calcium-dependent binding to acidic phosphoplipid membranes revealed that this process is not only mediated but also regulated by the environmental calcium concentration. Uniquely within the large family of annexins, α1-giardin disengages from the phospholipid membrane at high calcium concentrations possibly due to formation of a dimeric species. The observed behaviour is in line with changing calcium levels experienced by the parasite during excystation and may thus provide first insights into the molecular mechanisms underpinning the transformation and survival of the parasite in the host.

SBAL: a practical tool to generate and edit structure-based amino acid sequence alignments.

SUMMARY: Both alignment generation and visualization are important processes for producing biologically meaningful sequence alignments. Computational tools that combine reliable, automated and semi-automated approaches to produce secondary structure-based alignments with an appropriate visualization of the results are rare. We have developed SBAL, a tool to generate and edit secondary structure-based sequence alignments. It is easy to install and provides a user-friendly interface. Sequence alignments are displayed, with secondary structure assignments mapped to their corresponding regions in the sequence by using a simple colour scheme. The algorithm implemented for automated and semi-automated secondary structure-based alignment calculations shows a comparable performance to existing software. AVAILABILITY AND IMPLEMENTATION: SBAL has been implemented in Java to provide cross-platform compatibility. SBAL is freely available to academic users at http://www.structuralchemistry.org/pcsb/. Users will be asked for their name, institution and email address. A manual can also be downloaded from this site. The software, manual and test sets are also available as supplementary material. CONTACT: conan.wang@griffith.edu.au SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

DMAN: a Java tool for analysis of multi-well differential scanning fluorimetry experiments.

SUMMARY: Differential scanning fluorimetry (DSF) is a rapid technique that can be used in structural biology to study protein-ligand interactions. We have developed DMAN, a novel tool to analyse multi-well plate data obtained in DSF experiments. DMAN is easy to install and provides a user-friendly interface. Multi-well plate layouts can be designed by the user and experimental data can be annotated and analysed by DMAN according to the specified plate layout. Statistical tests for significance are performed automatically, and graphical tools are also provided to assist in data analysis. The modular concept of this software will allow easy development of other multi-well plate analysis applications in the future. AVAILABILITY AND IMPLEMENTATION: DMAN is implemented in Java to provide a cross-platform compatibility. It is freely available to academic users at http://www.structuralchemistry.org/pcsb/. To download DMAN, users will be asked for their name, institution and email address. A manual can also be downloaded from this site. CONTACT: conan.wang@griffith.edu.au; a.hofmann@griffith.edu.au.

Structural studies of bacterioferritin B from Pseudomonas aeruginosa suggest a gating mechanism for iron uptake via the ferroxidase center .

The structure of recombinant Pseudomonas aeruginosa bacterioferritin B (Pa BfrB) has been determined from crystals grown from protein devoid of core mineral iron (as-isolated) and from protein mineralized with approximately 600 iron atoms (mineralized). Structures were also obtained from crystals grown from mineralized BfrB after they had been soaked in an FeSO(4) solution (Fe soak) and in separate experiments after they had been soaked in an FeSO(4) solution followed by a soak in a crystallization solution (double soak). Although the structures consist of a typical bacterioferritin fold comprised of a nearly spherical 24-mer assembly that binds 12 heme molecules, comparison of microenvironments observed in the distinct structures provided interesting insights. The ferroxidase center in the as-isolated, mineralized, and double-soak structures is empty. The ferroxidase ligands (except His130) are poised to bind iron with minimal conformational changes. The His130 side chain, on the other hand, must rotate toward the ferroxidase center to coordinate iron. In comparison, the structure obtained from crystals soaked in an FeSO(4) solution displays a fully occupied ferroxidase center and iron bound to the internal, Fe((in)), and external, Fe((out)), surfaces of Pa BfrB. The conformation of His130 in this structure is rotated toward the ferroxidase center and coordinates an iron ion. The structures also revealed a pore on the surface of Pa BfrB that likely serves as a port of entry for Fe(2+) to the ferroxidase center. On its opposite end, the pore is capped by the side chain of His130 when it adopts its "gate-closed" conformation that enables coordination to a ferroxidase iron. A change to its "gate-open", noncoordinative conformation creates a path for the translocation of iron from the ferroxidase center to the interior cavity. These structural observations, together with findings obtained from iron incorporation measurements in solution, suggest that the ferroxidase pore is the dominant entry route for the uptake of iron by Pa BfrB. These findings, which are clearly distinct from those made with Escherichia coli Bfr [Crow, A. C., Lawson, T. L., Lewin, A., Moore, G. R., and Le Brun, N. E. (2009) J. Am. Chem. Soc. 131, 6808-6813], indicate that not all bacterioferritins operate in the same manner.

Binding of Pseudomonas aeruginosa apobacterioferritin-associated ferredoxin to bacterioferritin B promotes heme mediation of electron delivery and mobilization of core mineral iron.

The bfrB gene from Pseudomonas aeruginosa was cloned and expressed in Escherichia coli. The resultant protein (BfrB), which assembles into a 445.3 kDa complex from 24 identical subunits, binds 12 molecules of heme axially coordinated by two Met residues. BfrB, isolated with 5-10 iron atoms per protein molecule, was reconstituted with ferrous ions to prepare samples with a core mineral containing 600 +/- 40 ferric ions per BfrB molecule and approximately one phosphate molecule per iron atom. In the presence of sodium dithionite or in the presence of P. aeruginosa ferredoxin NADP reductase (FPR) and NADPH, the heme in BfrB remains oxidized, and the core iron mineral is mobilized sluggishly. In stark contrast, addition of NADPH to a solution containing BfrB, FPR, and the apo form of P. aeruginosa bacterioferritin-associated ferredoxin (apo-Bfd) results in rapid reduction of the heme in BfrB and in the efficient mobilization of the core iron mineral. Results from additional experimentation indicate that Bfd must bind to BfrB to promote heme mediation of electrons from the surface to the core to support the efficient mobilization of ferrous ions from BfrB. In this context, the thus far mysterious role of heme in bacterioferritins has been brought to the front by reconstituting BfrB with its physiological partner, apo-Bfd. These findings are discussed in the context of a model for the utilization of stored iron in which the significant upregulation of the bfd gene under low-iron conditions [Ochsner, U. A., Wilderman, P. J., Vasil, A. I., and Vasil, M. L. (2002) Mol. Microbiol. 45, 1277-1287] ensures sufficient concentrations of apo-Bfd to bind BfrB and unlock the iron stored in its core. Although these findings are in contrast to previous speculations suggesting redox mediation of electron transfer by holo-Bfd, the ability of apo-Bfd to promote iron mobilization is an economical strategy used by the cell because it obviates the need to further deplete cellular iron levels to assemble iron-sulfur clusters in Bfd before the iron stored in BfrB can be mobilized and utilized.

Gadolinium neutron capture in glioblastoma multiforme cells.

PURPOSE: A proof of principle for cell killing by Gadolinium (Gd) neutron capture in Magnevist preloaded Glioblastoma multiforme (GBM) cells is provided. MATERIALS AND METHODS: U87cells were pre-loaded with 5 mg/ml Magnevist (Gd containing compound) and irradiated using an enhanced neutron beam developed at NIU Institute for Neutron Therapy at Fermilab. These experiments were possible because of an enhanced fast neutron therapy assembly designed to use the fast neutron beam at Fermilab to deliver a neutron beam containing a greater fraction of thermal neutrons and because of the development of improved calculations for dose for the enhanced neutron beam. Clonogenic response was determined. RESULTS: U87 cell survival after gamma irradiation, fast neutron irradiation and irradiation with the enhanced neutron beam in the presence or absence of Magnevist were determined. CONCLUSIONS: U87 cells were the least sensitive to gamma radiation, and increasingly sensitive to fast neutron irradiation, irradiation with the enhanced neutron beam and finally a significant enhancement in cell killing was observed for U87 cells preloaded with Magnevist. The sensitivity of U87 cells pre-loaded with Magnevist and then irradiated with the enhanced neutron beam can at least in part be attributed to the Auger electrons emitted by the neutron capture event.

Trends in absenteeism rates following psychological intervention-preliminary results.

OBJECTIVE: To determine the relationship between absenteeism rates and psychological intervention over a 6-month period in an agency with 334 employees in Klang Valley, Malaysia, which had the highest absenteeism rates. METHODS: The absenteeism rates were measured every 6 months between January 1991 and December 1992. All employees were interviewed and those who scored one positive item on the diagnosis interviewed schedule (DIS) screening, were given counselling, referrals to appropriate agencies and treatment by 2 psychiatrists. RESULTS: Between July and December 1992, the frequency of absenteeism decreased, but the severity rate of absenteeism and the mean length (of a spell and lost time percentage) were higher compared to those in the preceding 6 months. We found that the percentage of absenteeism rate of less than 7 days was significantly reduced between July and December 1992, as compared to July and December 1991. CONCLUSION: Psychological interventions were likely to have contributed to the decreased absenteeism rates of less than 7 days but not for longer periods of absenteeism, which can be attributed to more serious illness or injuries.

Relationship of stress, experienced by rescue workers in the highland towers condominium collapse to probable risk factors--a preliminary report.

The aim of this study was to examine the relationship between the level of stress experienced by rescue workers after the collapse of a 13 story condominium in Kuala Lumpur, and other probable risk factors. Within a month of the incident, 123 firefighters filled up the Impact of Life Event score (Horowitz) and the General Health Questionnaire (GHQ). The results indicated that 7 (6%) firemen could be classified as possible 'cases' on the GHQ, and significantly 5 from this group also scored highly on the impact of events score. No other risk factors were identified in the firemen. On conclusion, the GHQ can be used to screen those with high impact scores to pick up possible cases early enough, so that intervention can be successful.

Neutron irradiation of human pelvic tissues yields a steep dose-response function for late sequelae.

PURPOSE: Analysis of the dose-response function in normal tissues following pelvic irradiation for carcinoma of the prostate. METHODS AND MATERIALS: A homogeneous group of 136 patients with locally advanced carcinoma of the prostate were treated with the Fermilab high-energy neutron beam at three dose levels: 19, 20.4, and 21 Gy, using the same treatment plan and fractionation scheme for all patients. RESULTS: Tumor control rates were about 83% at the three dose levels studied. However, the normal tissue complication rate (late sequelae) varied with dose: 0 out of 5 at 19 Gy, 5 out of 58 (8.6%) at 20.4 Gy, and 9 out of 73 (12.3%) at 21 Gy. CONCLUSIONS: Neutron therapy to the pelvis reveals a steep dose-response function for late effects with a coefficient of variation of only 11%. This is lower than that usually observed with photons or with less uniform clinical data sets, and may be characteristic for well-planned high-LET radiotherapy.

Neutron versus photon irradiation for unresectable salivary gland tumors: final report of an RTOG-MRC randomized clinical trial. Radiation Therapy Oncology Group. Medical Research Council.

PURPOSE: To compare the efficacy of fast neutron radiotherapy versus conventional photon and/or electron radiotherapy for unresectable, malignant salivary gland tumors a randomized clinical trial comparing was sponsored by the Radiation Therapy Oncology Group in the United States and the Medical Research Council in Great Britain. METHODS AND MATERIALS: Eligibility criteria included either inoperable primary or recurrent major or minor salivary gland tumors. Patients were stratified by surgical status (primary vs. recurrent), tumor size (less than or greater than 5 cm), and histology (squamous or malignant mixed versus other). After a total of 32 patients were entered onto this study, it appeared that the group receiving fast neutron radiotherapy had a significantly improved local/regional control rate and also a borderline improvement in survival and the study was stopped earlier than planned for ethical reasons. Twenty-five patients were study-eligible and analyzable. RESULTS: Ten-year follow-up data for this study is presented. On an actuarial basis, there continues to be a statistically-significant p = 0.009) but there is no improvement in overall survival (15% vs. 25%, p = n.s.). Patterns of failure are analyzed and it is shown that distant metastases account for the majority of failures on the neutron arm and local/regional failures account for the majority of failures on the photon arm. Long-term, treatment-related morbidity is analyzed and while the incidence of morbidity graded "severe" was greater on the neutron arm, there was no significant difference in "life-threatening" complications. This work is placed in the context of other series of malignant salivary gland tumors treated with definitive radiotherapy. CONCLUSIONS: Fast neutron radiotherapy appears to be the treatment-of-choice for patients with inoperable primary of recurrent malignant salivary gland tumors.

Prognostic implications of prostate-specific antigen in patients with locally advanced prostate cancer treated with high energy neutron beam therapy: preliminary results.

Serial serum prostate-specific antigen (PSA) levels were analyzed retrospectively for prognostic implications in 70 patients with locoregional (Stages B2, C, and D1) prostate cancer who were managed with high energy neutron beam therapy. Three groups of patients were identified. Group I included 30 patients whose serum PSA level decreased to the reference range (0-4 ng/mL) following neutron therapy and remained so subsequently: 28 (93%) remained disease-free and 2 (7%) have failed distantly. All 30 patients (100%) had no evidence of locally progressive disease. This group was categorized as having a good prognosis. The mean time for serum PSA value to decline to reference range was six months; calculated mean time to achieve a stable base-line PSA was 53 +/- 37 days. Follow-up period ranged from twelve to fifty-six months (median: 21 months). Group II consisted of 13 patients in whom there was an initial decrease in serum PSA to reference range followed by a subsequent increase: 6 of 13 (46%) have no overt clinical progression of disease; 7 (54%) have either persistent locoregional or distant metastatic disease. Follow-up period was from twelve to seventy-two months (median: 39 months). Calculated mean time to achieve stable baseline PSA for serum PSA in this group was 61 +/- 21 days. Group III patients had a persistently elevated or rising serum PSA concentration. Of 27 patients in this group, only 9 (33%) have no evidence of disease progression, while 18 patients (67%) have failed already, either locoregionally or distantly. Follow-up period ranged from twelve to sixty-nine months (median: 21 months). Mean time to achieve stable baseline of serum PSA in this cohort of patients with a poor prognosis was 108 +/- 76 days. We conclude that PSA has a predictable prognostic value in patients with locally advanced prostate cancer managed with high energy neutron beam therapy. Rapid normalization of PSA after therapy indicates a good prognosis. Persistent elevation signifies either presence of persistent locoregional disease or development of distant metastases. Subsequent elevation of the serum PSA concentration after definitive therapy signals progression of prostate cancer.

Pattern of alcoholism in the General Hospital, Kuala Lumpur.

This study establishes the prevalence rate for alcoholism among the inpatients of the General Hospital, Kuala Lumpur, as 11%, but as 25% among the drinking population. It also describes the demographic profile of the alcoholic as compared to the non-alcoholic drinker and the non-drinker and suggests that certain vulnerability factors could contribute to the development of alcoholism. A trend noted is also the changing racial trends in the use of alcohol.

A comparison of socio-economic profiles of mixed married and intracaste married working women.

PIP: Differences between women in caste marriages and mixed marriages in India were investigated based on data from 65 working women in caste marriages and 65 in intracaste marriages. The findings indicated that mixed marriage women were similar in age at marriage and educational levels. There were significant differences in income: women in mixed marriages had higher income at the time of marriage than women in intracaste marriages. During the marital period there was no evidence of significant changes in educational level among women in either type of marriage. There were significant changes in income during the marital period. There were also significant differences in duration of marriage between marriage types: 10.92 years for women in mixed marriages and 13.77 years for women in intracaste marriages. There were more women in mixed marriages than women in intracaste marriages who advanced in their occupations, which was commensurate with this group's older age, education, and income levels at the time of marriage. The number of women advancing in their occupations was comparatively small.^ieng