Frequency of Defecation and Form of Stool among General Bangladeshi Population.

Data on stool form and defecation frequency which are a prerequisite for defining normal bowel habit are lacking in Bangladesh. This observational cross sectional study was designed to find out defecation frequency and stool form among general population in Bangladesh. This study was performed in the Department of Gastroenterology, Shaheed Suhrawardy Medical College Hospital, Dhaka, Bangladesh from July 2017 to June 2018. Apparently healthy 1090 respondents were evaluated for predominant stool form (Bristol chart) and frequency. Data on demographic and life-style were collected. The study population consisted of 1090 respondents, among them, 65.13% male and 34.87% female and mean age of them was 40.20±12.39 years. Most of the people 874(80.2%) passed stool between 12-14 times per week followed by 111(10.2%) less than 3 stools per week, 95(8.7%) passed more than 14 stools per week and 10(0.9%) between 3-12 stools per week, p<0.001. Most people passed predominantly Bristol type IV stool- 610(56.0%); followed by type III- 274(25.1%). Other stool forms were: type I- 52(4.8%), type II- 59(5.4%), type V- 31(2.8%), type VI- 33(3.0%), type VII- 31(2.8%), p<0.001. In regard to the physical activity, most of the respondents (70.0%) are physically active whereas about 13.0% are sedentary and about 17.0% are physically intermediate between the two, p<0.001. In the case of dietary habit, most of the participants are non-vegetarian (82.5%) and the remaining are vegetarian (11.1%) and occasional non vegetarian (6.4%), p<0.001. Median stool frequency in the studied population was 14 per week and predominant form was Bristol type IV. Older age was associated with lesser stool frequency, particularly among female subjects.

An isothermal based recombinase polymerase amplification assay for rapid, sensitive and robust indexing of citrus yellow mosaic virus.

Management of viral diseases relies on definite and sensitive detection methods. Citrus yellow mosaic virus (CYMV), a double stranded DNA virus of the genus Badnavirus, causes yellow mosaic disease in citrus plants. CYMV is transmitted through budwood and requires a robust and simplified indexing protocol for budwood certification programme. The present study reports development and standardization of an isothermal based recombinase polymerase amplification (RPA) assay for a sensitive, rapid, easy, and cost-effective method for detection and diagnosis of CYMV. Two different oligonucleotide primer sets were designed from ORF III (coding for polyprotein) and ORF II (coding for virion associated protein) regions of CYMV to perform amplification assays. Comparative evaluation of RPA, PCR and immuno-capture recombinase polymerase amplification (IC-RPA) based assays were done using purified DNA and plant crude sap. CYMV infection was efficiently detected from the crude sap in RPA and IC-RPA assays. The primer set used in RPA was specific and did not show any cross-amplification with banana streak MY virus (BSMYV), another Badnavirus species. The results from the present study indicated that RPA assay can be used easily in routine indexing of citrus planting material. To the best of our knowledge, this is the first report on development of a rapid and simplified isothermal detection assay for CYMV and can be utilized as an effective technique in quarantine and budwood certification process.

Serum transaminase level changes in dengue fever and its correlation with disease severity.

It was observed that liver enzymes are elevated in dengue fever. In this study our aims were to determine the changes in serum transaminases in dengue fever, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) and to find out the relation of transaminase level changes with the disease severity. This cross sectional, prospective hospital based observational study was carried out in the department of Gastrointestinal Hepatobiliary and Pancreatic diseases and Internal Medicine department of BIRDEM Hospital, Dhaka. Patients are classified into 3 groups depending on clinical & laboratory findings: Group 1 dengue fever (DF), Group 2 dengue hemorrhagic fever & Group 3 dengue shock syndrome. A total of 240 cases were taken in this study who fulfilled the selection criteria. Out of whom 125 male and 115 female patients. DF was 157(65.4%) & DHF was 83(34.6%). Aminotransferases [aspartate aminotransferase (AST) and alanine aminotransferase (ALT)] were significantly raised in DHF cases compared to those of classical dengue fever (AST 84.5±42.4 in DF vs. 507±106.8 IU/L in DHF and ALT 59.9±31.3 in DF vs. 234±30.6 IU/L in DHF). The rise of AST is far greater than ALT in both DF and DHF. Dengue fever is usually associated with mild to moderate elevations of aminotransferase levels. The increase in aminotransferases, mainly AST has been associated with disease severity and serves as an early indicator of dengue infection.

Multi-frequency EPR and Mössbauer spectroscopic studies on freeze-quenched reaction intermediates of nitric oxide synthase.

It is believed by analogy to chloroperoxidase (CPO) from Caldariomyces fumago that the electronic structure of the intermediate iron-oxo species in the catalytic cycle of nitric oxide synthase (NOS) corresponds to an iron(IV) porphyrin-pi -cation radical. Such species can also be produced by the reaction of ferric NOS with external oxidants within the shunt pathway. We present multi-frequency EPR (9.6, 94, 285 GHz) and Mössbauer spectroscopic studies on freeze-quenched intermediates of the oxygenase domain of nitric oxide synthase which has reacted with peroxy acetic acid within 8-200 ms. The intermediates of the oxygenase domain of both the cytokine inducible NOS (iNOSox) and the neuronal NOS (nNOSox) show an organic radical signal in the 9.6-GHz spectrum overlapping with the spectrum of an unknown species with g-values of 2.24, 2.23 and 1.96. Using 94- and 285-GHz EPR the organic radical signal is assigned to a tyrosine radical on the basis of g-values (i.e. Tyr*562 in nNOSox and Tyr*341 in iNOSox). Mössbauer spectroscopy of (57)Fe-labeled unreacted nNOSox shows a ferric low-spin heme-iron (delta = 0.38 mms(-1), deltaE(Q) = 2.58 mms(-1)). The reaction of nNOSox with peroxy acetic acid for 8 ms leads to the disappearance of the magnetic background characteristic for native nNOSox and a new species with delta = 0.27 mms(-1) and deltaE(Q) = 2.41 mms(-1) is detected at 4.2 K which does not resemble the parameters typical for a Fe(IV) center. It is proposed that this intermediate species corresponds to a ferric low-spin species which magnetically couples to an amino acid radical (presumably Trp*409).

High pressure fourier transform infrared (FT-IR) spectroscopic studies on inducible nitric oxide (NO) synthase active site: a comparison to cytochrome p450CAM.

High pressure Fourier transform infrared (FT-IR) spectroscopy is performed for the first time to analyse the active site of inducible nitric oxide synthase (iNOSox) using the carbon monoxide (CO) heme iron ligand stretch mode (nuCO) as spectroscopic probe. A membrane-driven sapphire anvil high-pressure cell is used. Three major conformational substates exist in substrate-free iNOSox which are characterized by nuCO at approximately 1936, 1945 and 1952 cm(-1). High pressure favors the 1936 cm(-1) substate with a volume difference to the 1945 substate of approximately -21 cm3/mol. The pressure induced cytochrome P420 formation with a reaction volume of approximately -80 cm3/mol is observed. Arginine binding produces a very low nuCO at approximately 1905 cm(-1) caused by the H-bond from the substrate to CO. nuCO for the substates in the substrate-free and arginine-bound proteins shift linearly with pressure which is qualitatively similar to the observation on cytochrome P450cam. The slightly smaller positive slope of the shift in substrate-free iNOSox compared to substrate-free P450cam is interpreted as a slightly lesser compressible heme pocket. In contrast, the significant slower negative slope for arginine-bound iNOSox compared to camphor-bound P450cam results from the different kind of interactions to the CO ligand (electrostatic interaction in P450cam, H-bond in iNOSox).

Separation of thoraco-omphalopagus Siamese twin.

A pair of female thoraco-omphalopagus twins, with two pairs of lungs, common diaphragm and separate pleural cavities was separated at the age of 5(1/2) months after a parasitic relationship had developed between them. Before separation both the babies developed recurrent respiratory tract infection and frequent diarrhea. They were treated with medical therapy and made fit for anesthesia. The combined weight of the twins was 4(1/2) kg at birth and the combined weight at the time of separation was 9(1/2) kg. One baby was smaller in growth compared to the other baby. The closure of anterior abdominal wall was difficult in both babies. However, we could close it without use of any biodegradable patches or tissue expander. Both the babies survived and are doing well after 3 months of separation.

Particle size distribution and respiratory deposition estimates of beryllium aerosols in an extraction and processing plant.

The mass size distribution of beryllium aerosols generated in the various operational areas of a typical extraction and processing plant was studied using an eight-stage impactor sampler. The total concentration of beryllium in the plant was found to be well below the threshold limit value. The mean value of mass median aerodynamic diameter of beryllium particles observed for various operations ranged from 5.0-9.5 microm. The alveolar deposition for various operational areas was estimated to be 3-5% for nasal breathing and 9-13% for oral breathing based on the International Commission on Radiological Protection (ICRP) human respiratory tract model. Deposition during oral breathing was higher than during nasal breathing by approximately a factor of two to three. This study on exposure characterization was useful for reducing the respirable fraction of beryllium aerosol by optimizing the capture velocity and improving the quality of other control measures.

Beryllium: an easily scavenged metal ion in the environment.

This study was specifically aimed to determine the levels of beryllium in environmental samples near the vicinity of the beryllium metal plant (BMP). Air particulate samples collected at the BMP site, in the non-monsoon and monsoon seasons, showed an average beryllium concentration of 0.3 and 0.1 ng m(-3) respectively, where as rain water samples showed the beryllium values in the range of 0.01-0.2 ng ml(-1). The suspended particulate matter (dust load) at the site studied was 570 and 250 microg m(-3) in the non-monsoon and monsoon seasons respectively. The results obtained show that, in the environment, 80% of the total beryllium present is removed by rain.

Immobilisation of beryllium in solid waste (red-mud) by fixation and vitrification.

The objective of this study was to obtain information on the immobilization of beryllium (Be) in solid waste generated in the extraction process of beryllium from its ore, Beryl. This solid waste, termed red-mud, contains oxides of iron, aluminium, calcium, magnesium and beryllium. The red-mud waste contains beryllium at levels above the permissible limit, which prevents its disposal as solid waste. The total beryllium content in the red-mud analysed showed value ranging from 0.39 to 0.59% Be The studies showed that 50% of the total beryllium in red-mud can be extracted by water by repeated leaching over a period of 45 days. The cement mix, casting into cement blocks, was subjected to leachability studies over a period of 105 days and immobilization factor (IF factor) was determined. These IF values, of the order of 102, were compared with those obtained by performing leachability study on vitrified red-mud masses produced at different temperature conditions. Direct heating of the red-mud gave the gray coloured, non-transparent vitreous mass (as 'bad glass') showed effective immobilisation factor for beryllium in red-mud of the order of 10(4).

Host response to infection: the role of CpG DNA in induction of cyclooxygenase 2 and nitric oxide synthase 2 in murine macrophages.

Depending on sequence, bacterial and synthetic DNAs can activate the host immune system and influence the host response to infection. The purpose of this study was to determine the abilities of various phosphorothioate oligonucleotides with cytosine-guanosine-containing motifs (CpG DNA) to activate macrophages to produce nitric oxide (NO) and prostaglandin E(2) (PGE(2)) and to induce expression of NO synthase 2 (NOS2) and cyclooxygenase 2 (COX2). As little as 0.3 microg of CpG DNA/ml increased NO and PGE(2) production in a dose- and time-dependent fashion in cells of the mouse macrophage cell line J774. NO and PGE(2) production was noted by 4 to 8 h after initiation of cultures with the CpG DNA, with the kinetics of NO production induced by CpG DNA being comparable to that induced by a combination of lipopolysaccharide and gamma interferon. CpG DNA-treated J774 cells showed enhanced expression of NOS2 and COX2 proteins as determined by immunoblotting, with the relative potencies of the CpG DNAs generally corresponding to those noted for the induction of NO and PGE(2) production as well as to those noted for the induction of interleukin-6 (IL-6), IL-12, and tumor necrosis factor. Extracts from CpG DNA-treated cells converted L-arginine to L-citrulline, but the NOS inhibitor N(G)-monomethyl-L-arginine (NMMA) inhibited this reaction. The COX2-specific inhibitor NS398 inhibited CpG DNA-induced PGE(2) production and inhibited NO production to various degrees. The NOS inhibitors NMMA, 1400W, and N-iminoethyl-L-lysine effectively blocked NO production and increased the production of PGE(2) in a dose-dependent fashion. Thus, analogues of microbial DNA (i.e., CpG DNA) activate mouse macrophage lineage cells for the expression of NOS2 and COX2, with the production of NO and that of PGE(2) occurring in an interdependent manner.

Characterization of key residues in the subdomain encoded by exons 8 and 9 of human inducible nitric oxide synthase: a critical role for Asp-280 in substrate binding and subunit interactions.

Human inducible nitric oxide synthase (iNOS) is active as a dimer of two identical subunits. Each subunit has an amino-terminal oxygenase domain that binds the substrate l-Arg and the cofactors heme and tetrahydrobiopterin and a carboxyl-terminal reductase domain that binds FMN, FAD, and NADPH. We previously demonstrated that a subdomain in the oxygenase domain encoded by exons 8 and 9 is important for dimer formation and NO synthesis. Further, we identified Trp-260, Asn-261, Tyr-267, and Asp-280 as key residues in that subdomain. In this study, using an Escherichia coli expression system, we produced, purified, and characterized wild-type iNOS and iNOS-Ala mutants. Using H(2)O(2)-supported oxidation of N(omega)-hydroxy-l-Arg, we demonstrate that the iNOS mutants' inabilities to synthesize NO are due to selective defects in the oxygenase domain activity. Detailed characterization of the Asp-280-Ala mutant revealed that it retains a functional reductase domain, as measured by its ability to reduce cytochrome c. Gel permeation chromatography confirmed that the Asp-280-Ala mutant exists as a dimer, but, in contrast to wild-type iNOS, urea-generated monomers of the mutant fail to reassociate into dimers when incubated with l-Arg and tetrahydrobiopterin, suggesting inadequate subunit interaction. Spectral analysis reveals that the Asp-280-Ala mutant does not bind l-Arg. This indicates that, in addition to dimerization, proper subunit interaction is required for substrate binding. These data, by defining a critical role for Asp-280 in substrate binding and subunit interactions, give insights into the mechanisms of regulation of iNOS activity.

Beryllium concentrations in ambient air and its source identification. A case study.

Beryllium concentrations in atmospheric particulate and soil samples in and around a Beryllium Processing Facility (BPF) have been measured. The mean air concentration level of beryllium in and around the fence line of the BPF is 0.48 +/- 0.43 ng m(-3) (n = 397) and is mostly influenced by diurnal and seasonal changes. The observed air concentration levels were well below the prescribed ambient air quality (AAQ) standard of 10 ng m(-3). The soil concentration levels of beryllium in the study area were found to be in the range of 1.42-2.75 microg g(-1). The mass median aerodynamic diameter (MMAD) of beryllium aerosols in ambient air was found to be 6.9 microm. Source identification using the Enrichment Factor (EF) approach indicates soil as the predominant contributory source for air concentrations at the site.

FT-Infrared spectroscopic studies of the iron ligand CO stretch mode of iNOS oxygenase domain: effect of arginine and tetrahydrobiopterin.

The iron ligand CO stretch vibration mode of the inducible nitric oxide synthase oxygenase domain (iNOSox) has been studied from 20 to 298 K. iNOSox in the absence of arginine reveals a temperature-dependent equilibrium of two major conformational substates with CO stretch bands centered at about 1945 and 1954 cm(-)(1). This behavior is not qualitatively changed when tetrahydrobiopterin (H(4)B) is bound. Arginine binding changes significantly the spectrum by formation of a sharp CO stretch mode band at about 1905 cm(-)(1) and indicates the formation of a hydrogen bond to the CO ligand. For temperatures lower than 250 K, the stretch vibration frequency decreases almost linearly with decreasing temperature and indicates that the coupling between the CO ligand and the arginine/protein in the active site via the hydrogen bond is very strong. Flashphotolysis of the CO ligand carried out at 25 K revealed the CO stretch mode of the photodissociated CO ligand trapped in the heme pocket. There is a negative linear relation between the stretch vibration frequencies of the photodissociated and the iron-bound CO indicating that the photodissociated ligand stays near the heme.

Inducible nitric oxide synthase: role of the N-terminal beta-hairpin hook and pterin-binding segment in dimerization and tetrahydrobiopterin interaction.

The oxygenase domain of the inducible nitric oxide synthase (iNOSox; residues 1-498) is a dimer that binds heme, L-arginine and tetrahydrobiopterin (H(4)B) and is the site for nitric oxide synthesis. We examined an N-terminal segment that contains a beta-hairpin hook, a zinc ligation center and part of the H(4)B-binding site for its role in dimerization, catalysis, and H(4)B and substrate interactions. Deletion mutagenesis identified the minimum catalytic core and indicated that an intact N-terminal beta-hairpin hook is essential. Alanine screening mutagenesis of conserved residues in the hook revealed five positions (K82, N83, D92, T93 and H95) where native properties were perturbed. Mutants fell into two classes: (i) incorrigible mutants that disrupt side-chain hydrogen bonds and packing interactions with the iNOSox C-terminus (N83, D92 and H95) and cause permanent defects in homodimer formation, H(4)B binding and activity; and (ii) reformable mutants that destabilize interactions of the residue main chain (K82 and T93) with the C-terminus and cause similar defects that were reversible with high concentrations of H(4)B. Heterodimers comprised of a hook-defective iNOSox mutant subunit and a full-length iNOS subunit were active in almost all cases. This suggests a mechanism whereby N-terminal hooks exchange between subunits in solution to stabilize the dimer.

N-terminal domain swapping and metal ion binding in nitric oxide synthase dimerization.

Nitric oxide synthase oxygenase domains (NOS(ox)) must bind tetrahydrobiopterin and dimerize to be active. New crystallographic structures of inducible NOS(ox) reveal that conformational changes in a switch region (residues 103-111) preceding a pterin-binding segment exchange N-terminal beta-hairpin hooks between subunits of the dimer. N-terminal hooks interact primarily with their own subunits in the 'unswapped' structure, and two switch region cysteines (104 and 109) from each subunit ligate a single zinc ion at the dimer interface. N-terminal hooks rearrange from intra- to intersubunit interactions in the 'swapped structure', and Cys109 forms a self-symmetric disulfide bond across the dimer interface. Subunit association and activity are adversely affected by mutations in the N-terminal hook that disrupt interactions across the dimer interface only in the swapped structure. Residue conservation and electrostatic potential at the NOS(ox) molecular surface suggest likely interfaces outside the switch region for electron transfer from the NOS reductase domain. The correlation between three-dimensional domain swapping of the N-terminal hook and metal ion release with disulfide formation may impact inducible nitric oxide synthase (i)NOS stability and regulation in vivo.

Occurrence of Witches'-Broom, a New Phytoplasma Disease of Acid Lime (Citrus aurantifolia) in India.

In India, acid lime (Citrus aurantifolia (L.) Swingle) is one of the most important citrus fruits grown. It constitutes nearly 20% of the total citrus production. During 1995, an unusual type of disease was observed on a 6-year-old acid lime plant in an orchard in the Nagpur District in eastern Maharashtra. It was named witches'-broom disease (WBD) to reflect the most conspicuous symptom. Other symptoms included small chlorotic leaves, highly proliferated shoots, and shortened internodes. Leaves dropped prematurely and infected twigs were distorted. In advanced stages, infected branches had dieback symptoms. WBD of lime has been reported from Oman and UAE (1) and the causal phytoplasma was designated "Candidatus Phytoplasma aurantifolia" (2). Subsequent surveys in 1995-1998 revealed disease incidences as high as 5% in Maharashtra and in other major acid-lime-growing states-Andhra Pradesh, Tamilnadu, and Karnataka. After the grafting of infected acid lime shoots, disease symptoms developed on Troyer citrange, rough lemon, and Rangpur lime, but not on sweet orange (mosambi), mandarin (Nagpur), or trifoliate orange. The WBD agent was transmitted from infected acid lime to periwinkle (Catharanthus roseus) plants and vice versa by dodder (Cuscuta reflexa). Ultrathin sections of leaf midrib of infected acid lime plants were fixed on copper grids, stained with uranyl acetate and lead acetate, and examined in a JEM 100S transmission electron microscope. Numerous bodies having the characteristic morphology of phytoplasmas were observed in phloem sieve tubes of acid lime in diseased but not in healthy leaves. The phytoplasmal bodies ranged from 100 to 800 nm in diameter and were bounded by a poorly defined membrance. Freehand transverse sections of young internode regions of a WBD-infected periwinkle plant were stained in DAPI (4', 6 diamidino-2-phenylindole; 1.0 µg/ml) and were observed with a fluorescent microscope (Leica). An intense bluish-white fluorescence in the phloem elements of diseased periwinkle and its absence in healthy samples were consistent with the presence of phytoplasmas. This is the first report of phytoplasma-induced witches'-broom disease of acid lime in India. References: (1) M. Garnier et al. Plant Dis. 75:546, 1991. (2) L. Zreik et al. Int. J. Syst. Bacteriol. 45:449,1995.

Structure of nitric oxide synthase oxygenase dimer with pterin and substrate.

Crystal structures of the murine cytokine-inducible nitric oxide synthase oxygenase dimer with active-center water molecules, the substrate L-arginine (L-Arg), or product analog thiocitrulline reveal how dimerization, cofactor tetrahydrobiopterin, and L-Arg binding complete the catalytic center for synthesis of the essential biological signal and cytotoxin nitric oxide. Pterin binding refolds the central interface region, recruits new structural elements, creates a 30 angstrom deep active-center channel, and causes a 35 degrees helical tilt to expose a heme edge and the adjacent residue tryptophan-366 for likely reductase domain interactions and caveolin inhibition. Heme propionate interactions with pterin and L-Arg suggest that pterin has electronic influences on heme-bound oxygen. L-Arginine binds to glutamic acid-371 and stacks with heme in an otherwise hydrophobic pocket to aid activation of heme-bound oxygen by direct proton donation and thereby differentiate the two chemical steps of nitric oxide synthesis.