Accuracy and Reliability of Pallor for Detecting Anaemia: A Hospital-Based Diagnostic Accuracy Study

Background

Anaemia is a common disorder. Most health providers in resource poor settings rely on physical signs to diagnose anaemia. We aimed to determine the diagnostic accuracy of pallor for anaemia by using haemoglobin as the reference standard.

Methodology/Principal Findings

In May 2007, we enrolled consecutive patients over 12 years of age, able to consent and willing to participate and who had a haemoglobin measurement taken within a day of assessment of clinical pallor from outpatient and medicine inpatient department of a teaching hospital. We did a blind and independent comparison of physical signs (examination of conjunctivae, tongue, palms and nailbed for pallor) and the reference standard (haemoglobin estimation by an electronic cell counter). Diagnostic accuracy was measured by calculating likelihood ratio values and 95% confidence intervals (CI) at different haemoglobin thresholds and area under the receiver operating characteristic curve. Two observers examined a subset of patients (n = 128) to determine the inter-observer agreement, calculated by kappa statistics. We studied 390 patients (mean age 40.1 [SD 17.08] years); of whom 48% were women. The haemoglobin was <7 g/dL in 8% (95% confidence interval, 5, 10) patients; <9 g/dL in 21% (17, 26) patients and <12 g/dL in 64% (60, 70) patients. Among patients with haemoglobin <7 g/dL, presence of severe tongue pallor yielded a LR of 9.87 (2.81, 34.6) and its absence yielded a LR of 0. The tongue pallor outperformed other pallor sites and was also the best discriminator of anaemia at haemoglobin thresholds of 7 g/dL and 9 g/dL (area under the receiver operating characteristic curves (ROC area  = 0.84 [0.77, 0.90] and 0.71[0.64, 0.76]) respectively. The agreement between the two observers for detection of anaemia was poor (kappa values  = 0.07 for conjunctival pallor and 0.20 for tongue pallor).

Conclusions/Significance

Clinical assessment of pallor can rule out and modestly rule in severe anaemia.     

Relation between the mode of binding of nitrite and the energy distribution between the two photosystems

The reversibility of nitrite-induced inhibition in relation to energy distribution between the two photosystems was studied in spinach thylakoid membranes. Measurements of electron transfer rate catalyzed by photosystem I (PS I) and photosystem II (PS II), chlorophyll a (Chl a ) fluorescence induction kinetics, S₂ state multiline spectra, and room temperature electron paramagnetic resonance (EPR) signals indicated that nitrite anions bind PS II in two ways: dissociable (loose) and non-dissociable (tight). The inhibition caused by the dissociable binding was reversible in washed (nitrite-treated samples washed with nitrite-free medium) samples, while the inhibition caused by the non-dissociable binding was irreversible. At 77 K, an increase in absorption cross section of PS I (as inferred from the excitation spectra of Chl a fluorescence) and a decrease in absorption cross section of PS II in nitrite-treated sample when compared with sample washed with nitrite-free medium and control sample suggested that nitrite plays a role in regulating the distribution of absorbed excitation energy between the two photosystems. We propose, for the first time, that the removal of loosely bound nitrite leads to migration of light-harvesting complex II back to the PS II, and thus the mode of binding of nitrite regulates the extent of migration of antenna molecules between the two photosystems.     

Rice Pangenome Genotyping Array: an efficient genotyping solution for pangenome-based accelerated genetic improvement in rice

The advent of the pangenome era has unraveled previously unknown genetic variation existing within diverse crop plants, including rice. This untapped genetic variation is believed to account for a major portion of phenotypic variation existing in crop plants. However, the use of conventional single reference-guided genotyping often fails to capture a large portion of this genetic variation leading to a reference bias. This makes it difficult to identify and utilize novel population/cultivar-specific genes for crop improvement. Thus, we developed a Rice Pangenome Genotyping Array (RPGA) harboring probes assaying 80K single-nucleotide polymorphisms (SNPs) and presence–absence variants spanning the entire 3K rice pangenome. This array provides a simple, user-friendly and cost-effective (60–80 USD per sample) solution for rapid pangenome-based genotyping in rice. The genome-wide association study (GWAS) conducted using RPGA-SNP genotyping data of a rice diversity panel detected a total of 42 loci, including previously known as well as novel genomic loci regulating grain size/weight traits in rice. Eight of these identified trait-associated loci (dispensable loci) could not be detected with conventional single reference genome-based GWAS. A WD repeat-containing PROTEIN 12 gene underlying one of such dispensable locus on chromosome 7 (qLWR7) along with other non-dispensable loci were subsequently detected using high-resolution quantitative trait loci mapping confirming authenticity of RPGA-led GWAS. This demonstrates the potential of RPGA-based genotyping to overcome reference bias. The application of RPGA-based genotyping for population structure analysis, hybridity testing, ultra-high-density genetic map construction and chromosome-level genome assembly, and marker-assisted selection was also demonstrated. A web application ( http://www.rpgaweb.com ) was further developed to provide an easy to use platform for the imputation of RPGA-based genotyping data using 3K rice reference panel and subsequent GWAS.     

Metabolomic Profile by GC/MS and Antioxidant,Antidiabetic and Anti-Inflammatory Activities of Green Solvent Extracts of Heterospathe elata Leaves

Plants are the prime source of phytoconstituents that can act as potent agents for the prevention and treatment of various diseases. Heterospathe elata is a plant belonging to the Arecaceae family having numerous medicinal properties. The present study was undertaken to prepare crude extracts of Heterospathe elata leaves with solvents of different polarity dimethyl carbonate (DMC), isopropyl alcohol (IPA), hydro alcohol (HYA) and water (WTR) by using successive Soxhlet extraction method. Further, the antioxidant, antidiabetic, and anti-inflammatory activities were assessed by the spectrophotometric method and possible bioactive phytoconstituents from the hydro alcohol extract of Heterospathe elata leaves using GC/MS. In our study, it was found that the GC/MS analysis revealed the presence of nineteen bioactive phytoconstituents. The highest antioxidant activity was found in the water extract. In antidiabetic and anti-inflammatory activity highest potential was shown by hydro alcohol extract and the lowest was in the dimethyl carbonate extract. These findings support the Heterospathe elata leaves showed the high biological potential attributed to a high amount of bioactive phytoconstituents and could be utilized as value-added functional food and medicine.     

The telomeric 5′ end nucleotide is regulated in the budding yeast Naumovozyma castellii

The junction between the double-stranded and single-stranded telomeric DNA (ds–ss junction) is fundamental in the maintenance of the telomeric chromatin, as it directs the assembly of the telomere binding proteins. In budding yeast, multiple Rap1 proteins bind the telomeric dsDNA, while ssDNA repeats are bound by the Cdc13 protein. Here, we aimed to determine, for the first time, the telomeric 5′ end nucleotide in a budding yeast. To this end, we developed a permutation-specific PCR-based method directed towards the regular 8-mer telomeric repeats in Naumovozyma castellii. We find that, in logarithmically growing cells, the 320 ± 30 bp long telomeres mainly terminate in either of two specific 5′ end permutations of the repeat, both corresponding to a terminal adenine nucleotide. Strikingly, two permutations are completely absent at the 5′ end, indicating that not all ds‐ss junction structures would allow the establishment of the protective telomere chromatin cap structure. Using in vitro DNA end protection assays, we determined that binding of Rap1 and Cdc13 around the most abundant ds–ss junction ensures the protection of both 5′ ends and 3′ overhangs from exonucleolytic degradation. Our results provide mechanistic insights into telomere protection, and reveal that Rap1 and Cdc13 have complementary roles.     

Detection of native peptides as potent inhibitors of enzymes. Crystal structure of the complex formed between treated bovine alpha-chymotrypsin and an autocatalytically produced fragment, IIe-Val-Asn-Gly-Glu-Glu-Ala-Val-Pro-Gly-Ser-Trp-Pro-Trp, at 2.2 angstroms resolution

Chymotrypsin is a prominent member of the family of serine proteases. The present studies demonstrate the presence of a native fragment containing 14 residues from Ile16 to Trp29 in alpha-chymotrypsin that binds to chymotrypsin at the active site with an exceptionally high affinity of 2.7 +/- 0.3 x 10(-11) M and thus works as a highly potent competitive inhibitor. The commercially available alpha-chymotrypsin was processed through a three phase partitioning system (TPP). The treated enzyme showed considerably enhanced activity. The 14 residue fragment was produced by autodigestion of a TPP-treated alpha-chymotrypsin during a long crystallization process that lasted more than four months. The treated enzyme was purified and kept for crystallization using vapour the diffusion method at 295 K. Twenty milligrams of lyophilized protein were dissolved in 1 mL of 25 mM sodium acetate buffer, pH 4.8. It was equilibrated against the same buffer containing 1.2 M ammonium sulfate. The rectangular crystals of small dimensions of 0.24 x 0.15 x 0.10 mm(3) were obtained. The X-ray intensity data were collected at 2.2 angstroms resolution and the structure was refined to an R-factor of 0.192. An extra electron density was observed at the binding site of alpha-chymotrypsin, which was readily interpreted as a 14 residue fragment of alpha-chymotrypsin corresponding to Ile-Val-Asn-Gly-Glu-Glu-Ala-Val-Pro-Gly-Ser-Trp-Pro-Trp(16-29). The electron density for the eight residues of the C-terminus, i.e. Ala22-Trp29, which were completely buried in the binding cleft of the enzyme, was of excellent quality and all the side chains of these eight residues were clearly modeled into it. However, the remaining six residues from the N-terminus, Ile16-Glu21 were poorly defined although the backbone density was good. There was a continuous electron density at 3.0 sigma between the active site Ser195 Ogamma and the carbonyl carbon atom of Trp29 of the fragment. The final refined coordinates showed a distance of 1.35 angstroms between Ser195 Ogamma and Trp29 C indicating the presence of a covalent linkage between the enzyme and the native fragment. This meant that the enzyme formed an acyl intermediate with the autodigested fragment Ile16-Trp29. In addition to the O-C covalent bond, there were several hydrogen bonds and hydrophobic interactions between the enzyme and the native fragment. The fragment showed a high complementarity with the binding site of alpha-chymotrypsin and the buried part of the fragment matched excellently with the corresponding buried part of Turkey ovomucoid inhibitor of alpha-chymotrypsin.     

Folic acid-mediated inhibition of serum-induced activation of EGFR promoter in colon cancer cells

Although accumulating evidence suggests a chemopreventive role for folic acid (FA) in colorectal carcinogenesis, the underlying mechanisms are largely unknown. Previously, we reported that supplemental FA inhibits the expression and activation of epidermal growth factor receptor (EGFR) in colon cancer cell lines. To determine the mechanism(s) by which FA affects EGFR function, we have examined whether and to what extent supplemental FA or its metabolites 5-methyltetrahydrofolate (MTF), dihydrofolate (DF), and tetrahydrofolate (TF) will modulate basal and serum-induced activation of the EGFR promoter in the HCT-116 colon cancer cell line. HCT-116 cells were preincubated with or without (control) FA or one of its metabolites (10 microg/ml) for 48 h, transfected with the EGFR promoter luciferase reporter construct, and incubated for 48 h with FA, DF, TF, or 5-MTF in the absence or presence of 10% FBS. Supplemental FA as well as its metabolites markedly inhibited EGFR promoter activity and its methylation status. Exposure of the cells to 10% FBS caused a marked stimulation of EGFR promoter activity and its expression, both of which were greatly abrogated by supplemental FA and 5-MTF. In contrast, serum-induced activation of c-fos promoter activity was unaffected by 5-MTF. The 5-MTF-induced inhibition of serum-mediated stimulation of EGFR promoter activity and EGFR expression was reversed when methylation was inhibited by 5-aza-2'-deoxycytidine. Our data suggest that FA and its metabolite 5-MTF inhibit EGFR promoter activity in colon cancer cells by enhancing methylation. This could partly be responsible for FA-mediated inhibition of growth-related processes in colorectal neoplasia.     

Investigation of deleterious effects of chromium phytotoxicity and photosynthesis in wheat plant

Increasing human and industrial activities lead to heavy metal pollution. Heavy metal chromium (Cr) is considered to be a serious environmental contaminant for the biota. Phytotoxic effects of Cr were studied in wheat plants. Growth parameters were largely inhibited as a result of disturbances in the plant cell metabolism in response to Cr toxicity. Chromium toxicity led to decline in a number of active reaction centres of PSII, rate of electron transport, and change in PSII heterogeneity. Chromium did not cause any change in heterogeneity of the reducing side. A significant change in antenna size heterogeneity of PSII occurred in response to Cr toxicity. Chromium seems to have extensive effects on the light harvesting complex of PSII.     

Estimation of Nitrogen Pools in Irrigated Potato Production on Sandy Soil Using the Model SUBSTOR

Recent increases in nitrate concentrations in the Suwannee River and associated springs in northern Florida have raised concerns over the contributions of non-point sources. The Middle Suwannee River Basin (MSRB) is of special concern because of prevalent karst topography, unconfined aquifers and sandy soils which increase vulnerability of the ground water contamination from agricultural operations- a billion dollar industry in this region. Potato (Solanum tuberosum L.) production poses a challenge in the area due to the shallow root system of potato plants, and low water and nutrient holding capacity of the sandy soils. A four-year monitoring study for potato production on sandy soil was conducted on a commercial farm located in the MSRB to identify major nitrogen (N) loss pathways and determine their contribution to the total environmental N load, using a partial N budget approach and the potato model SUBSTOR. Model simulated environmental N loading rates were found to lie within one standard deviation of the observed values and identified leaching loss of N as the major sink representing 25 to 38% (or 85 to 138 kg ha^-1 N) of the total input N (310 to 349 kg ha^-1 N). The crop residues left in the field after tuber harvest represented a significant amount of N (64 to 110 kg ha^-1N) and posed potential for indirect leaching loss of N upon their mineralization and the absence of subsequent cover crops. Typically, two months of fallow period exits between harvest of tubers and planting of the fall row crop (silage corn). The fallow period is characterized by summer rains which pose a threat to N released from rapidly mineralizing potato vines. Strategies to reduce N loading into the groundwater from potato production must focus on development and adoption of best management practices aimed on reducing direct as well as indirect N leaching losses.